contract number: ica4-ct-2000-30002 final report · contract number: ica4-ct-2000-30002 final...

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INCO-DC: International Cooperation with Developing Countries

CONTRACT NUMBER: ICA4-CT-2000-30002

FINAL REPORT

Start date: 01 October 2000 Duration: 36 months

Increasing the Productivity of Bambara Groundnut (Vigna

subterranea (L.) Verdc.) for Sustainable Food Production

in Semi-Arid Africa

Project homepage: http://www.wzw.tu-muenchen.de/pbpz/bambara/html/

Keywords: Bambara groundnut, Vigna subterranea, Semi-Arid Africa

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Shared-Cost RTD Contract number: ICA4-CT-2000-30002

TITLE: Increasing the Productivity of Bambara groundnut (Vigna subterranea (L.) Verdc.) for Sustainable Food Production in Semi-arid Africa

COORDINATOR

UNIVERSITY OF NOTTINGHAM SUTTON BONINGTON CAMPUS SCHOOL OF BIOSCIENCES DIVISION OF AGRIC AND ENV SCIENCES LOUGHBOROUGH, LE12 5RD LEICESTERSHIRE, UK

DR SAYED AZAM-ALI Email: [email protected] Tel : +44-115-951-6049 Fax : +44-115-951-6060

CONTRACTORS

BOTSWANA COLLEGE OF AGRICULTURE DEPARTMENT OF CROP SCIENCE AND PRODUCTION PRIVATE B. 0027 GABORONE BOTSWANA

DR ELENIMO KHONGA Email: [email protected] Tel : 267-365-0333 Fax : 267-328-753

TECHNICAL UNIVERSITY MUNICH CHAIR FOR AGRONOMY AND PLANT BREEDING ALTE AKADEMIE 12 85350 FREISING-WEIHENSTEPHAN GERMANY

PROF. DR. G. WENZEL Email: [email protected] Tel. : +49-8161-713749 Fax : +49-8161-714419

MAHANENE RESEARCH STATION OMUSATI REGION UUTAPI CONSTITUENCY P.O. BOX 646 OMBALANTU NAMIBIA

MR KLAUS FLEISSNER Email: [email protected] Tel : 264-65-259057 Fax : none

UNIVERSITY OF SWAZILAND DEPARTMENT OF BIOLOGICAL SCIENCES FACULTY OF SCIENCE P/B4 KWALUSENI SWAZILAND

PROF ABU SESAY Email: [email protected] Tel : 268-518-4011 Fax : 268-518-5276 Or : 268-518-7378

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ABSTRACT

This report describes a multidisciplinary research project carried out by five academic and research institutions from Africa and Europe with the aim of increasing the productivity of bambara groundnut (Vigna subterranea (L.) Verdc.) for sustainable food production in semi-Arid Africa. Collaborating institutions were Botswana College of Agriculture, Botswana; the University of Nottingham, UK; the Ministry of Agriculture, Water and Rural Development, Namibia; Technical University of Munich, Germany and the University of Swaziland, Swaziland. A notable feature of this research project was the emphasis on incorporating indigenous knowledge and preferences of local growers and consumers of bambara groundnut in the research process. A significant finding of the surveys of local growers was that most of the preferences (high yield, large seeds, early maturity and fast-to-cook) expressed by farmers, for a bambara groundnut ideotype, were common among the African countries. One implication of this finding is that a single strategic breeding programme could meet the crop improvement needs of all three countries. Genetic and agronomic characterisation of nine bambara groundnut landraces in the field and in controlled-environment glasshouses showed that the performance of these landraces was mostly affected by the amount and distribution of rainfall. The best yields (647-1582 kg ha-1) were obtained in Swaziland where the average total rainfall ranged from 633 mm to 728 mm and lowest yields were in Botswana (69-160 kg ha-1) where the rainfall ranged from 389 mm to 433 mm. Genetic variations in the response to drought were evident, thereby offering an opportunity for crop improvement based on the existing genetic variation.

Investigations on the genetic diversity of bambara groundnut showed that landraces consisted of three to eight major different genotypes. Furthermore, accessions from neighbouring countries were clustered together but no pure clusters composed of accessions from a single country were observed. Participatory development of a simple and feasible marker technology for genetic diversity studies in collaboration with all partner countries and training for young African scientists on new and emerging modern technologies have enabled scientists from African partner countries to benefit from this approach and therefore reduce their isolation from the international community. Project partners were able to introduce, test and refine the molecular approaches in selected African partner institutions during the timeframe of the project.

For the first time, the project has successfully demonstrated that it is possible to hybridise bambara groundnut. The hybridisation technique was successfully transferred through training courses to all partner countries. The development of an operational method of crossbreeding, with the successful productivity of the first hybrids of bambara groundnut, is a significant achievement because it has opened up the possibility of breeding the first true varieties of this crop. This also provides an opportunity to position QTL (Quantitative Trait Loci) for physiological traits related to resource capture and use.

We are now at a critical stage for the future of bambara groundnut. Researchers in the EU INCO-DC funded institutions and elsewhere have significant new knowledge with which to assist farmers in how best to grow the crop. We have the basis for a major bambara groundnut breeding programme to produce novel varieties that best suit the demands of growers and consumers. There is enormous scope for further genetic, physiological and agronomic studies. However, perhaps the most important priority now is to establish promotional and marketing strategies that highlight the nutritional and economic potential of bambara groundnut as a basis to expanding demand for the crop.

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SUMMARY

This section summarises key outputs from the multidisciplinary study with the overall aims of increasing the productivity of bambara groundnut (Vigna subterranea (L.) Verdc.) for sustainable food production in semi-Arid Africa. Collaborating institutions were Botswana College of Agriculture (BCA), Botswana; the University of Nottingham (UNOTT), UK; the Ministry of Agriculture, Water and Rural Development (MAWRD), Namibia; Technical University of Munich (TUM), Germany and the University of Swaziland (UNISWA), Swaziland. Survey of growers and consumers Indigenous knowledge and information from local farmers and consumers on their preferences for an ideal bambara groundnut type was collated to guide the planning of a breeding programme for the crop. A total of 462 farmers and 115 consumers were interviewed in a series of surveys in Botswana, Namibia and Swaziland between 2001 and 2003. The preferences for a bambara groundnut ideotype were found to be common among the three African countries, and included: high yield, large pods, large seeds, spreading growth habit, early maturity and short cooking time. The ‘high yield’ trait was ranked above all other traits, followed by ‘large pods’. From a yield component analysis it was concluded that the achievable objectives for any bambara groundnut improvement programme in these countries should include development of varieties which, within the constraints set by local climate, achieved high biomass production, high harvest index, large pods (1.69 cm diameter and above) and early maturity (3 to 4 months). Agronomic and genetic characterisation of landraces The performance of bambara groundnut landraces was mostly affected by environmental factors, particularly amount and distribution of rainfall. The three seasons were characterised by low, erratic and poorly distributed below average rainfall especially in Botswana and Namibia. Drought tolerance and early maturity should be some of the traits to be incorporated into an ideotype especially for Botswana and Namibia. The best yields (649–1582 kg ha-1) were obtained in Swaziland where the average total rainfall ranged from 633 mm to 728 mm during the three seasons. The lowest yields were in Botswana (68.5-159.9 kg ha-

1) where the rainfall ranged from 389 mm to 433 mm. Three landraces of bambara groundnut were grown in glasshouses at UNOTT under two moisture regimes (fully irrigated and droughted) to investigate their growth response to soil moisture. The three landraces were DipC from Botswana, S19-3 from Namibia and Uniswa red from Swaziland. The drought treatment was imposed at 42 days after sowing (DAS), beyond which no irrigation was applied. Shoot and root growth was monitored between 21 and 140 DAS. Soil moisture had an effect on the growth of both the shoot and the root. Drought reduced leaf area, dry matter accumulation, seed weight and yield. Drought also significantly (p<0.05) reduced root length density, total root surface area, root diameter and root volume. Landrace variations were found in all the root-related parameters, except for surface area and root volume at 84 DAS. There was preferential allocation of dry matter to the roots with increase in the intensity of

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drought but this did not occur in Uniswa red. The water use efficiency of the three landraces was found to be 2.1 g m-2 mm-1, a value that is comparable to that of other legumes. Genetic variations in the response to drought seem to exist, thereby offering an opportunity for crop improvement based on the existing genetic variation. Landrace S19-3 in particular tended to show signs of exhibiting drought avoidance through a shortened life cycle. UNOTT has developed a non-destructive method of estimating leaf area in bambara groundnut for use in the field in Africa where most laboratories lack access to leaf-area meters. Molecular analysis of germplasm Investigations on the genetic diversity of bambara groundnut (Vigna subterranea) were conducted on 223 accessions from Botswana, Namibia and Swaziland using enzyme system EcoRI/MseI amplified fragment length polymorphism (AFLP) and the simple sequence repeat (SSR) marker techniques. In the AFLP approach, profiles were generated with 10 primer combinations namely: E32M47, E32M49, E33M49, E35M48, E38M53, E39M47, E40M47, E41M58, E44M49, and E46M49. The number of amplified fragments ranged from 55 to 186, with an average of 86. The level of polymorphism was 22%, while polymorphic fragments ranged from nine to 26 with an average of 18.9. Due to the non-availability of SSR primers specific for bambara groundnut, amplification was done using 14 heterologous primer pairs that are specific for other legume species. The primers used were for soybean (Glycine max), cowpea (Vigna unguiculata), mungbean (Vigna radiata) and common bean (Phaseolus vulgaris). PCR products were obtained with 10 primer pairs, with only one primer pair showing polymorphism. Primer pair AG81 from soybean revealed 18 different alleles at a size range of 136bp to 194bp. A dendrogram resulting from cluster analysis showed that the 223 landraces consisted of three to eight major different genotypes. Furthermore, accessions from neighbouring countries were clustered together but no pure clusters composed of accessions from a single country were observed. The analysis for intra-diversity showed that a landrace consists of three to eight very similar, but different genotypes, for example: AHM 753 6 genotypes Gap C 4 genotypes AHM 968 3 genotypes Nyakeni C1 5 genotypes AS 17 3 genotypes Nyakeni C2 8 genotypes Dip C 6 genotypes Uniswa red 3 genotypes OM1 3 genotypes Cibadak 7 genotypes Molecular technology transfer A clear description of the RAPD methodology was published in the Proceedings of a Mid-Project Workshop held in Swaziland in August 2001. In country implementation of the protocols at BCA, UNISWA and UNAM (University of Namibia) has been successful. The standardised protocol established at the UNOTT were tested and optimised to suit BCA and UNISWA laboratories. Experiences in different laboratories in relation to RAPD technology and with various thermocyclers are being collated as part of an evaluation programme. Microsatellite (SSR) analyses and the primer sequences that produce polymorphisms have been determined, in-country implementation of this marker system will be effected to compliment RAPD markers.

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Agro-ecological modelling A simulation model has been developed to predict dry matter production and yield of bambara groundnut in contrasting environments. In this model, crop yields are considered as (a) potential (i.e. limited only by temperature, solar radiation, photoperiod, CO2 level and landrace characteristics), and (b) water-limited (i.e. as for (a), but with the influence of water availability included). The model is sink-limited, this means that the number of pods produced by the plant determines its final yield. The parameters of the model have been determined with experiments in the field, glasshouses and laboratories. The model is a stand-alone computer program written in Delphi 6. It uses climate data, landrace specific parameters, soil data, and physiological relationships running on a daily timestep to determine the biomass production and yield of a landrace in a specific environment. The model will be used to define breeding objectives and provide accurate predictions of performance of a certain landrace in a specific environment. Cross breeding The potential of pure line selection breeding in self-pollinated crops, such as bambara groundnut is limited by the available genetic variability between and within landraces. In any bambara groundnut improvement programme aimed at developing improved cultivars with desirable traits, artificial hybridisation is essential. Cross breeding of selected parental lines allows for the controlled combination of traits, which were previously distributed between the parents, in one new stable line. Additionally, new genetic variability may be produced, possibly resulting in traits previously unknown in the parental lines. Prior to the BAMFOOD research project no success had been achieved in efforts to improve bambara groundnut through cross breeding. As part of the activities of the BAMFOOD research project we have successfully performed a number of crosses, thus demonstrating that it is possible to hybridise bambara groundnut. Success in artificial hybridisation in bambara groundnut depends on a number of factors, including a clear understanding of the floral biology, adoption of an appropriate hybridisation protocol, and careful environmental control during and after pollination. The hybridisation technique was successfully transferred via training courses to Swaziland and Namibia and was performed based on documentation in Botswana. Morphological and molecular markers were used to verify hybrid status of the F1 and F2 plants. The development of an operational method of crossbreeding, with the successful productivity of the first hybrids of bambara groundnut, is a significant achievement because it has opened up the possibility of breeding the first improved varieties of this crop. This has also provides an opportunity to position QTL (Quantitative Trait Loci) for novel physiological traits related to resource capture and use. Pure lines were also developed by single seeds descent method. Basic selection methods were transferred to local farmers to help them improve the germplasm available locally.

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Main publications

Sesay, A. Edje, O. T. Vilakati, B. Magagula, C. N. 2002. Working with farmers on the UNISWA jugo bean research project. In Proceedings of the Consultative Workshop on the University Involvement in Community Service. 4 June, 2002; University of Swaziland, Kwaluseni, Swaziland. (Ed. Keregero K.J.B) pp 81-86.

Massawe, F.J., Azam-Ali, S.N. and Roberts, J.A. (2002). Molecular technology transfer-RAPD markers. In: Sesay, A., Edje, O. T. and Cornelissen, R (eds) Increasing the productivity of bambara groundnut (Vigna subterranea (L) Verdc) for sustainable food production in semi-arid Africa. Proceedings of a Mid-Project Workshop held at the University of Swaziland 28 - 30 August, 2001. pp 123-149.

Fleissner, K. (2002). Management practices and preferences of bambara groundnut (Vigna subterranea) producers in Oshna region, North Central Namibia. In: Sesay, A., Edje, O. T. and Cornelissen, R (eds) Increasing the productivity of bambara groundnut (Vigna subterranea (L) Verdc) for sustainable food production in semi-arid Africa. Proceedings of a Mid-Project Workshop held at the University of Swaziland 28 - 30 August, 2001. pp 47-53.

Deswarte, J.C., Cornelissen, R. and Azam-Ali, S.N. (2002). Gas exchange analysis in bambara groundnut (Vigna subterranea) landraces from Botswana, Namibia and Swaziland. In: Sesay, A., Edje, O. T. and Cornelissen, R (eds) Increasing the productivity of bambara groundnut (Vigna subterranea (L) Verdc) for sustainable food production in semi-arid Africa. Proceedings of a Mid-Project Workshop held at the University of Swaziland 28 - 30 August, 2001. pp 93-112.

Cornelissen, R., Deswarte, J.C. and Azam-Ali, S.N. (2002). A preliminary strategy for model development in bambara groundnut. In: Sesay, A., Edje, O. T. and Cornelissen, R (eds) Increasing the productivity of bambara groundnut (Vigna subterranea (L) Verdc) for sustainable food production in semi-arid Africa. Proceedings of a Mid-Project Workshop held at the University of Swaziland 28 - 30 August, 2001. pp. 167-176.

Schenkel, W., E. Sticksel and G. Wenzel (2002): Development of a bambara groundnut core collection from IITA germplasm based on characterisation and evaluation data. In: Sesay, A., Edje, O. T. and Cornelissen, R (eds) Increasing the productivity of bambara groundnut (Vigna subterranea (L) Verdc) for sustainable food production in semi-arid Africa. Proceedings of a Mid-Project Workshop held at the University of Swaziland 28 - 30 August, 2001. pp. 115-122.

Mabuza, P. (2002). Taxonomic relationships in Vigna revealed by molecular markers. In: Sesay, A., Edje, O. T. and Cornelissen, R (eds) Increasing the productivity of bambara groundnut (Vigna subterranea (L) Verdc) for sustainable food production in semi-arid Africa. Proceedings of a Mid-Project Workshop held at the University of Swaziland 28 - 30 August, 2001. pp. 151-163.

Azam-Ali, S.N. (2002). Towards a methodological framework for underutilised crops: our experience with bambara groundnut. In: Sesay, A., Edje, O. T. and Cornelissen, R (eds) Increasing the productivity of bambara groundnut (Vigna subterranea (L) Verdc) for sustainable food production in semi-arid Africa.

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Proceedings of a Mid-Project Workshop held at the University of Swaziland 28 - 30 August, 2001. pp. 203-223.

Schenkel, W., E. Sticksel and G. Wenzel (2002): Development of a bambara groundnut core collection from IITA germplasm based on characterisation and evaluation data. In: Challenges to Organic Farming and Sustainable Land Use in the Tropics and Subtropics (A. Deininger Hrsg.). Kassel University Press GmbH, Kassel.

Fleissner, K., E. Sticksel and W. Schenkel (2002): Participatory breeding approach of neglected crops – experience with bambara groundnut (Vigna subterranea) in Northern Namibia. In: Challenges to Organic Farming and Sustainable Land Use in the Tropics and Subtropics (A. Deininger Hrsg.). Kassel University Press GmbH, Kassel.

Sesay, A. Edje, O.T., Vilakati, B. and Magagula, C.N. (2002). Working with farmers on the EU/UNISWA jugo bean research project. Consultative Workshop on the University of Swaziland’s s Involvement in Community Service.

Fleissner, K. (2003). The participatory breeding approach of the BAMFOOD project. AGRI-INFO, July, 2003.

Kwerepe, B.C., Khonga, E.B., Ramolemana, G. M. and Balole, T.V. (2003). Farmers’ perceptions of a bambara groundnut ideotype. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Sesay, A, Edje, O.T. and Magagula, C.N. (2003). Working with farmers on the BAMFOOD research project in Swaziland. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Sesay, A., Edje, O.T., Magagula, C.N. and Mansuetus, A.B. (2003). Agronomic performance and morphological traits of field grown bambara groundnut (Vigna subterranea) in Swaziland.

Khonga, E.B., Karikari, S.K., Balole, T.V. and Machacha, S. (2003). Agronomic performance of nine landraces of bambara groundnut in Botswana. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Singrün, C. and Schenkel, W. (2003). Fingerprinting of bambara groundnut germplasm with molecular markers. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Mine, M., Motlhabane, L.T. and Batlang, U. (2003). Preliminary assessment of genetic variation in bambara groundnut in Botswana using RAPD markers: A case of technology transfer. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Massawe, F.J., Schenkel, W. Temba, E. and Basu, S. (2003). Artificial hybridisation of bambara groundnut. In: Proceedings of the International

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Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Sesay, A. and Mabuza, P.E.Z. (2003). Bambara groundnut (Vigna subterranea) improvement by the BAMFOOD research project in Swaziland. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Basu, S., Roberts, J.A., Mithen, R, Davey, M, and Azam-Ali, S.N. (2003). Towards genetic linkage mapping in bambara groundnut. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Massawe, F.J., Azam-Ali, S.N., Roberts, J.A. and Mwale, S.S. (2003). Strategic breeding in bambara groundnut (Vigna subterranea (L.) Verdc). In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Cornelissen, R., Matthews, R.B. and Azam-Ali, S.N. (2003). Modelling dry matter production and yield in bambara groundnut (Vigna subterranea (L.) Verdc). In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Mwale, S. Azam-Ali, S.N., and Massawe, F.J. (2003) Effect of soil moisture on root and shoot growth of bambara groundnut. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Sticksel, E. and Schenkel, W. (2003). Approaches to improve the acceptance and quality of bambara groundnut (Vigna subterranea) - a literature review. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Azam-Ali, S.N., Massawe, F.J., Mwale, S., Basu, S. and Cornelissen, R. (2003). Can bambara groundnut become a major world crop? In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

MSc and PhD degrees resulting from work under the contract: 1. Deswarte, J.C. (2001). Photosynthesis of bambara groundnut landraces in

response to soil moisture. MSc thesis, University of Nottingham, UK. 2. Gonapa, M. (2002). Light interception and conversion in bambara

groundnut landraces in response to soil moisture. MSc thesis, University of Nottingham, UK.

3. Kijoji, A. (2003). Response of three bambara groundnut (Vigna subterranea (L.) Verdc) landraces to soil moisture stress. MSc thesis University of Nottingham, UK.

4. Boateng C.O. (2003). Photosynthesis of three bambara groundnut landraces in response to soil moisture stress. MSc thesis University of Nottingham, UK.

5. Mathengwane, G.L. (2003). Tissue culture and transformation of bambara groundnut. MSc thesis University of Nottingham, UK.

