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Mosquitoes (Diptera: Culicidae) pose a significant threat to public health worldwide, especially in tropical and subtropical regions, where they act as primary vectors in transmission of infectious agents. In Peru, 182 culicid species have been identified and several species of the genus Culex are known to transmit arboviruses. However, knowledge of mosquito diversity and distribution remains limited, with many studies focusing on specific regions only. Here, we describe a new morphological variation of Cx. (Culex) coronator Dyar and Knab, 1906, and report the presence of Culex (Carrollia) bonnei Dyar, 1921 in the central region of Peru, Huanuco. Specimens were obtained through larvae collections and identified through morphologic characterization, including dissection of male genitalia, and molecular analyses. In total, 17 mosquitoes were analyzed, and the genitalia of the male specimens allowed the identification of Cx. coronator and Cx. bonnei. Partial sequences of the CoxI gene corresponding to these two species were obtained (N = 10). Phylogenetic analysis revealed that the sequences of Cx. coronator grouped in a monophyletic clade with sequences ascribed to other species corresponding to the subgenus Carrollia, while Cx. bonnei specimens formed a monophyletic clade with homologous sequences from GenBank. This study underscores the importance of continued efforts to study the diversity and distribution of mosquitoes in Peru, including their potential role as vectors of human pathogens, to underpin effective disease control and prevention strategies, highlighting the importance of a complemented morphological and molecular analysis.

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Mosquitoes are commonly identified to species level using morphological traits, but complementary methods for identification are often necessary when specimens are collected as immature stages, stored inadequately, or when delineation of species complexes is problematic. DNA-barcoding using the mitochondrial cytochrome c oxidase subunit 1 (COI) gene is one such tool used for the morphological identification of species. A comprehensive entomological survey of mosquito species in Mexico State identified by COI DNA barcoding and morphology is documented in this paper. Specimens were collected from all the physiographic provinces in Mexico State between 2017 and 2019. Overall, 2,218 specimens were collected from 157 localities representing both subfamilies Anophelinae and Culicinae. A species checklist that consists of 6 tribes, 10 genera, 20 subgenera, and 51 species, 35 of which are new records for Mexico State, is provided. Three hundred and forty-two COI sequences of 46 species were analysed. Mean intraspecific and interspecific distances ranged between 0% to 3.9% and from 1.2% to 25.3%, respectively. All species groups were supported by high bootstraps values in a Neighbour-Joining analysis, and new COI sequences were generated for eight species: Aedes chionotum Zavortink, Ae. vargasi Schick, Ae. gabriel Schick, Ae. guerrero Berlin, Ae. ramirezi Vargas and Downs, Haemagogus mesodentatus Komp and Kumm, Culex restrictor Dyar and Knab, and Uranotaenia geometrica Theobald. This study provides a detailed inventory of the Culicidae from Mexico State and discusses the utility of DNA barcoding as a complementary tool for accurate mosquito species identification in Mexico.

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BACKGROUND: Despite the importance of some species of Culex (Melanoconion) (Diptera: Culicidae) as vectors of several arboviruses that cause diseases in humans and other animals, there are few taxonomic studies focusing on species of the subgenus, especially providing morphological keys for species identification. RESULTS: Thirteen species of the Atratus Group of Culex (Melanoconion) were reviewed, five new species are described, and two taxonomic changes are proposed: Cx. (Mel.) exedrus Root, 1927 and Cx. (Mel.) loturus Dyar, 1925 are resurrected from synonymy with Cx. (Mel.) dunni Dyar, 1918 and Cx. (Mel.) zeteki Dyar, 1918, respectively. The Atratus Group now includes fourteen species: Cx. (Mel.) atratus Theobald, 1901; Cx. (Mel.) caribeanus Galindo & Blanton, 1954; Cx. (Mel.) columnaris Sá & Hutchings n. sp.; Cx. (Mel.) commevynensis Bonne-Wepster & Bonne, 1919; Cx. (Mel.) comptus Sá & Sallum n. sp.; Cx. (Mel.) dunni; Cx. (Mel.) ensiformis Bonne-Wepster & Bonne, 1919; Cx. (Mel.) exedrus; Cx. (Mel.) longisetosus Sá & Sallum n. sp.; Cx. (Mel.) longistylus Sá & Sallum n. sp.; Cx. (Mel.) loturus; Cx. (Mel.) spinifer Sá & Sallum n. sp.; Cx. (Mel.) trigeminatus Clastrier, 1970; and Cx. (Mel.) zeteki. Keys, descriptions and illustrations for the identification of the male, female, pupal and fourth-instar larval stages of each species are provided. The treatment of each species includes a complete synonymy, descriptions of available life stages, a taxonomic discussion, updated bionomics and geographical distribution, and a list of material examined. CONCLUSIONS: The taxonomy of the Atratus Group of Culex (Melanoconion) is updated, including descriptions of five new species. The number of valid species is greater than the number recognized in the previous taxonomic study of the group, increasing from seven to 14 species. Distributional and bionomical data are updated. Morphology-based identification keys for females, males, fourth-instar larvae and pupae provided in this study will facilitate species identification.

