The alarming escalation of infectious diseases resistant to conventional antibiotics requires urgent global actions, including the development of new therapeutics. Antimicrobial peptides (AMPs) represent potential alternatives in the treatment of multi-drug resistant (MDR) infections. Here, we focus on Cecropins (Cecs), a group of naturally occurring AMPs in insects, and on synthetic Cec-analogs. We describe their action mechanisms and antimicrobial activity against MDR bacteria and other pathogens. We report several data suggesting that Cec and Cec-analog peptides are promising antibacterial therapeutic candidates, including their low toxicity against mammalian cells, and anti-inflammatory activity. We highlight limitations linked to the use of peptides as therapeutics and discuss methods overcoming these constraints, particularly regarding the introduction of nanotechnologies. New formulations based on natural Cecs would allow the development of drugs active against Gram-negative bacteria, and those based on Cec-analogs would give rise to therapeutics effective against both Gram-positive and Gram-negative pathogens. Cecs and Cec-analogs might be also employed to coat biomaterials for medical devices as an approach to prevent biomaterial-associated infections. The cost of large-scale production is discussed in comparison with the economic and social burden resulting from the progressive diffusion of MDR infectious diseases.

The mosquito microbiota impacts the physiology of its host and is essential for normal larval development, thereby influencing transmission of vector-borne pathogens. Germ-free mosquitoes generated with current methods show larval stunting and developmental deficits. Therefore, functional studies of the mosquito microbiota have so far mostly been limited to antibiotic treatments of emerging adults. In this study, we introduce a method to produce germ-free Aedes aegypti mosquitoes. It is based on reversible colonisation with bacteria genetically modified to allow complete decolonisation at any developmental stage. We show that, unlike germ-free mosquitoes previously produced using sterile diets, reversibly colonised mosquitoes show no developmental retardation and reach the same size as control adults. This allows us to uncouple the study of the microbiota in larvae and adults. In adults, we detect no impact of bacterial colonisation on mosquito fecundity or longevity. In larvae, data from our transcriptome analysis and diet supplementation experiments following decolonisation suggest that bacteria support larval development by contributing to folate biosynthesis and by enhancing energy storage. Our study establishes a tool to study the microbiota in insects and deepens our knowledge on the metabolic contribution of bacteria to mosquito development.

The microbiota of Anopheles mosquitoes interferes with mosquito infection by Plasmodium and influences mosquito fitness, therefore affecting vectorial capacity. This natural barrier to malaria transmission has been regarded with growing interest in the last 20 years, as it may be a source of new transmission-blocking strategies. The last decade has seen tremendous progress in the functional characterisation of the tripartite interactions between the mosquito, its microbiota and Plasmodium parasites. In this review, we provide insights into the effects of the mosquito microbiota on Plasmodium infection and on mosquito physiology, and on how these aspects together influence vectorial capacity. We also discuss three current challenges in the field, namely the need for a more relevant microbiota composition in experimental mosquitoes involved in vector biology studies, for a better characterisation of the non-bacterial microbiota, and for further functional studies of the microbiota present outside the gut.

The analysis of environmental microbial communities has largely relied on a PCR-dependent amplification of genes entailing species identity as 16S rRNA. This approach is susceptible to biases depending on the level of primer matching in different species. Moreover, possible yet-to-discover taxa whose rRNA could differ enough from known ones would not be revealed. DNA-based methods moreover do not provide information on the actual physiological relevance of each taxon within an environment and are affected by the variable number of rRNA operons in different genomes. To overcome these drawbacks we propose an approach of direct sequencing of 16S ribosomal RNA without any primer- or PCR-dependent step. The method was tested on a microbial community developing in an anammox bioreactor sampled at different time-points. A conventional PCR-based amplicon pyrosequencing was run in parallel. The community resulting from direct rRNA sequencing was highly consistent with the known biochemical processes operative in the reactor. As direct rRNA-seq is based not only on taxon abundance but also on physiological activity, no comparison between its results and those from PCR-based approaches can be applied. The novel principle is in this respect proposed not as an alternative but rather as a complementary methodology in microbial community studies.

Arthropod-borne viruses (arboviruses) transmitted by Aedes aegypti mosquitoes are an increasing threat to global health. The small interfering RNA (siRNA) pathway is considered the main antiviral immune pathway of insects, but its effective impact on arbovirus transmission is surprisingly poorly understood. Here, we use CRISPR-Cas9-mediated gene editing in vivo to mutate Dicer2, a gene encoding the RNA sensor and key component of the siRNA pathway. The loss of Dicer2 enhances early viral replication and systemic viral dissemination of four medically significant arboviruses (chikungunya, Mayaro, dengue, and Zika viruses) representing two viral families. However, Dicer2 mutants and wild-type mosquitoes display overall similar levels of vector competence. In addition, Dicer2 mutants undergo significant virus-induced mortality during infection with chikungunya virus. Together, our results define a multifaceted role for Dicer2 in the transmission of arboviruses by Ae. aegypti mosquitoes and pave the way for further mechanistic investigations.