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6. Mwale, S.S. (2005). Growth and development of bambara groundnut in response to soil moisture. PhD thesis, University of Nottingham, UK.

7. Cornelissen R. (2004). Modelling variation in the morphology and physiology of bambara groundnut. PhD thesis, University of Cranfield, UK.

8. Basu, S. (2005). Towards the construction of a genetic linkage map in bambara groundnut. PhD thesis, University of Nottingham, UK.

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ABSTRACT.............................................................................................3 SUMMARY .............................................................................................4

1.0 CONSOLIDATED SCIENTIFIC REPORT..................................... 14

1.1 Survey of growers .....................................................................15 1.1.1 Description of the work done......................................................... 15 1.1.2 Results ...................................................................................... 15 1.1.3 Discussion and Conclusions........................................................... 16

1.2 Characterisation of genetic and agronomic traits ......................16 1.2.1 Description of the work done......................................................... 16 1.2.2 Results ...................................................................................... 17 1.2.3 Discussion and Conclusions........................................................... 17 1.2.4 Problems.................................................................................... 18

1.3 Molecular analysis of germplasm...............................................18 1.3.1 Description of the work done......................................................... 18 1.3.2 Results ...................................................................................... 19 1.3.3 Discussion and Conclusions........................................................... 21

1.4 Molecular technology transfer ...................................................21 1.4.1 Description of the work done......................................................... 21 1.4.2 Results and Conclusions ............................................................... 21

1.5 Agro-ecological modelling .........................................................22 1.5.1 Description of the work done......................................................... 22 1.5.2 Results ...................................................................................... 22 1.5.3 Discussion and Conclusions........................................................... 23 1.5.4 Problems.................................................................................... 23

1.6 Crossbreeding ...........................................................................23 1.6.1 Description of the work done......................................................... 23 1.6.2 Results ...................................................................................... 24 1.6.3 Discussion and Conclusions........................................................... 28 1.6.4 Problems.................................................................................... 28 1.7 Overall Conclusions...................................................................... 28 1.8 List of publications....................................................................... 30 2.0 MANAGEMENT REPORT ........................................................ 36

2.1 Organisation of the collaboration ..............................................36 2.2 Meetings....................................................................................36 2.3 Exchange of staff.......................................................................37 2.4 Problems ...................................................................................37

3.0 INDIVIDUAL PARTNER FINAL REPORTS .................................. 38

University of Nottingham (UNOTT) .....................................................39 3.1 Survey of growers and consumers.............................................40

3.1.1 Description of the work done......................................................... 40 3.1.2 Results ...................................................................................... 40 3.1.3 Discussion and Conclusions........................................................... 40

3.2 Characterisation of genetic and agronomic traits ......................41 3.2.1 Description of the work done......................................................... 41 3.2.2 Results ...................................................................................... 42

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3.2.3 Discussion and Conclusions ...........................................................47 3.3 Molecular technology transfer................................................... 48

3.3.1 Description of the work done .........................................................48 3.3.2 Results .......................................................................................48

3.4 Agro-ecological modelling......................................................... 49 3.4.1 Description of the work done .........................................................49 3.4.2 Results .......................................................................................49 3.4.3 Discussion and Conclusions ...........................................................51 3.4.4 Problems ....................................................................................52

3.5 Crossbreeding ........................................................................... 52 3.5.1 Description of the work done .........................................................52 3.5.2 Results and Discussion..................................................................53 3.5.3 Conclusions.................................................................................53

Botswana College of Agriculture (BCA) .............................................. 55 4.1 Survey of growers..................................................................... 56

4.1.1 Description of the work done .........................................................56 4.1.2 Results, Discussion and Conclusions ...............................................56

4.2 Characterisation of genetic and agronomic traits...................... 56 4.2.1 Description of the work done .........................................................56 4.2.2 Results .......................................................................................57 4.2.3 Discussion and Conclusions ...........................................................63 4.2.4 Problems ....................................................................................64

4.3 Molecular technology transfer................................................... 64 4.3.1 Description of the work done .........................................................64 4.3.2 Results .......................................................................................64 4.3.3 Discussion and Conclusions ...........................................................64

4.4 Crossbreeding ........................................................................... 64 4.4.1 Description of the work done .........................................................64 4.4.2 Results .......................................................................................65 4.4.3 Discussion and Conclusions ...........................................................65 4.4.4 Problems ....................................................................................65

Technical University Munich (TUM) .................................................... 66 5.1 Characterisation of genetic and agronomic traits...................... 67

5.1.1 Description of the work done .........................................................67 5.1.2 Results .......................................................................................67 5.1.3 Discussion and Conclusions ...........................................................69 5.1.4 Problems ....................................................................................70

5.2 Molecular analysis of germplasm .............................................. 70 5.2.1 Description of the work done .........................................................70 5.2.2 Results .......................................................................................70 5.2.3 Discussion and Conclusions ...........................................................75

5.3 Molecular technology transfer................................................... 76 5.3.1 Description of the work done .........................................................76 5.3.2 Results .......................................................................................76

5.4 Crossbreeding ........................................................................... 76 5.4.1 Description of the work done .........................................................76 5.4.2 Results .......................................................................................77 5.4.3 Problems ....................................................................................77

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Ministry of Agriculture, Water and Rural Development (MAWRD).......78 6.1 Survey of growers .....................................................................79

6.1.1 Description of the work done......................................................... 79 6.1.2 Results and Discussion ................................................................. 79 6.1.3 Conclusions ................................................................................ 79 6.1.4 Problems.................................................................................... 79

6.2 Characterisation of genetic and agronomic traits ......................79 6.2.1 Description of the work done......................................................... 79 6.2.2 Results and Discussion ................................................................. 80 6.2.3 Conclusions ................................................................................ 80 6.2.4 Problems.................................................................................... 80

6.3 Crossbreeding ...........................................................................81 6.3.1 Description of the work done......................................................... 81 6.3.2 Future plans ............................................................................... 81

University of Swaziland (UNISWA).....................................................82 7.1 Survey of growers .....................................................................83

7.1.1 Description of the work done......................................................... 83 7.1.2 Results and Discussion ................................................................. 83

7.2 Characterisation of genetic and agronomic traits ......................84 7.2.1 Description of the work done......................................................... 84 7.2.2 Results and Discussion ................................................................. 84

7.3 Molecular technology transfer ...................................................88 7.3.1 Description of the work done......................................................... 89 7.3.2 Results ...................................................................................... 89

7.4 Agro-ecological modelling .........................................................89 7.4.1 Description of the work done......................................................... 89

7.5 Crossbreeding ...........................................................................90 7.5.1 Description of the work done......................................................... 90 7.5.2 Results ...................................................................................... 90

7.6 Strategic investigations.............................................................90 7.6.1 Description of the work done......................................................... 90 7.6.2 Results achieved ......................................................................... 90

7.7 Dissemination workshop ...........................................................91 Data sheet for final report ............................................................... 93

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1.0 CONSOLIDATED SCIENTIFIC REPORT

Evidence of bambara groundnut as an important African pulse crop dates back to the Middle Ages (Doku, 1997). However, the first recorded botanical reference appears to be Du Petit-Thours (1806) who describes the genus Voandzeia in which bambara groundnut was placed until Verdcourt (1980) redefined it as Vigna subterranea (L.) Verdc.). Until 1904, only 13 references to the species appear in the published literature. Hence, for most of its cultivated history, knowledge of the crop must have been transferred through successive generations in qualitative, anecdotal and unwritten forms. It is only 11 years since co-ordinated, multidisciplinary and international research has been directed to bambara groundnut. Since then, significant financial and human resources and physical facilities have been contributed by individual institutions and other sponsors but the bulk of support has been provided by the International Cooperation with Developing Countries Programme (INCO-DC) of the European Union (EU). This has been through two projects. The first between 1992 and 1996 (Evaluating the potential for bambara groundnut as a food crop in semi-arid Africa) involved collaboration between Botswana College of Agriculture (BCA), Botswana; Institute of Agricultural Research (IAR), Sierra Leone; University of Nottingham (UNOTT), UK; Sokoine University of Agriculture (SUA), Tanzania and Wageningen University (WU), the Netherlands. The second research project on bambara groundnut ‘BAMFOOD’ (Increasing the productivity of bambara groundnut for sustainable food production in semi-arid Africa) started in 2000. Partners again include BCA and UNOTT with new collaborators from the Ministry of Agriculture, Water and Rural Development (MAWRD), Namibia; Technical University of Munich (TUM), Germany and the University of Swaziland (UNISWA), Swaziland. Together, the EU funded projects have generated a unique research base on bambara groundnut centred around the above-mentioned institutions. In addition to EU funding, these ‘research nodes’ provide the critical mass around which additional, complementary activities on the crop can be built. Of course at the same time as this concentrated effort in the EU-funded institutions valuable work continues to be done in a number of African institutions, which have maintained research capacity often without the benefit of international support. For the past three years (2000-2003), the INCO-DC of the EU has funded a research project on bambara groundnut under Framework Programme 5 (FP5). The research involved laboratory, glasshouse, field and on-farm experiments, farmer survey and simulation modelling. In the following sections, a detailed analysis of the work done for the three years of the BAMFOOD research project is given. The synthesis is based on the specific scientific and technological objectives of the project, which are to:

1. Identify bambara groundnut ‘ideotypes’ for local conditions in Botswana, Namibia and Swaziland using a farmers survey.

2. Characterise the genetic and agronomic performance of bambara groundnut landraces from Botswana, Namibia and Swaziland in field, controlled glasshouse and on-farm environments.

3. Evaluate genetic diversity in bambara groundnut germplasm using simple, readily transferable molecular strategies.

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4. Produce a robust bambara groundnut model to match suitable ideotypes to contrasting environments and end users.

5. Establish an operational method of crossbreeding for intraspecific hybridisation in bambara groundnut.

6. Develop a strategic bambara groundnut breeding programme based on morphological and molecular considerations.

7. Provide a blueprint of how the methodology established for bambara groundnut can be applied to other underutilised crops.

1.1 Survey of growers

A notable feature of the BAMFOOD research project was the emphasis on incorporating indigenous knowledge and preferences of local growers of bambara groundnut in the research process. The objective of the surveys was to identify specific attributes and preferences for bambara groundnut ideotypes for each African partner country to support the improvement of the crop.

1.1.1 Description of the work done

African partner institutions conducted a series of extensive surveys of local farmers and consumers to collect information on local knowledge, perceptions and preferences for bambara groundnut, as a basis for identifying specific desirable attributes, and establishing a list of achievable objectives for breeders and agronomists. With the existence of information on local knowledge from a previous study, the focus of the surveys in Swaziland was on collecting information on farmer and consumer preferences for bambara groundnut and on ranking of desirable traits to define a minimum number of achievable objectives for improving the crop. Towards this end 95 farmers and 30 consumers were interviewed in focus group discussions using a checklist in 2001. Between 2002 and 2003 a total of 24 farmers were interviewed to rank preferences, using a structured questionnaire administered in association with plant samples. In 2001 50 farmers were surveyed in Botswana to collect information on farmers' knowledge and perceptions on the crop. Between 2002 and 2003 244 farmers, 108 sellers and 115 consumers were interviewed to collect further information on local knowledge and on preferences for the crop, using a semi-structured interview. Between 2001 and 2002 a total of 49 farmers were surveyed in Namibia to collect information on preferences for bambara groundnut. Data analysis was carried out using the QSR NUD*IST 4 software. Further details on the surveys are provided in the relevant Annual Reports.

1.1.2 Results

The results of the surveys on farmer and consumer preferences for a bambara groundnut ideotype indicate considerable overlap across the African countries (Table 1). According to the preference quantification or ranking survey, farmers would rank the "high yield" trait above all other preferred traits, followed by "large pods" and "pod number". Furthermore, in Swaziland farmers would consider pod yields of 2.0-3.0 t ha-1 as high yield (61.5%), pods of at least 1.69cm diameter as large pods (94.0%), and seeds of at least 58g per 100 seeds as large seeds (70.6%).

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Table 1: Farmers and consumer preferences for bambara groundnut in Botswana, Namibia and Swaziland. Traits Botswana Namibia Swaziland High yield _ + + Large pods _ _ + Large seeds + + + Rapid and uniform emergence

_ _ +

Spreading growth habit + + + Early maturity + + + Pod retention (strong pegs) + _ + Cream seeds + + + Easy-to-cook + + + Low anti-nutritional factors _ _ + Sweet taste + + _

1.1.3 Discussion and Conclusions

A significant feature of the findings of the surveys is that many of the preferences expressed by farmers, for a bambara groundnut ideotype, were common among the African countries. One implication of this finding is that a single strategic breeding programme could meet the crop improvement needs of all three countries. Although the farmers listed a range of preferences for a bambara groundnut ideotype, they also expressed a willingness to make trade-offs, and stressed that their priority was maximising yield, which is an integrative trait. Based on a yield component analysis therefore, the achievable objectives for bambara groundnut improvement programme should include the incorporation of the following traits: high biomass production, high harvest index, large pods (1.69 cm diameter and above) and early maturity (3 to 4 months).

1.2 Characterisation of genetic and agronomic traits

Genetic, agronomic and morphological traits of nine bambara groundnut landraces were evaluated in the field at BCA, MAWRD, UNISWA and in controlled-environment glasshouses at UNOTT. The objectives for these experiments were to: determine the yield potential of the nine landraces across a range of contrasting environments; determine variation in morphological, physiological and agronomic traits and determine genotype x environment interaction for the traits to assess the range of adaptation of landraces to different agro-ecosystems.


Nine bambara groundnut landraces, three from each African partner (Table 2) were planted at two sites between 2000 and 2003 in each country namely Notwane and Good Hope in Botswana; Mahanene and Mashare in Namibia and Malkerns and Luve in Swaziland. In addition, on-farm trials were conducted in Swaziland in order to assess the landraces under farmers’ conditions and also to collect information from the end-users on their preferences and concerns. The experiments at UNOTT were carried out in glasshouses using three landraces to assess their performance in response to soil moisture. The glasshouse experiments gave an opportunity to assess the landraces under

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drought and irrigated conditions with negligible confounding factors since the environment in the houses is controlled. Using a CIRAS-1 infrared gas exchange analyzer with an external light source, light response, transpiration and stomatal conductance were determined. In addition, the chlorophyll content of selected leaves was determined using a SPAD meter (Minolta 502). The SPAD readings were calibrated for actual chlorophyll content using a photospectrometer. Table 2: Landraces used by all partners in field experiments. Origin Landrace Source Description

NYAKENIC1 Farmer at Nyakeni Cream testa, black eye pattern NYAKENIC2 Farmer at Nyakeni Cream testa, brown eye

Swaziland

Uniswa red Manzini market Red testa, white hilum AHM753 Namibia

germplasm Red testa, early

AHM968 Namibia germplasm Tan colour, medium

Namibia

AS17 South Africa Late maturing DIPC BCA germplasm Cream testa, black eye GABC BCA germplasm Cream testa, brown eye

Botswana

OM1 BCA germplasm Cream colour, butterfly eye The trials in each of these countries adopted a common protocol with respect to experimental design, crop husbandry and data collection. Each field site comprised four replicate blocks of nine plots (landraces) giving a total of 36 plots. Each individual plot was 6 x 6 m, i.e. 36 m2. The inter- and intra-row spacing was 50 and 30 cm, respectively giving a plant population of 7 plants m-2 (252 plants per plot). Two seeds were sown per station and half intra-row spacing at a depth of 5 cm and thinned down to one per station and appropriate density 21 days after sowing (DAS).

1.2.2 Results

Results from the field trials showed that there were significant differences among landraces in traits such as numbers of flower, pods and leaves, days to physiological maturity and seed yield. Amount and distribution of rainfall highly influenced seed yield among countries and within each country (Table 3). The best yields (649–1582 kg ha-1) were obtained in Swaziland where the average total rainfall ranged from 633 mm to 728 mm during the three seasons and lowest yields were in Botswana (69-160 kg ha-1) where the rainfall ranged from 389 mm to 433 mm. In Namibia, yields were generally higher at Mashare than at Mahanene again because Mashare had higher rainfall than Mahenene. Yields from on farm trials followed similar trends. The results of the experiments at UNOTT showed effects of landrace, leaf age and water stress. Perhaps more importantly, UNOTT has developed a non-destructive method of estimating leaf area in bambara groundnut for use in the field in Africa where most laboratories lack access to leaf-area meters.


The performance of bambara groundnut landraces was mostly affected by environmental factors, particularly amount and distribution of rainfall. The three seasons were characterised by low, erratic and poorly distributed below

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average rainfall especially in Botswana and Namibia. Drought tolerance and early maturity should be some of the traits to be incorporated into an ideotype especially for Botswana and Namibia. Soil moisture had an effect on the growth of both the shoot and the root. Drought reduced leaf area, dry matter accumulation, seed weight and yield. Drought also significantly (p<0.05) reduced root length density, total root surface area, root diameter and root volume. Landrace variations were found in all the root-related parameters, except for surface area and root volume at 84 DAS. There was preferential allocation of dry matter to the roots with increase in the intensity of drought but this did not occur in Uniswa red. The water use efficiency of the three landraces was found to be 2.1 g m-2 mm-1, a value that is comparable to that of other legumes. Genetic variations in the response to drought seem to exist, thereby offering an opportunity for crop improvement based on the existing genetic variation. Landrace S19-3 in particular tended to show signs of exhibiting drought avoidance through a short life cycle. Table 3: Average seed yield (kg ha-1) of nine bambara groundnut landraces grown in Botswana, Namibia and Swaziland between 2000 and 2003. Swaziland Botswana Namibia

Landrace Malkerns (728 mm rainfall)

Luve (633 mm rainfall)

Good Hope (389 mm rainfall)

Notwane (433 mm rainfall)

Mahenene (413 mm rainfall (2000 only)*

Mashare (350 mm rainfall 2000 only)

AS17 1892.5 702.9 159.4 102.7 469.5 426.6 AHM968 953.0 446.2 126.9 76.8 496.1 598.6 AHM753 1671.4 682.7 188.1 73.1 311.3 590.8 GabC 1445.0 619.3 155.8 84.9 362.3 595.2 DipC 1533.9 738.5 142.7 42.5 449.3 673.1 OM1 1721.9 642.9 308.4 78.6 367.5 486.8 Uniswa red

1584.7 861.7 120.7 66.9 418.1 694.8

NkakC1 1791.7 666.4 134.4 45.2 269.8 645.8 NyakC2 1639.4 476.6 102.5 45.9 527.3 776.0 Mean 1581.5 648.6 159.9 68.5 407.9 609.7 * Namibia data averaged over two seasons.

1.2.4 Problems

The main problem was low and erratic rainfall in Botswana and Namibia. This adversely affected the experiments but at the same time assessed the tolerance of the landraces to what actually happens in the field from season to season. In Namibia there was a problem of staff shortage where two project technicians resigned during the project and the Team Leader was seriously ill during the last season.

1.3 Molecular analysis of germplasm


In total genomic DNA of over 400 individual bambara groundnut plants was isolated for molecular marker analysis. This included accessions from the IITA

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bambara groundnut genebank, the Namibian national collection and nine landraces three each from African partners in this project. For marker analysis Amplified Fragment Length Polymorphism (AFLP) markers were developed and Simple Sequence Repeats (SSRs) markers, also known as Microsatellite markers, established in other legume species, were adapted. The resulting molecular data were used to calculated genetic distances between accessions. A hierarchical cluster analysis was performed based on distance data. Within landrace diversity was estimated.

1.3.2 Results

The AFLP reactions were done according to Vos et al., 1995. After initial screening of AFLP primer combinations, ten different primer combinations were selected (Table 4). The number of amplified fragments ranged from 55 for E46M49 to 186 for E32M49 at an average of 86 fragments per primer pair. The average level of polymorphism per primer pair over all genotypes was 22.0%, ranging from 8.5% (9 of 103) for E39M47 to 37.5% (22 of 59) for E44M49. Table 4: An overview of AFLP primer combinations and level of polymorphism generated.