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Dengue, chikungunya and Zika are arboviruses transmitted by mosquitos of the genus Aedes and have caused several outbreaks in world over the past ten years. Morphological identification of mosquitos is currently restricted due to the small number of adequately trained professionals. We implemented a computational model based on a convolutional neural network (CNN) to extract features from mosquito images to identify adult mosquitoes from the species Aedes aegypti, Aedes albopictus and Culex quinquefasciatus. To train the CNN to perform automatic morphological classification of mosquitoes, we used a dataset that included 4,056 mosquito images. Three neural networks, including LeNet, AlexNet and GoogleNet, were used. During the validation phase, the accuracy of the mosquito classification was 57.5% using LeNet, 74.7% using AlexNet and 83.9% using GoogleNet. During the testing phase, the best result (76.2%) was obtained using GoogleNet; results of 52.4% and 51.2% were obtained using LeNet and AlexNet, respectively. Significantly, accuracies of 100% and 90% were achieved for the classification of Aedes and Culex, respectively. A classification accuracy of 82% was achieved for Aedes females. Our results provide information that is fundamental for the automatic morphological classification of adult mosquito species in field. The use of CNN's is an important method for autonomous identification and is a valuable and accessible resource for health workers and taxonomists for the identification of some insects that can transmit infectious agents to humans.

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Culex (Microculex) daumastocampa Dyar & Knab was originally described by Dyar and Knab (Proc US Nat Mus 35:53-70, 1908) from larvae collected at axils of bromeliads in Port San Felipe, Panama. Culex daumastocampa is found in Colombia, Venezuela, Panama, Suriname, Costa Rica, Nicaragua, Guatemala, and Mexico, although its presence had not been reported suggesting its northernmost distribution. In Mexico, the subgenus Microculex had included Cx. rejector Dyar and Knab, and Cx. imitator Theobald. However, after that collection specimens were re-examined along with other specimens collected during 2016 in Chiapas (all specimens are available in the Culicidae Collection of the Universidad Autonoma Agraria Antonio Narro Unidad Laguna [UAAAN-UL], Mexico). Culex daumastocampa is now reported for the first time in Mexico, Cx. rejector for Chiapas, and Cx. imitator removed from the checklist of previous reports as to be present in Mexico.

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Entre os múltiplos grupos que compõem as Seções Spissipes e Melanoconion do subgênero Melanoconion de Culex, Atratus e Educator apresentam espécies que estão envolvidas na transmissão de arbovírus ao ser humano e animais em ambientes rurais e silvestres. Porém, esses grupos exibem grande dificuldade na identificação das espécies, seja devido à grande semelhança morfológica entre seus representantes ou à falta de chaves de identificação atualizadas. Diante disso, objetivou-se realizar a revisão taxonômica das espécies pertencentes aos Grupos Atratus e Educator da Seção Melanoconion de Culex (Melanoconion). Para isso, foi realizado o estudo morfológico detalhado das diversas fases de desenvolvimento dos espécimes de interesse, verificada a existência de espécies a serem nomeadas e/ou espécies válidas colocadas na sinonímia de outras. Como resultado do estudo morfológico das espécies do Grupo Atratus, foram redefinidas oito espécies. Destas, duas foram validadas, pois estavam na sinonímia de outras e seis espécies desconhecidas da ciência foram descritas e serão formalmente nomeadas. Para o Grupo Educator, oito espécies foram descritas, sendo uma removida da sinonímia e três espécies desconhecidas da ciência foram descritas e serão formalmente nomeadas. Chaves de identificação ilustradas, com caracteres usados para o reconhecimento das espécies, foram elaboradas para todas as formas de desenvolvimento das espécies de ambos os grupos. Foram construídos mapas de distribuição geográfica das espécies, foram atualizados dados bionômicos, elaboradas ilustrações e pranchas fotográficas para comparação de estruturas morfológicas externas das formas imaturas, adultos e genitália masculina

