DNA Barcodes of Mansonia (Mansonia) Blanchard, 1901 (Diptera, Culicidae).

Females of the genus Mansonia feed on the blood of humans, livestock, and other vertebrates to develop their eggs. The females' biting behavior may cause severe disturbance to blood hosts, with a negative impact on public health and economics. Certain species have been identified as potential or effective disease vectors. The accurate species identification of field-collected specimens is of paramount importance for the success of monitoring and control strategies. Mansonia (Mansonia) morphological species boundaries are blurred by patterns of intraspecific heteromorphism and interspecific isomorphism. DNA barcodes can help to solve taxonomic controversies, especially if combined with other molecular tools. We used cytochrome c oxidase subunit I (COI) gene 5' end (DNA barcode) sequences to identify 327 field-collected specimens of Mansonia (Mansonia) spp. The sampling encompassed males and females collected from three Brazilian regions and previously assigned to species based on their morphological characteristics. Eleven GenBank and BOLD sequences were added to the DNA barcode analyses. Initial morphospecies assignments were mostly corroborated by the results of five clustering methods based on Kimura two-parameter distance and maximum likelihood phylogeny. Five to eight molecular operational taxonomic units may represent taxonomically unknown species. The first DNA barcode records for Mansonia fonsecai, Mansonia iguassuensis, and Mansonia pseudotitillans are presented.

Host Feeding Patterns of Mansonia (Diptera, Culicidae) in Rural Settlements near Porto Velho, State of Rondonia, Brazil.

Mosquito females of the genus Mansonia (Blanchard) can be a nuisance to humans and animals since they are voraciously hematophagous and feed on the blood of a variety of vertebrates. Despite their relevance, there is a lack of investigation into the blood-feeding patterns of the Mansonia species. Knowledge of the host preference is crucial in establishing the public health importance of a mosquito species and its potential to be involved in the transmission dynamics of pathogens. Species that are primarily anthropophilic can be more effective in spreading vector-borne pathogens to humans. In this study, we used an Illumina Nextera sequencing protocol and the QIIME2 workflow to assess the diversity of DNA sequences extracted in the ingested blood of mosquito species to evaluate the overall and local host choices for three species: Ma. titillans, Ma. Amazonensis, and Ma. humeralis, in rural areas alongside the Madeira River in the vicinities of the Santo Antonio Energia (SAE) reservoir in the municipality of Porto Velho, Rondônia, Western Brazil. By performing our analysis pipeline, we have found that host diversity per collection site showed a significant heterogeneity across the sample sites. In addition, in rural areas, Ma. amazonensis present a high affinity for B. taurus, Ma. humeralis shows an overall preference for C. familiaris and B. taurus, but also H. sapiens and E. caballus in urban areas, and Ma. titillans showed more opportunistic behavior in rural areas, feeding on wild animals and G. gallus, though with an overall preference for H. sapiens.

Biologia, ecologia e taxonomia molecular de Mansonia (Mansonia) Blanchard, 1901 (Diptera: Culicidae) / Biology, ecology and molecular taxonomy of Mansonia (Mansonia) Blanchard, 1901 (Diptera: Culicidae)

