Figures
Abstract
The Colombian Pacific region is second nationally in number of malaria cases reported. This zone presents great ecological heterogeneity and Anopheles species diversity. However, little is known about the current spatial and temporal distribution of vector species. This study, conducted in three ecologically different localities of the Pacific region, aimed to evaluate the composition and distribution of Anopheles species and characterize transmission intensity. A total of 4,016 Anopheles mosquitoes were collected representing seven species. The composition and dominant species differed in each locality. Three species were infected with malaria parasites: Anopheles darlingi and An. calderoni were infected with Plasmodium falciparum and An. nuneztovari with Plasmodium vivax VK210 and VK247. Annual EIRs varied from 3.5–7.2 infective bites per year. These results confirm the importance of the primary vector An. nuneztovari in areas disturbed by human interventions, of An. darlingi in deforested margins of humid tropical rainforest and An. albimanus and the suspected vector An. calderoni in areas impacted by urbanization and large-scale palm oil agriculture close to the coast. This constitutes the first report in the Colombia Pacific region of naturally infected An. darlingi, and in Colombia of naturally infected An. calderoni. Further studies should evaluate the epidemiological importance of An. calderoni in the Pacific region.
Citation: Naranjo-Díaz N, Altamiranda M, Luckhart S, Conn JE, Correa MM (2014) Malaria Vectors in Ecologically Heterogeneous Localities of the Colombian Pacific Region. PLoS ONE 9(8): e103769. https://doi.org/10.1371/journal.pone.0103769
Editor: Luciano A. Moreira, Centro de Pesquisas René Rachou, Brazil
Received: April 24, 2014; Accepted: July 3, 2014; Published: August 4, 2014
This is an open-access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.
Data Availability: The authors confirm that all data underlying the findings are fully available without restriction. Mosquito vouchers were deposited in the collection of “Grupo de Microbiología Molecular, Universidad de Antioquia”, and An. calderoni COI sequences were deposited in GenBank (Accesion numbers KF698816–KF698832).
Funding: This work received funding from the National Institutes of Health-United States of America, Grant No. R03-AI076710 to MMC. Additional support was received from Estrategia para la Sostenibilidad de Grupos 2013–2014, Universidad de Antioquia-Grant No. E01719 and Comité para el Desarrollo de la Investigación, UdeA, Grant No. 8700-074. NND received financial support for his doctoral studies from Departamento Administrativo de Ciencia, Tecnología e Innovación-COLCIENCIAS, Convocatoria 511-2010, Colombia. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Competing interests: The authors have declared that no competing interests exist.
Introduction
The Colombian Pacific (PAC) region is second in the number of national malaria cases reported [1], registering an average of 30% of the total cases [2]–[4] with the majority of cases due to infection with Plasmodium falciparum Welch (>60%) [1]. PAC is located in the biogeographic Chocó zone [5] and presents diverse ecological and climate conditions with five natural subregions [6] that are predicted to affect Anopheles vector species distribution and behavior [7].
Three species considered the main Colombian malaria vectors are present in the PAC region [7]–[9], with reports of natural Plasmodium infection for Anopheles (Nyssorhynchus) nuneztovari Gabaldon [10] and Anopheles (Nys.) albimanus Wiedemann [8]. Although Anopheles (Nys.) darlingi is considered the main vector in the Chocó rainforest [11], [12], there have been no published records of naturally infected An. darlingi in this area. Anopheles (Kerteszia) neivai Howard, Dyar & Knab, a species closely associated with forest bromeliads and mangroves [13], is a locally important vector in PAC and has been reported infected with P. falciparum [13] and P. vivax [8], [14]. Existing records also suggest an epidemiological association of Anopheles (Anopheles) punctimacula Dyar & Knab with malaria outbreaks in this area [15], [16]; this species was detected with salivary gland sporozoites of P. vivax in NW Colombia [17]. Various members of the Arribalzagia Series such as An. punctimacula, Anopheles malefactor Dyar & Knab, Anopheles calderoni Wilkerson and Anopheles guarao Anduze & Capdevielle, are characterized by a high degree of isomorphism that complicates their differentiation and accurate identification by morphological characters [18]–[20]. For example, in mosquito studies in PAC, An. punctimacula was the species recorded; however, An. calderoni was recently reported in various PAC localities, as supported by adult and larval diagnostic characters and mosquito barcoding [21]. In Piura Department in neighboring Peru, An. calderoni was infected with P. vivax and considered a locally important vector [22], [23]. Collectively, these observations suggest the need for further evaluation of the identity of these species and their involvement in malaria transmission.
Most vector biology studies in PAC have focused on distributions and parasite infection of the Anopheles species present, but little is known about variation in vector densities and transmission intensities at spatial and temporal scales. Therefore, a longitudinal survey was conducted from April 2009 to June 2010 in three ecologically diverse localities of the epidemiologically important PAC region to evaluate species composition and distributions at spatial and temporal levels and to determine important entomological parameters.
