algal growth potential and nutrient limitation in a tropical river-reservoir system of the central...

Algal growth potential and nutrient limitation in a tropicalriver-reservoir system of the Central Plateau, Mexico

Eugenia Lopez Lopeza,* , Laura Davalos-Lindb

aLab. Ictiologıa y Limnologıá, Depto. Zoologıá. Escuela Nacional de Ciencias Biolo´gicas, IPN. Carpio y Plan de Ayala, Col. Sto. Toma´s,Mexico, D.F. 11340, Mexico

bChapala Ecology Station. Universidad Autońoma de Guadalajara and Baylor University, PO Box 97388, Waco, TX 76798-7388, USA

Abstract

Algal bioassays to determine algal growth potential of waters and nutrient limitation were conducted using a standard testorganismSelenastrum capricornutumand native phytoplankton from the sites sampled. The study site was the river-reservoirsystem of the Rıó Grande de Morelia located on the Mexican Central Plateau. The US Environmental Protection Agency’sprotocol for algal bioassays was followed. In this river-reservoir system, urban and industrial impacts increase downstream. Thebioassays showed that water from sites impacted by rural settlements has a low algal growth potential and that nitrogen limitsgrowth. Sites located at urban and industrial settlements have a high algal growth potential and phosphorus limits algae growth.q 1999 Elsevier Science Ltd and AEHMS. All rights reserved.

Keywords:Bioassay;SelenastrunCapricornutum; Nutrient; Limitation

1. Introduction

This study assessed algal growth potential (AGP) ofwater and the nutrient limitation of a river-reservoirsystem of the Rıó Grande de Morelia using algalbioassays. The potential impact of external nutrientloads on aquatic ecosystems cannot adequately bedetermined on the basis of physical and chemicalfeatures alone. The measurement of P and N concen-trations in water gives static values and does notreflect processes taking place in the ecosystem(Nilssen, 1978; Box, 1983). Measurement of func-tional responses of the biota to nutrient additions bybioassays allows prediction of their impact on algalproduction and detection of synergistic effects that arenot possible to assess by other procedures (Miller etal., 1978).

Algal bioassay techniques for determining limitingnutrients were standardized by the National Eutrophi-cation Research Program (NAERP) and the US Envir-onmental Protection Agency (EPA) (NAERP andEPA, 1971; EPA, 1978). The standardized techniquemakes it possible to compare results among labora-tories. Several species were used as standard testorganisms, butSelenastrum capricornutumis widelyused as the test species in fresh water (Maloney andMiller, 1972; Couture et al., 1985). In our research,native phytoplankton was used in addition to the stan-dard test organism. The use of native organisms inbioassays was been found to give a more realisticresponse and is widely recommended (Munawar etal.,1983; Da´valos et al., 1989; Da´valos-Lind, 1996).The use of native algae in bioassays gives morerealistic responses as the manipulation of nutrientadditions will cause not only changes in the algaebiomass but also shifts in species dominance.

Aquatic Ecosystem Health and Management 1 (1998) 345–351

1463-4988/99/$19.00q 1999 Elsevier Science Ltd and AEHMS. All rights reserved.PII: S1463-4988(98)00028-1

* Corresponding author.

Bioassays conducted in temperate latitudes haveidentified P as the nutrient which is most oftenlimiting for algal growth (Maloney et al., 1972;Martin and Novotny, 1975; Golterman, 1983). Incontrast, few studies performed in tropical areasfound that N was the algal growth limiting nutrient,for example, at Lago Titicaca, Bolivia-Peru´ (Vincentet al., 1984; Wurtsbaugh et al., 1985, at Lobos Reser-voir in Brazil (Henry et al., 1984) and in some tropicalAfrican lakes (Kalff, 1983). In the only Mexicanbioassay determination of nutrient limitation to date,Davalos et al., 1989, who conducted bioassays usingSelenastrum capricornutumand native algae in LakeChapala, Mexico, found N to be the limiting nutrient.