Primer combination

No. of amplified fragments

No. of polymorphic fragments

Polymorphic fragments (%)

E32M47 85 27 31.8 E32M49 186 18 9.6 E33M49 62 19 30.5 E34M53 76 22 29.1 E35M48 87 26 29.8 E39M47 103 9 8.8 E40M47 79 9 11.5 E41M58 61 20 32.9 E44M49 59 22 37.5 E46M49 55 17 30.7 Mean 85.3 18.9 22.16

The SSR (Microsatellite) primers were chosen from crop species closely related to bambara groundnut (Table 5). Of 14 primer pairs, 10 amplified fragments in a size range corresponding to the original microsatellites. Nine primer pairs amplified monomorphic fragments, however primer pair AG81 from soybean (Table 5) amplified 18 different alleles in a size range from 136-194 bp. SSR allele of 136 bp was separated from the other alleles that spanned a range from 160 to 194 bp in steps of two base pairs (Figure 1). The distribution of the alleles corresponded to the Gaussian distribution with its maximum at a fragment size of 180 bp.

20

0

5

10

15

20

25

30

35

136 160 162 164 166 168 170 172 174 176 178 180 182 184 186 188 190 192 194

size [bp]

alle

le fr

eque

ncy

Figure 1: Allele frequencies of SSR AG81 in bambara groundnut. Table 5: Microsatellite markers, their motives, origin and amplified products generated in bambara groundnut. Mungbean (V. radiata) Kumar et al. 2002 LR7315A (AG)12 monomorphic LR738A (AG)11…(GA)6 monomorphic Soybean (G. max) Peakall et al. 1998 AG81 (AG)5 polymorphic, 18 alleles Cowpea (V. unguiculata) Li et al. 2001 VM5 (AG)32 monomorphic VM36 (CT)13 no amplification products VM31(CT)16 monomorphic VM39 (AC)13(AT)5(TACA)4 no amplification products VM35 (AG)11 (T)9 no amplification products Common bean (P. vulgaris) Yu et al. 1999, Gaitán-Solís et al. 2002 BM143 (GA)35 monomorphic BM200 (AG)10 monomorphic j04555 (CTT)3(T)3(CTT)6 no amplification products x61293(AT)18 monomorphic x04001(AG)8 monomorphic x80051(AT)12 monomorphic The clustering showed a good correlation to the origin of accessions. All Namibian accessions were clustered together (clusters 8 and 9). Accessions from neighbouring countries, Ghana and Nigeria, as well as those from Benin clustered together (clusters 4 and 15). Similarly, accessions from the neighbouring countries Zambia and Zimbabwe were clustered together (clusters 2, 5-7 and 17). Noticeable is the fact that the accessions from Madagascar and Indonesia neither were clustered together nor could they be associated with any other country. The developed markers were also used to analyse the intra landrace diversity. From each of the nine landraces 15 individual plants were randomly chosen for

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analysis. The number of identified different genotypes ranged from 3 to 8. Even the landrace AS 17, which was regarded as pure line prior to this analysis consisted of different lines.


Existing marker technologies were successfully adapted and applied on bambara groundnut. AFLP markers were not as easy to score as SSR markers but were available abundantly. The frequency of polymorphisms was near optimal in a range of approximately 20%. This results in a good efficiency of marker development with a high reliability due to a sufficient amount of common markers between each pair of genotypes. The application of SSR markers taken from related species was only partly successful. Only 10 out of 14 tested primers amplified fragments in bambara groundnut and of these only 1 produced polymorphic patterns. However, this result was expected, since SSR markers are known to be highly specific. For an efficient application of SSR markers these need to be developed for bambara groundnut. It is highly recommended to do so in the future since this marker type is most reliable and has the best prospect of being applied in laboratories with basic equipment. The benefit of molecular markers for analysis of genetic diversity was clearly demonstrated. The clustering of genotypes from the same origin into groups indicates that relationships can be identified based on genetic similarities. The information generated by this study allows scientists to make informed decisions in the development of breeding strategies.

1.4 Molecular technology transfer

The transfer of molecular technology from European to African partners was a component of the BAMFOOD research project. As part of this exercise, a standardised Random Amplified Polymorphic DNA (RAPD) protocol was established that could be used for genetic characterisation in laboratories with basic equipment.


Standardised DNA isolation and RAPD protocols were established in collaboration with African partners and were successfully adopted and used in laboratories of the three countries participating in the BAMFOOD research project. Detailed description of the protocols can be found in the Proceedings of a Mid-Project Workshop held at UNISWA in Swaziland (Massawe et al., 2002).

1.4.2 Results and Conclusions

A clear description of the RAPD methodology was published in the Proceedings of a Mid-Project Workshop held in Swaziland in August 2001 (Refs). In country implementation of the protocols at BCA, UNISWA and UNAM (University of Namibia) was successful. Details of results and other information related to this work package appear in the two BAMFOOD research project proceedings, Sesay et al. (2002 [eds.]) and Anonymous, (2003). Participatory development of a simple and feasible marker technology for genetic diversity studies in collaboration with all partner countries and training

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for young African scientists on new and emerging modern technologies have enabled scientists from African partner countries to benefit from this approach and therefore reduce their isolation from the international community. Project partners were able to introduce, test and refine the molecular approaches in selected African partner institutions during the timeframe of the project.

1.5 Agro-ecological modelling


A simulation model has been developed to predict dry matter production and yield of bambara groundnut in contrasting environments. The model is an adaptation of the SLM (Matthews, 2002) model for a leguminous crop. It is a sink-orientated model, i.e. the number of available sinks (pods) determines the final production. The model is a stand-alone computer program written in Delphi 6 (Borland®). The model divides the crop duration into five stages. These are Emerging, Vegetative, Flowering, Grain filling and Maturing. Crop phenology in this model is described in terms of phenochrons. The model uses environmental data (climate, soils, etc) and landrace specific parameters as inputs. The data is read into the model using two text files in a standard format. One file contains information on cropping methods while the other file contains climate data. A third input file is a database containing landrace specific parameters that the model uses to calculate the performance of a landrace. Bambara groundnut has an extrapolated theoretical base temperature of 9.90C for germination. In practice there is no germination below 120C and above 450C. The time between sowing and emergence varies between the different landraces and usually takes between 6 and 12 phenochrons. The relation between number of leaves and phenological age has been determined. From this relation the phyllochron interval can be determined. This is the number of phenochrons between two successive leaves. For modelling purposes this relation is divided into two linear relationships. The number of leaves increases until a maximum number, after which no further leaves are produced. Although the relationship is similar for the landraces, the maximum leaf number differs for each landrace. The model calculates the rate of increase of leaf area as a function of the current leaf number using a fitted relation. Analysis of available data indicated that this relation is similar for all landraces. The model estimates dry matter by an empirical equation calculating the increase in biomass per intercepted radiation, using radiation use efficiency (RUE), photosynthetically active radiation (PAR), leaf area index (LAI) and plant population. For bambara groundnut the model assumes a PAR of 0.5 and a RUE of 2.4 g/MJ PAR. The model calculates dry matter partitioning by calculating the fraction of the total dry matter for each different plant component at each developmental stage.

1.5.2 Results

The model has a user-friendly front end with a MS Windows based program, which will be easy to understand for those with basic computer skills. It will be fully compatible with the latest MS Windows versions. The model has been

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validated against glasshouse data and ongoing work is being done on the validation of the model against field data. The landrace database for nine landraces used in the project has been constructed and can be easily expanded when more landrace data becomes available.


The model is able to predict the performance of a landrace in its environment of origin or in an environment in which it has not yet been grown. In this way the model can help to choose an ideotype landrace for a specific environment. For the model to have any practical use the ideotype it generates must be based on physiologically sound principles, crop parameters that are from existing germplasm and environmental data. Unfortunately a model cannot invent data and can thus only predict the ideotype performance if it has source data either from an existing landrace or from a hypothetical plant, based upon physiologically sound information. The model will assist in a breeding programme by establishing criteria and breeding objectives based on the trade-offs between desirable attributes and resource limitations. Finally it should be mentioned that the model has been based on the SLM model for legumes (Matthews, 2002). This means that the model can easily be combined/added on other crop models built in this style. Instead of just functioning as a stand-alone bambara groundnut model, it could form part of a larger model where the production of bambara groundnut can be simulated in combination with other crops for which a SLM type model already exist. This leads to a larger integration of crop models and will increase the practical use of the model.

1.5.4 Problems

One of the main problems is the availability of full sets of climate data for every site for model validation. For some of the sites climate data were not collected or the sets are incomplete due to broken equipment. In order to enable the model to work for regions where weather data were unavailable, the model can be linked to a weather generator. A robust weather generator has been identified and efforts are under way to incorporate this in the model. Limited availability of data on photoperiod sensitivity of the landraces used in the current project in the literature has lead to the delay in the validation of the model using field data.

1.6 Crossbreeding


The floral biology of bambara groundnut was studied. An operational method for crossbreeding was developed and documented. The technique was successfully applied in glasshouse and growth room conditions at UNOTT and the Technical University of Munich (TUM). The technique was transferred via training courses to Swaziland and Namibia and was performed based on documentation in Botswana. During the project period bambara groundnut was for the first time artificially cross pollinated. Morphological markers were used to verify hybrid status of the F1 and F2 plants. Pure lines were developed by single seeds descent method. Basic selection methods were transferred to local farmers to help them improve the germplasm available locally.

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1.6.2 Results

Bambara groundnut is mainly self-pollinated and anthers dehisce and the stigma becomes receptive even before flowers open. In bambara groundnut, fertilisation takes place on the same day as anthesis (Linnemann, 1994). When fertilisation takes place the ovary covered by the calyx folds back at the flower base pointing towards the glandular apex (Figure 2 [a]). While the fertilised ovary stays unchanged for several days, the peduncle elongates to bring the fertilised ovary to or below the soil surface (Figure 2 [b]). The peduncle reaches its maximum length at the initiation of pod formation. The apical end, which bears ovary with the fertilised ovules, expands into a pod (Figure 2 [c]). After 10-18 d (depending on the landrace), pod development commences. The aim of artificial hybridisation in bambara groundnut is to produce new, improved cultivars that will be of interest to growers and consumers. There are a number of factors that limit successful hybridisation in grain legumes. These include: small sized flowers and twisted keels that make flower manipulation difficult. Rate of abortion is also high, particularly after mechanical manipulation of delicate floral organs. Receptivity is also selective and, therefore, some genotypes may be better parents than others. In bambara groundnut, selection of the right sized flowers for artificial hybridisation may dictate the rate of success. For example, a flower bud of greater than 7.0 mm in length may have started releasing pollen grains (Figure 3), therefore, there is a risk that self-pollination may have occurred. It is also important to note that, as in many grain legumes, the first developing flower buds on the plant tend to set pods more easily than later developing buds. Removing other flower buds on the same node and leaving only the one for crossing purposes ensures that assimilates are directed to one pod and avoids confusion in labelling. The choice of parents in most cases depends on the breeding objectives. Initially, our aim was to develop an operational method of crossbreeding, which would be applied to bambara groundnut to facilitate its improvement. To begin with, the female parents that were easy to manipulate (spreading type) and males with an easy to score marker (anthocyanin staining) were selected. After developing a technique, selection of the parents was based on the breeding objectives, as identified by the BAMFOOD scientists based on growers and consumers’ preferences. For example, in a survey of growers and consumers conducted in Botswana, Nambia and Swaziland, early maturity and high yield were identified as the main traits that farmers would want in their ideotypes.

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Figure 2: Bambara groundnut pod development. [a] fertilised bud, [b] elongated peduncle with developing pod, [c] pod development. 1 mm grid.

Figure 3: Bambara groundnut flowers just before and after pollen shedding (grid is 1 mm).

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Emasculation Emasculation protocol:

1. Grasp the selected flower bud at the base with the left hand tweezers (Figure 4a). Care must be taken not to bruise the flower.

2. Insert tip of right hand tweezers at base of keel petals (Figure 4b). 3. Move tweezers to the tip of flower splitting the keel petals. 4. Push tip of tweezers through the middle of standard petal and split to

the tip (Figure 4c). 5. Open the flower and locate stamens and style and stigma (Figure 4d).

Care should be taken to avoid any damage to the style and stigma. 6. Push the style with left tip of tweezers to the left side of the flower to

prevent damage (Figure 4e). 7. Hold accessible stamens with tweezers (Figure 4e). Remove stamens

(with their anthers) from the flower (Figure 4f). Repeat the procedure to get hold of the remaining stamens. Open flower to check for hidden anthers.

8. Clean the tip of right hand tweezers. Let flower go from left hand tweezers. Emasculation is completed.

Figure 4: Emasculation protocol for bambara groundnut. Pollination Pollination can be carried out immediately or the day after emasculation. However, in the field the best time is in the morning because the flower bud opens at sunrise therefore the stigma receptivity and pollen viability may be higher. Also, humidity is generally high in the morning, which helps to increase the success rate of hybridisation. For successful pollination, a healthy

27

flower from a pre-identified parent is chosen. The flower should be newly opened, and if the pollination is done later in the afternoon the flower should be collected in the morning and kept at low temperature (4 oC). Pollination protocol:

1. Get stamens and carpel from male parent flower. Ensure that anthers are freely shedding pollens.

2. Open the emasculated female flower with right hand tweezers exposing stigma (Figure 5a).

3. Pollinate stigma with stamens and carpel from male parent held by left hand tweezers by brushing the anthers over the stigma of the female flower (Figure 5a).

4. Close the pollinated female flower with right hand tweezers leaving stamens and anthers on the stigma (Figure 5b). Pollination is completed.

Figure 5: Artificial cross-pollination protocol for bambara groundnut. Successful fertilisation can be determined if ovaries fold back at the flower base, pointing towards glandular apex (Figure 2 a). Unfertilised flowers fall off without folding back. In most landraces pods are developed selectively with pods located underground being preferred over pods not able to penetrate the soil. If ovaries are folded back indicating successful fertilisation they should be covered with sand or fine gravel to promote pod development. Additionally, all surrounding buds, flowers and self-fertilised pods should be removed to support the development of the hybrid pod. For confirming hybridity the seeds obtained through artificial hybridisation should be planted with their male and female parents in marked pots/rows. All F1 plants should be carefully examined for various morphological traits and compared with both parents to confirm their hybridity. Anthocyan staining of plant organs is a very useful marker due to the generally dominant inheritance of dark colours over light colours. Sometimes confirmation of hybridity can be difficult due to close resemblance between male and female parents. In such cases, the seed resulting from F1 plants are planted and F2 generations are carefully observed for segregation. If the F2 plants still look uniform for both vegetative and reproductive characters, the result of hybridity should be rejected. With the advent of molecular technologies, molecular markers play a vital role in the confirmation of hybridity.

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The technique of crossbreeding was developed using genotypes that were mainly selected for best suitability for the crossing purpose. Since the technique is now operational, the next step will be that the parents are chosen based on the results of surveys of growers and consumers and in the analysis of agronomic traits. BAMFOOD scientists have already started the selection of the parents based on the breeding objectives, as identified by the growers and consumers’ preferences. For example, in a survey of growers and consumers conducted in Botswana, Nambia and Swaziland, early maturity and high yield were identified as the main traits that farmers would want in their ideotypes. The current breeding programme has taken these into account.

1.6.4 Problems

The main problem was and is the relatively low success rate with crossing of bambara groundnut.

1.7 Overall Conclusions

A notable feature of this research project was the emphasis on incorporating indigenous knowledge and preferences of local growers and consumers of bambara groundnut in the research process. A significant finding of the surveys was that most of the preferences (high yield, large seeds, early maturity and fast-to-cook) expressed by farmers, for a bambara groundnut ideotype, were common among the African countries. One implication of this finding is that a single strategic breeding programme could meet the crop improvement needs of all three countries. Genetic and agronomic characterisation of nine bambara groundnut landraces in the field and in controlled-environment glasshouses showed that the performance of these landraces was mostly affected by the amount and distribution of rainfall. The best yields (647-1582 kg ha-1) were obtained in Swaziland where the average total rainfall ranged from 633 mm to 728 mm and lowest yields were in Botswana (69-160 kg ha-1) where the rainfall ranged from 389 mm to 433 mm. Genetic variations in the response to drought seem to exist, thereby offering an opportunity for crop improvement based on the existing genetic variation.

Investigations on the genetic diversity of bambara groundnut showed that landraces consisted of three to eight major different genotypes. Furthermore, accessions from neighbouring countries were clustered together but no pure clusters composed of accessions from a single country were observed. Participatory development of a simple and feasible marker technology for genetic diversity studies in collaboration with all partner countries and training for young African scientists on new and emerging modern technologies have enabled scientists from African partner countries to benefit from this approach and therefore reduce their isolation from the international community. Project partners were able to introduce, test and refine the molecular approaches in selected African partner institutions during the timeframe of the project.

Perhaps more important, the project has successfully demonstrated that it is possible to hybridise bambara groundnut. The hybridisation technique was successfully transferred through training courses to all partner countries. The development of an operational method of crossbreeding, with the successful productivity of the first hybrids of bambara groundnut, is a significant

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achievement because it has opened up the possibility of breeding the first true varieties of this crop. This also provides an opportunity to position QTL (Quantitative Trait Loci) for physiological traits related to resource capture and use.

We are now at a critical stage for the future of bambara groundnut. Researchers in the EU INCO-DC funded institutions and elsewhere have significant new knowledge with which to assist farmers in how best to grow the crop. We have the basis for a major breeding programme to produce novel varieties that best suit the demands of growers and consumers. Systematic breeding activities have been initiated in three African partner countries. It is strongly recommended that these activities be followed up to achieve the final result of high yielding and locally adapted varieties of bambara groundnut. The expertise generated by this projects means institutions in Africa and Europe now have a critical mass of research on bambara groundnut that is probably unrivalled on any other underutilised species. Because these activities have been co-ordinated and multidisciplinary, outputs have been timely and cost-effective with minimal duplication of effort. Within the timeframe of this project, the successful hybridisation of existing landraces and wild accessions was achieved and a core collection of bambara groundnut germplasm established. In addition, significant progress in the agronomy, physiology and pest and disease characteristics of the crop and nutritional biochemistry of its seeds has been achieved. Farmers’ knowledge and perceptions of bambara groundnut in Botswana, Namibia and Swaziland have been evaluated and modelling and mapping techniques established to determine appropriate ideotypes and agro ecological potential. The overview provided by a team approach and frequent opportunities to review progress and revise priorities afforded by workshops and symposia have meant that momentum has been maintained across a broad front of activities. To this end, the EU INCO-DC and partners believe that this project is one of the most successful collaborative efforts on any underutilised species. There is enormous scope for further genetic, physiological and agronomic studies. However, perhaps the most important priority now is to establish promotional and marketing strategies that highlight the nutritional and economic potential of bambara groundnut as a basis to expanding demand for the crop. Central to this priority is the development of new post-harvest processing techniques and value-added products based on bambara groundnut seeds. Any future strategy needs to strengthen links between farmers, consumers and researchers within existing growing regions and establish links between them and stakeholders in new potential areas for cultivation. Mapping and modelling techniques allow us to predict where the crop is likely to grow best beyond its current areas of cultivation. To this end, the crop is already being evaluated across India, one of the largest regions of the semi-arid tropics.

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1.8 List of publications

Sesay, A. Edje, O. T. Vilakati, B. Magagula, C. N. 2002. Working with farmers on the UNISWA jugo bean research project. In Proceedings of the Consultative Workshop on the University Involvement in Community Service. 4 June, 2002; University of Swaziland, Kwaluseni, Swaziland. (Ed. Keregero K.J.B) pp 81-86.


Karikari, S. K., Aganga, A. A. and Fabi, F (2002). The potential of intercropping for sustainable crop and livestock production in Botswana: Some case studies with bambara groundnut (Vigna subterranea (L) Verdc. In: Proceedings, Workshop on Rangeland and Animal Production: Prospects and Challenges. BCA/CICE, 25 - 28 February, 2002.

Magagula, C.N. Sesay, A., Edje, O.T., Nkosi, B.S., Mamba, Z., Mabuza, K. and Dlamini, T. (2002). Farmer and consumer preferences for bambara groundnut ideotypes in Swaziland. In: Sesay, A., Edje, O. T. and Cornelissen, R (eds) Increasing the productivity of bambara groundnut (Vigna subterranea (L) Verdc) for sustainable food production in semi-arid Africa. Proceedings of a Mid-Project Workshop held at the University of Swaziland 28 - 30 August, 2001. pp 19-32

Edje, O.T., Sesay, A. and Magagula, C.N. (2002). Evaluation of bambara groundnut landraces by farmers in Swaziland. In: Sesay, A., Edje, O. T. and Cornelissen, R (eds) Increasing the productivity of bambara groundnut (Vigna subterranea (L) Verdc) for sustainable food production in semi-arid Africa.. Proceedings of a Mid-Project Workshop held at the University of Swaziland 28 - 30 August, 2001. pp 33-38.