Among the multiple groups that compose the Spissipes and Melanoconion Sections of the subgenus Melanoconion of Culex, the Atratus and Educator Groups present species that are involved in the transmission of arboviruses to man and animals in rural and wild environments. However, these groups present great difficulty in identifying the species, either because of the great morphological similarity between their representatives or the lack of updated identification keys. The aim of this study was to carry out a taxonomic review of the species belonging to the Atratus and Educator Groups of the Melanoconion Section of Culex (Melanoconion). Therefore, a detailed morphological study of the several stages of development of the specimens of interest was carried out, verifying the existence of species to be named and/or valid species placed in the synonymy of others. As a result of the morphological study of the Atratus Group species, eight species were redefined. Of these, two were resurrected from the synonymy of others and six species unknown to science were described and will be formally named. For the Educator Group, eight species have been described, one being removed from the synonymy and three species unknown to science have been described and will be formally named. Illustrated identification keys, with characters used for species recognition, have been created for all forms of development of species in both Groups. Geographic distribution maps of the species were created, illustrations and photographic plates for comparison of external morphological structures of the immatures forms, adults and male genitalia were provided, and bionomic data were updated

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BACKGROUND: Culex nigripalpus has a wide geographical distribution and is found in North and South America. Females are considered primary vectors for several arboviruses, including Saint Louis encephalitis virus, Venezuelan equine encephalitis virus and Eastern equine encephalitis virus, as well as a potential vector of West Nile virus. In view of the epidemiological importance of this mosquito and its high abundance, this study sought to investigate wing variation in Cx. nigripalpus populations from urban parks in the city of São Paulo, Brazil. METHODS: Female mosquitoes were collected in seven urban parks in the city of São Paulo between 2011 and 2013. Eighteen landmark coordinates from the right wing of each female mosquito were digitized, and the dissimilarities between populations were assessed by canonical variate analysis and cross-validated reclassification and by constructing a Neighbor-Joining (NJ) tree based on Mahalanobis distances. The centroid size was calculated to determine mean wing size in each population. RESULTS: Canonical variate analysis based on fixed landmarks of the wing revealed a pattern of segregation between urban and sylvatic Cx. nigripalpus, a similar result to that revealed by the NJ tree topology, in which the population from Shangrilá Park segregated into a distinct branch separate from the other more urban populations. CONCLUSION: Environmental heterogeneity may be affecting the wing shape variation of Cx. nigripalpus populations.

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Culex coronator Dyar and Knab and Culex usquatus Dyar belong to the Coronator Group of the subgenus Culex. Culex coronator and Cx. usquatus are widespread and sympatric throughout their distribution range, which includes Brazil. Morphological identification of these species is based primarily on the characteristics of the male genitalia; females are indistinguishable using the qualitative characteristics employed in identification keys. The primary objective of this study was to distinguish females of Cx. coronator from those of Cx. usquatus employing both wing geometric morphometrics, and DNA sequences (NADH5, COI, Hunchback, and CAD). Additionally, we employed the isolation with migration model (IMa) to evaluate: 1) the migration rates and 2) the divergence time, between Cx. coronator and Cx. usquatus. Specimens were captured in Pariquera-Açu and Cananéia south-eastern São Paulo, Ribeira Valley, Brazil. Canonical variate analysis (CVA) demonstrated two groups in the morphospace. The accuracy of species recognition was moderate (82.6%) for Cx. coronator and low (60.8%) for Cx. usquatus. Bayesian analyses of concatenated gene sequences recovered from specimens of Cx. coronator separated the species into three lineages (herein referred to as Culex coronator A, B, and C), whereas Cx. usquatus specimens clustered into a single lineage. Lineages A and B of Cx. coronator intermixed with specimens of Cx. usquatus, and one specimen of Cx. coronator clustered into the Cx. usquatus lineage. The IMa analysis indicated that the divergence of Cx. coronator and Cx. usquatus is a slow process, with some degree of gene flow between the two species.

