Host location by exophagic African Malaria vectors: evaluating the performance of a novel exposure-free human-baited mosquito trap and studies on the perception of indoors/outdoors by malaria vectors in southern Mozambique

A malária é transmitida por mosquitos que, preferencialmente, picam no interior (endofágicos) ou no exterior (exofágicos) das habitações. Atualmente, os métodos mais sucedidos no controlo vectorial são a pulverização residual intra-domiciliar (PIDOM) e as redes mosquiteiras tratadas com insecticidas de longa duração (REMILD), que têm como alvo principal os vectores endofágicos. No entanto, as espécies que picam no exterior (exófagicas) têm sido as mais difíceis de controlar e, atualmente, não existem métodos satisfatórios para o controlo de populações destes vectores. A transmissão da malária feita por vectores exofágicos é comum fora de África; contudo, o fenómeno vem, progressivamente, ganhando importância também no continente Africano. Daí que, para que os vectores exofágicos sejam controlados de forma efetiva, é necessário entender melhor como ou porquê alguns vectores preferem procurar refeições sanguíneas em ambientes fechados, enquanto os outros evitam entrar em tais compartimentos? Ou seja, como é que um vector localiza e reconhece um ambiente fechado (ou intra-domiciliar) ou aberto (extra-domiciliar)? Com vista a obter respostas à tais questões, varias experiências de campo foram realizadas na aldeia de Massavasse, localizada na região leste do distrito de Chóckwè, província de Gaza, sul de Moçambique. As experiências assentaram-se em dois objetivos principais: 1. Projetar, construir e avaliar a performance de uma armadilha de grelha elétrica, apelidada de armadilha Shock-wè (SHK-wè), que poderá ser utilizada como uma alternativa fiável às colheitas com isca humana, na amostragem de vectores da malária que procuram um hospedeiro humano em ambientes interiores ou exteriores. 2. Determinar a contribuição dos elementos estruturais básicos que compõe uma habitação (ou seja, tecto, paredes, pilares, etc.) para a ocorrência e entrada de mosquitos que procuram uma refeição sanguínea em casas experimentais. Para cumprir o primeiro objectivo, foram feitas colheitas emparelhadas de mosquitos através de armadilhas SHK-wè e colheitas com isca humana (CIH) durante 35 noites consecutivas. Os resultados indicam que a SHK-wè é uma ferramenta segura e fiável para a amostragem e vigilância das populações de vetores importantes da malária em África. SHK-wè mostrou um bom desempenho tanto no interior como no exterior e, portanto, é potencialmente um substituto prático, robusto e seguro para as capturas CIH convencionais. Para cumprir o segundo objetivo, foram igualmente utilizadas armadilhas SHK-wè numa série de experiências de tratamentos randomizados para determinar a resposta das populações de mosquitos locais à presença de componentes estruturais de uma casa experimental construída para o efeito. A experiência foi realizada durante 50 noites consecutivas e constituiu em montagens e desmontagens aleatórias da casa experimental nos seus elementos básicos (isto é, o suporte de metal, paredes inferiores e superiores e o tecto). Os resultados desta experiência sugerem que, contrariamente ao que se conhecia, o tecto da casa é um componente chave para o reconhecimento e entrada dos vectores endofágicos do complexo A. gambiae s.l em ambientes intra-domiciliares. Os dados indicaram que a probabilidade de A. gambiae s.l. entrar numa casa experimental aumenta 4.5 vezes [IRR = 1.50 (0.63 – 2.37; p = 0.001) quando o tecto está presente em comparação com outros tipos de tratamentosMalaria is, preferentially, transmitted by mosquitoes that bite indoors or outdoors. To date, the most successful vector control methods, namely: indoors residual spraying (IRS) and long lasting insecticide treated bed nets (LLINs) have targeted indoor species. Outdoor biters (exophagic) are difficult to target and no satisfactory control methods exist. Already common outside Africa, outdoor-transmitted malaria is growing in importance within Africa. To target exophagic mosquitoes we need to understand more how/why they choose to enter or to avoid houses, i.e. what do mosquitoes perceive as indoors or outdoors? Field experiments were undertaken in Massavasse, a village located in the eastern region of Chóckwè district, Gaza province, southern Mozambique, to investigate that question and addressing two objectives: 1. To design, build and evaluate an electric net trap (ENT), named the “Shock-wè (SHK-wè) trap” for reliable sampling of mosquitoes responding to human bait, indoors and outdoors; 2. To determine the contribution of simple elements of a house (i.e. roof, walls, partial walls) to occurrence and entry rates of host seeking mosquitoes. To accomplish the first objective, a series of paired field tests were performed to evaluate the performance of SHK-wè trap compared to Human landing catch (HLC) method. Results showed that SHK-wè trap is a safe and reliable sampling and surveillance tool for African malaria vector populations. The trap performed well indoors and outdoors and is potentially a robust and safe practical replacement for the conventional HLC method. To accomplish the second objective, SHK-wè trap was used in a series of randomized treatment-assigning experiments to determine the response of local mosquitoes populations to presence of structural components of a purpose-built experimental house. The experiment consisted on randomly dismantling or reassembling an experimental house into its basic elements (i.e. a frame on a base, lower walls, upper walls and roof). Results from this experiment suggest that, contrarily to current belief, the roof of the house was a key component that triggered endophagic malaria vectors, such as members of A. gambiae complex, to enter. Data indicated that the probability of A. gambiae s.l. entering an experimental house increased 4.5 [IRR = 1.50 (0.63 – 2.37); p = 0.001)] times when the roof was in place compared to other types of treatments