We report how AGP and the limiting nutrientchanges along the river that is impounded threetimes and that runs from a series of very small villages

at a forested headwaters to important urban areas suchas the city of Morelia and to important industrialdevelopments including a fertilizer factory. Thissystem presents the opportunity to evaluate changesin the health of the ecosystem using the headwaters asa reference point to compare with localized industrialand urban point sources of stressors to the environ-ment.

2. Study site

Rıo Grande de Morelia is located on the MexicanCentral Plateau (Fig. 1). The headwaters are located ina mountainous region known as ‘‘Mil Cumbres’’. Atthe headwaters a dam impounds the stream flow inLoma Caliente Reservoir at an elevation of 2220 m

E. Lopez Lopez, L. Da´valos-Lind / Aquatic Ecosystem Health and Management 1 (1998) 345–351346

Fig. 1. Location of sampling sites on the Rı´o Grande de Morelia river-reservoir system. Headwaters are Site 1, Loma Caliente Reservoir is Site2, Umecuaro Reservoir riverine section is Site 3 and lacustrine zone is Site 4, Rio Grande de Morelia Sites 5, 6 and 8 and Cointzio Reservoirlacustrine zone is Site 7. Loma Caliente, Ume´cuaro and Santiago Undameo settlements are also indicated.

above sea level. The discharge of this reservoir goesdirectly into a second reservoir called Ume´cuaro at anelevation of 2214 m above sea level. Both reservoirsare used as water supply by a few small villages andfor small scale agriculture, cattle-raising and forestry.The vegetation surrounding these reservoirs is aremnant forest ofPinus and ABIES. Down river,11 km from Umecuaro Reservoir the river receivesraw wastewaters from the village of SantiagoUndameo and from a fertilizer factory. In this areathe impact from the factory is substantial whencompared with the rest of the river-reservoir system.Further on, the river is impounded by Cointzio Reser-voir at an elevation of 1860 m above sea level. Thisreservoir supplies water for drinking and industrialuse to the city of Morelia. Downstream from CointzioReservoir the river flows through Morelia, where itreceives the city’s raw sewage. Finally, the riverflows into Lake Cuitzeo. Table 1. shows the morpho-metric and physico-chemical characteristics of theriver-reservoir system.

3. Methods

The Rıo Grande de Morelia was sampled during thesummer of 1993. Surface samples of 1000 ml weretaken from eight river and reservoir sites (Fig. 1).We used Hach spectrophotometric techniques toquantify nitrates (NO3), nitrites (NO2), ammonia

(NH3) and total P. Turbidity was measured asFormazin Turbidity Units with a spectrophotometer(Hach DREL/2000). Suspended solids were measuredby photometric techniques (non-filterable residue,Hach DREL/2000).

For the algal bioassays, we used EPA’S (1971)standardized protocol. Two types of bioassays wereconducted, one using the alga (Selenastrum capricor-nutum), recommended in the EPA protocol and theother using native algae from each site sampled(NAERP and EPA, 1971).

The water samples were filtered through 0.45mm(Gelman) membrane filters. The bioassays wereconducted in 30 ml scriw cap test tubes, a modifica-tion of the EPA protocol introduced by Davalos et al.(1989). The tubes were acid cleaned and rinsed withdeionized water. A ratio of 40:60 of sample and airwas used in the test tube. Three treatments (with fourreplicates) and a control (with three replicates) wereused. The first treatment was an addition of1000mg l21 N (as KNO3); the second treatment of1000mg l21 P (as KH2PO4); and the third treatmentwas the addition of both nutrients (1000mg l21 N andP). The controls consisted of filtered sample waterwith no nutrient additions. This control also servedto determine AGP.

