Cellular and Humoral Responses to Cte f 2, a Cat Flea Allergen, in Children with Papular Urticaria.

BACKGROUND: Papular urticaria (PU) is a common insect bite skin hypersensitivity in tropical countries. In order to gain insight into its causal allergens, we aimed to evaluate cellular and humoral immune responses to the recombinant salivary antigen Cte f 2 from the cat flea Ctenocephalides felis. METHOD: Sixty patients with PU and 27 healthy controls were included in this study. Specific IgE, IgG, IgG1, and IgG4 against Cte f 2 and C. felis extract were determined by ELISA. The T-cell response was analyzed using a carboxyfluorescein succinimidyl ester (CFSE)-based dilution assay and Th1/Th2/Th17 cytokine measurements. In addition, a proteomic analysis of IgG and IgE reactive spots of C. felis extract was performed. RESULTS: The frequency of IgE sensitization to Cte f 2 was similar between patients (36.7%) and controls (40.7%). The specific IgE, IgG1, and IgG4 responses to Cte f 2 and C. felis extract were not significantly different between patients and controls. Among the 3 conditions (i.e., Cte f 2, C. felis extract, and only medium) Cte f 2 was the strongest inducer of CD3+CD4+ proliferation in the patients; however, the mean response was not significantly different from those in controls (Cte f 2: 4.5 vs. 2.5%; p = 0.46). No salivary proteins were identified in C. felis, and most of the spots were identified as muscle-skeletal components (tropomyosin, actin, myosin, and ankirin). CONCLUSIONS: Cte f 2 induces IgE and IgG production as well as T-cell proliferation in children living in a geographical area where PU induced by a flea bite is common. The use of C. felis extract is not recommended for the study of bite-induced hypersensitivity disease since salivary antigens are not well represented.

Absence of knockdown mutations in pyrethroid and DDT resistant populations of the main malaria vectors in Colombia.

BACKGROUND: Knockdown resistance (kdr) is a well-characterized target-site insecticide resistance mechanism that is associated with DDT and pyrethroid resistance. Even though insecticide resistance to pyrethroids and DDT have been reported in Anopheles albimanus, Anopheles benarrochi sensu lato (s.l.), Anopheles darlingi, Anopheles nuneztovari s.l., and Anopheles pseudopunctipennis s.l. malaria vectors in Latin America, there is a knowledge gap on the role that kdr resistance mechanisms play in this resistance. The aim of this study was to establish the role that kdr mechanisms play in pyrethroid and DDT resistance in the main malaria vectors in Colombia, in addition to previously reported metabolic resistance mechanisms, such as mixed function oxidases (MFO) and nonspecific esterases (NSE) enzyme families. METHODS: Surviving (n = 62) and dead (n = 67) An. nuneztovari s.l., An. darlingi and An. albimanus mosquitoes exposed to diagnostic concentrations of DDT and pyrethroid insecticides were used to amplify and sequence a ~ 225 bp fragment of the voltage-gated sodium channels (VGSC) gene. This fragment spanning codons 1010, 1013 and 1014 at the S6 segment of domain II to identify point mutations, which have been associated with insecticide resistance in different species of Anopheles malaria vectors. RESULTS: No kdr mutations were detected in the coding sequence of this fragment in 129 samples, 62 surviving mosquitoes and 67 dead mosquitoes, of An. darlingi, An. nuneztovari s.l. and An. albimanus. CONCLUSION: Mutations in the VGSC gene, most frequently reported in other species of the genus Anopheles resistant to pyrethroid and DDT, are not associated with the low-intensity resistance detected to these insecticides in some populations of the main malaria vectors in Colombia. These results suggest that metabolic resistance mechanisms previously reported in these populations might be responsible for the resistance observed.

Spatial distributions of Anopheles species in relation to malaria incidence at 70 localities in the highly endemic Northwest and South Pacific coast regions of Colombia.