Characterisation of human exposure to nocturnal biting by malaria and arbovirus vectors in a rural community in Chókwè district, southern Mozambique

BACKGROUND : Understanding the magnitude of human exposure to mosquito biting is fundamental to reduce pathogen transmission. Here we report on a study quantifying the levels of mosquitoes attacking humans throughout the night in a rural area of Southern Mozambique. METHODS : Surveys were carried out in Massavasse village, southern Mozambique. The abundance and composition of host-seeking mosquito communities at night were assessed by human-landing catches (HLC) at one-hour intervals. Periods when people were located predominantly outdoors or indoors were used to estimate the amount of residents’ exposure to mosquito bites in either location, to explore the potential impact a bed net could have had in reducing biting by each vector species. RESULTS : A total of 69,758 host-seeking female mosquitoes comprising 23 species in four genera were collected. The exposure to biting by virtually all vector species was consistently high outdoors, typically at early evening and morning, with exception of An. gambiae s.l which was likely of biting a person with nearly same intensity indoors and outdoors throughout the night. Bed nets use could have reduced biting by An. gambiae s.l (dominated by An. arabiensis), Ma. africana, Ma. uniformis, Cx. pipiens, Cx. antennatus, and Cx. poicilipes by 53%, 47%, 46%, 38%, 31%, and 28% respectively, compared to non-users. Conversely, a bed net user would have had little protection against An. pharoensis, An. ziemanni, An. tenebrosus, and Cx. tritaeniorhynchus biting exposures. CONCLUSIONS : This study showed that Massavasse residents were exposed to high levels of outdoor biting by malaria and arbovirus vectors that abound in the village. The findings help to identify entomological drivers of persistent malaria transmission in Mozambique and identify a wide range of arbovirus vectors nocturnally active in rural areas, many with outbreak potential. The study highlights the need for a surveillance system for monitoring arboviral diseases vectors in Mozambique.Wellcome [098562] under the fellowship in Tropical Medicine and Public Health.https://wellcomeopenresearch.orgZoology and Entomolog

First confirmed occurrence of the yellow fever virus and dengue virus vector Aedes (Stegomyia) luteocephalus (Newstead, 1907) in Mozambique