In the bioassay usingSelenastrum capricornutum,1 ml containing 2.5× 103 algal cells ml21 was placedin each treatment test tube and in each control testtube. Incubation was carried out under controlled

E. Lopez Lopez, L. Da´valos-Lind / Aquatic Ecosystem Health and Management 1 (1998) 345–351 347

Table 1Morphometric and physico-chemical characteristics of the Rı´o Grande de Morelia reservoir system in June 1993. FTU� fromazin turbidityunits, na � not available or nor applicable

Characteristics Riverheadwaters

ReservoirLomaCaliente

ReservoirUmecuaroriverine zone

ReservoirUmecuarolacustrine zone

River River ReservoirCointzoilacustrine zone

River

Area (km2) na 0.48 na 1.42 na na 4.86 aMaximum length (km) na 1.26 na 3.5 na na 5 naMaximum width (km) na 0.86 na 1.5 na na 3 naShore line developmentindex (km)

na 2.75 na 10.25 na na 17 na

Maximum depth (m) na 6 na 6 na na 35 naHardness (mg l21 CaCo3) 8 6 4 14 14 27 27 27Total phosphates (mg l21

PO4)0.5 0.77 0.18 0.5 0. 0.28 0.58 0.3

Nitrates (mg l21NO3) 0.7 0.4 0.4 0.3 0.3 5.9 1.7 1Turbidity (FTU) 10 24 54 52 30 170 180 1200Suspended solids (mg l21) 0 0 10 11 24 104 117 285

light conditions following the EPA protocol (NEARPand EPA, l971). Temperature was maintained at 24^

28C.In the bioassay using native phytoplankton, 1 ml of

water was used as the inoculum after filtration througha 0.64mm mesh net to eliminate macrozooplankton.The native algal inoculum was prepared from waterfrom each site and inoculatedin situ into a second setof treatment and control test tubes. Incubation was asfor S. capricornutum. Algal growth changes weremeasured daily as chlorophyll fluorescence using aTurner Design AU-10 fluorometer.

We determined the growth limiting nutrient in twoways: 1) maximum fluorescence in the stationarygrowth phase, and 2) growth rate (K day21) calculatedaccording to the following equation (NEARP andEPA, l971):

K � Ln F2=F1

ÿ �=t2 2 t1

where t equals time at initiation (t1) and whenstationary phase was reached (t2), F2 � fluorescenceat t2 andF1 � fluorescence att1.

Maximum stationary phase fluorescence and K

values of each treatment were submitted to a factorialANOVA analysis to determine significance of thedifference among treatments and controls.

Algal growth potential was determined from thefluorescence of the controls from each site and isdefined according to Couture et al. (1985) as themaximum concentration of algae when the populationreaches stationary phase. For comparative purposeswe report AGP as a percent increase of the populationat stationary phase.

4. Results

4.1. Algal growth potential (AGP)

The AGP measured in the study is shown in Fig. 2.The AGP at all sites was greater forS. capricornutumthan for the native algae owing to species interactionexcept for Site 6 (Fig. 2). The AGP for native algaewas null at the headwaters (Site 1) but not forS.capricornutum(Fig. 2). We found a small decreasinggradient of AGP from Sites 1–5 (Fig. 2). Water from


Fig. 2. Algal growth potential (% growth) of Rı´o Grande de Morelia river-reservoir system water. Sampling sites: 1� headwaters, 2� LomaCaliente Reservoir, 3 and 4� Umeciarp Reservoir, 5� Rıo Grande de Morelia, 6� Rıo Grande de Morelia below fertilizer plant and urbandevelopments, 7� Cointzio Reservoir, 8� Rıo Grande de Morelia.

Site 6 (closest to the fertilizer factory) had the largestAGP of all sites. This high AGP decreased againalong the Cointzo reservoir at Sites 7 and 8.