BACKGROUND: A proper identification of malaria vectors is essential for any attempt to control this disease. Between 40 and 47 Anopheles species have been recorded in Colombia, and eight species complexes have been identified in the last decade. An update of Anopheles species distribution and its relationship with malaria is required, particularly for newly identified members of species complexes. METHODS: A cross-sectional entomological study was conducted at 70 localities in the highest malaria transmission areas in Colombia. In each locality, immature and adult mosquitoes were collected. All specimens were determined using morphological characters and confirmed used restriction profiles of Internal Transcribed Spacer 2 (PCR-RFLP-ITS2), and Cytochrome c Oxidase I (COI) sequence gene. To detect natural Plasmodium infections, enzyme-linked immunosorbent assay and nested PCR analysis were used. Distribution of Anopheles species was spatially associated with malaria incidence. RESULTS: A total of 1736 larvae and 12,052 adult mosquitoes were determined in the 70 localities. Thirteen Anopheles species were identified. COI sequence analysis suggested 4 new lineages for Colombia: for Anopheles albimanus (An. albimanus B), Anopheles pseudopunctipennis s.l., Anopheles neivai (An. neivai nr. neivai 4), and Anopheles apicimacula. Two members of species complexes were identified, as: Anopheles nuneztovari C, and Anopheles albitarsis I. Another seven species were confirmed. Four mosquitoes were infected with Plasmodium species, An. albimanus B and An. nuneztovari C. In Northwest of Colombia, An. nuneztovari C, An. albimanus, and Anopheles darlingi were present in the municipalities with highest annual parasitic index (API) (>35 cases/1000 inhabitants). In the north of South Pacific coast, with a similar API, An. nuneztovari C were widely distributed inland, and the main species in coastal regions were An. albimanus B and An. neivai s.l. In the South Pacific coast bordering with Ecuador, 3 Anopheles species were found in municipalities with high API (15-88 cases/1000 inhabitants): An. albimanus B, Anopheles calderoni and An. neivai s.l. CONCLUSIONS: In the highest malaria areas of Colombia, 13 Anopheles species and four new lineages were found, which highlights the need for updating the species distribution. A DNA barcode analysis allowed the taxonomic identification to be refined, particularly for species complexes, and to improve the further understanding of their relation with malaria transmission.

A multi-criteria decision analysis approach to assessing malaria risk in northern South America.

BACKGROUND: Malaria control in South America has vastly improved in the past decade, leading to a decrease in the malaria burden. Despite the progress, large parts of the continent continue to be at risk of malaria transmission, especially in northern South America. The objectives of this study were to assess the risk of malaria transmission and vector exposure in northern South America using multi-criteria decision analysis. METHODS: The risk of malaria transmission and vector exposure in northern South America was assessed using multi-criteria decision analysis, in which expert opinions were taken on the key environmental and population risk factors. RESULTS: Results from our risk maps indicated areas of moderate-to-high risk along rivers in the Amazon basin, along the coasts of the Guianas, the Pacific coast of Colombia and northern Colombia, in parts of Peru and Bolivia and within the Brazilian Amazon. When validated with occurrence records for malaria, An. darlingi, An. albimanus and An. nuneztovari s.l., t-test results indicated that risk scores at occurrence locations were significantly higher (p < 0.0001) than a control group of geographically random points. CONCLUSION: In this study, we produced risk maps based on expert opinion on the spatial representation of risk of potential vector exposure and malaria transmission. The findings provide information to the public health decision maker/policy makers to give additional attention to the spatial planning of effective vector control measures. Therefore, as the region tackles the challenge of malaria elimination, prioritizing areas for interventions by using spatially accurate, high-resolution (1 km or less) risk maps may guide targeted control and help reduce the disease burden in the region.

Human biting activity, spatial-temporal distribution and malaria vector role of Anopheles calderoni in the southwest of Colombia.