Background: Mozambique, same as many other tropical countries, is at high risk of arthropod-borne virus (arbovirus) diseases and recently two dengue virus (DENV) outbreaks occurred in the northern part of the country. The occurrence of some important vector species, such as Aedes (Stegomyia) aegypti (Linnaeus) and Ae. (Stg.) albopictus (Skuse), besides several other sylvatic vectors, have been reported in the country, which may indicate that the transmission of some arboviruses of public health importance may involve multiple-vector systems. Therefore, knowing the occurrence and distribution of existing and the new important vectors species, is crucial for devising systematic transmission surveillance and vector control approaches. The aim of this study was to map the occurrence and distribution of mosquito species with potential for transmitting arboviruses of human and veterinary relevance in Niassa Province, Northern Mozambique. Methods: Field entomological surveys were undertaken in April 2016 in Lago District, Niassa Province, northern Mozambique. Breeding sites of mosquitoes were inspected and immature stages were collected and reared into adult. Mosquitoes in the adult stages were morphologically identified using taxonomic keys. Morphological identification of Aedes (Stegomyia) luteocephalus (Newstead) were later confirmed using dissected male genitalia and molecular based on the phylogenetic analyses of the sequenced barcode (cox1 mtDNA) gene. Results: A total of 92 mosquito larvae collected developed into adults. Of these, 16 (17.39%) were morphologically identified as Ae. luteocephalus. The remaining specimens belonged to Ae. (Stg.) aegypti (n = 4, 4.35%), Ae. (Aedimorphus) vittatus (n = 24, 26.09%), Anopheles garnhami (n = 1, 1.09%), Culex (Culiciomyia) nebulosus (n = 28, 30.43%), Eretmapodites subsimplicipes (n = 18, 19.57%) and Toxorhynchites brevipalpis (n = 1, 1.09%), taxa already known to the country. Male genitalia and phylogenetic analyses confirmed the identity of Ae. luteocephalus specimens collected in this study. Conclusions: To our knowledge, this is the first detection of Ae. luteocephalus in Mozambican territory, a vector species of yellow fever virus (YFV), Zika virus (ZIKV) and dengue virus (DENV) in Africa. Further studies are encouraged to investigate the role of Ae. luteocephalus in the transmission of arboviral diseases in Mozambique.publishersversionpublishe

The Shockwè trap: a human-baited exposure-free device for surveillance and behaviour studies of anthropophilic vectors

Background: The human biting rate (MBR) and entomological inoculation rate (EIR) are common parameters routinely used to measure the risk of malaria transmission. Both parameters can be estimated using human landing catches (HLC). Although it is considered the gold-standard, HLC puts collectors at higher risk of infection with mosquito-transmitted pathogens. Methods: A novel exposure-free host-seeking mosquito electrocution trap, the Shockwè trap (SHK), was developed and its efficiency for monitoring mosquito community composition and abundance was compared with human landing catches (HLC) as the gold-standard. Field experiments were performed in Massavasse village, southern Mozambique. Simultaneous indoor and outdoor collections of nocturnal host-seeking mosquitoes were carried out using the SHK and HLC methods. The relative sampling efficiency of SHK was estimated as the ratio of the numbers of mosquitoes caught in SHK compared HLC. Proportionality and density-dependence between SHK and HLC catches were estimated by mean of Bayesian regression approaches. Results: A total of 69,758 and 27,359 host-seeking mosquitoes comprising nineteen species and four genera, were collected by HLC and SHK respectively. In general, SHK and HLC sampled similar numbers of mosquito species, with the exceptions of the least common species Aedes sudanensis, Ae. subargenteus, and Coquillettidia versicolor that were caught only by HLC. The relative sampling efficiency and proportionality between SHK and matched HLC catches varied greatly between species and collection site. However, all mosquitoes collected by SHK were unfed, confirming the Shockwè trap design’s performance and reliability as a successful mosquito exposure free sampling approach. Conclusions: Results demonstrate that SHK is a safe and reliable human-exposure free device for monitoring the occurrence of a wide range of mosquito, including major malaria and arboviruses vector species. However, improvements are needed to increase its sampling efficiency for less abundant mosquito species

Targeted elimination of species-rich larval habitats can rapidly collapse arbovirus vector mosquito populations at hotel compounds in Zanzibar

Understanding the dynamics of larval habitat utilization by mosquito communities is crucial for the design of efficient environmental control strategies. The authors investigated the structure of mosquito communities found at hotel compounds in Zanzibar, networks of mosquito interactions with larval habitats and robustness of mosquito communities to elimination of larval habitats. A total of 23 698 mosquitoes comprising 26 species in six genera were found. Aedes aegypti (n = 16 207), Aedes bromeliae/Aedes lillie (n = 1340), Culex quinquefasciatus (n = 1300) and Eretmapodites quinquevitattus (n = 659) were the most dominant species. Ecological network analyses revealed the presence of dominant, larval habitat generalist species (e.g., A. aegypti), exploiting virtually all types of water holding containers and few larval habitat specialist species (e.g., Aedes natalensis, Orthopodomyia spp). Simulations of mosquito community robustness to systematic elimination of larval habitats indicate that mosquito populations are highly sensitive to elimination of larval habitats sustaining higher mosquito species diversity. This study provides insights on potential foci of future mosquito-borne arboviral disease outbreaks in Zanzibar and underscores the need for detailed knowledge on the ecological function of larval habitats for effective mosquito control by larval sources management.Danida Fellowship Centrehttp://onlinelibrary.wiley.com/journal/10.1111/(ISSN)1365-2915am2022Zoology and Entomolog