Manthe, C. S., Ramolemana, G., Karikari, S. K., Khonga, E. B., Munthali, D. C and Motlhanka, D (2002). Preliminary survey of farmers perceptions of bambara groundnut (Vigna subterranea (L) Verdc ideotype in Botswana. In: Sesay, A., Edje, O. T. and Cornelissen, R (eds) Increasing the productivity of bambara groundnut (Vigna subterranea (L) Verdc) for sustainable food production in semi-arid Africa. Proceedings of a Mid-Project Workshop held at the University of Swaziland 28 - 30 August, 2001. pp 39-45.

Fleissner, K. (2002). Management practices and preferences of bambara groundnut (Vigna subterranea) producers in Oshna region, North Central Namibia. In: Sesay, A., Edje, O. T. and Cornelissen, R (eds) Increasing the productivity of bambara groundnut (Vigna subterranea (L) Verdc) for sustainable food production in semi-arid Africa. Proceedings of a Mid-Project Workshop held at the University of Swaziland 28 - 30 August, 2001. pp 47-53.

Kaulihowa, T. and Philander, A.J. (2002). Extracts of results of the Bamfood experiment at Mahanene Research Station and Mashare Agricultural Development Institute, Northern Namibia. In: Sesay, A., Edje, O. T. and Cornelissen, R (eds) Increasing the productivity of bambara groundnut (Vigna

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subterranea (L) Verdc) for sustainable food production in semi-arid Africa. Proceedings of a Mid-Project Workshop held at the University of Swaziland 28 - 30 August, 2001. pp 57-68.

Sesay, A., Edje, O.T., Magagula, C.N. and Mansuetus, A.B. (2002). Field performance of selected bambara groundnut (Vigna subterranea (L.) Verdc.) landraces in Swaziland. In: Sesay, A., Edje, O. T. and Cornelissen, R (eds) Increasing the productivity of bambara groundnut (Vigna subterranea (L) Verdc) for sustainable food production in semi-arid Africa. Proceedings of a Mid-Project Workshop held at the University of Swaziland 28 - 30 August, 2001. pp 69-82.

Karikari, S. K., Khonga, E. B., Manthe, C. S., Ramolemana, G., Munthali, D. C and Motlhanka, D (2002). A comparative study of the performance of nine bambara groundnut (Vigna subterranea (L) Verdc) landraces at Notwane and Good Hope, Botswana. In: Sesay, A., Edje, O. T. and Cornelissen, R (eds) Increasing the productivity of bambara groundnut (Vigna subterranea (L) Verdc) for sustainable food production in semi-arid Africa. Proceedings of a Mid-Project Workshop held at the University of Swaziland 28 - 30 August, 2001. pp 83-91.

Deswarte, J.C., Cornelissen, R. and Azam-Ali, S.N. (2002). Gas exchange analysis in bambara groundnut (Vigna subterranea) landraces from Botswana, Namibia and Swaziland. In: Sesay, A., Edje, O. T. and Cornelissen, R (eds) Increasing the productivity of bambara groundnut (Vigna subterranea (L) Verdc) for sustainable food production in semi-arid Africa. Proceedings of a Mid-Project Workshop held at the University of Swaziland 28 - 30 August, 2001. pp 93-112.

Schenkel, W., E. Sticksel and G. Wenzel (2002): Development of a bambara groundnut core collection from IITA germplasm based on characterisation and evaluation data. In: Sesay, A., Edje, O. T. and Cornelissen, R (eds) Increasing the productivity of bambara groundnut (Vigna subterranea (L) Verdc) for sustainable food production in semi-arid Africa. Proceedings of a Mid-Project Workshop held at the University of Swaziland 28 - 30 August, 2001. pp. 115-122.

Mabuza, P. (2002). Taxonomic relationships in Vigna revealed by molecular markers. In: Sesay, A., Edje, O. T. and Cornelissen, R (eds) Increasing the productivity of bambara groundnut (Vigna subterranea (L) Verdc) for sustainable food production in semi-arid Africa. Proceedings of a Mid-Project Workshop held at the University of Swaziland 28 - 30 August, 2001. pp. 151-163.

Cornelissen, R., Deswarte, J.C. and Azam-Ali, S.N. (2002). A preliminary strategy for model development in bambara groundnut. In: Sesay, A., Edje, O. T. and Cornelissen, R (eds) Increasing the productivity of bambara groundnut (Vigna subterranea (L) Verdc) for sustainable food production in semi-arid Africa. Proceedings of a Mid-Project Workshop held at the University of Swaziland 28 - 30 August, 2001. pp. 167-176.

Sticksel, E., Schenkel, W., Wolbing, G and Fleissner, K. (2002). Effect of sowing density on yield and yield components in two bambara groundnut (Vigna subterranea (L.). Verdc).landraces in Namibia. In: Sesay, A., Edje, O. T. and Cornelissen, R (eds) Increasing the productivity of bambara groundnut (Vigna subterranea (L) Verdc) for sustainable food production in semi-arid

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Africa. Proceedings of a Mid-Project Workshop held at the University of Swaziland 28 - 30 August, 2001. pp. 179-186.

Edje, O.T., Dlamini, B.S. and Sesay, A. (2002). Response of bambara groundnut to NPK fertiliser levels in fallow and non-fallow land in Swaziland. In: Sesay, A., Edje, O. T. and Cornelissen, R (eds) Increasing the productivity of bambara groundnut (Vigna subterranea (L) Verdc) for sustainable food production in semi-arid Africa. Proceedings of a Mid-Project Workshop held at the University of Swaziland 28 - 30 August, 2001. pp. 187-202.

Azam-Ali, S.N. (2002). Towards a methodological framework for underutilised crops: our experience with bambara groundnut. In: Sesay, A., Edje, O. T. and Cornelissen, R (eds) Increasing the productivity of bambara groundnut (Vigna subterranea (L) Verdc) for sustainable food production in semi-arid Africa. Proceedings of a Mid-Project Workshop held at the University of Swaziland 28 - 30 August, 2001. pp. 203-223.

Schenkel, W., E. Sticksel and G. Wenzel (2002): Development of a bambara groundnut core collection from IITA germplasm based on characterisation and evaluation data. In: Challenges to Organic Farming and Sustainable Land Use in the Tropics and Subtropics (A. Deininger Hrsg.). Kassel University Press GmbH, Kassel.

Fleissner, K., E. Sticksel and W. Schenkel (2002): Participatory breeding approach of neglected crops – experience with bambara groundnut (Vigna subterranea) in Northern Namibia. In: Challenges to Organic Farming and Sustainable Land Use in the Tropics and Subtropics (A. Deininger Hrsg.). Kassel University Press GmbH, Kassel.

Sesay, A. Edje, O.T., Vilakati, B. and Magagula, C.N. (2002). Working with farmers on the EU/UNISWA jugo bean research project. Consultative Workshop on the University of Swaziland’s s Involvement in Community Service.

Fleissner, K. (2002). Linking science and farmers’ needs. Proceedings of the 2002 Annual Agricultural Research Reporting and FSRE Implementation Conference.

Massawe, F.J. (2002). Phenotypic and genetic diversity in bambara groundnut (Vigna subterranea (L.) Verdc) landraces. Annual Meeting of the American Society of Plant Biologists on Plant Biology, Denver, CO.USA, August 3-7, 2002. P182.

Schenkel, W., Massawe, F.J., Sangwan, R.S. and Ng, Q. (2003). Promotion of bambara groundnut (Vigna subterranea): Latest developments of bambara groundnut research in improvement and breeding. In: Proceedings of the Second International Workshop of the International Bambara Groundnut Network (BAMNET), 23-25 September, 1998, CSIR, Accra, Ghana (in press).

Fleissner, K. (2003). The participatory breeding approach of the BAMFOOD project. AGRI-INFO, July, 2003.

Kwerepe, B.C., Khonga, E.B., Ramolemana, G. M. and Balole, T.V. (2003). Farmers’ perceptions of a bambara groundnut ideotype. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Sesay, A, Edje, O.T. and Magagula, C.N. (2003). Working with farmers on the BAMFOOD research project in Swaziland. In: Proceedings of the International

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Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Sesay, A., Edje, O.T., Magagula, C.N. and Mansuetus, A.B. (2003). Agronomic performance and morphological traits of field grown bambara groundnut (Vigna subterranea) in Swaziland.

Edje, O.T and Sesay, A. (2003) Plant density effects on development and yield of bambara groundnut (Vigna subterranea) in Swaziland. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Khonga, E.B., Karikari, S.K., Balole, T.V. and Machacha, S. (2003). Agronomic performance of nine landraces of bambara groundnut in Botswana. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Balole, T.V., Karikari, S.K., Khonga, E.B. and Ramolemana, G.M. (2003). Effect of earthing up on yield and yield parameters of nine bambara groundnut landraces in Botswana. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Munthali, D.C. and Ramorathudi, M. (2003). Relative susceptibility of nine bambara groundnut landraces to Hilda patruellis infestation. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Singrün, C. and Schenkel, W. (2003). Fingerprinting of bambara groundnut germplasm with molecular markers. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Mine, M., Motlhabane, L.T. and Batlang, U. (2003). Preliminary assessment of genetic variation in bambara groundnut in Botswana using RAPD markers: A case of technology transfer. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Massawe, F.J., Schenkel, W., Temba, E. and Basu, S. (2003). Artificial hybridisation of bambara groundnut. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Sesay, A. and Mabuza, P.E.Z. (2003). Bambara groundnut (Vigna subterranea) improvement by the BAMFOOD research project in Swaziland. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Basu, S., Roberts, J.A., Mithen, R, Davey, M, and Azam-Ali, S.N. (2003). Towards genetic linkage mapping in bambara groundnut. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Massawe, F.J., Azam-Ali, S.N., Roberts, J.A. and Mwale, S.S. (2003). Strategic breeding in bambara groundnut (Vigna subterranea (L.) Verdc). In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Cornelissen, R., Matthews, R.B. and Azam-Ali, S.N. (2003). Modelling dry matter production and yield in bambara groundnut (Vigna subterranea (L.)

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Verdc). In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Mwale, S. Azam-Ali, S.N., and Massawe, F.J. (2003) Effect of soil moisture on root and shoot growth of bambara groundnut. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Edje, O.T., Mavimbela, E.K. and Sesay, A. (2003). Effect of seed source on growth and yield of three landraces of bambara groundnut (Vigna subterranea) in Swaziland. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Magagula, C.N., Mansuetus, A.B. and Sesay, A. (2003). Yield loss associated with pests and diseases of bambara groundnut (Vigna subterranea) in Swaziland. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Khonga, E. and Kwerepe, B.C. (2003). Effect of Thiram and Fenamiphos on incidence of Fusarium wilt-root complex in nine bambara groundnut landraces in Botswana. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Munthali, D.C. and Sondashi, M.N. (2003). Evaluation of Vigna unguiculata and Combretum imberbe ashes, Bacillus thuringiensis and actellic powder for the control of cowpea bruchid,, Callosobruchus maculates (Fab) on stored bambara groundnut. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Edje, O.T., Mavimbela, E.K. and Sesay, A. (2003). Response of bambara groundnut to plant density. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Sticksel, E. and Schenkel, W. (2003). Approaches to improve the acceptance and quality of bambara groundnut (Vigna subterranea) - a literature review. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Azam-Ali, S.N., Massawe, F.J., Mwale, S., Basu, S. and Cornelissen, R. (2003). Can bambara groundnut become a major world crop? In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Senkondo, F.J., Sibuga, K.P., Lyimo, H.F., Misangu, R.N. and Rweyemamu, C.L. (2003). Crop loss assessment and requirements for management of weeds, insects and diseases in bambara groundnut (Vigna subterranea). In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Okatch, H., Torto, N. and Amarteifio, J.O. (2003). Characterisation of the monosaccharides in bambara groundnut. In: Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

Amarteifio, J.O. (2003). A review of the nutritional composition of bambara groundnut (Vigna subterreanea( L )Verdc) grown in Botswana. In:

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Proceedings of the International Symposium on Bambara Groundnut, Botswana College of Agriculture, 8-12 September, 2003.

MSc and PhD degrees resulting from work under the contract:

1. Deswarte, J.C. (2001). Photosynthesis of bambara groundnut landraces in response to soil moisture. MSc thesis, University of Nottingham, UK.

2. Gonapa, M. (2002). Light interception and conversion in bambara groundnut landraces in response to soil moisture. MSc thesis, University of Nottingham, UK.

3. Kijoji, A. (2003). Response of three bambara groundnut (Vigna subterranea (L.) Verdc) landraces to soil moisture stress. MSc thesis University of Nottingham, UK.

4. Boateng C.O. (2003). Photosynthesis of three bambara groundnut landraces in response to soil moisture stress. MSc thesis University of Nottingham, UK.

5. Mathengwane, G.L. (2003). Tissue culture and transformation of bambara groundnut. MSc thesis University of Nottingham, UK.

6. Mwale, S.S. (2005). Growth and development of bambara groundnut in response to soil moisture. PhD thesis, University of Nottingham, UK.

7. Cornelissen R. (2004). Modelling variation in the morphology and physiology of bambara groundnut. PhD thesis, University of Cranfield, UK.

8. Basu, S. (2005). Towards the construction of a genetic linkage map in bambara groundnut. PhD thesis, University of Nottingham, UK.

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2.0 MANAGEMENT REPORT

2.1 Organisation of the collaboration

Project leaders in each institution and the project co-ordinator were responsible for the overall management of the project and the attainment of milestones. There were regular contacts between those scientists who are most involved with day-to-day activities both in Africa and in Europe. These contacts were essential for the achievements that BAMFOOD project partners have attained. Informal reporting procedures ensured that each institution produced regular brief email communications that were circulated to all participating scientists and this has been successful in establishing continuous flow of information among partners. The end of project symposium held in Botswana included farmers from different geographic areas of Botswana and overall proved to be a very successful event to mark the end of the BAMFOOD research project. Project finances were carefully managed and where appropriate the transfer of activities and funds between partners were requested and agreed upon by the EU and implemented by partners. Strategic objectives and the development of common protocols or activities have been crucial for the success that the project has achieved. The project co-ordinator has been instrumental in ensuring consistency in experimental and analytical methods throughout the project timeframe. The annual meetings, workshops, symposium and co-ordination visits provided an opportunity to increase the local and international profile of the project. An essential outcome of these various exchanges is the production of informal and formal publications for circulation within the project team and more widely to the international scientific community. This has been and will continue to be a priority for all partners.

2.2 Meetings

All project partners attended the Mid-project workshop organised at the Kwaluseni campus of the University of Swaziland (UNISWA) between August 28 and 30, 2001. The Swaziland Minister of Agriculture and Cooperatives, Mr. Roy Fanourakis, performed the official opening of the workshop, and the Vice Chancellor, UNISWA Professor Lydia P. Makhubu welcomed participants. The EU Resident Technical Adviser Mr. Aloys Lorkeers also attended the event. A total of 74 people attended and the programme of the workshop included the presentation and discussion of experimental results and the discussion of future plans. The proceedings of the workshop held at the Kwaluseni campus of the University of Swaziland (UNISWA) between August 28 and 30, 2001 were published in July 2002 (Sesay et al., 2002) and copies have been distributed widely. The project co-ordinator attended the African partners meeting held in Namibia in August 2002 for discussions on common protocols, project management and budgetary control. A final project symposium was held at the Botswana College of Agriculture (BCA) from 8th to 12th September 2003. The Botswana Minister of Agriculture performed the official opening of the symposium and the Principal, BCA welcomed participants. Over 50 participants

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attended the symposium that included presentation and discussion of experimental results and information dissemination. The proceedings of the symposium have just been published and will be distributed to farmers and scientists around the globe.

2.3 Exchange of staff

From Swaziland, Dr P. Z. E. Mabuza visited the University of Nottingham, May to August 2001 to work on molecular technology transfer, under the Bursary For Young Researchers programme. UNISWA hosted Mr. Rutger Cornelissen of UNOTT for 12 weeks in 2002 and successfully facilitated his work to carry out additional field measurements to support the development of the crop simulation model. Dr Werner Schenkel of TUM visited UNISWA and MAWRD to initiate crossbreeding programme in 2002. Dr Schenkel trained Field Assistants in Namibia and Swaziland on crossbreeding and a number of crosses on both potted and field plants were done, some of which were successful. In April 2003 TUM team conducted a workshop on DNA isolation and the marker techniques RAPD (Random Amplified Polymorphic DNA) and SSR (Simple Sequence Repeats) markers in Windhoek, Namibia. Nine individuals from different departments of the University of Namibia and of the National Botanical Research Institute participated.

2.4 Problems

The wide range of activities that were carried out during the duration of this project required scientists to communicate across both geographical and disciplinary boundaries. Wherever possible, additional funds from other sources were directed to project activities e.g. molecular technology transfer to facilitate implementation of protocols and wider dissemination of experimental results. The production of reports such as this and the use of common nomenclature and definitions will assist in future research activities in bambara groundnut.

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3.0 INDIVIDUAL PARTNER FINAL REPORTS

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University of Nottingham (UNOTT)

Project Coordinator and Leader: Dr S N Azam-Ali

Project Scientists: Dr F J Massawe

Prof J A Roberts

Dr M Davey

Students: Mr Rutger Cornelissen

Mr Simon Mwale

Miss Shravani Basu

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3.1 Survey of growers and consumers

Using additional funding from the UK Department for International Development (DFID) UNOTT appointed Ms Karen Hampson to assist the BAMFOOD research project with participatory methodology and integration of qualitative and quantitative information. She visited the three African partner countries specifically to facilitate the development and dissemination of a common work plan and methodology for a farmers’ survey. This included, capacity building, workshops and training for local staff and affiliates. Training and software were provided specifically on the use of the NUD*IST 4 package for qualitative data analysis.


A training package for incorporation of participatory methods and QSR NUD*IST 4 analysis into quantitative formats was delivered to African collaborating institutions. Short workshops were held in each country to discuss and summarise use and capabilities of the research package. A common work plan, generic methodology and set of training modules were given to each team to contribute to future staff development. Visits were made to each institution collaborating in the BAMFOOD project, between 7 November 2000 and 19 January 2001. A joint meeting of all African partners was attended in Botswana at the beginning of November 2000. The methodology for integrating qualitative information, in the context of their proposed farmers survey on bambara groundnut, was presented. The checklists and methodology were modified according to comments and input from all the countries.

3.1.2 Results

All institutions were provided with their own copy of the QSR NUD*IST 4 software. This was installed in the relevant department at each institution. All team members had a one day training course in its usage, for use with the survey data and for future needs. A full set of training documents was left with each 'trainee' and a spare copy lodged with the institution. The documents contain references and links to support networks so teams have access to some back up later should it become necessary.


Short training workshops (two to three days) were held in each country for collaborators and other interested parties. These were intended to demonstrate and discuss the use and capabilities of the entire research package. The workshops incorporated a one day fieldwork, or a meeting with farmers to pre-test the content of the checklists and for the local research team to practice using them. Extra training in survey and participatory research methods was given as necessary. The teams were then, in follow-up evaluation sessions, able to adapt the methodology according to their local needs and cultural norms. A work plan and methodology documents were prepared in collaboration with each institution to provide a common outline for the farmer’s survey. The promotion of the integrated methodology included wider dissemination to those who could potentially benefit.

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The University of Nottingham Tropical Crops Research Unit (TCRU) owns a suite of controlled-environment glasshouses within which factors such as soil moisture, temperature and humidity can be controlled for crop stands growing in natural soil (Monteith et al., 1983 and Clifford et al., 1993). The system was used for detailed physiological experiments. The protocols used at UNOTT were similar to those used in partner countries in Africa so that results obtained under controlled environments can be compared to those obtained under field conditions. In addition to results on common agronomic traits that were also evaluated by African partner institutions UNOTT also carried out studies on photosynthesis activity (based on gas-exchange), chlorophyll content and a non-destructive method of estimating leaf area in bambara groundnut. Gas-exchange information on bambara groundnut will lead to a better understanding of the photosynthetic activity of different landraces, in particular the contribution of single leaves to resource use and conversion or the effect of drought on carbon assimilation and water economy.


