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Background: In Brazilian Amazon, deforestation and other anthropogenic activities as a consequence of human occupation have created new and artificial larval habitats for anopheline mosquitoes, providing conditions for oviposition, development and expansion of malaria vector populations. Objectives: This study aimed to structurally characterize and describe the entomological and limnological parameters of Anopheles larval habitats from a malaria region in northern Brazilian Amazon. Methods: Fifty-two larval habitats were investigated in the District of Ilha de Santana, in the Brazilian state of Amapá, comprising fish farming tanks, ponds, and streams. For entomological parameters, the immature larvae were collected monthly from July 2019 to June 2020. For limnological parameters, the water samples were collected from 20 larval habitats during the sampling period. The data were analyzed using Generalized Linear Models, Multivariate analyses, and Kruskal-Wallis tests. Results: Fifty habitats were positive for Anopheles larvae and a total of nine species were collected. The fish farming tanks had the highest abundance of larvae compared with ponds and streams. Anopheles darlingi, Anopheles nuneztovari s.l. and Anopheles triannulatus were collected in 94% of the larval habitats and showed the highest positivity index. The degree of shade and the type of water of the breeding sites were important factors for the presence of the main malaria vector, A. darlingi. This species was the most affected by pH, total dissolved solids, electrical conductivity, and nitrate. Conclusions: Our findings indicate that fish farming tanks are major contributors to vector anopheline abundance and malaria transmission. Vector control strategies focused on these habitats are urgently needed. [ABSTRACT FROM AUTHOR]

Risk of spillover and spillback of mosquito-borne viruses in the neotropics, including yellow fever, dengue, Zika (Flaviviridae: Flavivirus), chikungunya, and Mayaro (Togaviridae: Alphavirus) viruses, is highest at ecotones where humans, monkeys, and mosquitoes coexist. With a view to identifying potential bridge vectors, we investigated changes in mosquito community composition and environmental variables at ground level at distances of 0, 500, 1000, and 2000 m from the edge of a rainforest reserve bordering the city of Manaus in the central Brazilian Amazon. During two rainy seasons in 2019 and 2020, we sampled 9,467 mosquitoes at 244 unique sites using BG-Sentinel traps, hand-nets, and Prokopack aspirators. Species richness and diversity were generally higher at 0 m and 500 m than at 1000 m and 2000 m, while mosquito community composition changed considerably between the forest edge and 500 m before stabilizing by 1000 m. Shifts in environmental variables mainly occurred between the edge and 500 m, and the occurrence of key taxa (Aedes albopictus, Ae. scapularis, Limatus durhamii, Psorophora amazonica, Haemagogus, and Sabethes) was associated with one or more of these variables. Sites where Ae. aegypti and Ae. albopictus were detected had significantly higher surrounding mean NDBI (Normalized Difference Built-up Index) values than sites where they were not detected, while the opposite was true for Sabethes mosquitoes. Our findings suggest that major changes in mosquito communities and environmental variables occur within 500 m of the forest edge, where there is high risk for contact with both urban and sylvatic vectors. By 1000 m, conditions stabilize, species diversity decreases, and forest mosquitoes predominate. Environmental variables associated with the occurrence of key taxa may be leveraged to characterize suitable habitat and refine risk models for pathogen spillover and spillback. Author summary: Mosquito-borne viruses, including yellow fever, dengue, Zika, chikungunya, and Mayaro viruses, derive from sylvatic transmission cycles involving forest mosquitoes and wild animals. Occasionally, these sylvatic viruses spill over into humans, which can lead to limited disease outbreaks or sustained human transmission by urban mosquitoes. When translocated by humans to new geographic regions, these viruses have potential to spill back into novel sylvatic cycles, creating a lasting threat to human health. Ecotones, such as forest edges, where humans, mosquitoes, and monkeys coexist, are at high risk of spillover and spillback. We investigated changes in mosquito community composition at incremental distances up to 2000 m into rainforest bordering the Amazonian city of Manaus, where the above viruses circulate in urban and sylvatic cycles. We also characterized mosquito habitat and analyzed associations between key vector species and environmental variables like temperature and vegetation. Our results show that mosquito communities and environmental variables change abruptly within 500 m of the forest edge, where there is high risk for contact of both monkeys and humans with both urban and forest mosquitoes. By 1000 m, conditions stabilize, and forest mosquitoes predominate. Environmental variables associated with the presence of particular mosquito taxa will help characterize suitable habitat for key vector species and refine spillover and spillback risk models for these viruses. [ABSTRACT FROM AUTHOR]