BACKGROUND: Anopheles calderoni was first recognized in Colombia in 2010 as this species had been misidentified as Anopheles punctimacula due to morphological similarities. An. calderoni is considered a malaria vector in Peru and has been found naturally infected with Plasmodium falciparum in Colombia. However, its biting behaviour, population dynamics and epidemiological importance have not been well described for Colombia. METHODS: To assess the contribution of An. calderoni to malaria transmission and its human biting behaviour and spatial/temporal distribution in the southwest of Colombia, human landing catches (HLC) and larval collections were carried out in a cross-sectional, entomological study in 22 localities between 2011 and 2012, and a longitudinal study was performed in the Boca de Prieta locality in Olaya Herrera municipality between July 2012 and June 2013. All mosquitoes determined as An. calderoni were tested by ELISA to establish infection with Plasmodium spp. RESULTS: Larvae of An. calderoni were found in four localities in 12 out of 244 breeding sites inspected. An. calderoni adults were collected in 14 out of 22 localities during the cross-sectional study and represented 41.3% (459 of 1,111) of the collected adult specimens. Other species found were Anopheles albimanus (54.7%), Anopheles apicimacula (2.1%), Anopheles neivai (1.7%), and Anopheles argyritarsis (0.2%). In the localities that reported the highest malaria Annual Parasite Index (>10/1,000 inhabitants) during the year of sampling, An. calderoni was the predominant species (>90% of the specimens collected). In the longitudinal study, 1,528 An. calderoni were collected by HLC with highest biting rates in February, May and June 2013, periods of high precipitation. In general, the species showed a preference to bite outdoors (p < 0.001). In Boca de Prieta, two specimens of An. calderoni were ELISA positive for Plasmodium circumsporozoite protein: one for P. falciparum and one for Plasmodium vivax VK-210. This represents an overall sporozoite rate of 0.1% and an annual entomological inoculation rate of 2.84 infective bites/human/year. CONCLUSIONS: This study shows that An. calderoni is a primary malaria vector in the southwest of Colombia. Its observed preference for outdoor biting is a major challenge for malaria control.

Prospects and recommendations for risk mapping to improve strategies for effective malaria vector control interventions in Latin America.

With malaria control in Latin America firmly established in most countries and a growing number of these countries in the pre-elimination phase, malaria elimination appears feasible. A review of the literature indicates that malaria elimination in this region will be difficult without locally tailored strategies for vector control, which depend on more research on vector ecology, genetics and behavioural responses to environmental changes, such as those caused by land cover alterations, and human population movements. An essential way to bridge the knowledge gap and improve vector control is through risk mapping. Malaria risk maps based on statistical and knowledge-based modelling can elucidate the links between environmental factors and malaria vectors, explain interactions between environmental changes and vector dynamics, and provide a heuristic to demonstrate how the environment shapes malaria transmission. To increase the utility of risk mapping in guiding vector control activities, definitions of malaria risk for mapping purposes must be standardized. The maps must also possess appropriate scale and resolution in order to become essential tools in integrated vector management (IVM), so that planners can target areas in greatest need of control measures. Fully integrating risk mapping into vector control programmes will make interventions more evidence-based, making malaria elimination more attainable.

Larval habitat characteristics of the main malaria vectors in the most endemic regions of Colombia: potential implications for larval control.

BACKGROUND: Malaria incidence has recently decreased globally and, as malaria elimination is envisioned as a possibility by the health authorities, guidance is needed to strengthen malaria control strategies. Larval source treatment, which could complement routine vector control strategies, requires knowledge regarding the Anopheles larval habitats. METHODS: A cross-sectional study was conducted in three of the most malaria-endemic regions in Colombia. A total of 1116 potential larval habitats in 70 villages were sampled in three states located in western Colombia: Cordoba, Valle del Cauca and Nariño. RESULTS: Overall, 17.5 % (195) of the potential larval habitats were found positive for different Anopheles species. A total of 1683 larvae were identified belonging to seven species: Anopheles albimanus, Anopheles calderoni, Anopheles darlingi, Anopheles neomaculipalpus, Anopheles nuneztovari s.l., Anopheles pseudopunctipennis, and Anopheles triannulatus. The most widely distributed species was An. nuneztovari s.l., which was found mainly in human-made fishponds in Cordoba and temporary puddles in Valle del Cauca. Anopheles albimanus and An. calderoni were associated with human-made wells or excavation sites in Nariño. Cordoba displayed the greatest Anopheles species diversity with a total of six species (Shannon diversity index H': 1.063). Although Valle del Cauca had four species, one more than Nariño, the diversity was lower because only one species predominated, An. nuneztovari s.l. The larval habitats with the highest Shannon diversity index were lagoons (H': 1.079) and fishponds (H': 1.009) in Cordoba, excavation sites in Nariño (H': 0.620) and puddles in Valle del Cauca (H': 0.764). CONCLUSIONS: This study provides important information regarding the larval habitats of the main malaria vectors in the most malaria-endemic regions of Colombia, which will be useful in guiding larval control operations.