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Culex bidens and C. interfor, implicated in arbovirus transmission in Argentina, are sister species, only distinguishable by feature of the male genitalia; however, intermediate specimens of the species in sympatry have been found. Fourth-instar larvae and females of both species share apomorphic features, and this lack of clear distinction creates problems for specific identification. Geometric morphometric traits of these life stages also do not distinguish the species. The aim of the present study was to assess the taxonomic status of C. bidens and C. interfor using two mitochondrial genes and to determine the degree of their reproductive isolation using microsatellite loci. Sequences of the ND4 and COI genes were concatenated in a matrix of 993 nucleotides and used for phylogenetic and distance analyses. Bayesian and maximum parsimony inferences showed a well resolved and supported topology, enclosing sequences of individuals of C. bidens (0.83 BPP, 73 BSV) and C. interfor (0.98 BPP, 97 BSV) in a strong sister relationship. The mean K2P distance within C. bidens and C. interfor was 0.3% and 0.2%, respectively, and the interspecific variation was 2.3%. Bayesian clustering also showed two distinct mitochondrial lineages. All sequenced mosquitoes were successfully identified in accordance with the best close match algorithm. The low genetic distance values obtained indicate that the species diverged quite recently. Most morphologically intermediate specimens of C. bidens from Córdoba were heterozygous for the microsatellite locus GT51; the significant heterozygote excess observed suggests incomplete reproductive isolation. However, C. bidens and C. interfor should be considered good species: the ventral arm of the phallosome of the male genitalia and the ND4 and COI sequences are diagnostic characters.

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INTRODUCTION: Mass production of mosquitoes under laboratory conditions allows implementing methods to control vector mosquitoes. Colony development depends on mosquito size and weight. Body size can be estimated from its correlation with wing size, whereas weight is more difficult to determine. Our goal was to test whether wing size can predict the weight. METHODS: We compared dry weight and wing centroid size of Culex quinquefasciatus reared at different temperatures and four diets. RESULTS: Weight and wing size were strongly correlated. The diets did not influence wing size. CONCLUSIONS: Wing centroid size is a good predictor of Cx. quinquefasciatus body weight.

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The present study aimed to determine the effects of seasonal variation on phenotypic variation in Culex bastagarius, using geometric morphometric analysis based on wing shape and size. Samples were collected in the Guapiaçu Ecological Reserve, in Rio de Janeiro, Brazil. Mosquitoes were captured once every 2 months (December 2012-January 2014) using light traps. The results of geometric morphometric analysis revealed variations in wing size; however, variability was not evident in wing shape. The present study provides novel data regarding phenotypic plasticity of Cx. bastagarius that have been scarcely addressed in the past.

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Male and female adults of Culex (Culex) articularis Philippi are redescribed, and the 4th-instar larva and pupa are described and illustrated for the first time. Culex articularis is compared with other species of the subgenus Culex. Illustrations of diagnostic characters of the female, male genitalia, 4th-instar larva, and pupa are also provided.