Materials and Methods
Study sites
Adult Anopheles were collected from three ecologically diverse malaria endemic localities of the PAC region (Table 1, Figure 1); they were San Antonio de Padua (SAP), Zacarias (ZAC), and Carbonera and Pindales (CAR-PIN). SAP is in the PAC natural subregion Alluvial Valleys of the Atrato and San Juan rivers which is characterized by tropical forest [6]. The main economic activities in SAP are fishing, small-scale agriculture, especially rice and logging that is displacing the forest borders (Figure 2A–B). ZAC is in the PAC Coastal Plains subregion characterized by flooded plains, lakes and swamps [6] and presents one of the highest levels of precipitation in the world (>5,000 mm annually), with rainfalls occurring most of the year [6]. In ZAC, deforestation for human settlements, river extraction of building materials and pisciculture are the most important activities (Figures 2C–D). CAR and PIN are separated by 9 km, and, like ZAC, are located in the Coastal Plains subregion. Both localities are heavily impacted by anthropic changes. CAR is located approximately 3 km from the coast and is a periurban neighborhood of the municipality of San Andres de Tumaco, where urbanization has replaced wetlands and mangroves (Figure 2E). The first collection for these localities was performed in CAR. However, because of problems of public order, the following three collections were completed in PIN, located approximately 11 km from the coast and characterized by large-scale palm oil agriculture (Figure 2F).
San Antonio de Padua (SAP) in the municipality of Vigía del Fuerte, Antioquia Department; Zacarias (ZAC) in Buenaventura, Valle del Cauca Department and Carbonera-Pindales (CAR-PIN) in San Andres de Tumaco, Nariño Department. Spotted line defines Pacific Region boundaries.
San Antionio de Padua locality (SAP): Larval habitats A) originated from rice cultivation. B) pond in the forest border. Zacarias (ZAC): C) ZAC settlement and nearby river used for material extraction. D) Larval habitat - fishpond. E) Carbonera (CAR): Larval habitat - drainage channel. F) Pindales (PIN): oil-palm agriculture.
Mosquito collection
Each locality was visited four times, on average once every three months, from April 2009 to June 2010. Mosquitoes were collected indoors and outdoors (within ∼10 m of the house) by human landing catch under a protocol and informed consent agreement approved by a University of Antioquia Institutional Review Board (Comité de Bioética, Sede de Investigación Universitaria, CBEIH-SIU, approval number 07-41-082). Collections were conducted by two people in each setting, for five days, from 18∶00 to 24∶00 h and one additional night from 18∶00 to 06∶00 h. Adult mosquitoes were identified using morphology-based keys [24]–[26]. Species presenting difficulties during morphological identification were confirmed by PCR-RFLP-ITS2 [27]–[29] and specimens An. calderoni by COI barcoding [30] using primers by Folmer et al. [31] and standardized conditions [32]. In addition, potential larval habitats within approximately 1 km of adult collection sites were inspected to identify those positive for immature stages and third and fourth instar larvae were identified. Mosquito vouchers were deposited in the collection of “Grupo de Microbiología Molecular, Universidad de Antioquia”, and An. calderoni COI sequences were deposited in GenBank (Accesion numbers KF698816–KF698832).
Detection of natural Plasmodium infection
Natural infections with P. falciparum and P. vivax (VK210 and VK247) were determined by ELISA [8], [33], [34], using pools of heads and thoraces of up to five specimens of the same species. The ELISA was used as a screening test to evaluate a large number of mosquitoes. Positive pools in the ELISA were confirmed by a second ELISA. Nested genus specific PCR using DNA extracted from individual abdomens was performed to confirm ELISA results and to incriminate the individual specimen infected [35].
Data analysis
Infection and entomological risk parameter analyses were described previously [36]. Briefly, the infection rate (IR) was calculated as the percentage of Plasmodium positive mosquitoes out of the total number of mosquitoes analyzed by species and locality. The human biting rate (HBR) was expressed as the average of female Anopheles bites per person per night. The annual EIR for each site corresponds to the number of infective bites that a person may receive in one year. Spearman’s correlation coefficient in the SPSS program, version 18 (SPSS Inc., Chicago, IL) was used to estimate the relationships between rainfall data of the previous month to a collection and mosquito abundance; the monthly rainfall records for each locality were obtained from the Instituto de Hidrología, Meteorología y Estudios Ambientales (IDEAM).
Results
Mosquito abundance
A total of 4,015 Anopheles mosquitoes corresponding to seven species were collected during 467 h of sampling (Table 1). There was not a significant (p>0.05) correlation between mosquito abundance and rainfall for any species. Species composition and dominance varied in each locality. In SAP, An. darlingi dominated (98.6%), and its highest abundance was observed during a period of moderate rainfall (Figure 3A). The main vector, An. nuneztovari was collected in low abundance in two samplings coinciding with the increase of rainfall in this locality (Figure 3B). Anopheles punctimacula was detected in low abundance in two consecutive collections (Table 1), with peak abundance at the beginning of the rainy season (Table 1, Figure 3B). Anopheles costai/forattinii was also collected by human biting but only in the last sampling period. Anopheles-positive larval habitats were rice irrigation channels and small ponds near forest borders, both sustained by water from the Atrato’s river (Figure 2A and 2B, respectively). The species detected in these larval habitats was An. darlingi. In one pond near the forest border, An. malefactor, a species not detected as an adult, co-occurred with An. darlingi.
