2. Capítulo Bionomía de Anopheles spp De Puerto Gaitán, Meta Colombia 2009
2.1 Breeding sites of Anopheles spp (Diptera: Culicidae) in the municipality of Puerto
Meta, Colombia
[Paper written under Bulletin of Entomological Research guidelines]
Sebastián Durán
1, Luz Stella Buitrago
2, Jan E. Conn
3,4, Helena
Brochero
1*1
Facultad de Agronomía, Universidad Nacional de Colombia, Bogotá DC, Colombia 2
Unidad de Entomología, Secretaría Departamental de Salud del Meta, Villavicencio, Meta, Colombia
3
Griffin Laboratory, Wadsworth Center, New York State Department of Health, Slingerlands, NY, 12159 USA
4
Department of Biomedical Sciences, School of Public Health, State University of New York, Albany, NY, 12222 USA
*Corresponding author
RUNNING TITLE
Mal. vector breeding sites on Puerto Gaitán
ABSTRACT
Puerto Gaitán (Meta, Colombia), the 4th largest Colombian municipality, is located in the Colombian Orinoquia region, a savannah environment experiencing continuous land use change (Ramírez, 2012). During 2009, one fatal case of malaria was reported in this locality and it was hypothesized that this may have been due to urban transmission (Buitrago, Brochero, McKeon, Lainhart, & Conn, 2013). This work aimed to identify the Anopheles spp. in local (urban and periurban) breeding sites to provide baseline information on the local biology and ecology of immature stages. This study recorded five Anopheles species; An.
30 Determinación del papel en la transmisión de malaria de las especies de Anopheles
rangeli and An. albitarsis sl. The immature stages were distributed in 19 of 27
inspected breeding sites and both artificial and natural breeding sites were positive and included fishponds (44,4%, n=12), small streams (7,4%, n=2), and drainage ditches (7,4%, n=2). The remaining positive breeding site types (“Morichera”, river overflow, river margin) accounted for 3,7% each. Negative breeding site types (29,6%, n=8) were small stream (3,7%, n=1), fishponds (14,8%, n=4), flooded pastures (7,4%, n=2) and flooded forest (3,7% ,n=1). Factors such as pisciculture, cyclic flooding and distance of the breeding sites to human dwellings are likely to affect malaria transmission risk.
Keywords Anopheles immature breeding-sites biology ecology Colombia
Orinoquia
INTRODUCTION
Puerto Gaitán (149m), in the Orinoquia region, is the fourth largest municipality in Colombia (17424km2). It is located in a savannah environment, which has been transformed by urban growth. Is located on the Manacacías river Watershed (region), which periodically floods some local urban and periurban areas (Cicery, Gualtero, & Cicery, 2005). The average temperature is 26°C, annual rainfall is 2200mm, and there is a monomodal rainfall distribution (rainy season from March to November).
Economical activities range from livestock, agriculture, trade, oil exploitation, tourism, to fishing (artisanal). In the 2005 census there were 15475 inhabitants, with 37,7% self reporting as indigenous (DANE, 2005). Although, local government authorities argue that the population might be more above 31139 inhabitants, and native population could represent a bigger portion (43,21%). Factors such as internal armed conflict and increasing oil exploitation activities have resulted in unexpected immigration and local socioeconomic problems commonly associated with such events (Buitrago, Brochero, McKeon, Lainhart, & Conn, 2013).
An. darlingi is reported to be the main vector in Meta´s Department (Ahumada M. ,
2009; Olano, Brochero, Sáenz, Quiñones, & Molina, 2001; Ahumada, Pareja, Buitrago, & Quiñones, 2013), nevertheless, An. marajoara of the Albitarsis Complex has been hypothesized to be involved in local transmission in Villavicencio (Brochero, Rey, Buitrago, & Olano, 2005), Meta´s capital city. There is evidence of urban malaria transmission, and Plasmodium vivax is the prevalent parasite species (Buitrago, Brochero, McKeon, Lainhart, & Conn, 2013).