A survey of RNA viruses in mosquitoes from Mozambique reveals novel genetic lineages of flaviviruses and phenuiviruses, as well as frequent flavivirus-like viral DNA forms in Mansonia

Background: Mosquito-borne diseases involving arboviruses represent expanding threats to sub-Saharan Africa imposing as considerable burden to human and veterinary public health. In Mozambique over one hundred species of potential arbovirus mosquito vectors have been identified, although their precise role in maintaining such viruses in circulation in the country remains to be elucidated. The aim of this study was to screen for the presence of flaviviruses, alphaviruses and bunyaviruses in mosquitoes from different regions of Mozambique. Results: Our survey analyzed 14,519 mosquitoes, and the results obtained revealed genetically distinct insect-specific flaviviruses, detected in multiple species of mosquitoes from different genera. In addition, smaller flavivirus-like NS5 sequences, frequently detected in Mansonia seemed to correspond to defective viral sequences, present as viral DNA forms. Furthermore, three lineages of putative members of the Phenuiviridae family were also detected, two of which apparently corresponding to novel viral genetic lineages. Conclusion: This study reports for the first-time novel insect-specific flaviviruses and novel phenuiviruses, as well as frequent flavivirus-like viral DNA forms in several widely known vector species. This unique work represents recent investigation of virus screening conducted in mosquitoes from Mozambique and an important contribution to inform the establishment of a vector control program for arbovirus in the country and in the region.publishersversionpublishe

Risk factors for occurrence and abundance of Aedes aegypti and Aedes bromeliae at hotel compounds in Zanzibar

BACKGROUND : A field survey was performed to investigate local environmental factors promoting occurrence and abundance of Aedes aegypti and Ae. bromeliae mosquitoes at hotel compounds in the south-east coastal region of Zanzibar Island. METHODS : The potential risk factors were determined using generalized linear mixed models. Aedes (Stegomyia) spp. indices such as container index (CI) and pupae per container (PPC) index were also estimated. RESULTS : Aedes aegypti and Ae. bromeliae were the most abundant vector species, accounting for 70.8% of all Aedes mosquitoes collected. The highest CI was observed for plastic containers irrespective of the season, whereas the highest PPC was observed for coconut shells and aluminium containers in the rainy and dry seasons, respectively. The risk of Aedes mosquito occurrence and abundance were significantly associated with presence of plastic containers, coconut shells, used tyres and steel containers. These were discarded in shaded places, in the open and gardens, or found in plant nurseries. CONCLUSION : This study shows that Aedes species of global health significance occur at hotel compounds on this part of Zanzibar Island. The occurrence and abundance are sustained by the presence of abundant and poorly managed solid wastes and containers used for gardening tasks. This highlights an urgent need for the adoption of area-wide environmentally sustainable Aedes mosquito management interventions that also integrate solid waste management and ornamental plant production practices for reducing the risk of arboviral disease epidemics.Additional file 1: Table S1. Relative abundance of Aedes mosquitos found at hotel compounds stratified according to larval habitat characteristics, that is, type of habitat, location, function, size, presence of vegetation, presence of organic matter and season.AK, MA, KS, CWW and PF conceived the study; AK, FS, FK, CWW, MA, KS and PF designed the study; AK, FS, KH and DLS performed and supervised field surveys and data collection; AK performed data analysis and wrote the first draft of the manuscript.Danida Fellowship Centre (Denmark’s Development Cooperation), as part of the EnSuZa project.http://www.parasitesandvectors.comam2022Zoology and Entomolog