Localities: San Antonio de Padua (SAP), Zacarias (ZAC), Carbonera-Pindales (CAR-PIN). Left axis: human biting rate (HBR) or number of mosquitoes/person/night, right axis: rainfall expressed in mm. The underlined months indicate collection periods and ★ periods when infected Anopheles were detected.
In ZAC, An. nuneztovari was the dominant species (90%), detected in all collections; peak abundance was at the onset of a rainy period, when it was the only species collected (Table 1, Figure 3C). Anopheles neivai was only collected in this PAC locality, during the three initial samplings, always in low abundance. It was not detected in the fourth collection when there was a decrease in rainfall (average 900 to 300 mm) (Table 1, Figure 3D). The Anopheles-positive larval habitats in ZAC were fishponds (Figure 2D), swamps and temporal flooded ponds and the only larvae collected corresponded to An. nuneztovari; bromeliads were not sampled.
The dominant species in CAR was An. albimanus (97.3%), and in PIN, An. calderoni (87.21%). In the three collections in PIN, An. calderoni always outnumbered An. albimanus and its peak abundance corresponded to the highest rainy period in this locality (Table 1, Figure 3E). In the analyses of species distribution and peak abundance performed independently or together for specimens from CAR and PIN there were no significant differences, therefore these localities were presented as one (CAR-PIN). In CAR, An. albimanus-positive larval habitats were wetland drainage channels near houses (Figure 2E), and in PIN, irrigation channels of oil-palm agriculture (Figure 2F), and water wells where An. albimanus larvae were detected.
The total number of Anopheles and individual species collected did not show normal distributions (All: Kolmogorov-Smirnov Z = 4.3 p<0.001), except for An. neivai (Z = 1.96 p>0.05). In SAP, the number of An. darlingi ranged between 0 and 452 (Mean = 121.24, SD±121.26), An. nuneztovari between 0 and 23 (Mean = 1.62, SD±5.05), and An. punctimacula between 0 and 2 (Mean = 0.2, SD±0.51). In ZAC, An. nuneztovari ranged between 6 and 50 (Mean = 20.17, SD±11.28) and An. neivai between 0 and 3 (Mean = 0.5, SD±0.84). In CAR-PIN, An. calderoni ranged between 0 and 103 (Mean = 20.78, SD±28.67) and An. albimanus between 0 and 30 (Mean = 7.91, SD±9.97).
Biting activity peak
Mosquito biting activity, expressed as the mean proportion of mosquitoes collected per hour and per site, from 18∶00–24∶00 h, recorded for the most abundant species, varied by species (Figure 4). In SAP, An. darlingi showed a slight preference for biting outdoors, mostly after 20 h (61.3%) (Figure 4A), although no significant differences were detected (t = −1.61 p>0.05, n = 44) (not shown); its biting peak was between 19∶00–22∶00 h (Figure 3A). In ZAC, An. nuneztovari showed significant endophagic activity (t = 3.95 p<0.05, n = 23), with a main peak between 21∶00–23∶00 h (Figure 4B). In CAR-PIN, An. calderoni did not show a significant biting preference (t = −1.27 p>0.05, n = 23), the highest biting peak was between 20∶00–24∶00 h (Figure 4C). In contrast, An. albimanus showed a significant preference for biting outdoors (t = −2.4 p<0.05, n = 23), and was more active after sunset with its highest peak from 18∶00–21∶00 h (Fig. 4D).
Biting activity is expressed as percentage of bites per human per hour. A. An. darlingi in San Antonio de Padua (SAP), B. An. nuneztovari in Zacarias (ZAC), C. An. calderoni in Carbonera-Pindales (CAR-PIN), D. An. albimanus in Carbonera-Pindales (CAR-PIN).
Human biting rate (HBR)
During the collection of April 2009 in SAP, An. darlingi exhibited the highest HBR among all species collected in any locality, 89.9 bites/person/night (b/p/n; Table 1). HBRs for An. nuneztovari were less than 10% of the highest HBR for An. darlingi; for example in ZAC this ranged between 2.7 and 8.6 b/p/n; in SAP, HBRs were between 0.2 and 1.3 b/p/n (Table 1). Particularly, An. nuneztovari was not detected in CAR-PIN where An. calderoni and An. albimanus were the only species collected in the four visits; furthermore, these species were exclusive to CAR-PIN. Anopheles calderoni exhibited its highest HBR in April 2010 (14.7 b/p/n), and An. albimanus in June 2009 collection (4.4 b/p/n) (Table 1). In general in PAC, species at low densities exhibited HBRs ≤0.2 b/p/n (Table 1).