Human intervention in the environment plays an important role in malaria vector ecology, particularly in the generation of new breeding sites able to be colonized by anophelines (Conn, et al., 2002; Gil, Tada, Katsuragawa, Ribolla, & da Silva, 2007). Such breeding sites could interact with cyclic floods (Ahern, Kovats, Wilkinson, Few, & Matthies, 2005; Osorio S. , 2011) to alter species
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dynamics such as relative abundances, composition, seasonality and malaria transmission (Conn, et al., 2002; Gil, Tada, Katsuragawa, Ribolla, & da Silva, 2007). The study objectives were 1) to identify the Anopheles spp. and their breeding sites in Puerto Gaitán; and 2) provide fundamental data on the biology and the ecology of Anopheles spp. immature stages in this locality. Additionally, this study seeks to provide data to support decision-making related to local control strategies of the immature forms.
MATERIALS AND METHODS
Study area
This study was performed in 2009 (March-May, July-August, October, December), in urban and periurban areas of Puerto Gaitán-Meta, Colombia (04°19′N, 72°05′W).
Figura 2.1-1. Puerto Gaitán (Meta, Colombia) (taken from Wikimedia Commons)
Immature collection and breeding site characterization
A standard dipper method was used to collect Anopheles spp. immatures (Service M. , 1993; WHO, 1975) in the breeding sites previously reported by local public health authorities, each site was completely covered taking 20 dives every 5 m. Breeding sites were classified into 21 main categories, as seen on Tabla 2.1-1.
32 Determinación del papel en la transmisión de malaria de las especies de Anopheles
Collections were carried out monthly, and alive collected individuals were transported to the Laboratorio de Genética de Insectos insectary at Universidad Nacional de Colombia (Bogotá DC, Colombia- temperature from 24°C-27°C, relative humidity from 65% -72%) or to the Unidad de Entomología (Laboratorio de Salud Pública, Secretaría Departamental de Salud del Meta, Villavicencio, Meta, Colombia), and were fed with ground dog food to obtain adults from entomological series (Belkin, Schick, Galindo, & Aitken, 1967; WHO, 1975) for taxonomic identification. Adult individuals were coded and individually stored in 1.5ml eppendorf tubes; which were tagged, grouped and maintained in bags with silica gel desiccant in a -20°C freezer. Dead immature forms, exuviae of fourth instar larvae and pupae were stored in individually labeled plastic vials with 70%V/V ethanol.
Categories of the Breeding Sites
1. Lake, lagoon 8. Tree hole 15. Fallen fruit 2. Fishpond 9. Rock hole 16. Stem internode 3. River margin 10. Small stream 17. Water sump 4. Flooded pasture 11. Animal foot-print 18. Swamp 5. Flooded forest. 12. Car tire-print 19. Puddle
6. Drainage ditch 13. Crab hole 20. Artificial reservoir 7. Well, tank 14. Ricefield 21. “Jagüey”
22. Others (p.ej. “Morichera”, River overflow)
Tabla 2.1-1. Established breeding site categories for Puerto Gaitán (Meta, Colombia), 2009.
Georeferencing was carried out using a Garmin eTrex® GPS device (WGS84 coordinate system, degrees and minutes with three decimal positions). Whenever possible, variables such as estimated area [width(m) X length(m); then categorized depending upon the whole dataset], sunlight exposure (full sunlight exposure, partial and total shade), presence of some associated vegetation types (emerging, surrounding and/or floating vegetation), water movement (fast, slow and null stream), water turbidity (clear, turbid), presence of organic material (WHO, 1975) and general observations.
Taxonomic identification
Taxonomic identification was conducted with dichotomous keys and based on characteristics of the immature stages and general adult morphology (Faran M. , 1980; Faran & Linthicum, 1981; González & Carrejo, 2009; Rubio-Palis, 2000).
Ecological and statistical analyses
The breeding sites localization map was overlaid onto a thematic ecosystem map of the Orinoquia region [IaVH, WMS service; (Romero, Galindo, Otero, & Armenteras, 2004)], and the data generated were added to each breeding site characterization and considered for further analyses. Interactions between
Capítulo 2 33
and visweb found on the R (R Development Core Team, 2011) package bipartite (Dormann et.al. 2009).
Correspondence Analysis (CA) was carried out to evaluate the association between Anopheles species and the breeding sites and types of positive breeding sites. Results were interpreted using the different environmental variables and observations taken for this study, i.e., indirect gradient analysis (ter Brak, 1986). CAs were obtained with vegan (Oksanen, et al., 2011) for the R software environment for statistical computing and graphics (R Development Core Team, 2011).
