Vertical Transmission of Zika Virus (Flaviviridae, Flavivirus) in Amazonian Aedes aegypti (Diptera: Culicidae) Delays Egg Hatching and Larval Development of Progeny.

Zika virus (ZIKV) has emerged as a globally important arbovirus and has been reported from all states of Brazil. The virus is primarily transmitted to humans through the bite of an infective Aedes aegypti (Linnaeus, 1762) or Aedes albopictus (Skuse, 1895). However, it is important to know if ZIKV transmission also occurs from Ae. aegypti through infected eggs to her offspring. Therefore, a ZIKV and dengue virus (DENV) free colony was established from eggs collected in Manaus and maintained until the third-fourth generation in order to conduct ZIKV vertical transmission (VT) experiments which used an infectious bloodmeal as the route of virus exposure. The eggs from ZIKV-infected females were allowed to hatch. The resulting F1 progeny (larvae, pupae, and adults) were quantitative polymerase chain reaction (qPCR) assayed for ZIKV. The viability of ZIKV vertically transmitted to F1 progeny was evaluated by cultivation in C6/36 cells. The effects of ZIKV on immature development of Ae. aegypti was assessed and compared with noninfected mosquitoes. AmazonianAe. aegypti were highly susceptible to ZIKV infection (96.7%), and viable virus passed to their progeny via VT. Moreover, eggs from the ZIKV-infected mosquitoes had a significantly lower hatch rate and the slowest hatching. In addition, the larval development period was slower when compared to noninfected, control mosquitoes. This is the first study to illustrate VT initiated by oral infection of the parental population by using mosquitoes, which originated from the field and a ZIKV strain that is naturally circulating in-country. Additionally, this study suggests that ZIKV present in the Ae. aegypti can modify the mosquito life cycle. The data reported here suggest that VT of ZIKV to progeny from naturally infected females may have a critical epidemiological role in the dissemination and maintenance of the virus circulating in the vector.

Zika virus transmission to mouse ear by mosquito bite: a laboratory model that replicates the natural transmission process.

BACKGROUND: Zika disease has transformed into a serious global health problem due to the rapid spread of the arbovirus and alarming severity including congenital complications, microcephaly and Guillain-Barré syndrome. Zika virus (ZIKV) is primarily transmitted to humans through the bite of an infective mosquito, with Aedes aegypti being the main vector. METHODS: We successfully developed a ZIKV experimental transmission model by single infectious Ae. aegypti bite to a laboratory mouse using circulating Brazilian strains of both arbovirus and vector. Mosquitoes were orally infected and single Ae. aegypti were allowed to feed on mouse ears 14 days post-infection. Additionally, salivary gland (SG) homogenates from infected mosquitoes were intrathoracically inoculated into naïve Ae. aegypti. Mosquito and mouse tissue samples were cultured in C6/36 cells and processed by quantitative real-time PCR. RESULTS: A total of 26 Ae. aegypti were allowed to feed individually on mouse ears. Of these, 17 mosquitoes fed, all to full engorgement. The transmission rate of ZIKV by bite from these engorged mosquitoes to mouse ears was 100%. The amount of virus inoculated into the ears by bites ranged from 2 × 102-2.1 × 1010 ZIKV cDNA copies and was positively correlated with ZIKV cDNA quantified from SGs dissected from mosquitoes post-feeding. Replicating ZIKV was confirmed in macerated SGs (2.45 × 107 cDNA copies), mouse ear tissue (1.15 × 103 cDNA copies, and mosquitoes 14 days post-intrathoracic inoculation (1.49 × 107 cDNA copies) by cytopathic effect in C6/36 cell culture and qPCR. CONCLUSIONS: Our model illustrates successful transmission of ZIKV by an infectious mosquito bite to a live vertebrate host. This approach offers a comprehensive tool for evaluating the development of infection in and transmission from mosquitoes, and the vertebrate-ZIKV interaction and progression of infection following a natural transmission process.

A competência vetorial do Aedes aegyptiassociada à genética populacional na cidade de Belo Horizonte, Minas Gerais

A Dengue é uma doença emergente, causada por quatro sorotipos do DENV (DENV-1, DENV-2, DENV-3 e DENV-4) pertencentes ao gênero Flavivirus e à família Flaviviridae. O vírus é transmitido pela picada de mosquitos fêmeas do gênero Aedes, sendo o Aedes aegypti o vetor primário. Estima-se 390 milhões de infecções anuais e 22.000 mortes, principalmente em crianças no mundo (WHO, 2016). A presença e a alta densidade dos vetores intensifica a necessidade de ações de controle. No entanto, um importante fato precisa ser considerado: as populações de mosquitos variam na sua permissividade para o desenvolvimento do flavivírus. A cidade de Belo Horizonte, alvo desse estudo, passou por 6 epidemias da doença nos últimos 20 anos (PBH, 2016). Os ovos dos Ae. aegypti de cada regional administrativa foram coletados pela Secretaria Municipal de Saúde separadamente e colonizados no insetário do LEM. Os adultos foram submetidos à extração de DNA, o qual foi amplificado para a sequência total de ITS. As análises filogenéticas foram realizadas com o software MEGA6. Em nosso estudo filogenético com as populações de Ae. aegypti de Belo Horizonte, observamos clusters melhores definidos e valores mais altos de bootstrap para três regionais: Oeste (O), Leste (L) e Nordeste (ND)

Essas foram selecionadas para as demais análises. Mosquitos fêmeas dessas regionais foram infectadas experimentalmente com DENV-2 e o vírus foi detectado através da RT-qPCR nas amostras dissecadas dos corpos e cabeças contendo as glândulas salivares. As análises das regionais ND, L e O revelou, em 7dpi taxas de infecção de 70, 80 e 70%, respectivamente. Em 14 dpi, as taxas foram de 80% para a regional ND e de 90% para as regionais L e O. A competência vetorial das três populações em estudo variou de 50 à 70%. Não observamos diferenças estatísticas entre nenhum os grupos Realizamos a análise do ITS dos indivíduos submetidos à infecção experimental. Não observamos o padrão de clusters encontrado anteriormente. No entanto, foi possível observar dois indivíduos negativos agrupados, ambos provenientes da regional nordeste, o que pode ser um fraco indicativo (devido ao número de amostras) de fenótipo característico para as amostras negativas