Serotype Specificity of Antibodies against FMDV in Cattle in Selected

Districts in Uganda

FRANK NORBERT MWIINE

Department of Biomolecular Resources & Biolab Sciences (BBS)

School of Biosecurity, Biotechnical & Laboratory Sciences (SBLS)

College of Veterinary Medicine, Animal Resources and Biosecurity (COVAB)

Ph.D dissemination of research findings

Introduction

• Foot-and-mouth disease (FMD) is one of the most important livestock diseases globally due to its economic impact

• Affects cattle, pigs, sheep, goats, buffalos, and all cloven hoofed wildlife species.

• FMD virus (FMDV): classified within the Aphthovirus genus as a member of the Picornaviridae family

• Extremely contagious, difficult and expensive to control and eradicate

Introduction

• FMDV: 7 immunologically distinct serotypes:-O, A, C, Asia 1, SAT 1, SAT 2 & SAT 3

• In Africa, 6 serotypes exist apart from Asia 1 that has never been reported

• In Uganda FMD is endemic, first confirmed in 1953

• Control measures based on vaccination and quarantine of livestock

Clinical signs of FMDCharacterized by fever and vesicles in the mouth, muzzle, teats, and feet. Excess salivation (drooling) is obvious

Clinical signs of FMD

Severe erosions on the dental pad and gums. (mouth lessions)

Raw tongue

FMD, Ruptured tongue

FMD lesion on the tongue

Classical FMD, raw tongue, flothing

Clear FMD wounds on coronary band

FMD lesion on the gum

FMD, Raw gum

FMD classical sign

FMD, lesion on nostril

FMD Lesion on the tongue

Healing tongue, FMD

Burning FMD infected cattle

Study objective• To determine the serotype-specificity of the circulating

FMDV antibodies in cattle in selected districts in Ugandaduring the major FMD outbreak in 2006.

Map of Uganda showing FMD post-outbreak study districts, 2006.

study carried out in 7 districts;

Bushenyi, Isingoro, Mbarara, Kasese, Mpigi, Kiboga and Kiruhura

A total of 349 cattle sera were collected from 28 herds

30 OP collected from 9 herds (Mb,Ks)

Methods• Serum antibody assays

i. Screening for antibodies to FMDV Non Structural Proteins (NSP) & Structural Proteins (SP) was performed using Ceditest® FMDV-NS & FMDV type O test kits (Cedi Diagnostics BV, Lelystad, The Netherlands).

ii. Solid Phase Blocking ELISA (SPBE) an in-house system set up at DTU, National Veterinary Institute, Lindholm Denmark/MAAIF-Uganda

• FMD Virus isolation /Ag ELISA

ELISA TEST

ELISA TESTColour development, FMD serotype O

After addition of stop solution FMD serotype O

Results From 22 cattle herds with visible typical clinical signs

to FMD, 285 sera were obtained; NSP= 78%, SP-O=82%

From 6 cattle herds that had no visible c/s to FMD,64 sera were obtained; NSP= 5%, SP-O=6%

SPBE; serotypes O (38%, 22/58), SAT 1 (11%, 5/45),SAT 2 (2%, 1/49) and SAT 3 (9%, 7/79) at cut-off of≥160

Only FMD serotype O virus was isolated(GBACN:EF611987)

District Village Herd no. No. of samp. tested NSP Positive (%) SP-O Positive (%)Bushenyi Kitwe Bs6 9 9 (100) 9 (100)

Kitwe Bs5 10 9 (90) 10 (100)Rwenjeru Bs3 11 10 (91) 11 (100)Kashozi Bs1 10 0 (0) 0 (0)Busheregyenyi Bs2 10 0 (0) 0 (0)Mashonga Bs4 9 0 (0) 0 (0)

Isingiro Kagogo Is1 5 5 (100) 5 (100)Kasese Kahendero Ks2* 6 5 (83) 6 (100)

Kahendero Ks3* 8 7 (88) 7 (88)Kahendero Ks4 27 22 (81) 25 (93)Kahendero Ks13 2 1 (50) 1 (50)Kahendero Ks15 3 1 (33) 3 (100)Kasomoro II Ks5* 21 15 (71) 14 (67)Kasomoro II Ks6 16 7 (44) 15 (94)Nyabubale Ks7* 4 4 (100) 4 (100)Nyabubale Ks8 14 13 (93) 11 (79)Nyabubale Ks9 13 13 (100) 13 (100)Nyabubale Ks10* 9 9 (100) 9 (100)Nyabubale Ks11*π 19 16 (84) 18 (95)National park Ks12 30 19 (63) 2 (7)

Kiboga Butembe Kb1 6 4 (67) 4 (67)Mbarara Ishanyu Mb1 8 8 (100) 8 (100)

Kafunjo Mb2* 21 11 (52) 18 (86)Muko Mb3* 12 10 (83) 12 (100)Muko Mb4*ß 10 7 (70) 10 (100)

