Journal of the American Contol Association, 2O(4):4O1-404,2OO4 Copyright A 2OO4 by the American Association, Inc.

THE USE OF THE MESOCYCLOPSLONGISEZUS AS A BIOLOGICAL CONTROL AGENT FOR IN CALI, COLOMBIA

MARCELA SUAREZ-RUBIOI,z EIIU MARCO E SUAREZ3

ABSTRACT. We present data on the efficacy of longisetus as a biocontrol agent in controlling Aedes aegypti larvae in catch basins in Cali, Colombia. Additionally, we determined some of the features that facilitated the establishment ofthe in catch basins. Between June 1999 and February 200O,201 catch basins were treated with an average of 500 adult copepods. The copepods had established in 49.2Vo of all the basins and they maintained Ae. aegypti larvae at low densities until the end of the 8-month study. The corrected efficacy percent was 9O.44o. The copepods established in basins located in a flat area as opposed to those in steep areas, exposed to sunlight and with 0-7ovo of floating organic matter. when the catch basins were con- taminated with synthetic washing agents, like detergents, the copepods did not survive. The copepod M. lon- gisetus cottld be incorporated as a biological control agent in an integrated Ae. aegypti control program.

KEY WORDS Biological control, Aedes aegypti, copepods, , Colombia

INTRODUCTION vae (Su6rez et al. 1991) and can suppress larval populations to very low levels (Marten et al. 1994). Many strategies, such as vector eradication pro- Some cyclopoid copepods have been observed grams, chemical control measures, environmental to control Ae. aegypti larvae in domestic and peri- sanitation with community participation, and bio- domestic containers (Marten 1990, Marten logical control agents, have been used to prevent or et al. 1994, Schreiber et al. 1996). For control dengue outbreaks. An early approach to example, Macro- cyclops albidus (Cocal), Mesocyclops longisetus control and was to erad- (Thi6baund), and Mesocyclops (Da- icate the dengue vector, Aedes aegyprl (L.). The aspericornis day) are highly effective in controlling eradication programs failed due to financial, polit- Aedes al- bopictus (Skuse) (Marten l99O) and Ae. aegyptilar- ical, technical, and administrative problems. As a vae in abandoned tires (Su6rez 1991. consequence, dengue vector populations returned to et al. Marten et al. 1993, Marten et al. 1994) and in other previous or higher densities, and the mosquito dis- artifi- cial habitats (e.g., persed into new areas (Halstead 1984, Nelson 1986, barrels, water tanks). In other studies, M. aspericornis eliminated Clark 1995, Gratz and Knudsen 1995, PAHO 1995, 99Vo of the lst- stage larvae of Ae. aegypti and Aedes polynesiensis Gubler and Clark 1996). Chemical control was also (Marks) in artificial habitats (Riviere used to eliminate the vector, but the reduction of et al. 1987; Brown et al. l99ta,1991b). In addition, female mosquitoes was transitory (Leontsini et al. copepods may help in controlling mosquito 1993), and the mosquito developed resistance to larvae in natural habitats (e.g., (Lardeaux many insecticides (PAHO 1995). Environmental tree holes, crab holes) 1992). Mesocyclops sanitation with community participation involved aspericornis reduced larval populations of Ae. polynesiensis andAe. the elimination of all artificial breeding sites of the aegyptiby 9l-99%o (Riviere vector. Community-based approaches provide et al. 1987), demonstrating that the copepods have potential short-term success, but this control strategy has yet a value as a biological control agent (Kay to show a significant impact on vector populations et al. 1992). In Cali, Colombia, the principal in operational levels (Gubler and Clark 1996). Bi- breeding sites of Ae. aegypti (Fig. ological control is a safe strategy and does not have are catch basins 1), located along streets in residential some of the negative environmental drawbacks of areas. Catch basins have been primary insecticides (Howarth 1991). The main drawbacks described as a breeding site in Cali because Ae. aegypti larvae were present of biological control are that the predator has to in 57.2Vo of the city's basins (Gonz6lez colonize the habitats of its prey in urban environ- et al. 1993). Due to their design, the basins ments, there is a lag time for its establishment contain water throughout the year, thus providing appropriate (Rawlins et al. 1997), and repeated applications are conditions for the needed for effective control (Laird 1981). development of mosquito larvae. Additionally, Copepods attack and kill lst-stage mosquito lar- catch basins in Cali produce 27 times more pupae than all other breeding sites combined within a city block (GonzLlez and Sudrez, unpublished data), rUniversidad del Valle, Departamento de Biologfa, suggesting that catch basins are mosquito factories. Apartado 2536O, Cali, Colombia. The characteristics 2 Present address: Institute For Tropical Ecosvstem of catch basins make it dif- Studies, University of Puerto Rico, PO-Box 2334i, San ficult to incorporate traditional control strategies Juan, Puerto Rico 00931-3341. that are effective in other places. For example, 3 Secretaria de Salud Municipal, Calle 48 36-00, Cali, chemical control, especially liquid or granular for- Colombia. mulations, require continued applications to control