Three TCRU glasshouses at UNOTT were used to evaluate genetic and agronomic potential of three bambara groundnut landraces (DipC from Botswana; Uniswa red from Swaziland and S19-3 from Namibia). The crops were sown at a spacing of 35 cm between the rows and 10 cm between plants within each row. At 21 days after sowing (DAS) the crops were thinned to an intra-row spacing of 20 cm leaving a population of 180,000 plants ha-1. Prior to each planting, soil samples were taken in each bay to a depth of 60 cm and analysed for nitrogen (N), phosphorus (P) and potassium (K). Based on the soil analysis results, appropriate quantities of N, P, and K were added to each bay of the glasshouses. In addition to nutrient analysis, the soil was sterilised by fumigation with methyl bromide in order to kill any soil-borne disease pathogens and also to control weeds. There was no chemical control of either diseases or pests during the growth of the crops. Instead, biological pest control was employed by introduction of Phytoseilus persimilis in each glasshouse every fortnight at the supplier’s recommended rate. The experiments were based on two factors: landrace (genotype) and soil moisture. Growth and yield of the three landraces were studied under conditions of full irrigation and drought. In each glasshouse, one of the bays was designated as the fully irrigated bay while the other was the droughted bay. All the plots were given similar amounts of water from sowing until 42 DAS, after which no further irrigation was supplied to the droughted plots until harvesting. At 42 DAS, the crops had reached 50% flowering and the profile had at least 270 mm of water, which is considered the field capacity level for this soil as reported by Shamudzarira (1996). Recording of emergence was done everyday at 0900 h between 3 and 21 DAS, using the central five rows in the plot. In these experiments, a seedling was considered to have emerged when it had two fully expanded leaves that had broken free from the soil. Leaf number was counted twice per week using 10 randomly chosen plants in each plot. Dry matter production and partitioning was measured through growth analysis that was conducted at 3 week intervals starting at 21 DAS. Ten randomly chosen plants were harvested from each plot at each occasion and dried in an oven at 80oC for 48 h. Prior to drying, each plant was partitioned into leaves, stem and pods. In 2002, root growth

42

was measured by taking soil cores and washing the cores in order to obtain the roots. Soil coring was done on three occasions in 2002: at 42 DAS, 105 DAS and at harvest. The cores were taken at 10 cm intervals up to a maximum depth of 100 cm using a Jarrat auger with a diameter of 10 cm. A total of four positions were cored at each occasion, two of which were taken on the plant rows and the other two between the rows. Root front velocity, root length density, root diameter, root surface area and root dry weight were measured on these samples. To determine the light response of individual leaves an IRGA (CIRAS-1, PP-systems, UK) was attached to the middle leaflet of a chosen leaf, using a cuvette with a measuring area of 2.5 cm2. Different light intensities were obtained by attaching an artificial light source on top of the cuvette. The use of different combination of filters and a diffuser enabled the use of light intensities between 20 and 2000 mol PAR m-2 s-1. The CO2 concentration in the cuvette was set at 350 ppm. The measurements were taken every two weeks on the same leaflet to determine the changes in gas exchange over time. At the same time as the gas exchange measurement a SPAD (Minolta 502) was used to measure the leaf greenness of the leaflet. Measurements were taken on both sides of the midrib in the sampling area of the CIRAS cuvette and averaged.

3.2.2 Results

Results from the three years of studies show that seedling emergence under the glasshouse conditions used start to emerge at 8 DAS. Although the onset of seedling emergence is delayed, the synchrony of emergence is high as shown by the fact that the majority of the seedlings emerged within 4 to 5 days (Figure 6 A). The average seedling emergence of the three landraces did not exceed 80%. While the seedling emergence of Uniswa red and DipC was close to 80% (Figure 6 A), the corresponding value for S19-3 was only 63%. The failure to achieve higher seedling emergence in the three landraces was not investigated in these studies but the possible causes could be linked to seed dormancy and low viability.

Leaf number in the three landraces was almost linear at the beginning, after which it reduced and finally tailing off (Figure 6 B). Leaf number was, however, influenced by soil moisture content. The rate of leaf production in the irrigated treatments only showed a reduction after 84 DAS while the droughted plots showed slow leaf accumulation as early as 70 DAS. Moisture stress caused both low rate of leaf production and lower final leaf number of the plants. The effect of soil moisture on final leaf number was more severe in DipC and Uniswa red than S19-3.

Dry matter accumulation was similar in all the treatments between 21 and 63 DAS, after which the droughted treatments had a lower total dry matter than the irrigated treatments (Figure 7 A). Although the drought treatment was imposed at 42 DAS, the effect on dry matter accumulation was only evident at 63 DAS and beyond. There was a huge reduction in total dry matter due to soil moisture stress.

43

Figure 6: [A] The influence of landrace on seedling emergence and [B] the effect of landrace and soil moisture on leaf number in bambara groundnut. Data average for three years.

[A]

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44

Whereas the average dry weight of plants from the irrigated treatments was almost 40 g plant-1, the dry weight of the droughted plants was below 30 g plant-1. Figure 7 B shows the pod yield of three bambara groundnut landraces grown under irrigation and drought. Under no moisture stress, S19-3 out-yielded both Uniswa red and DipC. The yield of S19-3 under irrigation was 350 gm-2, while DipC and Uniswa red gave 310 and 296 gm-2, respectively. Soil moisture stress resulted in significant reduction in pod yield in all the landraces. The yield of both DipC and Uniswa red under drought was about 100 g m-2 while S19-3 gave close to 200 g m-1. Chlorophyll content For the relationship between SPAD values and chlorophyll contents (Figure 8) both linear and second-degree polynomials were tested. The linear model showed a good precision (95%), and also the advantage of complete linearity between SPAD value and chlorophyll content, which is important in case of averaged data. Nevertheless, this model was biased. A comparison of regression between the linear and the polynomial model showed that the polynomial model was significantly better (F=14.75*). During this experiment, the three landraces analysed exhibited the same relationship between the SPAD values and the chlorophyll content. As a consequence the actual difference in chlorophyll content between plants of these landraces can be quickly estimated using a simple, non-destructive method. The average chlorophyll content increased in the leaves with time, stabilising around two weeks after emergence. In the irrigated treatment, S19-3 exhibited the lowest chlorophyll content, whereas DipC showed the highest. Strong differences appeared between landraces in terms of chlorophyll concentration depending on the water treatment: S19-3 increased the chlorophyll content in its leaves in the droughted situation and showed the highest chlorophyll content of the three landraces, whereas DipC decreased it and exhibited the lowest pigment concentration; chlorophyll content in Uniswa red was unaffected by the water treatment. Gas exchange Gas exchange results indicated that the drought effect appeared at 70 DAS, i.e. one month after the last watering on the droughted plots. The most evident effect is a considerable drop in the maximum net assimilation, from 24 to 10 molCO2m-2s-1, in the case of the droughted plants, whereas irrigated plants maintained their maximum assimilation rate. The value of the saturating irradiance was also reduced for droughted plants. The mitochondrial respiration was similar for irrigated and droughted plants. No significant landrace effect can be seen, neither in terms of maximum assimilation rate in irrigated situation, nor in the resistance to drought.

45

Figure 7: The effect of soil moisture on [A] total dry matter production (g) per plant and [B] pod yield in three bambara groundnut landraces. Data average for three years.

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46

Figure 8: Relationship between SPAD value and chlorophyll content (area basis). Differences between landraces were more pronounced in terms of transpiration and stomatal conductance, but were less important than the water stress or the date effect. In the irrigated treatment, there was little ageing effect (no constant trend in time, but some variation between the different dates, probably caused by different meteorological conditions). However, the landraces seemed to show different transpiration rates, with S19-3 from Namibia, transpiring more than the other two landraces. In the case of the dry treatment, there was a similar sequence as that observed for the carbon assimilation: there was a significant drop one month after the end of the water supplied (70 DAS). Then, there were 2 to 3-fold reduction in the transpiration rate compared to the irrigated plants. In terms of the interaction between date and water treatment, stomata conductance remained stable in the irrigated treatment (values around 450-650 mmol m-2 s-1), reflecting no leaf ageing effect or landrace effect. The effect of drought appeared at 70 DAS, with a large reduction of stomatal conductance, from 500-100 mmol m-2 s-1. All the landraces were similarly affected by drought in terms of stomatal conductance. Non-destructive method to estimate leaf area in bambara groundnut Leaf area is an important parameter in the description of plant growth, light interception and all other photosynthetic relations, in particular for gas-exchange. The common method of estimating leaf area is that of harvesting the leaves and measuring the area using a leaf-area meter. This method is time consuming and area meters are not always available in laboratories in Africa. A simple way of estimating total leaf area is described below. A welcome bonus of this method is the possibility of following the leaf area in time, without destroying the plant. A good estimate of the leaf area of bambara groundnut is based on the assumptions that:

y = 1.77x - 18.28R2 = 0.95

y = 0.02x2 + 0.60x + 1.80R2 = 0.96

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47

i) bambara groundnut has leaves whose shape is very close to an ellipse Area for the ellipse:

A = Length x Width x π /4 (1) Equation to estimate the Leaflet area:

A = α x Length x Width x.π /4 (2) were π = 3.1416 and α = correction factor (difference between actual shape and an ellipse) ii) the size of the lateral leaflets are usually closely related to the size of the middle leaflet iii) the plant leaf-area is dependent on the leaf number and the single-leaf area. Bambara groundnut has trifoliate leaves; although these three leaflets have the same shape, they do not always exhibit the same dimensions or leaf area. In order to compensate for these differences an extra parameter is added to equation 2: A = β x 3 (α Length x Width x.π /4) (3) The step to leaf area of the whole plant is simply multiplying equation 3 with the total number of leaves. However, an extra parameter needs to be added to compensate for inaccuracy in sample methods. Young, not fully unfolded, leaves and leaves that look significantly smaller than others are rejected. This can lead to under or overestimation of the leaf area. The final equation becomes: Aplant = γ x leaf number [β x 3 (α x Length x Width x π /4)] (4) Landrace independent values for , and are 0.95, 0.91 and 0.86, respectively. R2 = 90.90 %.


Bambara groundnut showed low seedling emergence and also variations among landraces with respect to seedling emergence The studies seem to suggest that the agronomic and physiological performance of bambara groundnut landraces are strongly influenced by soil moisture. Factors such as leaf number, rate of leaf production, dry matter production and yield all showed strong dependence on the soil moisture status. Under low soil moisture, these factors were significantly depressed. However, this study appears to support the general view that the crop is drought tolerant. Although soil moisture stress led to reduction in yield, the droughted treatments still gave at least one third of the yield obtained under favourable soil moisture conditions. There was also some genetic variation among the landraces in their response to soil moisture stress, a factor that could be exploited in breeding. The different landraces didn't exhibit different photosynthetic potentials in terms of maximum net assimilation, quantum yield and saturating irradiance.

48

The transpiration rate and the Instantaneous Transpiration Efficiency (ITE), in absence of water shortage, were similar for the three landraces. The leaves showed a short period with higher respiration rate, higher light compensation point and lower chlorophyll content, just after unfolding. A logarithmic-shaped relationship seemed to link assimilation rate to stomatal conductance, which could demonstrate a form of stomatal limitation in the assimilation rate.


Developments in molecular methods for assessing genetic diversity have made it possible to estimate intra- and inter-specific variations in crop plant species. However, most of these techniques require well-equipped laboratories and trained personnel. As part of BAMFOOD research project we established a standardised RAPD protocol that can be used for genetic characterisation in laboratories with basic equipment. UNOTT has also tested cowpea-based microsatellite (SSR) primers in search for suitable primers that can be used for the same purposes in partner countries. Microsatellites are simple sequence repeats which are highly variable with regard to repeat number, show co-dominant inheritance (as opposed to dominant inheritance shown by RAPDs) and are highly efficient in the fingerprinting of crops. As with RAPDs, microsatellite analyses require only a small amount of DNA since they are a PCR-based marker system.


Standardised DNA isolation and RAPD protocols were established in collaboration with African partners and have been adopted and used successfully in three partner institutions in Africa. Detailed description of the protocols was reported in the first year annual report and appears in the Proceedings of a Mid-Project Workshop held at UNISWA, Kwaluseni, Swaziland (Sesay et al., 2002).

3.3.2 Results

Clear description of the RAPD methodology was published in the Proceedings of a Mid-Project Workshop held in Swaziland in August 2001 (Sesay et al., 2002). In country implementation of the protocols at BCA, UNISWA and UNAM (University of Namibia) was successful. Details of results and other information related to this work package appear in the two BAMFOOD research project proceedings Sesay et al. (2002) and Anonymous (2003). Participatory development of a simple and feasible marker technology for genetic diversity studies in collaboration with all partner countries and training for young African scientists on new and emerging modern technologies have enabled scientists from African partner countries to benefit from this approach and therefore reduce their isolation from the international community. Project partners were therefore able to introduce, test and refine the molecular approaches in selected African partner institutions during the timeframe of the project. Experiences in different laboratories in relation to RAPD technology and with various thermocyclers will be collected as part of an evaluation.

49



A model predicting the growth and production of bambara groundnut has been developed. The model is an adaptation of the SLM (Matthews, 2002) model for a leguminous crop. It is a sink-orientated model, i.e. the number of available sinks (pods) determines the final production. The model is a stand-alone computer program written in Delphi 6 (Borland®). It uses climate data, landrace specific parameters and physiological relationships and runs on a daily time-step to determine the biomass production and yield of a landrace in a specific environment. The basic structure of the model is demonstrated in Figure 9.

Figure 9: General overview of the bambara groundnut model. The model is built up of three separate parts: input, engine and output. The input consists of weather data, landrace dependent coefficients, soil data and questions generated from a socio-economic database. The engine is physiological based equations simulating the growth and production of bambara groundnut. The output can be linked to a GIS-system.

3.4.2 Results

A user-friendly front end for the model has been constructed. This is a MS Windows based program, which will be easy to understand with basic computer skills. It will be fully compatible with the latest MS Windows versions. Figure 10 shows the layout of the front end of the model. The front end can be connected to the main engine of the model and can then be used as the user interface.

C ro p s im u la tio n m o d e lP h ys io lo g ic a lly -b a s e d c ro p s im u la tio n m o d e l a b le to s im u la te y ie ld d iffe re n c e s b e tw e e n la n d ra c e g ro u p s in d iffe re n t e n v iro n m e n ts

G IS s ys te m

P ro d u c tio n o f a g lo b a l d is tr ib u tio n m a p fo r e a c h la n d ra c e g ro u p

O U T P U T

L a n d ra c e d a ta b a s eT o c o n ta in d a ta o n e a c h la n d ra c e g ro u p (e .g . s e e d s iz e , c o lo u r, ta s te , g e n o typ e c o e ffic ie n ts fo r s im u la tio n m o d e l, e tc ) .

S o c io -e c o n o m ic d a ta b a s eR e s u lts o f F a rm e r S u rv e y

W e a th e r D a taIN P U T

E N G IN E


G IS s ys te m


O U T P U T





G IS s ys te m


O U T P U T

G IS s ys te m


G IS s ys te m


O U T P U T







E N G IN E

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Figure 10: The front end of the bambara groundnut model. The model has been validated against glasshouse data and currently being validated against field data. Examples of output for observed and calculated values for pod production and leaf number are given in Figures 11 and 12.

05

101520253035

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observedestimated

Figure 11: Simulated and observed potential pod number for Uniswa red in controlled environment. A landrace database, containing all parameters necessary for the model engine, has been constructed in MS Excel. This database contains information for ten landraces used in the project, but can easily be expanded with information on other landraces or new varieties, when the information becomes available.

51

0

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Figure 12: Simulated and observed potential leaf number for Uniswa Red in controlled environment.


The model tends to underestimate leaf and biomass production in the glasshouses. An explanation for this can be the relatively low radiation values measured. A solution could be adding a correction factor to correct for the radiation levels in the glasshouses. In contrast the model seems to be overestimating the biomass production in the field. A problem for validating the model in the field is that there is no detailed data on photoperiod sensitivity of the landraces used in the current project, and the consequences on production are not yet fully understood. This should not give any problems for regions where daylength does not exceed 12.1 h, but it could be a serious constraint in areas were the days are longer. The model is able to predict the performance of a landrace in its environment of origin or in an environment in which it has not yet been grown. In this way the model can help to choose an ideotype landrace for a specific environment. For the model to have any practical use the ideotype it generates must be based on physiologically sound principles, crop parameters that are from existing germplasm and environmental data. Unfortunately a model cannot invent data and can thus only predict the ideotype performance if it has source data either from an existing landrace or from a hypothetical plant, based upon physiologically sound information. Secondly, the model could assist in a breeding programme. It will establish criteria or breeding objectives based on the trade-offs between desirable attributes and resource limitations. Finally, it should be mentioned that the model has been based on the SLM model for legumes (Matthews, 2002). This means that the model can easily be combined/added on other crop models built in this style. Instead of just functioning as a stand-alone bambara groundnut model, it could form part of a larger model where the production of bambara groundnut can be simulated in combination with other crops for which a SLM type model already exist. This leads to a larger integration of crop models and will increase the practical use of the model.

52

3.4.4 Problems

Non-availability of data on photoperiod sensitivity of the landraces has delayed the validation of the model using field data. Current efforts are aimed at validating the model against field data.

3.5 Crossbreeding

As part of the BAMFOOD research programme, project partners have hybridised bambara groundnut and a number of F1 hybrids have been developed for the first time. The F1 hybrids were created from crosses between contrasting cultivated and wild genotypes, selected on the basis of yield potential, variation in molecular and morphological markers, maturity and growth habit. The development of the first hybrids of bambara groundnut is a significant achievement because it has opened up the possibility of breeding the first ever improved varieties of this crop. This also provides an opportunity to position QTL (Quantitative Trait Loci) for novel physiological traits related to resource capture and use. In order to construct a genetic linkage map of V. subterranea two crosses were made: one between a wild and a cultivated accession and the second between two cultivated accessions of bambara groundnut. The F1 progeny from these crosses was then selfed. The resultant F2 progeny is being used to study a range of traits of agronomic interest. Polymorphic AFLP markers, using different primer combinations, are being identified between the parental lines and their segregation studied in the F2 population, the latter comprising more than 100 plants for each F2 population. The data obtained will be analysed in 2 stages. Firstly, the markers will be used for developing a linkage map. An attempt will also be made to correlate the genes for the traits of interest with the markers located on the map. UNOTT has also been involved in the development of a number of pure lines using the Single Seed Descent method. These pure lines have been evaluated under controlled environments and are ready to be tested and evaluated in the field. It is envisaged that these pure lines will be grown under typical farmers’ conditions and farmers will be invited to assess and make their own selection of desired lines.


Artificial hybridisation between cultivated landraces and between cultivated and wild accessions of bambara groundnut was carried out in controlled-environment growth rooms and the TCRU glasshouses at UNOTT. Although the success rate was low (~ 5 %), success was achieved in a number of crosses including those in which wild and cultivated accessions were used as parents. For detailed description of the hybridisation protocol see section 1.6. Molecular and phenotypic markers were used to confirm the success in hybridisation. Based on the success in a cross breeding programme, selection was made of 13 landraces, 2 wild accessions, 10 putative F1 hybrids between different landraces, and 1 hybrid between a bambara groundnut landrace (DipC) and the wild accession, VSSP11. These were established in a glasshouse at UNOTT during 2002. Subsequently, F2 seeds were obtained by selfing the F1 plants in the glasshouse during 2002. The focus of the 2003 glasshouse experiment has been the F2 population obtained by selfing the two single F1 plants obtained

53

from the cross between DipC and VSSP11 and Tiga nicuru and DipC. One hundred and eight F2 seeds, along with 6 seeds each of DipC and VSSP11 and 144 F2 seeds, along with 6 seeds each of Tiga nicuru and DipC, were sown on 28 April 2003. Some wild accessions failed to set seeds naturally (through selfing). Experiments were conducted to micropropagate cultivated and wild accessions using a range of culture media based on Murashige and Skoog (1962) formulation, supplemented with growth regulators known to promote shoot multiplication and root development in other legumes.

3.5.2 Results and Discussion

Several contrasting traits in plant morphology and growth habit have been identified between the parents of the hybrids. For example, VSSP11, the wild accession from Cameroon and the pollen/male parent, is of spreading habit with branches trailing over several metres. In contrast, DipC, the female parent from Botswana, is of erect habit. VSSP11 is more-or-less perennial in nature and is late maturing and has poor yields (15-20 seeds/plant). Flower initiation and pod formation (in the event of successful self fertilisation) takes place at every node along the entire length of the stems. The seed vary in size and have reddish brown testa with dark brown marks. DipC is characterised by being an annual and high yields (150-200 seeds/plant). Flower and pod formation is localized around the base of the plant; the seeds are consistent in size with cream coloured testa, each with a dark eye. The F1 hybrid seed, produced by crossing these 2 accessions, had the same testa colour as the female parent (DipC), i.e. cream testa with a dark eye. However, on selfing this single F1 plant, 147 F2 seeds were obtained which were nearly black or dark tan in colour. The inheritance of this trait and the recovery of the parental traits will be studied in the F2 and subsequent generations. Success has been achieved in multiplication of the wild accessions of bambara groundnut using stem nodal cuttings on the Murashige and Skoog media (Plate 1). Micropropagation is crucial for increasing existing stock plants prior to their incorporation into glasshouse trials and sexual hybridisation programmes.