Mpigi Madu Mp1 23 18 (78) 21 (91)Kiruhura Rurambira Kr3 15 0 (0) 1 (7)

Kazo Kr4Ω 18 2 (11) 2 (11)Total 349 225 (65) 239 (69)Total for herds with clinical signs of FMD 285 222 (78) 235 (82)Total for herds without clinical signs of FMD 64 3 (5) 4 (6)

Res. T2: Titration of serotype-specific antibodies against seven Foot -and-mouth disease (FMDV) serotypes

Titre* O A C Asia 1 SAT 1 SAT 2 SAT 3

< 1.90 19/58 15/15 8/8 2/2 37/45 44/49 67/79

≥1.9, < 2.20 17/58 0/15 0/8 0/2 3/45 4/49 5/79

≥ 2.20 22/58 0/15 0/8 0/2 5/45 1/49 7/79

*Expressed on log10 and cut-off ≥160.

serotypes O (38%, 22/58), SAT 1 (11%, 5/45), SAT 2 (2%, 1/49) and SAT 3 (9%, 7/79)

Conclusions

• Study shows that majority of the FMD outbreaks in 2006 were caused by FMDV serotype O

• Evidence of antibodies to both SAT 1 and SAT 3 in one FMD outbreak in a non-vaccinated herd (ks12) inside Queen Elizabeth national park area

© 2010 Blackwell Verlag GmbH • Transboundary and Emerging Diseases. 57 (2010) 365–374

Serotype-specificity of antibodies towards FMD in cattle herds surrounding Lake Mburo National Park in

2008

Study areaLake Mburo National Park (LMNP) in the South-Western part of Uganda-Kiruhura District

211 sera samples from 23 herds of long horned Ankole breeds aged more than four years were collected with a history of previous FMD outbreaks

Methods• Antibody assays

i. Screening for antibodies to FMDV Non Structural Proteins (NSP) & Structural Proteins (SP) was performed using Ceditest® FMDV-NS & FMDV type O test kits (Cedi Diagnostics BV, Lelystad, The Netherlands).

ii. Solid Phase Blocking ELISA (SPBE) an in-house system set up at DTU, National Veterinary Institute, Lindholm Denmark/MAAIF-Uganda

Results• Out of 211 cattle sera samples:-

– 42.7% (90/211) were positive for antibodies against NSP of FMDV

– 75.4% (159/211) were positive for SP of FMDV serotype O

FMDV Titres ≥ 1:160 in SPBEs were:-

– Serotype O, 61% (19/31),

– Serotype A, 33% (5/15),

– SAT 1, 67% (20/30),

– SAT 2, 37% (10/27)

– SAT 3, 2% (4/33)

conclusions

• FMD outbreaks in cattle herds around LMNP were probably caused by FMDV serotype O, A and/ or SAT-serotype(s).

• Usage of non-purified, multivalent vaccines obscures the serological diagnosis of FMDV outbreaks

General Recommendations

• Regional studies should be carried out to establishFMDV serotypes and/or strains involved in diseaseoutbreaks and emphasis should be on virus recovery.

• For future serological work, circulating field viruses inUganda should be obtained and homologous reagentbe made for SPBE.

• Post outbreak sampling for serological diagnosisshould be focused on young unvaccinated stock of 6-12-months of age.

• Carrier status should be evaluated in districts whereFMD outbreaks are frequent.

FMD Control

• Vaccination of animals –cattle and sheep goats and pigs

• Strict quarantine of animals

• Establish FMD free zones in Uganda/regions

• Observe all-in-all out rule of animals

• Restriction of persons and trucks and proper sanitation(animal owners, vets, trucks)

Acknowledgements

• This study was funded by DANIDA under the Livestock Wildlife Diseases in East Africa Project (LWDEA), grant number: P104.Dan.8.1.316.

1. National Animal Disease Diagnostics and Epidemiology Centre, Ministry of Agriculture Animal Industry and Fisheries, Entebbe, Uganda

2. Makerere University Institute of Environment and Natural Resources, Kampala, Uganda

3. National Veterinary Institute, Danish Technical University, Kalvehave, Denmark

4. Department of Biology, University of Copenhagen, Denmark

• My former supervisors,

William Olaho-Mukani, Kirsten Tjørnehøj

• Project coordinators and scientists/advisors

Anna Rose Ademun Okurut, Prof. Søren Alexandersen, Vincent Muwanika,, Laurids Siig Christensen, Hans Redlev Siegismund, Charles Masembe , Karl Johan Sørensen, Graham Belsham, Jorn Klien, Bøtner Annete

• Former PhD scientists on LWDEA project : Chrisostom Ayebazibwe, Sheila Nina Balinda, Abraham Kiprotich Sangula

• Technical team:

Esau Martin, Eugene Arinaitwe, Zac Duluga (deceased), Patrick Atim Nedi, Preben Norman, Jani Christensen, Jane Borch and Tina Fredricksen

Thank you