4.2. Limiting nutrient

Maximum growth measurements and growth ratedata showed the same results. The limiting nutrientvaried along the river-reservoir system and nutrientco-limitation was frequent (Table 2). Both bioassays(S.capricornutumand native algae) consistently gavethe same results, but as expected, the magnitude ofresponse varied owing to species interaction (Table2). At the headwaters (Site 1), P was the limitingnutrient for S. capricornutumand at this site nativealgae showed strong co-limitation, that is, the additionof both nutrients highly stimulated the growth of thealgae, much more than the addition of P along. At thissite native algae showed strong co-limitation, this is,the addition of both nutrients highly stimulated thegrowth of the algae, much more than the addition ofP only. Sites 2 through 5 showed co-limitation forboth S. capricornutumand native algae (Table 2).Site 6 (closest to the fertilizer factory) did not showany nutrient as limitingS. capricornutumgrowth, butdid show co-limitation for the native algae. Sites 7 and8 showed P as the growth limiting nutrient forS.capricornutum but strong colimitation was foundagain for native algae.

5. Discussion

The algae growth promoting potential of the watervaried along the river-reservoir system. It was verysurprising that in the headwaters where anthropogenicimpact is apparently low, NO3 and PO4 were high, 700and 500mg l21, respectively. Also surprising was thatnative algae did not grow (Table 2 and Fig. 2). Never-theless, when P was added, native algae grew and thegrowth was the highest recorded for all sites when Nand P were added.

We found that the reservoirs and their biota areserving as processors of nutrients. The AGP of thewater both forS. capricornutumand native algaediminishes through the reservoir long axis (Fig. 2).Rich waters enter the reservoir and the nutrients aretaken up by organisms. When the water leaves thereservoir the AGP was diminished by half at LomaCaliente – Ume´cuaro and by a fourth at Cointzio. TheAGP at the less impacted reservoirs Loma Calienteand Umecuaro (Sites 2–5) was one third of the mostimpacted areas where the fertilizer factory (Site 6) andthe urban developments are located. This study showsclearly that water from Site 6 entering Cointzio Reser-voir provides major nutrient enrichment. It waspossible to identify Site 6 as the point with majornutrient input according to AGP results and the detec-tion of no nutrient limitation. The AGP of the waterentering the reservoir (Site 6) was nine times largerfor S. capricornutumthan at Site 5 and 51 times larger


Table 2Mean (n� 4) percent maximum growth response and growth rate per day relative to controls (max and K, respectively) ofS.capricornutumandNative algae. N indicates nitrogen addition, P phosphorus addition and NP nitrogen plus phosphorus addition. Same letter indicates nosignificant difference.p , 05

Sites 1 2 3 4 5 6 7 8

S. CapricornutumN (max) 120 a 142 a 136 a 260 a 205 a 114 a 110 a 91 aS. capcricornutumP (max) 184b 61 b 90 b 90 b 118 b 115 a 269b 338bS. CapricornutumNP (max) 183 b 243c 334c 545c 529c 116 a 255 b 323 bNative algae N (max) 40 a 226 a 132 a 310 a 158 a 99 a 343 a 151 aNative algae P (max) 834 b 110 b 8 b 81 b 74 b 112 a 405 a 586 bNative algae NP (max) 15400c 1504c 1100c 8776c 10016c 243b 1414b 1184cS. CapricornutumN (K) 0.11 a 0.04 a 2 0.09 a 0.15 a 0.13 a 0.02 a 0.02 a 2 0.01 aS. CapricornutumP (K) 0.34 b 0.00 b 2 0.05 b 0.02 b 0.03 b 0.03 a 0.24 b 0.23 bS. CapricornutumNP (K) 0.16 b 0.19 c 0.23 c 0.35 c 0.25 c 0.02 a 0.14 b 0.20 bNative algae N (K) 0.00 a 0.07 a 0.05 a 0.07 a 0.14 a 0.02 a 0.12 a 0.07 aNative algae P (K) 0.44 b 0.00 b 2 0.03 b 0.02 b 0.27 b 0.06 a 0.17 a 0.39 bNative algae NP (K) 0.85 c 0.44 c 0.37 c 0.45 c 0.55 c 0.12 b 0.26 b 0.26 c

for the native algae than at Site 5. It should be notedthat the fertilizer factory claims to have no dischargewith a board displayed for the public. Table 2 showsthe large input of nitrates at Site 6; the nutrient back-ground concentration measured along the river-reser-voir sites before the fertilizer factory does not recovereven at Site 7.