Infectivity and entomological inoculation rates
Four infected specimens were detected in the three PAC localities (Table 1). Anopheles darlingi in SAP and An. calderoni in CAR-PIN were infected with P. falciparum (IR = 0.036% and 0.194%, respectively). In ZAC, two An. nuneztovari were infected, one with P. vivax VK247 and the other with P. vivax VK210 (IR = 0.203% each). The annual EIR ranged from 3.5 infective bites per year (CAR-PIN) to 7.2 infective bites per year (ZAC), the highest recorded for this study (Table 1). Transmission intensity peaks or periods where Anopheles mosquitoes were detected infected with Plasmodium usually corresponded to low rainfall periods, except when An. calderoni was detected infected in PIN, during a rainy period (Table 1, Figure 3), that was also the period of its peak abundance.
Discussion
Anopheles species dominance and abundance are influenced by environmental factors such as rainfall, land cover and topography [37]–[39]. In this longitudinal study in ecologically different localities of the malaria epidemiologically important PAC region, the influence of rainfall on Anopheles species abundance and diversity was analyzed and the main ecological characteristics of the localities described. In addition, the occurrence of the three main Colombia malaria vectors, An. darlingi, An. nuneztovari and An. albimanus was confirmed in PAC. However, the distribution and importance of these species as malaria vectors varied among localities.
For San Antonio de Padua locality, in a subregion characterized by tropical forest [6], An. darlingi was the most abundant and dominant species. In this locality An. darlingi-positive breeding sites were flooded, deforested forest margins and terrains used by the villagers for personal rice cultivation. Such ecological conditions and type of larval habitats favor the presence of this important vector [40]–[42]; i.e., in the Peruvian Amazon, An. darlingi adults dominate in deforested environments [43]. In SAP, the highest HBR for An. darlingi was during a moderate rainy period, but it is notable that this locality is characterized by persistent rain throughout the year. The HBR indicates that a person in SAP might receive 2,697 An. darlingi bites in a month, suggesting a higher likelihood of malaria transmission by this important vector. In a NW Colombian endemic area where malaria incidence is twice that of PAC, An. darlingi entomological parameters varied according to the locality and higher HBRs were observed at the beginning or end of rainy periods [36]. Data from this and other studies suggest that rainfall is a key factor affecting An. darlingi presence and dominance. For example, high HBRs have been correlated with low rainfall in Brazil [44], [45] and Venezuela [46], but also with rainy periods in the Amazonian forest of French Guiana [47] and Brazil [48]. These different responses to rainfall regimes combined with An. darlingi-positive larval habitats that had specific ecological conditions, suggest the importance of land cover [43], [49] and probable interaction effects among rainfall, ecological and land cover parameters on abundance and distribution of this main malaria vector in endemic regions of Colombia.
Annual EIR indicated that one person may receive one An. darlingi infective bite every 2.3 months. This EIR was higher than previously detected for this vector in the most important Colombian malaria endemic region, the Urabá Bajo Cauca and Alto Sinú (UCS) [36], indicating that An. darlingi is also important in malaria transmission in impacted habitats near rainforest. The infected An. darlingi specimen was detected when the rains were increasing after very low rainfall, in agreement with reports of high malaria transmission in Colombia, usually related to low or moderate rainfall periods [50]. In SAP, An. darlingi showed a biting peak between 19∶00–22∶00 h with a slight tendency for biting outdoors after 20 h. This behavior would be expected to increase infection risk since residents are active outdoors after work (N. Naranjo, personal communication), increasing human-vector contact [46]–[48].
In contrast, in SAP, the main Colombian vector An. nuneztovari was detected in low abundance, with HBRs ≤1. The highest density for this species was observed at the beginning of a rainy period when larval habitats are more stable compared to during heavy rainfall [51]. In Colombia, An. nuneztovari is characterized as being more tolerant of human impacted zones than An. darlingi, and it was previously detected in higher abundances in localities of the epidemiologically important UCS, NW Colombia [36], [52]. Also in SAP, An. punctimacula, a species of local importance, was present when the rain began to increase and showed even lower HBR values than An. nuneztovari. The presence of An. punctimacula during periods of low rains was also observed in NW Colombian localities [36], and its low abundance may be related to the collection methodology used in relation to its reported zoophilic tendency [53], [54].
In ZAC, in the Coastal Plains subregion of central PAC, An. nuneztovari and An. neivai were the only anthropophilic species detected. This locality is characterized by flooded plains and strongly influenced by deforestation and pisciculture. These human activities provide optimal larval habitats for An. nuneztovari [48], the dominant species during the entire sampling in ZAC. However, An. nuneztovari abundances were lower during the most intense rainfall period, coinciding with a decrease of Anopheles-positive larval habitats. Rainfall in this region is very high, with 6,980 mm annual average [6], which may cause flooding of the breeding sites affecting this species’ abundance. HBRs for An. nuneztovari ranged from 2.7 to 8.6 bites per person per night. Even lower HBRs have been previously reported for this species in Buenaventura municipality in PAC [55] and in other malaria endemic Colombian regions [36], [52], [56]; regardless of these observations, An. nuneztovari has previously been incriminated as a vector [36], [52].