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BACKGROUND: The Coronator Group encompasses Culex coronator Dyar & Knab, Culex camposi Dyar, Culex covagarciai Forattini, Culex ousqua Dyar, Culex usquatissimus Dyar, Culex usquatus Dyar and Culex yojoae Strickman. Culex coronator has the largest geographic distribution, occurring in North, Central and South America. Moreover, it is a potential vector-borne mosquito species because females have been found naturally infected with several arboviruses, i.e., Saint Louis Encephalitis Virus, Venezuelan Equine Encephalitis Virus and West Nile Virus. Considering the epidemiological importance of Cx. coronator, we investigated the wing shape diversity of Cx. coronator from South and Southeast Brazil, a method to preliminarily estimate population diversity. METHODS: Field-collected immature stages of seven populations from a large geographical area in Brazil were maintained in the laboratory to obtain both females and males linked with pupal and/or larval exuviae. For each individual female, 18 landmarks of left wings were marked and digitalized. After Procrustes superimposition, discriminant analysis of shape was employed to quantify wing shape variation among populations. The isometric estimator centroid size was calculated to assess the overall wing size and allometry. RESULTS: Wing shape was polymorphic among populations of Cx. coronator. However, dissimilarities among populations were higher than those observed within each population, suggesting populational differentiation in Cx. coronator. Morphological distances between populations were not correlated to geographical distances, indicating that other factors may act on wing shape and thus, determining microevolutionary patterns in Cx. coronator. Despite the population differentiation, intrapopulational wing shape variability was equivalent among all seven populations. CONCLUSION: The wing variability found in Cx. coronator populations brings to light a new biological problem to be investigated: the population genetics of Cx. coronator. Because of differences in the male genitalia, we also transferred Cx. yojoae to the Apicinus Subgroup.

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Culex coronator Dyar & Knab e Culex usquatus Dyar são duas espécies irmãs, que fazem parte do Complexo Coronator, composto por mais quatro espécies (Culex usquatissimos Dyar, Culex ousqua Dyar, Culex camposi Dyar, Culex covagarciai Forattini). As fêmeas deste grupo são indistinguíveis por caracteres morfológicos, sendo a identificação possível somente através da distribuição e tamanho das cerdas apicais que ornamentam o gonocoxito da genitália masculina. Cx. coronator, é a espécie com maior distribuição geográfica, ocupando as Américas do Norte, Central e Sul. Já Cx. usquatus só foi registrado nas Américas Central e do Sul, ocorrendo em simpatria no Brasil com Cx. coronator. Apesar da semelhança morfológica das fêmeas das duas espécies, até o momento, somente Cx. coronator foi encontrado naturalmente infectado por diversas arboviroses. Considerando que estudos populacionais são importantes para compreender a evolução e a dinâmica de populações de pontencias vetores, e que a correta identificação de fêmeas é fundamental para estudos de competência vetorial, os objetivos deste trabalho foram: (1) distinguir fêmeas adultas de Cx. coronator de Cx. usquatus (2) obter conhecimento da estrutura populacional de Culex coronator nas regiões Sul e Sudeste (3) examinar a possível existência de outras espécies não descritas e/ou incorretamente identificadas sob o epíteto de Cx. coronator. Para tanto foram utilizadas duas ferramentas: uma morfológica (morfometria de asa), e outra genética...

Culex coronator and Culex usquatus are sibling species belonging to Coronator Group, which comprises five other species (Culex usquatissimos Dyar, Culex ousqua Dyar, Culex camposi Dyar, Culex covagarciai Forattini). Except by Cx. yojae, the females of this group are indistinguishable, being the identification only possible by the analysis of the arrangement and number of appendicles on the apical lobe of the gonocoxite of the male genitalia. Cx. coronator is the most widely distributed species in the complex, occupying North, Central and South America, while Cx. usquatus was recorded only in Central and South America. Therefore, these species are sympatric in Brazil. Despite the morphological similarity of the females of both species, only Cx. coronator has epidemiological importance, being found infected with many arboviruses. Since studies focusing in population structure are important to understand the evolution and dynamics of potencial vectors and that the correct female identification is critical for development of vectorial competence studies, the aims of this study were to: (1) distinguish adult females of Cx. coronator from Cx. usquatus (2) obtain knowledge of the population structure of Culex coronator in Southern and Southeastern areas (3) examine the presence of undescribed and/or incorrectly identified species under Cx. coronator. Thereby, a survey was carried out using morphological (wing geometry) and genetic (4 microsatellite loci and barcode region...

