DNA Barcoding of Morphologically Characterized Mosquitoes Belonging to the Genus Mansonia from the Atlantic Forest and Brazilian Savanna.

The identification of mosquito species is necessary for determining the entomological components of disease transmission. However, identification can be difficult in species that are morphologically similar. The cytochrome c oxidase subunit I (COI) DNA barcode region is considered a valuable and reliable diagnostic tool for mosquito species recognition, including those that belong to species complexes. Mansonia mosquitoes are found in forests near swampy areas. They are nocturnal and are highly attracted to light. Hematophagous adult females exhibit aggressive biting behavior and can become infected with and transmit pathogens during their feeding, including some epizootic viruses and avian malaria. In Brazil, twelve Mansonia species have been reported. In a recent study from the São Paulo Zoo in Brazil, three morphologically distinct species were collected and identified, namely: Mansonia (Mansonia) indubitans, Ma. (Man.) pseudotitillans and Ma. (Man.) titillans. However, confirmation of these species by molecular identification was unsuccessful due to a lack of COI sequences in the GenBank database. Thus, this research aimed to describe the COI DNA barcode sequences of some morphologically characterized Mansonia (Man.) species from Brazil and to determine their utility in delimiting species collected from the Atlantic Forest and Brazilian Savanna. Accordingly, we provide tools for the genetic identification of species that play a significant role in pathogen transmission in wildlife and potentially humans. We show that the delimitation of Mansonia species via five different approaches based on COI DNA sequences (BI, NJ, ASAP, bPTP and GMYC) yield basically the same groups identified by traditional taxonomy, and we provide the identification of specimens that were previously identified only up to the subgenus level. We also provide COI sequences from two Mansonia species that were not previously available in sequence databases, Ma. wilsoni and Ma. pseudotitillans, and thus contribute to the ongoing global effort to standardize DNA barcoding as a molecular means of species identification.

Mapping environmental suitability of Haemagogus and Sabethes spp. mosquitoes to understand sylvatic transmission risk of yellow fever virus in Brazil.

BACKGROUND: Yellow fever (YF) is an arboviral disease which is endemic to Brazil due to a sylvatic transmission cycle maintained by infected mosquito vectors, non-human primate (NHP) hosts, and humans. Despite the existence of an effective vaccine, recent sporadic YF epidemics have underscored concerns about sylvatic vector surveillance, as very little is known about their spatial distribution. Here, we model and map the environmental suitability of YF's main vectors in Brazil, Haemagogus spp. and Sabethes spp., and use human population and NHP data to identify locations prone to transmission and spillover risk. METHODOLOGY/PRINCIPAL FINDINGS: We compiled a comprehensive set of occurrence records on Hg. janthinomys, Hg. leucocelaenus, and Sabethes spp. from 1991-2019 using primary and secondary data sources. Linking these data with selected environmental and land-cover variables, we adopted a stacked regression ensemble modelling approach (elastic-net regularized GLM, extreme gradient boosted regression trees, and random forest) to predict the environmental suitability of these species across Brazil at a 1 km x 1 km resolution. We show that while suitability for each species varies spatially, high suitability for all species was predicted in the Southeastern region where recent outbreaks have occurred. By integrating data on NHP host reservoirs and human populations, our risk maps further highlight municipalities within the region that are prone to transmission and spillover. CONCLUSIONS/SIGNIFICANCE: Our maps of sylvatic vector suitability can help elucidate potential locations of sylvatic reservoirs and be used as a tool to help mitigate risk of future YF outbreaks and assist in vector surveillance. Furthermore, at-risk regions identified from our work could help disease control and elucidate gaps in vaccination coverage and NHP host surveillance.

Genomic Surveillance of Yellow Fever Virus Epizootic in São Paulo, Brazil, 2016 - 2018.

São Paulo, a densely inhabited state in southeast Brazil that contains the fourth most populated city in the world, recently experienced its largest yellow fever virus (YFV) outbreak in decades. YFV does not normally circulate extensively in São Paulo, so most people were unvaccinated when the outbreak began. Surveillance in non-human primates (NHPs) is important for determining the magnitude and geographic extent of an epizootic, thereby helping to evaluate the risk of YFV spillover to humans. Data from infected NHPs can give more accurate insights into YFV spread than when using data from human cases alone. To contextualise human cases, identify epizootic foci and uncover the rate and direction of YFV spread in São Paulo, we generated and analysed virus genomic data and epizootic case data from NHPs in São Paulo. We report the occurrence of three spatiotemporally distinct phases of the outbreak in São Paulo prior to February 2018. We generated 51 new virus genomes from YFV positive cases identified in 23 different municipalities in São Paulo, mostly sampled from NHPs between October 2016 and January 2018. Although we observe substantial heterogeneity in lineage dispersal velocities between phylogenetic branches, continuous phylogeographic analyses of generated YFV genomes suggest that YFV lineages spread in São Paulo at a mean rate of approximately 1km per day during all phases of the outbreak. Viral lineages from the first epizootic phase in northern São Paulo subsequently dispersed towards the south of the state to cause the second and third epizootic phases there. This alters our understanding of how YFV was introduced into the densely populated south of São Paulo state. Our results shed light on the sylvatic transmission of YFV in highly fragmented forested regions in São Paulo state and highlight the importance of continued surveillance of zoonotic pathogens in sentinel species.

Paridade e desenvolvimento ovariano de Ochlerotatus scapularis (Diptera Culicidae) em condições de laboratório e de campo na região do Vale do Ribeira, São Paulo, Brasil / Parity and ovarian development of Ochlerotatus scapularis (Diptera: Culicidae) in laboratory and field conditions in Vale do Ribeira region. São Paulo, Brazil

Objetivo. Identificar o estado de paridade em fêmeas de Ochlerotatus scapularis em condições de laboratório e campo. Das fêmeas coletadas em campo, determinar o grau de paridade considerando sua importância epidemiológica como possível transmissor de patógenos. Métodos. O trabalho foi dividido em duas partes: no campo foram realizadas coletas quinzenais no Vale do Ribeira de abril de 2003 a março de 2004, utilizando-se a técnica da aspiração manual. No laboratório, 100 fêmeas (F1) de Ochlerotatus scapularis foram criadas e observadas individualmente anotando-se o número de repastos por elas realizados, duração do ciclo gonotrófico e número de ovos colocados. A observação do estado de paridade e desenvolvimento ovariano foi feita através da dissecação dos ovários de 90 fêmeas, por coleta, e de todas as fêmeas criadas. Resultados. Das 100 fêmeas mantidas em laboratório e dissecadas, o diagnóstico de condição de paridade conferiu com os resultados constatados em 55 por cento dos casos, sendo subestimados em 37 por cento e superestimados em 2 por cento. Ainda em laboratório, de 106 ciclos gonotróficos complementados, cercade 55 por cento das fêmeas necessitaram de mais de um repasto sangüíneo antes de ovipor, indicando uma possível discordância gonotrófica para a espécie. Sob condições laboratoriais observou-se uma sobrevivência de até 26 dias para a espécie. Foram dissecadas 1180 fêmeas de Oc. scapularis do campo: 418 (35,4 por cento) foram consideradas nulíparas, 655 (55,5 por cento) uníparas, 46 (3,9 por cento) como bíparas e 61 (5,2 por cento) não puderam ser avaliadas. Foi observado que 90 fêmeas apresentavam-se na fase III-V de Christopher e Mer, reforçando a discordância gonotrófica. A variável estado de paridade variou ao longo do ano. Conclusões: Observando fêmeas de Oc. scapularis no campo e laboratório, confirmamos a hipótese de discordância gonotrófica.