implication for assessing the risk of arbovirus outbreaks

Funding Information: This work was also supported by the U.S. Centers for Disease Control and Prevention through a cooperative agreement number 5NU14GH001237-03-00. The views expressed in this written publication do not necessarily reflect the official policies of the U.S. Department of Health and Human Services. European Foundation Initiative into Neglected Tropical Disease also supported this study. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. We thank all community leaders, household heads staff from the provincial health directorates, staff from the Medical Entomology Laboratory from the National Institute of Health, staff from National Malaria Control Programme, and staff from the Laboratório de Transmissores de Hematozoários of the Oswaldo Cruz Institute for their support during field work, laboratory testing and identification of Aedes spp. We also thank José Feriano Américo who produced the maps of this work. Ours special thanks goes to Professor J. Dereck Charlwood who revised the English grammar and typos of the entire manuscript. Consent for publication Our manuscript does not present any individual person's data. Publisher Copyright: © 2018 Abílio et al. http://creativecommons.org/licenses/by/4.0/.Background: Aedes-borne arboviruses have emerged as an important public health problem worldwide and, in Mozambique, the number of cases and its geographical spread have been growing. However, information on the occurrence, distribution and ecology of Aedes aegypti and Ae. albopictus mosquitoes remain poorly known in the country. Methods: Between March and April 2016, a cross-sectional study was conducted in 32 districts in Mozambique to determine the distribution and breeding sites of Ae. aegypti and Ae. albopictus. Larvae and pupae were collected from a total of 2,807 water-holding containers using pipette, dipper, funnel and sweeping procedures, depending on the container type and location. Both outdoor and indoor water-holding containers were inspected. The immature forms were reared to adults and the identifications of the mosquito species was carried out with a stereomicroscope using a taxonomic key. Results: Aedes aegypti was found in every district sampled, while Ae. albopictus was only found in Moatize district, situated in Tete Province in the central part of the country. Six hundred and twenty-eight of 2,807 (22.4%) containers were positive for Ae. aegypti but only one (0.03%) was positive for Ae. albopictus. The Container Index (CI) of Aedes was highest in densely populated suburban areas of the central region (260/604; 43.0%), followed by suburban areas in northern areas (228/617; 36.9%) whilst the lowest proportion was found in urbanized southern areas (140/1586; 8.8%). The highest CI of Aedes was found in used tires (448/1268; 35.3%), cement tanks (20/62; 32.3%) and drums (21/95; 22.1%). Conclusion: Data from our study showed that Ae. aegypti is present nation-wide, since it occurred in every sampled district, whilst Ae. albopictus had a limited distribution. Therefore, the risk of transmission of dengue and chikungunya is likely to have been underestimated in Mozambique. This study highlights the need for the establishment of a national entomological surveillance program for Aedes spp. in Mozambique in order to gain a better understanding about vector bionomics and to support the development of informed effective vector control strategies.publishersversionpublishe

High DDT resistance without apparent association to kdr and Glutathione-Stransferase (GST) gene mutations in Aedes aegypti population at hotel compounds in Zanzibar

Global efforts to control Aedes mosquito-transmitted pathogens still rely heavily on insecticides. However, available information on vector resistance is mainly restricted to mosquito populations located in residential and public areas, whereas commercial settings, such as hotels are overlooked. This may obscure the real magnitude of the insecticide resistance problem and lead to ineffective vector control and resistance management. We investigated the profile of insecticide susceptibility of Aedes aegypti mosquitoes occurring at selected hotel compounds on Zanzibar Island. At least 100 adults Ae. aegypti females from larvae collected at four hotel compounds were exposed to papers impregnated with discriminant concentrations of DDT (4%), permethrin (0.75%), 0.05 deltamethrin (0.05%), propoxur (0.1%) and bendiocarb (0.1%) to determine their susceptibility profile. Allele-specific qPCR and sequencing analysis were applied to determine the possible association between observed resistance and presence of single nucleotide polymorphisms (SNPs) in the voltage-gated sodium channel gene (VGSC) linked to DDT/pyrethroid cross-resistance. Additionally, we explored the possible involvement of Glutathione-S-Transferase gene (GSTe2) mutations for the observed resistance profile. In vivo resistance bioassay indicated that Ae. aegypti at studied sites were highly resistant to DDT, mortality rate ranged from 26.3% to 55.3% and, moderately resistant to deltamethrin with a mortality rate between 79% to and 100%. However, genotyping of kdr mutations affecting the voltage-gated sodium channel only showed a low frequency of the V1016G mutation (n = 5; 0.97%). Moreover, for GSTe2, seven non-synonymous SNPs were detected (L111S, C115F, P117S, E132A, I150V, E178A and A198E) across two distinct haplotypes, but none of these were significantly associated with the observed resistance to DDT. Our findings suggest that cross-resistance to DDT/deltamethrin at hotel compounds in Zanzibar is not primarily mediated by mutations in VGSC. Moreover, the role of identified GSTe2 mutations in the resistance against DDT remains inconclusive. We encourage further studies to investigate the role of other potential insecticide resistance markers.This study forms a part of the EnSuZa project funded by the Ministry of Foreign Affairs of Denmark.https://journals.plos.org/plosntdsdm2022Zoology and Entomolog

