First report of an Onchocercidae worm infecting Psychodopygus carrerai carrerai sandfly, a putative vector of Leishmania braziliensis in the Amazon.

Sandflies are insects of public health interest due to their role as vectors of parasites of the genus Leishmania, as well as other pathogens. Psychodopygus carrerai carrerai is considered an important sylvatic vector of Leishmania (Viannia) braziliensis in Amazonia. In this study, sandflies were collected in a forested area in the Xapuri municipality, in the State of Acre (Northern Brazil). Two Ps. carrerai carrerai females were found parasitized with a larval form of a filarial worm, one in the labium of the proboscis, the other after the head was squashed, suggesting they were infective larvae. Sandflies were identified through morphological characters as well as amplification and sequencing of the cytochrome oxidase gene (COI). This was the first sequence obtained for Ps. carrerai carrerai for this marker. The obtained nematodes were also characterized through direct sequencing of a fragment of COI and 12S genes, both mitochondrial, and ITS1, a nuclear marker. Phylogenetic analyses revealed that the filarial nematodes belong to a species without sequences for these markers in the database, part of family Onchocercidade and closely related to genus Onchocerca (12S tree). Although sandfly infection with nematodes including members of the Onchocercidae has been reported in the Old World, this is the first report of sandfly infection by a member of the Onchocercidae family in the New World, to the best of our knowledge. Considering that the phylogenetic relationships and location in the insect, it can be expected that this is a parasite of mammals and the transmission cycle should be clarified.

Development of a molecular xenomonitoring protocol to assess filariasis transmission.

According to the World Health Organization, lymphatic filariasis (LF), a mosquito-borne neglected tropical disease (NTD), should be eliminated as a public health concern by the end of 2020. To this end, the goals of the Global Programme to Eliminate Lymphatic Filariasis (GPELF) include interrupting transmission through mass drug administration (MDA). After two decades, several countries have implemented MDA and are now ready to confirm whether transmission has been interrupted. The method for detecting the parasites in mosquito vectors known as xenomonitoring is a non-invasive tool for assessing the current transmission status of the filarial nematode Wuchereria bancrofti (which is responsible for 90% of cases) by their vectors. There are several methods available for detection of the worm in mosquito samples, such as dissection or polymerase chain reaction (PCR). However, most of these techniques still produce a considerable number of false-negative results. The present study describes a new duplex PCR protocol, which is an improvement on the traditional PCR methodology, enhanced by introducing the actin gene as an endogenous control gene. After adjusting the mosquito pool size, DNA extraction, and WbCx PCR duplex design, we achieved a reliable and sensitive molecular xenomonitoring protocol. This assay was able to eliminate 5% of false negative samples and detected less than one Wb larvae. This high sensitivity is particularly valuable after MDA, when prevalence declines. This new method could reduce the number of false-negative samples, which will enable us to improve our ability to generate accurate results and aid the monitoring strategies used by LF elimination programmes.

High prevalence and lack of diversity of Wolbachia pipientis in Aedes albopictus populations from Northeast Brazil.

The use of Wolbachia as a tool to control insect vectors has recently been suggested. In this context, studies on the prevalence and diversity of this bacterium in wild populations are relevant. Here, we evaluated the diversity of two Wolbachiagenes (ftsZ and wsp) and the prevalence of this endosymbiont in wild Aedes albopictus. Using semi-nested polymerase chain reaction, our results showed that 99.3% of the individuals were superinfected with Wolbachia. In regards to genetic diversity, the two genes showed no variation within or among mosquito populations. An analysis of other Wolbachia markers may help to clarify the relationship between insect and endosymbiont.

High prevalence and lack of diversity of Wolbachia pipientis in Aedes albopictus populations from Northeast Brazil

The use of Wolbachia as a tool to control insect vectors has recently been suggested. In this context, studies on the prevalence and diversity of this bacterium in wild populations are relevant. Here, we evaluated the diversity of two Wolbachiagenes (ftsZ and wsp) and the prevalence of this endosymbiont in wild Aedes albopictus. Using semi-nested polymerase chain reaction, our results showed that 99.3 percent of the individuals were superinfected with Wolbachia. In regards to genetic diversity, the two genes showed no variation within or among mosquito populations. An analysis of other Wolbachia markers may help to clarify the relationship between insect and endosymbiont.

Diversidade genética de Wolbachia Pipientis em populações de culicídeos vetores e no parasito Wuchereria Bancrofti / Genetic diversity of Wolbachia pipientis in populations of vector mosquitoes and the parasite Wuchereria bancrofti

A bactéria endossimbionte Wolbachia tem sido sugerida como mecanismo de controle de insetos vetores e alvo para o tratamento das filarioses. Nesse contexto, estudos sobre a prevalência de Wolbachia em populações de vetores de campo, a análise da sua densidade em insetos sob diferentes condições fisiológicas e a investigação da diversidade genética da bactéria dentro e entre diferentes populações de vetores e em pacientes filarêmicos são relevantes. O presente estudo teve como objetivos: determinar o percentual de infecção em Culex quinquefasciatus e Aedes albopictus coletados em diferentes áreas da RMR - PE - BR; descrever a diversidade dos genes ftsZ e wsp nas linhagens de Wolbachia encontradas tanto nos vetores citados quanto nos vermes filariais coletados de pacientes microfilarêmicos da região; como também comparar a densidade da bactéria em duas populações de C. quinquefasciatus do estado de PE, uma susceptível e outra resistente a temefós. Nossos resultados mostraram que o simbionte Wolbachia está presente em 100 por cento dos vetores analisados quando diagnosticados pelo método de PCR seminested, o qual se mostrou mais eficiente que a PCR convencional. Quanto à densidade do endossimbionte, as fêmeas de C. quinquefasciatus resistentes a organofosforados de Santa Cruz do Capibaribe, Sertão - PE, apresentaram a densidade média, cerca de sete vezes maior que às fêmeas susceptíveis de Peixinhos, RMR - PE, confirmando dados encontrados na literatura. Isto sugere que mosquitos resistentes apresentam maior dificuldade para controlar a densidade do endossimbionte provavelmente pelo custo biológico associado à resistência. Em relação à diversidade genética, os dois genes estudados não mostraram nenhuma variação nas populações observadas. A análise de outros marcadores pode ajudar a esclarecer a relação entre hospedeiros e endossimbionte dando maior suporte ao uso de Wolbachia no desenvolvimento de novas estratégias de controle e tratamento de doenças transmitidas por vetores.