Plate 1: Multiplication (using stem nodal cuttings) of the wild bambara groundnut accessions using tissue culture techniques.

3.5.3 Conclusions

Prior to the BAMFOOD research project, bambara groundnut never received any appreciable research effort, especially for its genetic improvement. Until then, only selection breeding was practised in which existing landraces were

54

evaluated and their seeds multiplied. However, no new combinations resulting from hybridisation had ever been produced. As a result of the BAMFOOD research project, the first ever hybrids of bambara groundnut have been developed. The development of these hybrids is a significant scientific and practical achievement and opens up the possibility of breeding true varieties of this crop.

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Botswana College of Agriculture (BCA)

Team Leader: Dr E.B. Khonga

Scientists: Prof S.K. Karikari

Dr B.C. Kwerepe

Dr G. Ramolemana

Dr T.TV. Balole

Prof D.C. Munthali

Mr S. Machacha

Dr O.M. Mine

Mr G. Legwaila

Dr J.O. Amarteifio

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In order to define an ideotype of bambara groundnut with desirable traits for farmers and consumers in Botswana, farmer and consumer surveys were conducted in each of the three seasons of the BAMFOD research project as follows: Farmer preference survey The first survey was conducted between March and August 2001 involving 50 farmers from Kgatleng (20 farmers), Southern (10 farmers), Central (8 farmers) and North East (12 farmers) districts of Botswana. Data were analysed using QSR NUD*IST 4 software.

Farmer, trader and consumer preference survey The second survey involved 467 people consisting of 244 farmers, 108 sellers and 115 consumers from seven administrative districts. In addition, 15 surveyed farmers were provided with seeds of GabC and OM1 for farmer managed landrace evaluation trials. Farmer preference quantification survey A total of 20 farmers (10 from Southern, 5 from Kweneng and 5 from Central districts) were provided with seed of OM1 and GabC for farmer managed trials. Of these 15 were interviewed in June 2003 on bambara ideotype and quantitative yield expectations using a questionnaire agreed by the African partners.

4.1.2 Results, Discussion and Conclusions

Farmers identified the following as preferred traits: bunch or spreading, high yielding, large cream seeds, early emergence and maturity (Table 6). Sellers and consumers preferred cream seed colour because it cooks faster and has good taste. For the majority of the farmers (90%), high yield meant 250-400 kg ha-1. Early maturity meant that bambara groundnut matured within 60 days after sowing while large seeds were at least 1.5 cm in diameter and 50 g per 100 seeds.



In order to characterise the genetic and agronomic traits of bambara groundnut, nine landraces, three each from Botswana, Namibia and Swaziland were evaluated in the field at two sites during 2000/2001, 2001/2002 and 2002/2003 growing seasons. The two sites were Notwane (24o33’S; 25o54’E, 994 m a.s.l.) and Good Hope (25o28’S, 25o26’E, 1245 m a.s.l.). Fields were ploughed, fertilised, and treated with nemacur and weeded regularly and seeds were dressed with Thiram. At Good Hope the crop was rainfed during the three cropping seasons while at Notwane supplementary irrigation was applied to save plants from drying out during the 2000/2001 cropping season.

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Sowing dates for 2000, 2001 and 2002 at Notwane were 13th December, 27th November and 18th December, respectively, while at Good hope the dates were 12th December, 20th November and 12th December, respectively. The experiment was arranged in a randomised complete block design with 6m x 6m plots with inter- and intra-row spacing of 50 cm and 30 cm, (6.7 plants m-

2). Climatic and agronomic data were collected. Table 6: Farmer and consumer preference for a bambara groundnut ideotype for Botswana. Ideotype trait Preference Percentage Yield High yielding 100 Seed Colour Cream 90 Pod and seed size Large 100 Seeds per pod 2-4 90 Emergence Early and uniform 60 Pod retention at harvest High 60 Plant size Large and spreading 90 Crop duration Early maturing 60 Cooking quality Cooks fast 100 Management requirement

Drought tolerant, no earthing up

100

4.2.2 Results

The total rainfall in 2000/2001, 2001/2002 and 2002/2003 cropping seasons for Notwane and Good Hope were 429 mm, 530.2 mm and 342 mm; and 395.7 mm, 467.7 mm and 303.9 mm, respectively. The rainfall distribution during the three seasons varied at both sites (Figure 13) but 2002/2003 was the poorest resulting in crop failure. The landrace effects on agronomic characteristics of the nine landraces varied with season and site (Tables 7-9). Landrace effects were significant (p<0.05) for numbers of flowers and nodules at Notwane and for stem dry weight, leaf area and number of flowers at Good Hope in 2000 / 2001 (Table 7). In 2001/ 2002 season, landrace effects were significant for root dry weight and number of nodules at Notwane and for leaf dry weight, leaf area and number of flowers (Table 8). In 2002/2003 significant differences were observed for dry weights of roots, leaf blades, petioles and stems, and in the number of flowers at both sites (Table 9). OM1 consistently performed better in most growth and reproductive parameters at both sites than the other landraces. Dry matter accumulation pattern in the nine landraces was variable within and between sites and seasons (Figures 14-16). In 2000/2001, most landraces at Good Hope reached optimal dry weight 116 days after sowing while at Notwane, the optimal weight was reached 133 days after sowing. There were no significant differences in seed yield among the nine landraces in each season. The seed yield was very low in 2002/2003 due to severe drought. There were no significant differences in overall yield of the landraces averaged across the season (Figure 17) but the trend (p=5.8%) showed that OM1 yielded highest while Nyakeni C2 yielded the lowest.

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From the findings, OM1 is closer to the ideotype requirement for Botswana in that is cream coloured, it has large seeds and yielded closer to range preferred by the farmers of 250-500 kg ha-1.

Figure 13: Rainfall distribution at Notwane (A) and Good Hope during 2000/2001, 2001/2002 and 2002/2003 growing seasons. Table 7: Morphological and yield parameters of nine landraces in 2000/2001crop season.

Site Parameters¶

DIPC

AHM 753

GABC

N C2 AHM 968

Un red

N C1 AS17 OM1 LSD

1 Rtdw (g) 0.36 0.34 0.35 0.31 0.25 0.35 0.34 0.32 0.50 ns Lfdw (g) 6.33 5.40 5.18 5.15 4.51 5.86 6.08 8.58 5.63 ns Stdw (g) 1.80 1.95 1.69 1.65 1.71 1.65 2.30 2.32 1.67 ns Pddw 0.03 0.01 0.05 0.25 0.21 0.75 0.71 0.36 0.75 ns Nlvs 45.3 37.6 48.7 44.1 38.1 36.1 42.4 47.7 55.9 ns LA (cm2) 235 201 192 191 167 217 232 318 209 ns Nfl 2.18 1.07 1.04 2.8 1.54 1.36 1.21 1.57 7.29 4.35 Nndl 8.00 6.96 6.43 6.25 6.50 2.14 8.79 6.14 7.50 5.50 Npg 14.4 9.75 10.7 13.0 10.5 12.0 12.0 11.0 1.46 ns Npd 0.79 0.07 0.86 2.64 1.79 0.75 0.71 0.36 0.75 ns 2 Rtdw (g) 1.10 1.56 1.31 1.14 1.36 1.89 1.07 1.49 1.47 ns Lfdw (g) 21.4 23.1 16.7 24.3 28.4 20.0 18.7 35.8 32.8 11.0 Stdw (g) 9.02 7.69 7.39 10.4 10.3 6.12 6.64 13.2 12.2 4.10 Pdw 3.57 6.26 2.47 3.50 6.18 2.51 4.56 11.0 11.6 6.05 Nlvs 86.3 81.9 75.8 88.6 97.1 67.7 74.4 92.0 87.5 ns LA (cm2) 795 829 632 946 961 733 1099 1159 1216 ns Nfl 7.04 10.5

7 7.54 8.46 7.21 4.75 3.11 9.71 5.14 ns

Nndl 11.1 10.0 9.29 7.93 9.14 9.79 13.5 8.86 13.3 ns Npg 22.9 38.7 25.1 52.5 26.9 17.1 37.6 51.0 44.5 21.0 Npd 10.0 25.4 11.9 17.7 24.0 6.43 13.3 25.1 27.5 13.4

¶ Rtdw = Root dry weight; Lfdw = Leaf dry weight of leaves; Stdw = Stem dry weight; Pddw = Pod dry weight; Nlvs = Number of leaves; LA = Leaf area; Nfl = Number of flowers; Nndl = Number of nodules; Npg = Number of pegs; Npd = Number of pods. ns = Not significant (p=0.05).

0

50

100

150

200

250

Oct Nov Dec Jan Feb Mar Apr

A

0

50

100

150

Oct Nov Dec Jan Feb Mar Apr

B

200020012002

59

Table 8: Means of morphological and yield parameters of nine landraces grown at Notwane (Site 1) and Good Hope (Site 2) in 2001/ 2002 crop season.

Site Parameter¶

Bambara groundnut landraces

DIPC AHM 753

GABC N C2 AHM 968

Un red

N C1 AS17 OM1 LSD

1 Rtdw (g) 0.17 0.21 0.25 0.11 0.17 0.19 0.15 0.16 0.27 0.10 Lfdw (g) 11.2 14.2 17.6 10.4 15.2 13.1 13.5 13.1 22.1 ns Stdw (g) 0.178 0.135 0.198 0.092 0.145 0.433 0.130 0.140 0.212 ns Nlvs 39.8 59.4 54.1 33.3 63.5 46.8 53.0 40.4 72.0 ns LA (cm2) 418.0 537.1 657.6 386.8 568.8 493.0 504.5 487.6 825.2 ns Nfl 7.5 6.6 9.2 5.4 6.0 6.9 8.5 5.6 10.8 ns Nndl 1.5 6.0 2.9 0.6 3.3 4.3 2.4 2.8 3.0 2.43 Npg 16.3 37.2 27.8 35.9 38.7 26.8 33.1 28.5 57.1 ns Npd 4.5 11.0 7.4 5.3 8.2 4.8 5.8 2.3 8.9 ns 2 Rtdw (g) 0.278 0.228 0.279 0.215 0.289 0.285 0.284 0.268 0.322 ns Lfdw (g) 33.0 21.1 32.6 19.0 36.0 27.9 38.4 33.0 33.2 12.4 Stdw (g) 0.24 0.21 0.32 0.16 0.30 0.28 0.30 0.34 0.26 0.10 Nlvs 77.0 55.0 78.0 45.5 92.6 66.2 80.4 103.1 75.6 ns LA (cm2) 1274 729 1215 664 1341 1039 1430 1267 1228 462 Nfl 13.5 12 19.0 12.5 29.9 15.5 16.1 19.4 22.0 11.2 Nndl 4.7 4.1 4.6 3.6 4.7 4.3 5.3 5.1 4.8 ns Npg 32.8 30.7 39.0 16.6 30.5 23.4 32.0 35.6 39.1 ns Npd 14.5 27.3 14.7 4.3 17.4 7.0 21.9 10.1 18.3 ns

¶ Rtdw = Root dry weight; Lfdw = Leaf dry weight of leaves; Stdw = Stem dry weight; Pddw = Pod dry weight; Nlvs = Number of leaves; LA = Leaf area; Nfl = Number of flowers; Nndl = Number of nodules; Npg = Number of pegs; Npd = Number of pods ; ns = Not significant (p=0.05).

60

Table 9: Morphological and yield parameters of nine landraces in 2002/2003 crop season.

Site

Parameter¶

Bambara groundnut landraces

DIPC AHM 753

GABC N C2 AHM 968

Un red

N C1 AS17 OM1 LSD

1 Rtdw (g) 0.23 0.28 0.17 0.19 0.17 0.41 0.23 0.21 0.19 0.11 Lfdw (g) 8.74 6.52 8.62 8.19 7.70 7.63 6.63 8.18 8.89 3.16 Ptdw 2.00 1.54 2.03 2.20 2.17 1.79 2.02 2.07 1.84 1.11 Stdw (g) 2.34 1.95 2.32 2.46 2.46 2.15 2.06 2.43 2.47 1.26 Nlvs 64.00 56.81 64.86 67.75 69.61 63.94 55.75 69.81 71.31 ns LA (cm2) 323.8 232.9 306.1 303.9 285.3 283.1 245.9 303.7 329.8 ns Nfl 0.00 0.56 10.17 3.89 3.42 4.78 13.97 6.17 15.56 10.1 Nndl 0.78 1.03 0.89 1.22 1.39 1.22 1.42 1.67 1.64 ns Npg 15.31 7.94 9.64 17.61 17.72 13.61 11.89 17.53 18.28 ns Npd 0.25 0.06 0.00 0.03 0.17 0.14 0.00 0.00 0.53 ns 2 Rtdw (g) 0.40 0.45 0.56 0.32 0.39 0.39 0.33 0.36 0.30 0.11 Lfdw (g) 15.21 15.21 16.06 13.93 16.43 16.36 19.58 19.65 13.81 3.16 Ptdw (g) 4.30 4.02 5.56 3.90 4.68 5.58 6.37 5.66 3.83 1.11 Stdw (g) 6.08 5.71 6.15 4.66 5.89 5.97 7.23 7.22 4.87 1.26 Nlvs 82.67 81.03 77.76 70.03 91.75 96.44 88.17 89.25 75.56 ns LA (cm2) 564.3 514.4 611.9 515.0 609.5 588.7 666.6 729.2 511.5 ns Nfl 0.64 0.01 9.40 11.17 26.86 29.44 39.64 24.78 26.11 10.1 Nndl 1.64 1.22 1.38 2.25 1.22 1.92 1.08 1.25 1.67 ns Npg 21.67 27.17 19.64 29.36 29.53 19.58 23.64 24.69 22.22 ns Npd 3.99 4.31 2.81 2.92 5.86 2.03 3.42 2.22 2.08 ns

¶ Rtdw = Root dry weight; Lfdw = Leaf dry weight of leaves; Stdw = Stem dry weight; Pddw = Pod dry weight; Nlvs = Number of leaves; LA = Leaf area; Nfl = Number of flowers; Nndl = Number of nodules; Npg = Number of pegs; Npd = Number of pods ; ns = Not significant (p=0.05).

61

NOTWANE

0

50

100

150

46 74 101

132

Days after sowing

Dry

mat

ter/p

lant

(g) DIPC

AHM753GABCNyakeni C1HM 968Uniswa RedNyakeni C2AS17OM1

GOOD HOPE

0

10

20

30

40

47 61 71 89 103 117 133

Days after sowing

Ddr

y m

atte

r/pla

nt (g

) DIPCAHM753GABCNyakeni C1HM 968Uniswa RedNyakeni C2AS17OM1

Figure 14: Total dry matter accumulation in nine bambara groundnut landraces grown at Notwane and Good Hope in 2000/2001.

62

NOTWANE

0

20

40

60

80

47 61 71 89 103 117 133

Days after sowing

Plan

t mas

s/pl

ant (

g) DIPCAHM753GABCNyakeni C2HM 968Uniswa RedNyakeni C1AS17OM1

GOOD HOPE

0

20

40

60

80

100

50 64 78 93 107 122

Days after sowing

DIPCAHM753GABCNyakeni C2HM 968Uniswa RedNyakeni C1AS17OM1


63

NOTWANE

05

1015202530

21 63 84 98 119

Days after sowing

Dry

mat

ter/p

lant

(g)


GOOD HOPE

0

20

40

60

80

21 63 84 98 119

Days after sowing

Plan

t mas

s/pl

ant (

g)



0

50

100

150

200

Seed

yie

ld (K

g/ha

)

DC A753 GC NC2 A968 UR NC1 As17 OM1

BGN Landrace Figure 17: Seed yield of nine landraces averaged over three seasons.


Environmental factors, especially rainfall had the greatest effect on performance of the nine landraces. In general, OM1 and GabC were closest to the ideotype of bambara groundnut for Botswana conditions due to its cream colour and relatively high yield compared to the other landraces.

64

4.2.4 Problems

The main problem experienced during the study was the erratic and low rainfall especially during the 2002/2003 growing season.



Seeds of the nine landraces were sown in pots using vermiculite as growth medium. After 14-21 days DNA was extracted from seedlings using the protocol adapted from Massawe et al. (2002). RAPD reactions were performed as described by Massawe et al. (2002).

4.3.2 Results

High molecular weight genomic DNA was successfully isolated from 90 individual plants, 10 individuals per landrace. We are currently carrying out PCR amplifications on the 90 DNA samples from 10 individual plants of each landrace using primers from kits C and D. Preliminary results suggest that there are variation among landraces. Further work and data analysis are currently underway. Capacity in the use of RAPD technology has been developed at BCA, and institutional capacity in the use of molecular technology will continue to be developed and strengthened.


The transfer of molecular technology from European to African partners was a component of the BAMFOOD research project. As part of this exercise, a preliminary assessment of genetic diversity among nine landraces of bambara groundnut was carried out using RAPD markers. RAPD markers were generated by two sets of 20 arbitrary 10-mer primers. Preliminary results suggest the existence of genetic variation among the bambara groundnut landraces.

4.4 Crossbreeding


Fifty seeds of each of the nine landraces were germinated in Petri dishes in the laboratory and in pots in the greenhouse and examined for anthocyanin in the primary leaflets, stems and the pulvini. Plants with anthocyanin (females) were crossed with anthocyanin free plants (males). About 150 flowers of AS17, NyakeniC1 and OM1 were emasculated and pollinated with pollen of DipC, GabC, Uniswa red, OM1 and NTSR in the greenhouse and in the field. After emasculation all other flowers present and any subsequent flowers developed were removed. Single seed descent and pure lines Ten plants of each landrace in the main agronomic plots were selected and tagged. Seeds from each plant were harvested and kept separately for growing during next season in order to characterise them as first generation selfed seeds to produce SSD lines.

65

4.4.2 Results

Nyakeni C1, AS17 and OM1 were found to be anthocyanin-free and were used as females in cross breeding. Four flowers in the greenhouse (two each from Nyakeni C1 x AS17 and Nyakeni C1 x OM1) and nine (two from Nyakeni C1 x AS17; four C1 x OM1 and three OM1 x NTSR) in the field developed pods. These were harvested and the F1 seeds stored for future research. Single seed descent Two generations of SSD lines have been produced for all the landraces and evaluated for pod and seed characteristics. The SSD selection will be continued to purify the landraces.


Crossbreeding of bambara groundnut has been accomplished and it is hoped that the techniques developed during the BAMFOOD research project will be utilised by other researchers working on the crop. The development of single seed descent lines will continue for further purification.

4.4.4 Problems

Poor germination of the F1 seed was the main problem.

66

Technical University Munich (TUM)

Team Leader: Prof Dr G Wenzel

Scientists: Dr W Schenkel

Dr E Sticksel

Dr C Singruen

67



Seed protein, P and K analysis To obtain additional information on the landraces that were investigated under BAMFOOD research project seeds were analysed for crude protein, potassium and phosphorus contents. Seeds of 154 genebank accessions were also analysed alongside the landraces for comparison. Definition of a core collection The aim of this work was to classify genebank accessions into several groups or clusters based on their genetic distance, in order to retrieve from each cluster one member as entry into a core collection. Thus, a high genetic diversity can be maintained within the core collection. Since no information on the diversity based on molecular data was available, phenotypic data had to be used to estimate genetic distances of accessions. Passport descriptors were excluded from the analysis, since it cannot be assumed that genetic characters influence them. Characterisation descriptors are generally highly heritable and can be easily seen by eye and are equally expressed in all environments and are therefore highly informative in terms of revealing genetic diversity. Evaluation descriptors are more prone to environmental variability and are therefore less informative than the previous category. Since evaluation descriptors are generally the most interesting traits for crop improvement, and are strongly influenced by genetic factors, they were included in this analysis with the same weight as the characterisation descriptors. Based on characterisation and evaluation data from the IITA germplasm, genetic distances were calculated and a cluster analysis performed to establish a core collection. After suitable transformation and standardisation genetic distances between all accessions were calculated. Based on the resulting distance matrix, a hierarchical cluster analysis using Ward’s minimal variance method was performed resulting in a dendrogram and an agglomeration schedule for 1013 accessions. From the selected cluster levels a hierarchical nested core collection was defined.