In this study, the highest EIR was registered in ZAC for An. nuneztovari (7.5 infective bites per year), indicating that a person may receive one infective bite every one and a half months. This information, the fact that the two infected An. nuneztovari were detected in the period of its maximum abundance, together with the significant preference for biting indoors, suggest an increase in malaria risk for humans when they are resting at home (21∶00 to 24∶00 h). This information is of importance to reduce exposure and risk, especially inside the house, for example by the use of insecticide-treated mosquito nets, one of the main tools for vector control [57]. Various characteristics in ZAC favor the presence of An. neivai, a local vector in PAC [13], [58]. The presence of this species has been correlated to high humidity, lowland tropical forest and a variety of epiphyte plants that serve as its breeding sites [13]. However, in ZAC An. neivai was detected in low abundance (HBRs <1). It is possible that in-progress deforestation in this locality is reducing or eliminating preferred breeding sites of An. neivai. The low HBRs detected for An. neivai are similar to those previously reported in PAC for disturbed areas, compared with sylvatic conditions [7].
In CAR-PIN, localities impacted by anthropic changes, An. albimanus and An. calderoni were the only anthropophilic species detected. In particular, the dominant species in CAR was An. albimanus whereas in PIN it was An. calderoni. These differences may be related to their preferred larval habitats [7], [59], [60]. In CAR, mangroves and wetland drainage channels provide appropriate breeding sites for An. albimanus. The highest HBR for An. albimanus in CAR may be influenced by the proximity of this locality to the coast. Previous reports also show HBRs for An. albimanus increasing with proximity to the coast because of availability of its preferred larval habitats [7], [55]. Variation in An. albimanus abundance was not related to rainfall; however, HBRs decreased with increasing rainfall, and probably floods affected the stability of larval habitats. In PIN, small irrigation channels of oil-palm agriculture and wells favored the dominance of An. calderoni, previously associated with extensive palm agriculture near the coast [20], [61]. The highest HBR for An. calderoni was observed during the period of heaviest rainfall. Comparable HBRs have been reported for An. calderoni in other PAC localities, also during rainfall periods [62]. In PIN, high human-vector contact occurs from 18∶00 to 24∶00 h, with An. albimanus more active around sunset followed by An. calderoni from 21∶00 h. This specific information should help direct local vector control measures. In PIN, the annual EIR indicated that a person might receive one An. calderoni infective bite every three months. This species was recently confirmed in Buga, a PAC municipality, where it was initially identified as An. punctimacula [21], a species of local importance in some PAC localities [58]. However, An. calderoni has not yet been incriminated as a vector in Colombia, although it is of some importance in Peru [22], [23]. In this study, An. calderoni was detected infected with P. falciparum by the first ELISA and nested PCR but could not be confirmed by the second ELISA [63]. Given the difficulties still encountered in the tests to determine parasite infected mosquitoes, this result is reported here according to recommendation of defining a positive infected mosquito with the result of at least two positive tests [63]. Nevertheless, the status of An. calderoni as a local vector in PAC should be further evaluated. Even though An. albimanus is considered the main malaria vector in coastal PAC [8], [9], [59], it was not detected infected by Plasmodium spp. The predominant exophilic behavior of these two anthropophilic species suggest that application of residual insecticides on the outside walls of houses may be appropriate vector control.
Conclusions
The ecological heterogeneity of the PAC localities was reflected in the variation of Anopheles species dominating in each one. Of concern is the fact that these species are among the main Colombian malaria vectors. Their importance in malaria transmission in different human impacted settings was confirmed. Infected An. darlingi was detected in deforested margins of humid tropical forest and An. nuneztovari in an area affected by various human interventions. The main PAC vector An. albimanus and the suspected vector An. calderoni were in areas impacted by urbanization and large-scale palm oil agriculture. Knowledge of their contribution to local malaria transmission and their spatial/temporal distributions helps to understand the dynamics of malaria transmission in this region and will provide the basis for the design and evaluation of more focused control measures.
Acknowledgments
We are grateful to personnel at the Instituto Departamental de Salud de Nariño for their logistic support during this work; to L. Garcia, I. Zea, J. Marin and A. Yate for mosquito collection and N. Alvárez and L.M. Jaramillo for laboratory technical support.
Author Contributions
Conceived and designed the experiments: SL JEC MMC NND. Performed the experiments: NND MA. Analyzed the data: NND MMC. Contributed to the writing of the manuscript: NND MMC SL JEC. Revised and approved final manuscript: NND MA SL JEC MMC.
References
- 1.
Organización Panamericana de la Salud (2011) Informe de la situación del paludismo en las Americas. Available: http://www.paho.org/hq/index.php?option=com_content&view=article&id=2459%3Areport-on-the-situation-of-malaria-in-the-americas-2008&catid=1617%3Ahsd0107g-malaria-statistics-and-maps&Itemid=2049&lang=es. Accessed 2013 January 23.
- 2.