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determine the proportion of basins with and without Wets €ntEn6 copepods IJ and Ae. aegypti larvae. In each visit, 4 ::.:...f}---.., samples of water were taken from each basin using a2OO-cm3 ladle. The samples were taken from each corner of the basin at 5-min intervals to allow the larvae and the copepods to return to the organic matter, and each sample was considered individu- ally for further analysis. In addition, to identify the characteristics correlated with copepod establish- 2/ Exit tub€ ment in the basins, we evaluated 3 physical features of catch basins (i.e., percentage shade, percentage cover of floating organic mattel and the topography around the basin). The topography was Fig. 1. Schematicdrawing of catch basins.The basins measured constantly contain standing water, and they have various as a function of the inclination of the road where amounts of organic matter (OM). the basin was located. Finally, the presence of any synthetic washing agent, such as detergent, inside catch basins was recorded. mosquito populations because its effectiveness lasts for 8-15 days. Slow-release formulations are more Statistical analysis expensive and the government does not support Chi-square tests were performed to evaluate dif- them, and they may cause environmental damage ferences in presence of Ae. aegypti larvae and co- (Schreiber et al. 1996). Additionally, Ae. aegypti pepods in catch basins over time. The corrected ef- populations in Cali are resistant to temephos, one ficacy percent was calculated using the Henderson- of the most commonly used larvicide for their con- Tilton's formula (Henderson and Tilton 1955). A trol (Gonziilez et al. 1993). Thus, it became nec- principal component analysis (PCA) was performed essary to evaluate biological control as an alterna- to analyze the effect of the percent shade, percent tive to the chemical control of Ae. aegypti. cover of organic matter, and the topography on co- This article presents data on the efficacy of M. pepods establishment. Afterward, multiresponse longisetus as a biocontrol agent in controlling Ae. permutation procedures (MRPP) were used to de- aegypti larvae in catch basins and the appropriate termine if there were statistical differences between characteristics of catch basins for its establishment. the groups in the PCA. Moreover, an association We hypothesized that, after the introduction of M. test was used to determine differences in copepod longisetus, the proportion of catch basins with Ae. presence between catch basins with and without de- aegypti larvae would be reduced, whereas the pro- tergent. The level of significance for all tests was portion of catch basins with copepods would in- 0.05. crease.