The growth limiting nutrient was different fordifferent reaches of this river-reservoir system. Wefound P to be the limiting nutrient in the headwatersof the system (Site 1). The limiting nutrient changeswhen the river is impounded into the Loma Calienteand Umecuaro Reservoirs (Sites 2–5) from P to coli-mitation in these reservoirs. In this area humanactivity is composed of small-scale farming, cattleraising and forestry from inhabitants of two smallvillages. The limiting nutrient changes back to P atSites 7 and 8. These sites are the most impacted byhuman activity, being influenced by a fertilizerfactory and the sewage of two major cities.

In tropical and subtropical regions, N has beenidentified as the limiting nutrient caused by theyoung volcanic terrain where P can be exceptionallyabundant (Carignan and Planas, 1994). Studies byHenry et al., 1984Vincent et al., 1984, Wurtsbaugh,1985, and Davalos et al., 1989 showed N as thelimiting nutrient for algal biomass production intropical lakes. We did not find N to be the limitingnutrient.

It was surprising for us to find P as the limitingnutrient at the headwater riverine component of thesystem forS. capricornutum,whereas the native algaeshowed co-limitation. We found no co-limitation forS. capricornutumand strong co-limitation for thenative algae in most cases. This may be becasusephytoplankton in rivers is not abundant and does notestablish as stable a community as in the reservoir,and may have very specific nutritional requirements.As soon as the water is impounded, the phytoplanktoncommunity develops and the nutrient limitationswitches to co-limitation.

The measured changes in AGP and in nutrientlimitation through the riverine-reservoir systemwhere human impact varies from rural to industrialand urban, show the severity of the impact of diversehuman activities on the health of the wholeecosystem. This study measured a limited functionalresponse of a small component of the biota of the

river-reservoir system. In order to assess the healthof the area and changes owing to the various impacts,other biotic factors need to be studied. Nevertheless,data on functional responses at the base of the foodweb are generally the most revealing. An importantconsideration for proposed further studies is thatcontinuous disturbances are taking place. The RıóGrande de Morelia is an important area of endemismin fish fauna (Miller, 1982). The changes experiencedin the health of this river-reservoir system havealready caused the loss of some native fish species(Dıaz-Pardo et al., 1993). Environmental deteriorationhas left the headwaters as the only refuge for nativeaquatic biota in the entire drainage area (Soto-Galeraet al., in press).

Assuming that the environmental deterioration is atleast a continuing if not growing process, strategies torestore the health of this aquatic ecosystem should beimplemented. The establishment of nutrient-removingwaste water treatment plants for the fertilizer factoryand the village of Undameo should be mandatory.Even if the stress is less at the headwaters, a reforesta-tion program in the upper portion of the basin tocontrol erosion will help to preserve the only refugefor native aquatic biota left in the drainage area.

Acknowledgements

The authors acknowledge the team of biologiststhat collaborated to obtain water samples: Bio´l. Jaque-lina Calderoń from Universidad Michoacana de SanNicolas de Hidalgo., Bio´l. Paul Vallejo de Aquino, M.en C. Eduardo Soto Galera, M. en C. Joel Paulo Mayaand Biol. JoseAngel Serna Hernańdez from EscuelaNacional de Ciencias Biolo´gicas del IPN. We alsothank Dr. Owen Lind for comments on the design,development of bioassays and manuscript, and toBiol. Oscar Polaco and all the referees for their valu-able commentaries on the manuscript.

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