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Carbon dioxide (CO2) is an important component for activating and attracting host-seeking mosquitoes. The BG-Sentinel® trap is a well-established monitoring tool for capturing Culicidae, but CO2 role for the trap effectiveness has not been evaluated in highly urbanised areas. The objective was to evaluate the effectiveness of BG-Sentinel traps baited with and without CO2 for capturing urban mosquitoes. Fifteen areas were selected within the city of Manaus, Brazil, where four BG-Sentinels were operated for 24 h, two of them with CO2 and two without CO2. Captured Aedes aegypti females were dissected for the determination of their parity status. A significantly higher proportion of traps (from 32-79%) were positive for female Ae. aegypti when using the BG-Sentinel with CO2 (χ2 = 11.0271, p ≤ 0.001). Catches of female Culex spp were six times higher in CO2 traps (Mann-Whitney U test = 190.5; p = 0.001). Parity rates were similar for both traps. This study showed that CO2 has primarily an enhancing effect on the efficacy of BG-Sentinel for capturing Culex spp in Manaus. For Ae. aegypti, the positivity rate of the trap was increased, when CO2 was added.

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The purpose of this study was for the first time to establish phylogenetic hypotheses among Culex (Culex) species from Argentina, and to evaluate the relationships of this subgenus with subgenera Phenacomyia and Phytotelmatomyia. We employed maximum parsimony to analyse morphological characters derived from the morphology of the adult, pupal and larval stages. More than 30 figures, including photographs, illustrating 87 characters are provided. The cladistic analysis of 61 terminal taxa and 95 characters was undertaken, under implied weighting method, with a range of K values from 6 to 9. Three unstable taxa were detected therefore a reduced strict consensus tree was created. The subgenus Culex appears polyphyletic relative to subgenus Oculeomyia. The internal classification of subgenus Culex is shown to be chaotic; the Pipiens Group is polyphyletic relative to the Sitiens and Coronator Groups, the latter being polyphyletic with respect to the Pipiens Group. Phylogenetic analysis combining characters provided by morphology and molecular sources are needed to interpret more fully the relationships in the group.

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Sequences of the cytochrome c oxidase subunit I (COI) mitochondrial gene from adults of 22 Culex (Culex) species from Argentina and Brazil were employed to assess species identification and to test the usefulness of COI for barcoding using the best close match (BCM) algorithm. A pairwise Kimura two-parameter distance matrix including the mean intra and interspecific distances for 71 COI barcode sequences was constructed. Of the 12 COI lineages recovered in the Neighbour-joining topology, five confirmed recognised morphological species (Cx. acharistus, Cx. chidesteri, Cx. dolosus, Cx. lygrus and Cx. saltanensis) with intraspecific divergences lower than 1.75%. Cx. bilineatus is formally resurrected from the synonymy of Cx. dolosus. Cx. maxi , Cx. surinamensis and the Coronator group species included were clustered into an unresolved lineage. The intraspecific distance of Cx. pipiens (3%) was almost twice the interspecific between it and Cx. quinquefasciatus (1.6%). Regarding the BCM criteria, the COI barcode successfully identified 69% of all species. The rest of the sequences, approximately 10%, 18% and 3%, remained as ambiguously, mis and unidentified, respectively. The COI barcode does not contain enough information to distinguish Culex (Cux.) species.

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Sequences of the cytochrome c oxidase subunit I (COI) mitochondrial gene from adults of 22 Culex ( Culex ) species from Argentina and Brazil were employed to assess species identification and to test the usefulness of COI for barcoding using the best close match (BCM) algorithm. A pairwise Kimura two-parameter distance matrix including the mean intra and interspecific distances for 71 COI barcode sequences was constructed. Of the 12 COI lineages recovered in the Neighbour-joining topology, five confirmed recognised morphological species ( Cx. acharistus , Cx. chidesteri , Cx. dolosus , Cx. lygrus and Cx. saltanensis ) with intraspecific divergences lower than 1.75%. Cx. bilineatus is formally resurrected from the synonymy of Cx. dolosus . Cx. maxi , Cx. surinamensis and the Coronator group species included were clustered into an unresolved lineage. The intraspecific distance of Cx. pipiens (3%) was almost twice the interspecific between it and Cx. quinquefasciatus (1.6%). Regarding the BCM criteria, the COI barcode successfully identified 69% of all species. The rest of the sequences, approximately 10%, 18% and 3%, remained as ambiguously, mis and unidentified, respectively. The COI barcode does not contain enough information to distinguish Culex ( Cux. ) species.

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