RESULTS
Immature collections
Five species were recorded: An. braziliensis, An. darlingi, An. albitarsis sl., An.
oswaldoi sl. and An. rangeli. Individuals that did not develop to the 4th instar or those that were damaged could not been identified to species level (data not shown).
Figura 2.1-2. Seasonality of Anopheles spp. immature individuals in Puerto Gaitán (Meta, Colombia)- 2009. N=178
34 Determinación del papel en la transmisión de malaria de las especies de Anopheles
Two breeding sites were not geo-referenced, a flooded pasture type and a river margin type. Small streams (n=3) were permanent water bodies, sizes from 2000- 5000m2, partial shade, with slow water movement and always with circundant vegetation but a variable composition of the other two defined types of vegetation (floating and emerging).
Figura 2.1-3. Anopheles spp. breeding sites on Puerto Gaitán (Meta, Colombia)-2009 (IDEAM, WMS services)
Fishponds (n=16) were devoted to raising “cachama” (Colossoma macropomum); were permanent (n=10) and temporal (n=6) breeding sites, with sizes between 18m2 and 800m2, ranging from shallow (water level=5cm) to 150cm, with total or partial sunlight exposure, clear or turbid water, always stagnant, and with different combinations of vegetation types.
Morichera breeding site type (n=1, natural breeding site) was in a flooded area
dominated by the presence of “moriche” palm-trees (Mauritia flexuosa). Its area was about 200m2, it was temporal with full sunlight exposure, clear water, surrounding and emergent vegetation, and with a sandy texture on the bottom. On one sampling occasion it was found polluted with sewage water because it was connected to the rainwater drainage system.
River overflow (n=1) was located on the opposite margin of the Manacacías river. The georeference information for this breeding site was taken from previous studies carried out by local health authorities and was not fully sampled during
Capítulo 2 35
2009 due to safety issues. Flooded pasture (n=2) were temporal flooded grassland patches, with a combination of partial shade and full sunlight exposure, clear and turbid water, with a slow current and stagnant water, circundant vegetation, and muddy bottom material. One pasture was reported with human fecal contamination.
Flooded forest (n=1), area above 100m2, was temporal, with all three vegetation types. It was completely dry in the December collection. Drainage ditches (n=2), were temporal narrow water strips, with full sunlight exposure, slow water movement, three vegetation types and with sandy and muddy bottom textures.
Ecological and statistical analyses
CA was carried out to assess Anopheles spp. association with breeding sites (dimensions accounted for a cumulative percentage of variance of 70,6%) (data not shown), after performing clustering (Fig2.1-4) of the breeding sites based on
Anopheles spp. distribution. As most of the fishpond sites clustered together, an
additional CA was carried out.
Figura 2.1- 4. Breeding site clustering after CA. *temporal breeding site type; DrainD=Drainage ditch, SmStr=Small stream, FishP=Fishpond, RivOF=Rivero overflow, Mori=”Morichera”, RivMa=River Margin. (Breeding sites with abundances equal to 1 Anopheles spp. individual were not taken in
36 Determinación del papel en la transmisión de malaria de las especies de Anopheles
For the new CA, each one of the aforementioned breeding sites types (Tabla 2.1-1) was subdivided into permanent and temporal, thus doubling the amount of types of breeding sites. This was done in order to assess the association between
Anopheles spp. and types of breeding sites taking in count their persistence thru
time as an important factor. CA factor map plotting dimensions accounted for a cumulative percentage of variance of 83.63%.
Figura 2.1- 5. Correspondence Analyses factor map; Anopheles spp. VS breeding sites types.*temporal breeding site type; DrainD=Drainage ditch, SmStr=Small stream, FishP=Fishpond,
RivOF=Rivero overflow, Mori=”Morichera”, RivMa=River Margin (Breeding sites with abundances equal to 1 individual were not taken in count)
According to ecosystem map [IaVH, 2013 WMS service; (Romero, Galindo, Otero, & Armenteras, 2004)] overlaying breeding sites (BS) were distributed among five ecosystems; Urban zones-infrastructure (3 small stream BS´s, 13 fishpond BS´s, 1 drainage ditch BS), Water-body (1 flooded forest BS), Overflow savannah on aeolian-influenced floodplain (3 fishpond BS´s, 1 drainage ditch BS), Dune savannah on aeolian-influenced floodplain (1 flooded pasture BS, 1 “Morichera” BS); a river overflow BS was found between the transition of Overflow savannah
-1 0 1 2 -1 .0 -0 .5 0 .0 0 .5 1 .0 1 .5 2 .0
CA factor map
Dim 1 (57.86%) D im 2 ( 2 5 .7 7 % ) SmStr FishP FishP* Mori* RivOf* RivMa* DrainD* An_bra An_dar An_alb An_osw An_ranCapítulo 2 37
on aeolian-influenced floodplain ecosystem and Medium (altitude) dense forest on flooding plane of aeolian-influenced floodplain ecosystem.