Instituto Nacional de Salud (2011) Boletín epidemiológico Semanal. Estadísticas del sistema de vigilancia en salud pública- SIVIGILA, Casos totales en la Semana Epidemiológica 52 y acumulados del año, Subdirección de Vigilancia y Control en Salud Pública. 2011. Available: http://www.ins.gov.co/lineas-de-accion/Subdireccion-Vigilancia/sivigila/Estadsticas%20SIVIGILA/2011%20MALARIA%20VIVAX.pdf. Accessed 2013 March 14.
- 3.
Instituto Nacional de Salud (2010) Boletín epidemiológico Semanal. Estadísticas del sistema de vigilancia en salud pública- SIVIGILA, Casos totales en la Semana Epidemiológica 52 y acumulados del año, Subdirección de Vigilancia y Control en Salud Pública. 2010. Available: http://www.ins.gov.co/lineas-de-accion/Subdireccion-Vigilancia/sivigila/Estadsticas%20SIVIGILA/SEMANA%2052%20DE%202010.pdf. Accessed 2013 March 14.
- 4.
Instituto Nacional de Salud (2012) Boletín epidemiológico Semanal. Estadísticas del sistema de vigilancia en salud pública- SIVIGILA, Casos totales en la Semana Epidemiológica 52 y acumulados del año, Subdirección de Vigilancia y Control en Salud Pública. 2012. Available: http://www.ins.gov.co/lineas-de-accion/Subdireccion-Vigilancia/sivigila/Estadsticas%20SIVIGILA/CASOS%20POR%20EVENTO%202012%20FINAL.pdf. Accessed 2013 May 27.
- 5. Morrone JJ (2006) Biogeographic areas and transition zones of Latin America and the Caribbean islands based on panbiogeographic and cladistic analyses of the entomofauna. Annu Rev Entomol 51: 467–494.
- 6.
Instituto Geografico Agustin Codazzi (2002) Atlas de Colombia. 5th ed. Imprenta Nacional de Colombia. 342 p.
- 7. Solarte Y, Hurtado C, Gonzalez R, Alexander B (1996) Man-biting activity of Anopheles (Nyssorhynchus) albimanus and An. (Kerteszia) neivai (Diptera: Culicidae) in the Pacific lowlands of Colombia. Mem Inst Oswaldo Cruz 91: 141–146.
- 8. Gutiérrez LA, Naranjo N, Jaramillo LM, Muskus C, Luckhart S, et al. (2008) Natural infectivity of Anopheles species from the Pacific and Atlantic Regions of Colombia. Acta Trop 107: 99–105.
- 9. Herrera S, Suarez M, Sanchez G, Quiñones M, Herrera M (1987) Uso de la técnica inmunoradiométrica (IRMA) en Anopheles de Colombia para la identificación de esporozoitos de Plasmodium. Colomb Med 18: 2–6.
- 10. Fajardo P, Alzate A (1987) Anopheles nuneztovari como vector de malaria en el bajo Calima, Buenaventura, Colombia. Colomb Med 18: 14–18.
- 11. Suarez MF, Quiñones ML, Palacios JD, Carrillo A (1990) First record of DDT resistance in Anopheles darlingi. J Am Mosq Control Assoc 6: 72–74.
- 12. Ochoa J, Osorio L (2006) Epidemiología de malaria urbana en Quibdó, Chocó. Biomedica 26: 278–285.
- 13. Carvajal H, de Herrera MA, Quintero J, Alzate A, Herrera S (1989) Anopheles neivai: a vector of malaria in the Pacific lowlands of Colombia. Trans R Soc Trop Med Hyg 83: 609.
- 14. Escovar JE, González R, Quiñones ML (2013) Anthropophilic biting behaviour of Anopheles (Kerteszia) neivai Howard, Dyar & Knab associated with Fishermen’s activities in a malaria-endemic area in the Colombian Pacific. Mem Inst Oswaldo Cruz 108: 1057–1064.
- 15. Ferro V (1979) Revisión de los recursos aplicables a la lucha contra el paludismo. Rev ENSP 5: 11–18.
- 16.
Servicio de Erradicación de la Malaria (1957) Plan para la erradicación de la malaria en Colombia: 365 p.
- 17. Huffaker CB, Soto H, REy H (1945) Additional wild-caught Anopheles punctimacula D. and K. Infected with Malaria Plasmodia in Colombia, South America. Am J Epidemiol 42: 107–110.
- 18. Harbach RE (2004) The classification of genus Anopheles (Diptera: Culicidae): a working hypothesis of phylogenetic relationships. Bull Entomol Res 94: 537–553.
- 19. Wilkerson R (1990) Redescriptions of Anopheles punctimacula and An. malefactor. J Med Entomol 27: 225–247.
- 20. Wilkerson R (1991) Anopheles (Anopheles) calderoni n. sp., a malaria vector of the Arribalzagia Series from Peru (Diptera: Culicidae). Mosq Syst 23: 25–38.