Introdução - As fêmeas do gênero Mansonia Blanchard, 1901 depositam seus ovos na face inferior das folhas de plantas aquáticas presentes em coleções de água doce. Após a eclosão, as larvas e, depois, as pupas se mantêm fixadas aos tecidos submersos das plantas. Assim que emergem, as fêmeas adultas hematófagas buscam, voraz e agressivamente, vertebrados que possam lhes servir como fontes de repasto sanguíneo. Esse comportamento pode prejudicar severamente a Saúde Pública e a economia em áreas com altas densidades populacionais de Mansonia spp. Para além do incômodo causado pelas picadas, algumas espécies transmitem patógenos aos humanos em certas regiões geográficas. Para que sejam eficazes, as medidas de monitoramento e controle das populações de Mansonia spp. devem ser planejadas com base na biologia de cada espécie e seus padrões de interação com fatores bióticos e abióticos nos ecossistemas. Dessa forma, a identificação acurada dos exemplares coletados em pesquisas de campo é fundamental, mas é dificultada pelos padrões de heteromorfismo intraespecífico e isomorfismo interespecífico historicamente observados. Objetivo - Adquirir conhecimentos inéditos para o planejamento de ações de monitoramento e controle populacional de Mansonia (Mansonia) spp. Métodos - Realizaram-se: ampla revisão bibliográfica narrativa; coletas de espécimes imaturos e adultos em três regiões do Brasil; identificação morfológica das amostras em nível específico; sequenciamento da região código de barras do gene mitocondrial codificador da subunidade I da enzima oxidase do citocromo c (COI); e análise das sequências por meio de cinco métodos diferentes, baseados em distâncias genéticas estimadas com o modelo Kimura dois-parâmetros (Neighbor joining, ABGD, ASAP e RESL) e em inferência filogenética de máxima verossimilhança (mPTP). Resultados - Os conteúdos bibliográficos examinados foram enfeixados em um único manuscrito que apresenta, em síntese, dados de Mansonia spp. sobre desenvolvimento, interações com hospedeiros, padrões alimentares, importância epidemiológica e adaptabilidade a ambientes antropogênicos, entre outros. A análise das amostras de imaturos do lago da Usina Hidrelétrica Santo Antônio revela evidências de alternância na associação com as diferentes espécies de macrófitas do local e o registro inédito de larvas de Mansonia humeralis associadas às raízes de Limnobium laevigatum. Trezentas e vinte e sete novas sequências da região código de barras do gene COI foram geradas a partir de exemplares de Mansonia (Mansonia) spp. A identificação baseada na morfologia apontou oito espécies na amostra de culicídeos. Os resultados dos cinco métodos implementados para a segregação das sequências em unidades taxonômicas operacionais (UTOs) foram majoritariamente congruentes. Os padrões de polimorfismo das sequências permitiram distinguir as oito espécies, corroborando a prévia identificação morfológica. No entanto, há evidências de que ao menos três dos táxons morfologicamente definidos podem representar complexos de espécies taxonomicamente desconhecidas. Foram obtidos os primeiros registros da região código de barras do gene COI para Mansonia fonsecai, Mansonia iguassuensis e Mansonia pseudotitillans. Conclusões - A reunião de informações referenciadas no manuscrito resultante da revisão bibliográfica facilitará a consulta para pesquisas futuras. A publicação dos dados inéditos sobre taxonomia molecular e associação com macrófitas aquáticas beneficiará o planejamento para o monitoramento e controle populacional de Mansonia spp.

Introduction - Females of the genus Mansonia Blanchard, 1901 lay their eggs on the underside of aquatic plant leaves in freshwater collections. After hatching, the larvae and later the pupae remain attached to the submerged plant tissues. As soon as they emerge, the hematophagous adult females search, voraciously and aggressively, for vertebrate sources of blood meal. In areas with high population densities of Mansonia spp. this behavior can severely harm Public Health and economics. In addition to the nuisance caused by bites, some species transmit pathogens to humans in certain geographic regions. To be effective, Mansonia spp. monitoring and control should be planned on the basis of the biology each species and its patterns of interaction with biotic and abiotic factors. Therefore, accurate field-collected specimens identification is essential, but it is hampered by historically observed patterns of intraspecific heteromorphism and interspecific isomorphism. Objective - To acquire unprecedented knowledge for planning actions to monitor and control population of Mansonia (Mansonia) spp. Methods - The following activities were carried out: a broad narrative bibliographical review; collections of immature and adult specimens in three Brazilian regions; morphological identification of samples at the species level; cytochrome c oxidase subunit I (COI) mitochondrial gene barcode region sequencing; and sequence analysis using five different methods, based on Kimura two-parameter genetic distances (Neighbor joining, ABGD, ASAP and RESL) and maximum likelihood phylogenetic inference (mPTP). Results - The bibliographic contents examined were assembled into a single manuscript that summarizes data on Mansonia spp. development, hosts, feeding patterns, epidemiological importance and adaptability to anthropogenic environments, among others. The analysis of immature samples from the Santo Antônio Hydroelectric Power Plant dam shows evidence of alternation in the attachment to host plant species and the unprecedented record of Mansonia humeralis larvae attached to Limnobium laevigatum roots. Three hundred and twenty-seven new COI barcode sequences were generated from Mansonia (Mansonia) spp. specimens. Morphology-based identification found eight species in the sample. The results for sequence segregation into operational taxonomic units (OTUs) were mostly congruent among five analysis methods. Sequence polymorphism patterns allowed the identification of eight species, corroborating the previous morphological identification. However, there is evidence that at least three of the morphology-based taxa may represent taxonomically unknown species complexes. The first records of COI barcodes for Mansonia fonsecai, Mansonia iguassuensis and Mansonia pseudotitillans were obtained. Conclusions - The referenced information gathered into the resulting manuscript from bibliographic review will contribute to future research. The publication of unprecedent data on molecular taxonomy and attachment to aquatic macrophytes will benefit the planning for Mansonia spp. monitoring and population control.