Figura 2.1- 6. Interaction network between Anopheles spp. and their breeding sites, Puerto Gaitán- Meta (Colombia, 2009) .*temporal breeding site type; DrainD=Drainage ditch, SmStr=Small stream, FishP=Fishpond, RivOF=Rivero overflow, Mori=”Morichera”, RivMa=River Margin. Thickness of the
38 Determinación del papel en la transmisión de malaria de las especies de Anopheles
connecting line represents connected Anopheles spp. abundance to a specific breeding site, breeding site box thickness represents the amount of all Anopheles spp. found there, and species box thickness represents a species total abundance (Breeding sites with abundances equal to 1 individual
were not taken in count, n=17)
DISCUSSION
Observed presence and abundance of An. darlingi and the presence of An.
albitarsis sl support the ecoregion sabana-subregion llanos classification of Puerto
Gaitán (Rubio-Palis & Zimmerman, 1997). As observed in the clustering (Figura 2.1- 4), fishponds seem to provide a very suitable environment (water quality, sunlight conditions, temperature, pH) and may provide certain resources (food, shelter) for An. darlingi immature stages. Although this was not fully evaluated, the CA interpretation (Figura 2.1- 5) is suggestive, and should be considered in studies to evaluate environmental and ecological factors using a finer resolution than the present study. This study is important because An. darlingi has not usually been reported occupying this type of breeding sites (Sinka, et al., 2010) and only few examples of this exist (Montoya-Lerma, et al., 2011; Vittor, et al., 2009). As these breeding sites were located in highly anthropic environments, this species seems to demonstrate adaptation to local land-use changes (Montoya-Lerma, et al., 2011). An. darlingi high abundances in these breeding sites were associated with the rainy season (Rubio-Palis, Menare, Quinto, Magris, & Amarista, 2005), although there were not enough sampled months in the dry season during this study (March and December) to fully evaluate this. In addition, An. darlingi abundance drops in August for no apparent reason.
Cyclic flooding represents an additional risk because it provides new breeding sites and increases the sizes of the existing ones. This could be assessed more completely by additional breeding site analysis in Puerto Gaitán to fully place this assertion. Remote sensing has also been a useful tool for detecting potential new breeding sites (Mushinzimana, et al., 2006) and could also serve to evaluate future land use changes in this Orinoquia locality.
Distance of An. darlingi positive breeding sites to human habitation increases risk (Barros, Honorio, & Arruda, 2010) and in our study neighborhoods with high malaria incidence (local malaria hotspots) such as Trampolín and Manacacías accounted for most of the fishponds. Although, as An. darlingi flight range is estimated at 2-7 km(Charlwood & Alecrim, 1989) these fishponds may also be a source of this human-biting species for the whole urban area, and represent an important factor in local malaria transmission dynamics and epidemiology (Buitrago, Brochero, McKeon, Lainhart, & Conn, 2013).
Entomological control strategies focusing on immature stages, such as biolarvicide application (Fillinger & Lindsay, 2011; Fillinger, Sonye, Killeen, Knols, & Becker, 2004) and weeding should considered as a local strategy for small and medium sized artificial sites like fishponds (Brochero, Rey, Buitrago, & Olano, 2005). We also suggest carrying out a study to project the construction of engineering works to mitigate cyclic flooding occurrence nearby human dwellings.
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ACKNOWLEDGEMENTS
Secretaría Seccional de Salud del Meta (Colombia), Secretaria de Salud de Puerto Gaitán - Meta (Colombia), Humberto Mosquera, Jimena Molano, Luz Stella Buitrago and Puerto Gaitán community. Funding sources; Universidad Nacional de Colombia (QuiPu 201010012197), USA NIH AI 2R01AIO54139.
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