5.1.2 Results

Seed protein, P and K analysis Among the landraces closely investigated under BAMFOOD research project there was a wide range of protein contents spanning from 14.5% in Nyakeni C2 to 20.1% in DipC (Table 10). The average P and K content in this group was 0.34 and 1.40%, respectively. The protein content of the seeds obtained from the genebank accessions showed a wide range of protein contents from 13.9% to 29.6% with a mean of 20.1 % in a normal distribution (Figure 18). The Potassium content showed a distribution that seems to split the genotypes in two groups with an average percentage of approximately 1.5% K and 2.2% P, respectively (Figure 19).

68

Table 10: Percentage protein, potassium and phosphorus in the seed of bambara groundnut landraces. Landrace % Protein % P % K Nyakeni C2 14.5 0.16 1.49 Uniswa red 15.6 0.23 1.14 AS17 15.8 0.38 1.40 Nyakeni C1 16.5 0.22 1.41 OM1 16.6 0.36 1.30 Gab C 17.0 0.48 1.58 AHM968 17.5 0.38 1.35 AHM753 18.2 0.46 1.69 Dip C 20.1 0.39 1.24

Protein%29,027,025,023,021,019,017,015,013,0

30

20

10

0

Std.abw. = 2,81 Mittel = 20,1N = 154,00

Protein%29,027,025,023,021,019,017,015,013,0

30

20

10

0

Std.abw. = 2,81 Mittel = 20,1N = 154,00

Figure 18: Distribution of seed protein content in bambara groundnut genotypes.

P %,65,60,55,50,45,40,35,30,25,20,15

30

20

10

0

Std.dev. = ,09

Mean = ,43

N = 154,00

P %,65,60,55,50,45,40,35,30,25,20,15

30

20

10

0

Std.dev. = ,09

Mean = ,43

N = 154,00

K %

2,302,10

1,901,70

1,501,30

1,10

50

40

30

20

10

0

Std.dev. = ,22 Mean = 1,53N = 154,00

K %

2,302,10

1,901,70

1,501,30

1,10

50

40

30

20

10

0

Std.dev. = ,22 Mean = 1,53N = 154,00

Figure 19: Distribution of seed phosphorus and potassium content in bambara groundnut genotypes. Definition of a core collection Since the statistical procedure for calculating distances with SPSS software does not allow missing data in the data matrix, accession numbers and descriptors were selected in order to optimise the use of the data set. The exclusion of 23 descriptors with missing data resulted in a set of 33 remaining descriptors with complete data for 1013 out of 1463 accessions. Thus, 69.0 %

69

of the characterised germplasm was included in the analysis. In this study the size of the core collection was not predefined but deduced from the structure of the data, thus resulting in a core collection that represents the genetic structure of the main collection. To achieve this, cluster levels with a higher stability than surrounding cluster levels were identified. The stability of a cluster level was defined by the increment in coefficient of distance from one cluster level to the next and was retrieved from the agglomeration schedule. A high increment in coefficient of distance indicated a stable and "natural" cluster level. Out of the identified stable cluster level 5 levels with 8, 16, 32, 70 and 102 clusters, respectively were selected, based on stability information. From each of the selected cluster levels a core collection was defined in such a way that the total number of entries for all five core collections does not exceed 102. The most typical accession in each cluster was identified and chosen as representative for the cluster. Thus at the highest cluster level a core collection of eight entries was defined representing eight groups of accessions (Figure 20). These eight accessions were fixed to represent their respective cluster at each lower level. Therefore at the next lower level with 16 clusters eight additional representatives were identified to form a core collection of 16 entries. This procedure was continued down to the 102 cluster level, forming a hierarchical nested core collection. All IITA accession numbers of the core collection are presented in Figure 20.

Figure 20: Dendrogram of hierarchical cluster analysis and accession numbers of core collection entries of bambara groundnut.


All of the landraces investigated under the BAMFOOD research project have lower protein content compared to the average of all investigated genotypes, with exception of DipC. The wide range of protein contents in genebank material indicates that there is substantial variation that can be exploited in breeding programmes. Since bambara groundnut can contribute substantially

70

to the supply of high value protein to rural farmers, the existing variability should be exploited in breeding programmes. A completely new type of core collection was developed. The advantages of this type of core collection are, an adjustable size, a facilitated genebank management, an optimised genetic diversity in the core collection, a strong linkage to the main collection. Besides the information on the genetic structure of the germplasm, information on agronomic and physiological traits can be linked with entries to the core collection and groups of accessions. These cluster profiles can be used to identify groups of accessions with features corresponding to ideotypes identified by agro-ecological modelling or farmer’s surveys. The combination of information on genetic structure and information on agronomic traits will support the development of breeding strategies for bambara groundnut.

5.1.4 Problems

The major drawback of this method is that the descriptors that are influenced by environmental variance give only a crude estimation of genetic distances. This problem can be circumvented by the use of molecular marker data, which are not subject to environmental influences. For this reason a molecular marker analysis was performed (See section 5.2).

5.2 Molecular analysis of germplasm


In total, genomic DNA of over 400 individual bambara groundnut plants was isolated for molecular marker analysis. The accessions included those from the IITA bambara groundnut genebank, the Namibian national collection and the nine landraces specifically studied under BAMFOOD research project. For marker analysis AFLP markers were used and SSR markers established in other legume species were adapted. The resulting molecular data were used to calculated genetic distances between accessions. A hierarchical cluster analysis was performed based on distance data. Within landrace diversity was investigated.

5.2.2 Results

The AFLP reactions were performed with ten primer combinations (Table 11). The number of amplified fragments ranged from 55 for E46M49 to 186 for E32M49. The average level of polymorphism per primer pair over all genotypes was 22% resulting in a total number of 189 AFLP -markers developed for bambara groundnut. Of 14 SSR primer pairs, 10 amplified fragments in a size range corresponding to the original microsatellites. Of these only primer pair AG81 from soybean amplified 18 polymorphic markers. The analysis for intra-landrace-diversity showed that the analysed landraces consist of three to eight very similar, but different genotypes: AHM 753 6 genotypes Gap C 4 genotypes AHM 968 3 genotypes Nyakeni C1 5 genotypes AS 17 3 genotypes Nyakeni C2 8 genotypes Dip C 6 genotypes Uniswa Red 3 genotypes OM1 3 genotypes Cibadak 7 genotypes

71

Table 11: AFLP primer combinations and level of polymorphism generated in bambara groundnut.

Primer combination

No. of amplified fragments

No. of polymorphic fragments

Polymorphic fragments (%)

E32M47 85 27 31.8 E32M49 186 18 9.6 E33M49 62 19 30.5 E34M53 76 22 29.1 E35M48 87 26 29.8 E39M47 103 9 8.8 E40M47 79 9 11.5 E41M58 61 20 32.9 E44M49 59 22 37.5 E46M49 55 17 30.7 Mean 85.3 18.9 22.16

The marker data were used to construct a dendrogram using Ward method. The resulting dendrogram describes the genetic relationship among the different bambara groundnut landraces (Figure 21). The cluster levels for interpretation of the dendrogram were chosen based on stability information from fusion level graph (Figure 22). The 17 cluster level was the most interesting for analysing the grouping. The clustering showed a good correlation to the origin of accessions. All Namibian accessions were clustered together (clusters 8 and 9). Accessions from neighbouring countries, Ghana and Nigeria, as well as those from Benin clustered together (clusters 4 and 15). Similarly, accessions from the neighbouring countries Zambia and Zimbabwe were clustered together (clusters 2, 5-7 and 17). Noticeable is the fact that the accessions from Madagascar and Indonesia neither were clustered together nor could they be associated with any other country. Table 12 presents the cluster membership of the investigated bambara groundnut accessions.

72

Figure 21: Dendrogram showing genetic relationships among 223 bambara groundnut accessions generated using AFLP marker data.

8

11 12

14

13

15

16

17

1

9

10

4 5

6

7

3

2

73

Figure 22: Fusion level diagram generated from marker data in bambara groundnut. Table 12: Cluster membership of investigated Germplasm. Cluster 1: Cluster 2: Cluster 4: Cluster 6: BEN TVsu193 GHA TVsu157 GHA TVsu164 ZIM TVsu1052 NGA TVsu620 TZA Dodoma

cream BEN Tvsu207 ZIM TVsu1072

GHA TVsu142 ZMB TVsu733 GHA Tvsu216 NGA Tvsu610 ZMB TVsu850 ZMB TVsu762 NGA Tvsu586 ZIM TVsu1056 ZMB Tvsu712 ZMB TVsu790 NGA Tvsu577 NGA Tvsu573 GHA TVsu150 ZMB TVsu892 GHA Tvsu215 ZMB Tvsu740 ZMB TVsu922 NGA Tvsu588 GHA Tvsu220 ZIM TVsu972 ZMB TVsu927 NGA Tvsu631 Cluster 7: TZA TVsu183 MDG TVsu810 GHA Tvsu210 ZMB Tvsu747 GHA TVsu139 CMR Tvsu560 NGA Tvsu673 ZMB TVsu788 NAM Sb2-1 NGA TVsu838 GHA Tvsu221 ZIM TVsu1064 IDN Parung black NGA Tvsu599 NGA Tvsu608 ZIM TVsu974 IDN Ramayana black

ZIM TVsu1035 NGA Tvsu653 ZIM TVsu1017

IDN Parung red ZIM TVsu1006 CMR Tvsu541 ZMB TVsu955 TZA Dodoma red ZMB TVsu857 CMR Tvsu569 ZMB TVsu854 SWZ Manzini tan ZIM TVsu1000 NGA Tvsu597 ZMB TVsu884 SWZ Manzini black ZIM TVsu992 MDG TVsu815 ZMB TVsu932 ZMB TVsu735 Cluster 5: ZIM TVsu1025 SWZ Manzini butterfly

MDG TVsu817 KEN TVsu793 ZIM TVsu1034

ZMB Tvsu730 ZMB Tvsu787 ZIM Tvsu1011 NGA TVsu590 ZMB Tvsu742 ZMB TVsu791 ZIM TVsu1050 ZIM TVsu1008 ZMB Tvsu712 ZIM TVsu1060 ZMB TVsu875 ZMB TVsu935 ZIM TVsu1061 ZMB Tvsu692 MDG TVsu814 ZMB Tvsu739

0

0,5

1

1,5

2

2,5

3

3,5

25 24 23 22 21 20 19 18 17 16 15 14 13 12 11 10 9 8 7 6 5 4 3 2

Number of clusters

Dis

tanc

e co

effic

ient

74

SWZ Manzini star Cluster 3: ZMB Tvsu695 SWZ Manzini shade star

BEN TVsu189 ZMB Tvsu686

SWZ Manzini shade eye

NGA TVsu614 ZMB Tvsu691

SWZ Manzini blackring

CMR TVsu551 ZMB Tvsu688

ZMB Tvsu986 NGA TVsu581 ZIM TVsu1016 ZIM TVsu1024 NGA TVsu841 ZIM TVsu1029 NAM Sb16-5a ZIM TVsu1032 MDG Tvsu822 ZIM TVsu1033 Cluster 8: Cluster 8

continued Cluster 10 Cluster 12

ZMB Tvsu852 ZIM TVsu1026 ZMB TVsu866 BWA Dip C NAM 1761/3 ZIM TVsu1013 ZMB TVsu927 BWA Dip C NAM 1603/2 NAM Sb4-2 ZMB TVsu784 BWA Dip C NAM 1689/2 MDGTVsu829 BWA Dip C NAM 1756/3 NAM Sb19-3 Cluster 11 BWA Dip C NAM 1143/2 NAM 235/3 NAM AHM 753 BWA Dip C NAM 1689/2 NAM Sb16-5a NAM AHM 753 MDG TVsu801 NAM 1690/2 NAM KFBN 9501 NAM AHM 753 ZIM TVsu1049 NAM 1690/2 IDN Parung brown NAM AHM 753 BWA Gab C NAM 1756/3 NAM 1749/3 NAM AHM 753 BWA Gab C NAM 1144/3 NAM 1143/2 NAM AHM 753 BWA Gab C NAM 1144/3 NAM AHM 968 BWA Gab C NAM 1690/2 Cluster 9: NAM AHM 968 BWA OM 1 NAM 481/3 NAM 1691/2 NAM AHM 968 BWA OM 1 NAM 1144/3 NAM 426/3 ZAF AS17 BWA OM 1 NAM 1689/2 NAM 235/3 ZAF AS17 NAM 1143/2 NAM 1749/3 ZAF AS17 NGA Tvsu130 NAM 481/3 NAM Sb2-1 NAM 235/3 Cluster 13 Cluster 15 Cluster 16 Cluster 17 SWZ Nyakeni C1 GHA TVsu134 TZA TVsu184 GHA TVsu161 SWZ Nyakeni C2 GHA TVsu160 TZA TVsu379 ZMB TVsu942 SWZ Nyakeni C1 GHA TVsu158 MDG TVsu811 ZIM TVsu999 SWZ Nyakeni C1 ZMB TVsu845 ZMB TVsu889 ZIM TVsu1018 SWZ Nyakeni C1 ZMB TVsu846 ZMB TVsu853 ZIM TVsu1045 SWZ Nyakeni C1 GHA TVsu166 ZMB TVsu736 ZIM TVsu966 SWZ Nyakeni C2 NGA TVsu591 ZMB TVsu743 ZIM TVsu990 SWZ Nyakeni C2 NGA TVsu601 ZMB TVsu716 ZIM TVsu988 SWZ Nyakeni C2 NGA TVsu173 ZMB TVsu721 ZIM TVsu989 SWZ Nyakeni C2 NGA TVsu174 ZMB TVsu881 ZIM TVsu1009 SWZ Nyakeni C2 GHA TVsu136 ZMB TVsu719 ZIM TVsu1038 SWZ Nyakeni C2 GHA TVsu140 ZMB TVsu727 ZMB TVsu891 SWZ Nyakeni C2 GHA TVsu138 MDG TVsu796 ZIM TVsu996 GHA TVsu144 ZIM TVsu1015 Cluster 14 ZMB TVsu950 SWZ Swazi red ZMB TVsu954 SWZ Swazi red ZIM TVsu1004 SWZ Swazi red ZIM TVsu1005

75

IDN Cibadak ZIM TVsu1023 IDN Cibadak ZIM TVsu1014 IDN Cibadak ZMB TVsu930 IDN Cibadak ZMB TVsu948 IDN Cibadak ZMB TVsu896 IDN Cibadak ZIM TVsu1043 IDN Cibadak ZMB TVsu921 ZMB TVsu925 ZMB TVsu934 ZMB TVsu941 Intra landrace diversity analysis The developed markers were also used to analyse the intra landrace diversity. None of the investigated landraces was genetically pure. From each of the nine landraces 15 individual plants were randomly chosen for analysis. The number of identified different genotypes ranged from 3 to 8. Even the landrace “AS 17” which was regarded as pure line prior to this analysis consisted of different lines. Despite these findings within landrace diversity was lower in landraces from partner countries compared to the genebank accessions (Figure 23).


Existing marker technologies were successfully adapted and applied on bambara groundnut. AFLP markers were not as easy scorable as SSR markers but were available abundantly. The frequency of polymorphisms was near optimal in a range of approximately 20.0%. This results in a good efficiency of marker development with a high reliability due to a sufficient amount of common markers between each pair of genotypes. The application of SSR markers adapted from related species was only partly successful. Only 10 out of 14 tested primers amplified fragments in bambara groundnut and of these only 1 produced polymorphic patterns. This result was expected, since SSR markers are known to be highly specific. For an efficient application of SSR markers these need to be developed for bambara groundnut. It is highly recommended since this marker type is most reliable and has the best prospect of being applied in laboratories with basic equipment. The benefit of molecular markers for analysis of genetic diversity was clearly demonstrated. The clustering of genotypes from the same origin into groups indicates that relationships can be identified based on genetic similarities. Each group of three landraces from the three African partner countries was clearly separated from each other (Figure 23). The Namibian landraces “ AS 17”, “AHM 968” and “AHM 753” clustered close together as can be seen from the dendrogram and Figure 23. This information can be directly converted into practical breeding recommendations. When selecting parents for crossing the Namibian lines should be crossed with foreign lines rather than with each other in order to maximise variability in the resulting population and thus selection success. The landraces from Botswana “OM1”, “DipC” and “GabC” showed a higher diversity than the Namibian landraces, indicating a broader genetic base. In the Swaziland landraces “Uniswa red”, “NyakeniC1” and “NyakeniC2” there was a surprisingly high distance between “Uniswa red” and the “NyakeniC” group, indicating a generally great diversity in Swaziland landraces.

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Figure 23: Two-dimensional scaling of genetic distances between accessions. Countries of origin are coded by symbol and investigated landraces from partner countries are named.



In April 2003 a workshop on DNA isolation and the marker techniques RAPD and SSR took place in Windhoek, Namibia. Nine individuals from different departments of the University of Namibia (UNAM) and of the National Botanical Research Institute participated.

5.3.2 Results

Nine individuals from different departments of the University of Namibia (UNAM) and of the National Botanical Research Institute participated were successfully trained in techniques for DNA extraction, SSR and RAPD analysis.

5.4 Crossbreeding


Plants were raised under glasshouse conditions for flower structure, floral biology studies and cross breeding. A procedure for emasculation of bambara groundnut flowers was developed and applied on glasshouse grown plants. The procedure was documented and transferred to African partners. See Annual report 1 and 2 and the proceedings of the Mid-project workshop held in Kwaluseni, Swaziland (Massawe et al., 2002).

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5.4.2 Results

Emasculated flowers were retained for at least 3 d demonstrating that peduncles and pedicels were not mechanically damaged. The procedure of emasculation and cross-pollination was documented by digital photography for further dissemination of the technique among partners. In TUM glasshouse facilities, 272 flowers were emasculated and pollinated. Folding back of the ovaries and sepals at the top end of the peduncle about 2-4 d after pollination was identified as marker for successful fertilisation. Based on this marker the success rate for cross fertilisation was at approximately 30 %. Unfortunately in bambara groundnut development of pods depends strongly on their position on the plant. Due to better accessibility, most flowers located on higher or outer stems of the plant were cross- pollinated, resulting in many cross-fertilised flowers not developing into pods. The first successful cross hybridisation in bambara groundnut was performed in September 2001 with the Namibian landrace "AS17" as a female parent and Tanzanian landrace "Dodoma red" as pollen donor. Flowers with numbers 11201 and 11501 developed pods, which were harvested in December 2001. Seed colour and size was typical for "AS17". Thus the hybrid status of plants “11201” and “11501” were confirmed by the transfer of a dominant paternal trait, namely anthocyanin staining of primary leaf to the F1 generation. At later stages five additional hybrid seeds were harvested. The success rate for fully developed hybrid pods was 2.6%. The hybrid plants 11201 and 11501 yielded 14 and 9 F2 seeds, respectively. Both hybrid plants showed clear signs of heterosis effects. The plants were superior in leaf number, stem number, petiole length and leaf area to their parental lines ‘AS17’ and ‘Dodoma red’. The F2 seeds were germinated in August 2002 to achieve optimal climatic conditions with respect to day length, temperature and light. A total of 21 F2 plants developed. This F2 generation demonstrates a segregating pattern in petiole and leaf colouration. To transfer the technology for crossing of bambara groundnut effectively to the African partners Dr Schenkel visited Swaziland and Namibian partners in March 2002. In both countries colleagues were trained successfully on the technique and crossings were done in field as well as pot grown plants. During this visit about 200 flowers were cross-pollinated.

5.4.3 Problems

Despite the first successful cross hybridisation of bambara groundnut was performed the success rate is low. This can be attributed to morphological and physiological properties of bambara groundnut. With properly trained personnel and optimised conditions of cultivation it should be possible to raise the success rate in a breeding programme to an acceptable level of 5-10%.

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Ministry of Agriculture, Water and Rural Development (MAWRD)

Team Leader: Mr K Fleissner

Scientists: Miss T Kaulihowa

Miss E Namwandi

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Farmer and consumer surveys were conducted during the three growing seasons (2000 – 2002) in order to define an ideotype of bambara groundnut with desirable traits for farmers and consumers in Namibia. In 2000/2001 season, 29 farmers from Uukwangula and Ompundja Agricultural Development Centres were interviewed. In 2001/2002 20 farmers from Okahao-Kangala and Omakange villages, 16 consumers and 28 traders from Rundu, Oshakati, Windhoek, Walvis Bay and Swakopmund were inteviewed. A semi-structure questionaire agreed by African partners was used and the data analysed using QSR NUD*IST 4 software.