- 21. González R, Carrejo N, Wilkerson RC, Alarcon J, Alarcon-Ormasa J, et al. (2010) Confirmation of Anopheles (Anopheles) calderoni Wilkerson, 1991 (Diptera: Culicidae) in Colombia and Ecuador through molecular and morphological correlation with topotypic material. Mem Inst Oswaldo Cruz 105: 1001–1009.
- 22. Calderón G, Fernández R (1995) Especies de la fauna anofelina, su distribución y algunas consideraciones sobre suabundancia e infectividad en Perú. Rev Peru Epidem 8: 5–23.
- 23.
Kroeger A, Alarcón J (1993) Malaria en Ecuador y Peru y estrategias alternativas de control: 316.
- 24. Faran M, Linthicum L (1981) handbook of the Amazonian species of Anopheles (Nyssorhynchus) (Diptera: Culicidae). Mosq Syst 13: 1–81.
- 25.
Gonzalez R, Carrejo N (2007) Introducción al estudio taxonomico de Anopheles de Colombia Claves y notas de distribución. Cali: Universidad del Valle. 260 p.
- 26.
Gonzalez R, Carrejo N (2009) Introducción al estudio taxonómico de Anopheles de Colombia Claves y notas de distribución. 2nd ed. Cali: Universidad del Valle. 260 p.
- 27. Cienfuegos AV, Gómez GF, Córdoba LA, Luckhart S, Conn JE, et al. (2008) Diseño y evaluación de metodologías basadas en PCR–RFLP de ITS2 para la identificación molecular de mosquitos Anopheles spp. (Diptera: Culicidae) de la Costa Pacífica de Colombia. Rev Biomed 19: 35–44.
- 28. Cienfuegos AV, Rosero DA, Naranjo N, Luckhart S, Conn JE, et al. (2011) Evaluation of a PCR-RFLP-ITS2 assay for discrimination of Anopheles species in northern and western Colombia. Acta Trop 118: 128–135.
- 29. Zapata MA, Cienfuegos AV, Quirós OI, Quiñones ML, Luckhart S, et al. (2007) Discrimination of seven Anopheles species from San Pedro de Uraba, Antioquia, Colombia, by polymerase chain reaction-restriction fragment length polymorphism analysis of its sequences. Am J Trop Med Hyg 77: 67–72.
- 30. Hebert PDN, Cywinska A, Ball SL, de WaardJR (2003) Biological identifications through DNA barcodes. Proc Biol Sci 270: 313–321.
- 31. Folmer O, Black M, Hoeh W, Lutz R, Vrijenhoek R (1994) DNA primers for amplification of mitochondrial cytochrome c oxidase subunit I from diverse metazoan invertebrates. Mol Mar Biol Biotechnol 3: 294–299.
- 32. Gómez G, Jaramillo L, Correa MM (2013) Wing geometric morphometrics and molecular assessment of members in the Albitarsis Complex from Colombia. Mol Ecol Resour 13: 1082–1092.
- 33. Wirtz RA, Sattabongkot J, Hall T, Burkot TR, Rosenberg R (1992) Development and evaluation of an enzyme-linked immunosorbent assay for Plasmodium vivax-VK247 sporozoites. J Med Entomol 29: 854–857.
- 34. Wirtz RA, Zavala F, Charoenvit Y, Campbell GH, Burkot TR, et al. (1987) Comparative testing of monoclonal antibodies against Plasmodium falciparum sporozoites for ELISA development. Bull World Health Organ 65: 39–45.
- 35. Rosero DA, Gutiérrez LA, Cienfuegos AV, Jaramillo LM, Correa MM (2010) Optimización de un procedimiento de extracción de ADN para mosquitos anofelinos. Rev Col Ent 36: 260–263.
- 36. Naranjo-Diaz N, Rosero DA, Rua-Uribe G, Luckhart S, Correa MM (2013) Abundance, behavior and entomological inoculation rates of anthropophilic anophelines from a primary Colombian malaria endemic area. Parasit Vectors 6: 61.
- 37. Githeko AK, Lindsay SW, Confalonieri UE, Patz JA (2000) Climate change and vector-borne diseases: a regional analysis. Bull World Health Organ 78: 1136–1147.
- 38. Kovats RS, Bouma MJ, Hajat S, Worrall E, Haines A (2003) El Niño and health. Lancet 362: 1481–1489.
- 39. Lindblade KA, Walker ED, Onapa AW, Katungu J, Wilson ML (2000) Land use change alters malaria transmission parameters by modifying temperature in a highland area of Uganda. Trop Med Int Health 5: 263–274.
- 40. Rubio-Palis Y, Menare C, Quinto A, Magris M, Amarista M (2005) Caracterización de criaderos de anofelinos (Diptera: Culicidae) vectores de malaria del Alto Orinoco, Amazonas, Venezuela. Entomotropica 20: 29–38.
- 41. Hudson JE (1984) Anopheles darlingi Root (Diptera: Culicidae) in the Suriname rain forest. Bull Entomol Res 74: 129–142.
- 42. Rozendaal JA (1992) Relations between Anopheles darlingi breeding habitats, rainfall, river level and malaria transmission rates in the rain forest of Suriname. Med Vet Entomol 6: 16–22.