Mansonia spp. population genetics based on mitochondrion whole-genome sequencing alongside the Madeira River near Porto Velho, Rondonia, Brazil.

In high abundance, females of the genus Mansonia (Blanchard) can be a nuisance to humans and animals because they are voraciously hematophagous and feed on the blood of a myriad of vertebrates. The spatial-temporal distribution pattern of Mansonia species is associated with the presence of their host plants, usually Eichhornia crassipes, E. azurea, Ceratopteris pteridoides, Limnobium laevigatum, Pistia stratiotes, and Salvinia sp. Despite their importance, there is a lack of investigation on the dispersion and population genetics of Mansonia species. Such studies are pivotal to evaluating the genetic structuring, which ultimately reflects populational expansion-retraction patterns and dispersal dynamics of the mosquito, particularly in areas with a history of recent introduction and establishment. The knowledge obtained could lead to better understanding of how anthropogenic changes to the environment can modulate the population structure of Mansonia species, which in turn impacts mosquito population density, disturbance to humans and domestic animals, and putative vector-borne disease transmission patterns. In this study, we present an Illumina NGS sequencing protocol to obtain whole-mitogenome sequences of Mansonia spp. to assess the microgeographic genetic diversity and dispersion of field-collected adults. The specimens were collected in rural environments in the vicinities of the Santo Antônio Energia (SAE) hydroelectric reservoir on the Madeira River.

Efficient Monitoring of Adult and Immature Mosquitoes Through Metabarcoding of Bulk Samples: A Case Study for Non-Model Culicids With Unique Ecologies.

The rapid and economical monitoring of mosquitos is imperative to understanding the dynamics of both disease vectors and nuisance species. In light of technological advances in mosquito sampling and DNA sequencing, health agencies can now utilize the full potential of metabarcoding pipelines for rapid and standardizable surveillance. Here, we describe mosquito spatial and temporal variation, with particular focus on Mansonia Blanchard species, in the Madeira (Rondônia State) and the Ribeira (São Paulo) watersheds, Brazil using metabarcoding of the D2 rDNA marker. Sampling and molecular pipelines were used to evaluate the taxonomic contribution of mosquitos in pools of culicids collected en masse from macrophyte-roots (immatures) and from Mosquito Magnet traps and protected human landings (adults). Results for adult captures are comparable to morphological diagnoses and clarify previously unknown temporal and spatial species turnover. Metabarcoding of immature stages also confirmed the extent of the geographical distribution of some species and each taxon's association with macrophyte species. Given the benefits of metabarcoding, such as taxonomic acuity, high throughput processing, and objectivity, we suggest such techniques should be more fully incorporated into culicid monitoring schemes. The metabarcoding protocol described herein paired with standardized field sampling schemes, when used by mosquito monitoring professionals, offers substantial improvements in terms of practicality, speed and cost.