Recent studies have shown that Anopheles oswaldoi sensu lato comprises a cryptic species complex in South America. Anopheles konderi, which was previously raised to synonymy with An. oswaldoi, has also been suggested to form a species complex. An. oswaldoi has been incriminated as a malaria vector in some areas of the Brazilian Amazon, Colombia, Peru and Venezuela, but was not recognized as a vector in the remaining regions in its geographic distribution. The role of An. konderi as a malaria vector is unknown or has been misattributed to An. oswaldoi. The focus of this study was molecular identification to infer the evolutionary relationships and preliminarily delimit the geographic distribution of the members of these complexes in the Brazilian Amazon region. The specimens were sampled from 18 localities belonging to five states in the Brazilian Amazon and sequenced for two molecular markers: the DNA barcode region (COI gene of mitochondrial DNA) and Internal Transcribed Spacer 2 (ITS2 ribosomal DNA). COI (83 sequences) and ITS2 (27 sequences) datasets generated 43 and 10 haplotypes, respectively. Haplotype networks and phylogenetic analyses generated with the barcode region (COI gene) recovered five groups corresponding to An. oswaldoi s.s., An. oswaldoi B, An. oswaldoi A, An. konderi and An. sp. nr. konderi; all pairwise genetic distances were greater than 3%. The group represented by An. oswaldoi A exhibited three strongly supported lineages. The molecular dating indicated that the diversification process in these complexes started approximately 2.8 Mya, in the Pliocene. These findings confirm five very closely related species and present new records for these species in the Brazilian Amazon region. The paraphyly observed for the An. oswaldoi complex suggests that An. oswaldoi and An. konderi complexes may comprise a unique species complex named Oswaldoi-Konderi. Anopheles oswaldoi B may be a potential malaria vector in the extreme north of the Brazilian Amazon, whereas evidence of sympatry for the remaining species in other parts of the Brazilian Amazon (Acre, Amazonas, Pará and Rondônia) precluded identification of probable vectors in those areas. [ABSTRACT FROM AUTHOR]

Background: Lutzomyia umbratilis is an important Leishmania guyanensis vector in South America. Previous studies have suggested differences in the vector competence between L. umbratilis populations situated on opposite banks of the Amazonas and Negro Rivers in the central Amazonian Brazil region, likely indicating a species complex. However, few studies have been performed on these populations and the taxonomic status of L. umbratilis remains unclear. Methodology/Principal Findings: Phylogeographic structure was estimated for six L. umbratilis samples from the central Amazonian region in Brazil by analyzing mtDNA using 1181 bp of the COI gene to assess whether the populations on opposite banks of these rivers consist of incipient or distinct species. The genetic diversity was fairly high and the results revealed two distinct clades ( = lineages) with 1% sequence divergence. Clade I consisted of four samples from the left bank of the Amazonas and Negro Rivers, whereas clade II comprised two samples from the right bank of Negro River. No haplotypes were shared between samples of two clades. Samples within clades exhibited low to moderate genetic differentiation (FST =20.0390-0.1841), whereas samples between clades exhibited very high differentiation (FST = 0.7100- 0.8497) and fixed differences. These lineages have diverged approximately 0.22 Mya in the middle Pleistocene. Demographic expansion was detected for the lineages I and II approximately 30,448 and 15,859 years ago, respectively, in the late Pleistocene. Conclusions/Significance: The two genetic lineages may represent an advanced speciation stage suggestive of incipient or distinct species within L. umbratilis. These findings suggest that the Amazonas and Negro Rivers may be acting as effective barriers, thus preventing gene flow between populations on opposite sides. Such findings have important implications for epidemiological studies, especially those related to vector competence and anthropophily, and for vector control strategies. In addition, L. umbratilis represents an interesting example in speciation studies. [ABSTRACT FROM AUTHOR]