Morphological and Molecular Characterization Using Genitalia and CoxI Barcode Sequence Analysis of Afrotropical Mosquitoes with Arbovirus Vector Potential

Funding Information: Eddyson Montalvo-Sabino was recipient of a grant from “Programa Nacional de Becas y Crédito Educativo” (PRONABEC), 2019—Beca Generacion del Bicentenario, from the “Ministerio de Educación” of Peru. A.P. Abilio was a recipient of a grant from Wellcome Trust (Grant WT087546MA) through SACIDS RVF and NPHI-Phase-II from the National Institute for Health of Mozambique through a cooperative agreement number [5NU14GH001237-03-00]. Marietjie Venter was a recipient of a sub-award from the Global Disease Detection Program, US-CDC award 5U19GH000571-02 with the NICD and University of Pretoria that funded vector surveillance in South Africa (2012–2015) and by the Cooperative Agreement Number (5 NU2GGH001874-02-00) with the University of Pretoria (2014–2017). Milehna M. Guarido received a studentship through this grant. A.P.G. Almeida has been a recipient of the Visiting Professor Programme by the University of Pretoria for the work in South Africa. This work received financial support from the Global Health and Tropical Medicine Center (GHTM|IHMT|NOVA), which is funded through FCT contract UID/Multi/04413/2013, Portugal. The findings and conclusions expressed in this manuscript are those of the author(s) and do not necessarily represent the official position of the funding agencies. Publisher Copyright: © 2022 by the authors.Potential arboviral Afrotropical mosquito vectors are underrepresented in public databases of CoxI barcode sequences. Furthermore, available CoxI sequences for many species are often not associated with voucher specimens to match the corresponding fine morphological characterization of specimens. Hence, this study focused on the characterization of Culicine mosquitoes from South Africa, Mozambique, and Angola and their classification using a complementary approach including a morphological analysis of specimens’ genitalia and phylogenetic study based on the analysis of CoxI barcode sequences using maximum likelihood and Bayesian phylogenetic inference methods, alongside Median-Joining Network and PCOORD analyses. Overall, 800 mosquitoes (652 males and 148 females) from 67 species, were analyzed. Genitalia from 663 specimens allowed the identification of 55 species of 10 genera. A total of 247 CoxI partial gene sequences corresponding to 65 species were obtained, 11 of which (Aedes capensis, Ae. mucidus, Culex andersoni, Cx. telesilla, Cx. inconspicuosus, Eretmapodites subsimplicipes, Er. quinquevittatus, Ficalbia uniformis, Mimomyia hispida, Uranotaenia alboabdominalis, and Ur. mashonaensis) are, to the best of our knowledge, provided here for the first time. The presence of Cx. pipiens ecotypes molestus and pipiens and their hybrids, as well as Cx. infula, is newly reported in the Afrotropical region. The rates of correct sequence identification using BOLD and BLASTn (≥95% identity) were 64% and 53%, respectively. Phylogenetic analysis revealed that, except for subgenus Eumelanomyia of Culex, there was support for tribes Aedini, Culicini, Ficalbiini, and Mansoniini. A divergence >2% was observed in conspecific sequences, e.g., Aedeomyia africana, Ae. cumminsii, Ae. unilineatus, Ae. metallicus, Ae. furcifer, Ae. caballus, and Mansonia uniformis. Conversely, sequences from groups and species complexes, namely, Ae. simpsoni, Ae. mcintoshi, Cx. bitaeniorhynchus, Cx. simpsoni, and Cx. pipiens were insufficiently separated. A contribution has been made to the barcode library of Afrotropical mosquitoes with associated genitalia morphological identifications.publishersversionpublishe