RESULTS MATERIALS AND METHODS The number of catch basins infested with Ae. ae- Study site gypti larvae significantly decreased after copepods We evaluated 2Ol catch basins along streets in were introduced (1' -- 36.48, df : 1, P < 0.000), residential areas in southern sectors of Cali but the number of catch basins with copepods did (3"27'N, 76"32'W), Colombia (elevation 995 m). not differ through time (1' : 0.3368, df = 1, P : The annual precipitation is 1,153 mm, with rainy 0.562). The number of catch basins with larvae de- peaks in April and October, and dry peaks in July creased significantly over time, but the basins and December, and the average temperature is 23"C where copepods established remained positive for (rGAC 1980). copepods (49.2Vo), and in these basins, the cope- pods were present for 8 months (Fig. 2). The cor- rected efficacy percent was 9O.47o and the mean of Field surveys larvae density per sample was 0.53. Between June 1999 and February 2OOO, 2Ol From the 201 catch basins, 68Vo were located in catch basins were evaluated on a monthly basis for places in which they were exposed to sunlight, 877o presence and absence of M. longisetus and Ae. ae- were located in roads with low slope, and 9l%o re- gypti larvae and 70 catch basins were used as con- ceived less than 5O7o of floating organic matter trols. In June, we sampled the Ae. aegypti larvae in (Fig. 3). The amount of shade and the nature of the catch basins to define the proportion of infested local topography were the most important features basins. In July, we introduced approximately 500 contributing to the establishment of copepod pop- adult copepods per basin. The copepods were taken ulations. In the PCA, axis I was correlated with from laboratory culture and counted using a plastic shade and topography, explaining 48.67o of thevar- dropper. In August, an 8-month survey began to iance with an eigenvalue of 1.458; while axis 2 was DECEMBER2004 Copepoos As A BIoLocIcAL CoNTRoL AGENT 403