Anopheles species composition explains differences in Plasmodium transmission in La Guajira, northern Colombia.

Malaria in La Guajira, the most northern state of Colombia, shows two different epidemiological patterns. Malaria is endemic in the municipality of Dibulla whereas in Riohacha it is characterised by sporadic outbreaks. This study aimed to establish whether differences in transmission patterns could be attributed to different vector species. The most abundant adult female species were Anopheles aquasalis, exclusive to Riohacha, and Anopheles darlingi, restricted to Dibulla. Anopheles mosquitoes were identified using morphology and the molecular markers internal transcribed spacer 2 and cytochrome c oxidase I. All specimens (n = 1,393) were tested by ELISA to determine natural infection rates with Plasmodium falciparum and Plasmodium vivax. An. darlingi was positive for P. vivax 210, with an infection rate of 0.355% and an entomological inoculation rate of 15.87 infective bites/person/year. Anopheles albimanus larvae were the most common species in Riohacha, found in temporary swamps; in contrast, in Dibulla An. darlingi were detected mainly in permanent streams. Distinctive species composition and larval habitats in each municipality may explain the differences in Plasmodium transmission and suggest different local strategies should be used for vector control.

Morphology of the larvae, male genitalia and DNA sequences of Anopheles (Kerteszia) pholidotus (Diptera: Culicidae) from Colombia.

Since 1984, Anopheles (Kerteszia) lepidotus has been considered a mosquito species that is involved in the transmission of malaria in Colombia, after having been incriminated as such with epidemiological evidence from a malaria outbreak in Cunday-Villarrica, Tolima. Subsequent morphological analyses of females captured in the same place and at the time of the outbreak showed that the species responsible for the transmission was not An. lepidotus, but rather Anopheles pholidotus. However, the associated morphological stages and DNA sequences of An. pholidotus from the foci of Cunday-Villarrica had not been analysed. Using samples that were caught recently from the outbreak region, the purpose of this study was to provide updated and additional information by analysing the morphology of female mosquitoes, the genitalia of male mosquitoes and fourth instar larvae of An. pholidotus, which was confirmed with DNA sequences of cytochrome oxidase I and rDNA internal transcribed spacer. A total of 1,596 adult females were collected in addition to 37 larval collections in bromeliads. Furthermore, 141 adult females, which were captured from the same area in the years 1981-1982, were analysed morphologically. Ninety-five DNA sequences were analysed for this study. Morphological and molecular analyses showed that the species present in this region corresponds to An. pholidotus. Given the absence of An. lepidotus, even in recent years, we consider that the species of mosquitoes that was previously incriminated as the malaria vector during the outbreak was indeed An. pholidotus, thus ending the controversy.

High Levels of Diversity in Anopheles Subgenus Kerteszia Revealed by Species Delimitation Analyses.

The Anopheles subgenus Kerteszia is a poorly understood group of mosquitoes that includes several species of medical importance. Although there are currently twelve recognized species in the subgenus, previous studies have shown that this is likely to be an underestimate of species diversity. Here, we undertake a baseline study of species delimitation using the barcode region of the mtDNA COI gene to explore species diversity among a geographically and taxonomically diverse range of Kerteszia specimens. Beginning with 10 of 12 morphologically identified Kerteszia species spanning eight countries, species delimitation analyses indicated a high degree of cryptic diversity. Overall, our analyses found support for at least 28 species clusters within the subgenus Kerteszia. The most diverse taxon was Anopheles neivai, a known malaria vector, with eight species clusters. Five other species taxa showed strong signatures of species complex structure, among them Anopheles bellator, which is also considered a malaria vector. There was some evidence for species structure within An. homunculus, although the results were equivocal across delimitation analyses. The current study, therefore, suggests that species diversity within the subgenus Kerteszia has been grossly underestimated. Further work will be required to build on this molecular characterization of species diversity and will rely on genomic level approaches and additional morphological data to test these species hypotheses.