From the farmers survey, the ideal bambara groundnut landrace should be: early maturing, high yielding with large pods and seeds, attractive seed colour mainly cream, sweet taste and short cooking time. The consumers and sellers preferred attractive colour (cream), large seeds, sweet taste. For breeding purposes, early maturity, high yield and attractive seed colour were the most important traits for incorporation into a bambara groundnut variety.

6.1.3 Conclusions

Through successful crossbreeding efforts, it may be possible to develop bambara groundnut varieties with desired traits for farmers and consumers in Namibia.

6.1.4 Problems

Human and financial resources limited surveys. The 2002/2003 in-depth quantitative surveys of farmers were not conducted because of poor health of the Team Leader.



In order to characterise the genetic and agronomic traits of bambara groundnut, nine landraces, three each from Botswana, Namibia and Swaziland were evaluated in the field at two sites during 2000/2001, 2001/2002 and 2002/2003 growing seasons. The two sites were Mahenene Research Station in North Central Region and Mashare Agricultural Development Institute in Kavango Region. Fields were ploughed and plots were prepared soon after adequate rains had been received. The experiment was arranged in a randomised complete block design with 6m x 6m plots with inter- and intra-row spacing of 50cm and 30 cm, (6.7 plants m-2 respectively) as agreed by African partners. Climatic and agronomic data were collected. Farmer-managed trials were also conducted during the three seasons using four landraces from the BAMFOOD research project and one from the farmers.

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Farmers were provided with seed and stationery for recording data but managed the plots using their practices. Only final grain yield was assessed at the end of the season.


The three seasons were characterised by low, and poorly distributed rainfall and high temperatures. These affected agronomic parameters such as emergence, number of flowers and pod development at the two sites. Seed yield among the nine landraces were not significantly different at each site and during each season (Table 13). In general, yields at Mahanene and Mashare were higher in 2000/2001 season than in 2001/2002 season because of low rainfall in 2001/2002 compared to 2000/2001. Based on yield and other agronomic traits the bambara ideotypes for Namibia are AHM 968, NyakeniC2 and DipC for areas with low rainfall like Mahanene and NyakeniC2, Uniswa red and DipC for areas with relatively higher rainfall like Mashare. Results from farmer-managed trials showed that AS17 and NyakeniC2 yielded lower than the farmers’ landraces while AHM 968 was at par with the farmers’ landraces (Figure 24).

6.2.3 Conclusions

Rainfall had the greatest impact on the performance of the nine landraces. In general, AHM 968, NyakeniC2, Uniswa red and DipC were promising landraces close to the ideotype of bambara groundnut for Namibia. Under farm-managed trials, however, AHM968 was the best landrace in terms of yield.

6.2.4 Problems

Erratic rainfall was the main problem during the study. During the 2002/2003 season, the resignation of the project technician Ms T Kaulihowa and the poor health of the team Leader adversely affected the collection and analysis of data. Table 13: Seed yield of nine bambara groundnut landraces during two cropping seasons at Mashare and Mahanene in Namibia. Season

Location

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AS 17

AHM 968

AHM 753

GABC DIPC OM1 UNIWA

RED NYAKC 1 NYAKC2

2000 Mahanene 548.5 720 390.2 363.7 603 519.4 472.3 308 683.9 Mashare 699.7 990 1055 968.1 1070 806.5 1166 1018.4 1269.7 2001 Mahanene 390.5 272.1 232.4 360.8 296.3 215.6 363.8 231.5 370.7 Mashare 153.5 207.2 126.4 222.2 276.1 167.1 223.5 273.2 282.3

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Figure 24: Performance of AS17, Nakeni C2, AHM 968 and DipC compared to farmers’ own landraces.

6.3 Crossbreeding


Single plant descents, covering the whole spectrum of genotype diversity of each landrace, have been collected from the Mahanene experiment according to the prescribed protocol. Dr. Schenkel from TUM spent two weeks in Namibia in March 2002, to train the BAMFOOD assistant, Ms Kaulihowa, in crossbreeding technologies for bambara groundnut. There were a number of initially successful crosses (from close to 100 executed crosses), which however failed to develop pods. Reasons for that are not known.

6.3.2 Future plans

Single seed descent lines have been selected and will continue to be purified.

Performance of landraces on-farm

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University of Swaziland (UNISWA)

Team Leader: Prof A Sesay

Scientists: Prof O T Edje

Dr C N Magagula

Dr A B Mansuetus

Dr B S Nkosi

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To objectives were to define a local bambara groundnut ideotype, identify specific desirable bambara groundnut attributes, and establish a list of achievable objectives to guide the improvement of the crop, through the collection of information on farmers’ knowledge, perception and preferences for bambara groundnut.


Building upon the findings of a baseline survey carried out among 124 subsistence farmers during the 1997/98 cropping season as a prelude to the commencement of the BAMFOOD project, and funded by the Research Board of the University of Swaziland, three surveys were conducted in series as follows: Farmer and consumer preference survey This survey was conducted between February and July 2001, involving a total of 95 farmers across the four administrative regions of the country. They were interviewed in 11 focus group discussion sessions, using a checklist. Additionally, a total of 30 consumers were interviewed individually in three cities (Mbabane, Manzini and Nhlangano). Data analysis was carried out using the QSR NUD*IST 4 (Non-numerical Unstructured Data* Indexing Searching and Theorising) software. Farmer preference quantification surveys In other to quantify some of the qualitative data obtained from the farmer preference survey and to rank preferences expressed by the farmers for an ideotype, a total of 24 farmers, were interviewed (7 in March 2002 and 17 in April-May 2003), using a structured questionnaire and administered in association with samples of seed, pods, etc.


From the baseline survey conducted before the commencement of the BAMFOOD project information had been collected on: growers of the crop, where and how it is cultivated, problems associated with growing the crop, and local knowledge existing among the farmers regarding the crop. In this survey farmers identified, among other characteristics, high yield, large pods, large seed size, pod retention (strong pegs) and early maturity, as desirable traits for a bambara groundnut ideotype for Swaziland (Table 14). Consumers stressed the easy-to-cook characteristic. Farmer preference quantification survey To the majority of farmers (61.5%), “high yield” means pod yields of 2.0 to 3.0 t ha-1. Large pods are pods of at least 1.69 cm diameter, and “large seeds” refer to seed of at least 58g/100 mass. An early maturing bambara groundnut variety is one that would complete its life cycle in three to four months. Thus the achievable objectives for any bambara groundnut improvement programme aimed at attaining high yield in Swaziland should include:

- high biomass production or crop growth rate - high harvest index or partitioning coefficient

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- large pods (1.69 cm diameter and above) - early maturity (3 to 4 months)

Table 14: Farmer and consumer preference for a bambara groundnut ideotype for Swaziland. Crop Trait

Preference and (% of respondents)

Yield

High yielding (100)

Seed colour Cream (50) Emergence rate Early and uniform Growth habit / plant size Spreading / large (83.2) Crop duration Early maturing Number of seeds per pod 2 to 4 (76.6) Pod and seed size Large (100) Pod retention at harvest High (100) Cooking quality Easy to cook (100) Management requirement No requirement for earthing-up


To characterize genetic and agronomic traits of bambara groundnut landraces from African partner countries.


Nine landraces, three each from Botswana, Namibia and Swaziland, obtained through exchanges of seed between the African partners, were evaluated in three field experiments conducted in the 2000/2001, 2001/2002 and 2002/2003 crop seasons. The trials were conducted at two sites, Malkerns (26º30´ S, 31º 13´ E; 700 m a.s.l.) and Luve (26º 20´ S, 31º 14´ E; 580 m a.s.l.). Sowing dates were November 15, 21, and 25 at Malkerns and December 6, January 7 and December 9 at Luve, for the three crop seasons, respectively, within each location. The trials were conducted in a randomized complete block design with four replications. Individual plot size was 6 m x 6 m. Plots consisted of twelve rows spaced 50 cm apart, with 30 cm between plants in the row (6.7 plants m-2). All plots were fertilised, and insect, weed and disease control measure were applied. Farmers were invited to participate in the evaluation of the landraces at the Malkerns location. In 2001/2002 and 2002/2003 landraces exhibiting positive agronomic traits and matching farmers’ needs, as defined in the surveys, were evaluated in farmer-managed on-farm trials.


Morphological development Bambara groundnut landraces varied significantly (P 0.01) in leaf production and maximum leaf area index (LAI) (P 0.05) (Tables 15-17). There were highly significant differences between locations for all vegetative traits measured. However, there was no significant location x landrace interaction for any of the vegetative traits.

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Table 15: Range and means of morphological, phenological and yield characteristics of nine bambara groundnut landraces grown in two locations in Swaziland, 2000/2001.

Malkerns Luve Traits Range Mean S.E. Range Mean S.E. Days to physiological maturity (PM)

140-152 146 0.75 112-119 116 0.50

Thermal time to PM ( C d) 1651-1762 1699 6.83 1547-1709 1661 11.34 Leaf no. plant-1 117-155 128 2.37 82-109 96 2.39 Reproductive period (d) 95-101 96 0.75 67-72 70 0.55 Seed yield ( kg ha-1) 1015-1749 1211 72.16 276-831 577 62.74 Dry matter (t ha-1) 5-7 5.8 0.18 2-3 3 0.19 100-seed mass (g) 36-60 50 1.76 35-52 45 1.56 Pods plant-1 40-73 57 3.77 25-43 32 2.61 Harvest index 8-30 21 0.01 14- 37 23 0.02 Partition. coefficient 0.2-0.7 0.5 0.03 0.3-0.8 0.6 0.05 CGR kg ha-1 d-1 35-52 43 2.39 26-39 31 2.25

* Thermal time (ºC d) to PM; thermal times based on air temperature above Tb = 10 ºC. Table 16: Range and means of morphological, phenological and yield characteristics of nine bambara groundnut landraces grown in two locations in Swaziland, 2001/2002.

Malkerns Luve Traits Range Mean S.E. Range Mean S.E.

Days to physiological maturity (PM)

126-135 131 0.53 122-126 125 0.39

Thermal time to PM ( C d) 1577-1999 1748 60.2 1743-1778 1763 3.34 Leaf no. plant-1 130-158 40 2.33 51-91 62.8 1.52 Maximum leaf area index 5-6 5.3 0.21 1-2 1.4 0.10 Reproductive period (d) 87-89 88 0.53 59-75 68 0.98 Seed yield (kg ha-1) 965-1743 1493 75.9 319-468 92 18.6 Dry matter (t ha-1) 5-7 6 0.18 1.5-2.2 2 0.83 100-seed mass (g) 66-81 62 2.20 43-63 64 1.88 Pods plant-1 51-96 63 3.19 15-27 20 1.19 Harvest index 19-30 25 0.01 18-31 25 0.01

* Thermal time (ºC d) to PM; thermal times based on air temperature above Tb = 10 ºC.

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Table 17: Range and means of morphological, phenological and yield characteristics of nine bambara groundnut landraces grown in two locations in Swaziland, 2002/2003.

Malkerns Luve Traits Range Mean S.E. Range Mean S.E. Days to physiological maturity (PM)

1245-131 128 0.36 99-105 101 0.38

Thermal time to PM ( C d)

1651-1762 1699 6.83 1691-1797 1760 7.41

Leaf no. plant -1 133-158 146 2.54 NA NA NA Maximum leaf area index

3-5 4 0.15 1.2-1.4 1.3 0.06

Reproductive period (d)

69-75 73 0.40 55-61 57 0.47

Seed yield (kg ha-1) 1348-2548 2042 92.6 744-1299 977 50.0 Dry matter (t ha-1) 4.4-5.6 4.9 0.17 2.6-4.0 3.4 0.14 100-seed mass (g) 47-78 60 1.80 32-55 45 1.34 Pods plant-1 35-81 52 2.93 32-57 38 1.84 Harvest index 32-52 42 0.02 24-44 31 0.02 Partitioning coefficient 0.6-0.9 0.8 0.02 0.4-0.6 0.5 0.07 CGR kg ha-1 d-1 36-58 50 2.88 35-48 41 1.43 Total abortion (%)¶ 36-74 60 2.36 33.4-64.0 56 2.38

Thermal time (ºC d) to PM; thermal times based on air temperature above Tb=10 ºC. ¶ Flower + ovule + seed abortion.

Reproductive development and phenology Time to physiological maturity varied significantly (P 0.01) among landraces (Tables 15-17). Flowering and maturity occurred significantly (P 0.01) earlier at Luve than at Malkerns, with year-to-year variations due to early season drought. Dry matter production and seed yield Dry matter accumulation was similar among the nine landraces at both locations (Table 15-17; Figure 25), but was significantly affected by location, averaging 5.9 t ha-1 and 2.7 t ha-1 of total dry matter at Malkerns and Luve, respectively, in the 2000/2001 cropping season. The trends in seasonal accumulation of dry matter in the component plant parts (Figure 26), suggest substantial dependence of pod-filling on partitioning of assimilates from current photosynthesis than on remobilisation of stored assimilates from vegetative organs. Seed yield varied significantly (P 0.01) among landraces in each of the three years (Tables 15-17). However, the relative performance of the landraces was not consistent between locations and among years. Seed yield was consistently and significantly (P 0.01) higher at Malkerns than at Luve across the three years. Seed yield components Number of pods per plant and 100-seed mass differed significantly (P 0.01) among landraces in each of the three years (Tables 15-17). Location, years and landraces had highly significant (P 0.01) effects on harvest index (HI), but no effect on crop growth rate (CGR). However, partitioning coefficient, estimating the proportion of post-flowering dry matter partitioned into pods, varied significantly (P<0.01) among landraces in 2000, and between locations in 2002. The landrace AS 17, with its high LAI, large seed size, high harvest

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index and high partition coefficient, was the highest yielding landrace overall, followed by Uniswa red and NyakeniC1.

Figure 25: Dry matter production against days after sowing for nine bambara groundnut landraces grown at Malkerns (A) and Luve (B) in Swaziland, 2002/2003. Vertical bar denotes S.E. mean.

Figure 26: Seasonal dry matter accumulation in component parts of bambara groundnut landraces at Malkerns 2002/2003: A = NyakeniC1, B = AS 17.

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Flower production was not a limiting factor to yield in any of the locations (Figure 27). In 2002/2003 only 39.2% of flowers produced at Malkerns developed into fertile pods, compared to 44.9% at Luve. Total abortion per plant (flower + ovule + seed) ranged from 35.8% to 74.4% at Malkerns, and from 33.4 to 64% at Luve.

Figure 27: Cumulative number of flowers per plant against days after sowing for nine bambara groundnut landraces grown at Malkerns (B) and Luve (A), in 2002/03. Relationship between seed yield and agronomic traits Seed yield was significantly correlated with number of pods per plant (r=0.73**), seed mass (r=0.27**), total dry matter at PM (r=0.69**) and harvest index (r=0.54**), across landraces, locations and years. Regression analysis indicated that 91% of the total variation in seed yield of the nine landraces was accounted for by a linear function involving number of pods per plant, total dry matter at maturity and harvest index. Thus the improvement of pod production, HI and total dry matter could lead to an increase in bambara groundnut yields. Farmer-managed on-farm trials of selected landraces Averaged across farmers, the test landraces out-yielded the farmers’ control both in the 2001/2002 (Figure 28) and the 2002/2003 crop seasons, but the differences were not significant. The feedback indicates a general preference among farmers for the test landraces, largely because of their large seed size.


To develop a “standardized RAPD Kit” with African partners to facilitate genetic characterization of local bambara groundnut landraces.

To assist in building a molecular technology base in partner countries To provide training to young scientists from African country partners on the

practicality of molecular technology.

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Figure 28: Performance of selected bambara groundnut landraces in On-farm trials in the Manzini region, Swaziland, 2001/2002.


Dr. P. Z. E. Mabuza visited the University of Nottingham, May to August 2001, on EU fellowship under the Bursary for Young Researchers Programme, to enhance her skills in the use of molecular markers. She developed a simple protocol for DNA extraction. She then used this protocol in association with RAPD and AFLP analyses to study taxonomic relations in Vigna. A PCR machine was bought with funds transferred from BCA. Accessory units and consumables have been procured and a laboratory established. Preparations are being made for testing of the RAPD kit developed at UNOTT in characterizing local bambara groundnut landraces.

7.3.2 Results

Dr Mabuza’s results provided further confirmation that the current taxonomy of Vigna, which is based mainly on morphology, does not reflect true genetic relationships within the genus. Through the BAMFOOD research project molecular technology capacity is being strengthened at UNISWA.


Aimed at incorporating farmer’s survey, experimental and molecular information into a quantitative database, which can be used for selection criteria and crop simulation modelling.


We hosted Mr Cornelissen from UNOTT for 12 weeks and successfully facilitated his work to carry out additional field measurements to support the development of the crop simulation model. We have provided survey, field

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data and weather data to support the development of the crop simulation model.

7.5 Crossbreeding

Aimed at establishing an operational method of crossbreeding in bambara groundnut.


Between 2001 and 2003 a total of 92 crosses were made between bambara groundnut landraces employing the artificial hybridization technique developed at our European partner institutions. Additionally, the single-seed descent (SSD) method was used to advance single plant selections, made on field experimental plots, through three selfing generations.

7.5.2 Results

Three successful crosses were achieved in which pods developed to maturity. Also, through three selfing generations of single plant selections made during the 2000/2001 cropping season ten SSD lines were produced, with their seed mass ranging from 57.6 to 136.5 g/100 air-dried seeds. F3 seeds have been obtained from the 2001/2002 crosses. The hybrid status of these seeds is yet to be determined, using molecular marker technology.

7.6 Strategic investigations


The following additional field studies were performed to include local problems and practices:

Yield loss associated with insect pests and diseases Mineral nutrient content of landraces Performance of bambara groundnut on fallow and non-fallow land Residual effect of bambara groundnut on subsequent maize crop Influence of plant density on yield-forming traits Effect of seed source on the performance of bambara groundnut

7.6.2 Results achieved

1. Pests and diseases are not major factors contributing to low yields in bambara groundnut in Swaziland

2. Significant differences exist among landraces in iron, zinc and calcium content of seeds

3. Bambara groundnut yielded significantly higher on fallow than on non-fallow land

4. Bambara groundnut did not significantly affect the yield of the subsequent maize crop

5. Plant density had no significant effect on seed yield 6. Environmental conditions during seed production can influence the yield

performance of the progeny.

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7.7 Dissemination workshop

A one-day workshop was planned for end of November 2003 to disseminate the results to NGOs, farmers, Extension Division of the Ministry of Agriculture, Senior research and administrative personnel of the Ministry of Agriculture, the University community etc. End of November is the earliest date that would not conflict with planting operations for farmers.

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References Anonymous (2003). Proceedings of the International Symposium on Bambara groundnut, 8-12 August 2003. Botswana College of Agriculture, Botswana. Clifford, S.C., Stronach, I.M., Mohamed, A.D., Azam-Ali, S.N. and Crout, N.M.J. (1993) The effects of elevated at atmospheric carbon dioxide and water stress on light interception, dry matter production and yield in stands of groundnut (Arachis hypogaea L). Journal of Experimental Botany, 44:1763-1770.

Doku, V.E. (1997). Problems and prospects for the improvement of bambara groundnut. In: Proceedings of the First International Symposium on Bambara Groundnut, University of Nottingham UK, July 1996. pp 3-19.

Du Petit-Thouars, L. (1806). Genera nova Madagascariensia, Paris, France. P23.

FAO (1981). Legumes in human nutrition. FAO, Food and Nutrition Paper 20. FAO,Rome, Italy.

Hodges, T. (1991). Predicting crop phenology. CRC Press, Boca Raton, FA, USA. 248 pp.


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Data sheet for final report

Contract number: ICA4-CT-2000-30002

Final report

1. Dissemination activities

Total (cumulative)

Number of communications in conferences (published) 50 Number of communications in other media (internet,

video) 30

Number of publications in refereed journal (published) 0 Number of articles/books (published) 0 Number of other publications 2 2. Training Number of PhDs 3 Number of MScs 5 Number of visiting scientists 2 Number of exchanges of scientists (stay longer than 3

months) 2

3. Achieved results Number of patent applications 0 Number of patents granted 0 Number of companies created 0 Number of new prototypes/products developed 0 Number of new tests/methods developed 30 Number of new norms/standards developed 2 Number of new software/codes developed 11 Number of production processes 0 4. Industrial aspects Industrial contacts No Financial contribution by industry No Industrial partners: Large No SME No

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