- 43. Vittor AY, Pan W, Gilman RH, Tielsch J, Glass G, et al. (2009) Linking deforestation to malaria in the Amazon: characterization of the breeding habitat of the principal malaria vector, Anopheles darlingi. Am J Trop Med Hyg 81: 5–12.
- 44. Póvoa M, Wirtz R, Lacerda R, Miles M, Warhurst D (2001) Malaria vectors in the municipality of Serra do Navio, State of Amapá, Amazon Region, Brazil. Mem Inst Oswaldo Cruz 96: 179–184.
- 45. Souza-Santos R (2002) Distribuição sazonal de vetores da malária Amazônica, Brasil Seasonal distribution of malaria vectors in Machadinho d’ Oeste, Rondônia State, Amazon Region, Brazil. Cad Saude Publica 18: 1813–1818.
- 46. Moreno JE, Rubio-Palis Y, Páez E, Pérez E, Sánchez V (2007) Abundance, biting behaviour and parous rate of anopheline mosquito species in relation to malaria incidence in gold-mining areas of southern Venezuela. Med Vet Entomol 21: 339–349.
- 47. Girod R, Gaborit P, Carinci R, Issaly J, Fouque F (2008) Anopheles darlingi bionomics and transmission of Plasmodium falciparum, Plasmodium vivax and Plasmodium malariae in Amerindian villages of the Upper-Maroni Amazonian forest, French Guiana. Mem Inst Oswaldo Cruz 103: 702–710.
- 48. Tadei WP, Dutary Thatcher B (2000) Malaria vectors in the Brazilian amazon: Anopheles of the subgenus Nyssorhynchus. Rev Inst Med Trop Sao Paulo 42: 87–94.
- 49. Hiwat H, Bretas G (2011) Ecology of Anopheles darlingi Root with respect to vector importance: a review. Parasit Vectors 4: 177.
- 50. Poveda G, Rojas W, Quiñones ML, Vélez ID, Mantilla RI, et al. (2001) Coupling between annual and ENSO timescales in the malaria-climate association in Colombia. Environ Health Perspect 109: 489–493.
- 51. Rozendaal JA (1987) Observationos on the biology and behaviouro of Anophelines in the Suriname rainforest with special reference to Anopheles darlingi Root. Cah ORSTOM sér Entomol Méd Parasitol 25: 33–43.
- 52. Gutiérrez LA, González JJ, Gómez GF, Castro MI, Rosero DA, et al. (2009) Species composition and natural infectivity of anthropophilic Anopheles (Diptera: Culicidae) in the states of Córdoba and Antioquia, Northwestern Colombia. Mem Inst Oswaldo Cruz 104: 1117–1124.
- 53. Elliott R (1972) The influence of vector behavior on malaria transmission. Am J Trop Med Hyg 21: 755–763.
- 54. Ulloa A, Gonzalez-Cerón L, Rodríguez MH (2006) Host selection and gonotrophic cycle length of Anopheles punctimacula in southern Mexico. J Am Mosq Control Assoc 22: 648–653.
- 55. Olano V, Carrasquilla G, Méndez F (1997) Transmisión de la malaria urbana en Buenaventura, Colombia: aspectos entomológicos. Pan Am J PublicHealth 1: 287–294.
- 56. Brochero H, Pareja P, Ortiz G, Olano V (2006) Breeding places and biting activity of Anopheles species in the municipality of Cimitarra, Santander, Colombia. Biomédica 26: 269–277.
- 57.
World Health Organization (2010) World Malaria Report 2010. Genove: World Health Organization. 204 p.
- 58. Olano V, Brochero H, Sáenz R, Quiñones M, Molina J (2001) Mapas preliminares de la distribución de especies de Anopheles vectores de malaria en Colombia. Biomédica 21: 402–408.
- 59.
Frederickson E (1993) Bionomía y control de Anopheles albimanus. Washington, D.C.: Pan American Health Organization. 76 p.
- 60. Faran M (1980) Mosquitos Studies (Díptera, Culicidae) XXXIV. A revision of the Albimanus Section of the subgenus Nyssorhynchus of Anopheles. Contrib Amer Ent Inst 15: 1–215.
- 61. Cruz CG, Valle JT, Ruiz AM (2004) Determinación de los habitos de An. pseudopunctipennis y An. calderoni en dos localidades del valle de Chao. La Libertad, Péru. Rev Peru Med Exp Salud Publica 21: 223–230.
- 62. Lucumi-Aragón D, González RO, Salas-Quinchucua C (2011) Actividad de picadura de Anopheles calderoni (Diptera: Culicidae) en dos localidades del Valle del Cauca, Colombia. Rev Colomb Entomol 37: 256–261.
- 63. Rosero DA, Naranjo-Diaz N, Alvarez N, Cienfuegos AV, Torres C, et al. (2013) Colombian Anopheles triannulatus (Diptera: Culicidae) Naturally Infected with Plasmodium spp. ISRN Parasitol 2013: 10.