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suppress Ae. aegypti larvae. Our results are in agreement with those of Rawlins et al. (1997), who found that, in Trinidad, M. longisetus was effective s as a biological control tool against Ae. aegypti lar- ;60 .: vae because the copepods almost completely sup- gro pressed the larvae in drums and tires. However, M. .E 40 longisetus showed low predatory capacity against o (Say) o because M. longisetus c30 g is benthic and preys on bottom-feeding Ae. aegypti o oo20 but have little effect in reducing surface-feeding Cx. quinquefasciatus populations (Riviere et al. 10 1987). In contrast, Kay et al. (1992) found that larg- 0 er individuals of M. longisetus were effective pred- Juna October February ators of both Ae. aegypti and Cx. quinquefasciatus Monthof suiloy larvae in the field. The number of catch basins positive for M. lon- Fig. 2. (a) Proportion of catch basins infested with gisetus in Cali did not increase throughout time. It Aedes aegypti larvae before and after the introduction of successfully got established in 49.2Vo the copepod Mesocyclops longisetus, and (b) proportion of the catch of catch basinswith establishedM. longisetuspopulations. basins and maintained its numbers during the study period (8 months) in these basins. The survival of copepods in catch basins is consistent with results correlated with organic matter, explaining 32.6Vo of of studies in Honduras. in which 79Vo of the sam- the variance with an eigenvalue of O.977. The co- pled drums had copepods 7 months after their in- pepods did not establish in catch basins located in troduction (Marten et al. 1994). The difference in areas with steep slopes and under complete shade. the magnitude of copepods established in Honduras However, they did establish in basins located in flat and Cali could be due to the characteristics of the areas, exposed to sunlight, and with O-l}Vo of or- artificial containers. Drums are used to store water ganic matter. The MRPP found significant differ- and are exposed to continuous sunlight, while catch ences between these two groups (T = -12.38, P : basins may be in shaded areas but may flush during 0.000). Nevertheless, copepods were absent from rains if they are located in areas with steep slopes. 2OVoof catch basins with suitable characteristics if Because the copepods feed on algae and protozoa detergent that came from drainage of the neighbor- (Sudrez et al. 1992), the sunlight allows the fast hood was present (Yate's corrected X, : 8.83, P : growth of these photosynthetic groups. It was noted 0.003). that the copepods do not tolerate water contami- nated with detergents, possibly due to the toxic ac- tion that kills the copepods but did not kill the DISCUSSION Ae. aegypti larvae. Thus, copepods should not be used After 8 months, the percent of catch basins in- in catch basins where the drainage may be contam- fested with Ae. aegypti larvae dropped from 7l to inated with any synthetic washing agent. 3Vo. These data demonstrate the well-known pred- The copepod M. longisetus is a good candidate atory activity of the copepods (Marten et al. 1994). as a biological control agent because it serves as a As long as the copepod populations are established sustainable control strategy. It suppresses Ae. ae- and prevailing in the catch basins, they are able to gypti popnlations and tolerates organophosphate in- 404 JounNar-op rHB AlrsnrcAN Moseurro CoNrnol Assocrarutr,r Vor-. 20, No. 4 secticides used in mosquito control (Tietze et al. Henderson CE Tilton EW. 1955. Tests with acaricides 1994, Rawlins et al. 1997). Additionally, M. lon- against the brown wheat mite. J Econ Entomol 48:157- gisetus thrives in the presence of Bacillus thurin- 161. giensis van israeliensis (Bti), a commonly used mi- Howarth FG. 1991. Environmental impacts of classical biological control. Annu crobiological insecticide (Riviere et al. 1987, Rev Entornol 36:485-509. IGAC. 1980. Diccionario Geogrdfico de Colombia. Tomo Marten et al. 1993). Thus, application of Bti to the L Segunda Edici6n. BogotS, Colombia: Instituto Geo- habitat of this copepod does not impact copepods grdfi co Agustin Codazzi. adversely. However, there are some disadvantages Kay BH, Cabral CP, Sleigh AC, Brown MD, Ribeiro ZM, in using M. longisetus as a biological control agent. Vasconcelos AW. 1992. Laboratory evaluation of Bra- One major disadvantage is that, after introduction, zilian Mesocyclops (Copepoda: ) for mos- the predatory pressure on mosquito larvae lags until quito control. J Med Entomol 29:599-6O2. the copepods reach high population densities (Tiet- Laird M. 1981. Biocontrol of medical and veterinary ze et al. 1994). In addition, they cannot survive Pests. New York: Praeger. Lardeux F. 1992. Biological control of Culicidae with desiccation (Riviere et al. 1987) and detergents. the copepod and larvivorous fish Hence, the disadvantages may be solved if cope- (Poeciliidae) in a village of French Polynesia. J Med pods are used along with a safe microbial control Entomol 6:9-15. agent and in catch basins without inflow of deter- Leontsini E, Gil E, Kendall C, Clark GG. 1993. Effect of gents. Because of their short life cycle, their high a community-based Aedes aegypti control program on fecundity, their protozoa-based diet, and their tol- mosquito larval production sites in El Progreso, Hon- erance to Bti, M. longisetus could be incorporated dtt:l:as.Trans Roy Soc Trop Med Hyg 87:267-271. as a biological control agent in an integrated control Marten GG. 1990. Evaluation of cyclopoid copepods for Aedes albopiclus control in tires. J Am Mosq program of Ae. aegypti. Control Assoc 6:681-693. Marten GG, Borjas G, Cush M, Ferndndez E, Reid J. 1994. Control of larval Aedes aegypti (Diptera: Culic- ACKNOWLEDGMENTS idae) by cyclopoid copepods in peridomestic breeding We thank Adolfo Le6n Montoya and Orlando containers. J Med Entomol 3l:36-44. Marten GG, Che Bordes ES. 1993. Compatibility of Perafdn for their assistance in the field, Martin Sa- W cyclopoid copepods with mosquito insecticides. "/ Az tizabal for his recommendations in the sampling Mosq Control Assoc 9:150-154. method, Maria Fernanda Barberena for her assis- Nelson MJ (Pan American Health Organization). 1986. tance in statistical analyses, Gary G. Clark, Mitch Aedes aegypti.' Biology and ecology. Washington, DC: Aide and Carolina Monmany for their comments PAHO PNSP/86_64. on the manuscript, and Maria Eugenia Cuadros and PAHO [Pan American Health Organization]. t995. Den- Ranulfo Gonzdlez for their support and advice. 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