The small non-coding RNA response to virus infection in the Leishmania vector Lutzomyia longipalpis

Abstract

Sandflies are well known vectors for Leishmania but also transmit a number of arthropod-borne viruses (arboviruses). Few studies have addressed the interaction between sandflies and arboviruses. RNA interference (RNAi) mechanisms utilize small non-coding RNAs to regulate different aspects of host-pathogen interactions. The small interfering RNA (siRNA) pathway is a broad antiviral mechanism in insects. In addition, at least in mosquitoes, another RNAi mechanism mediated by PIWI interacting RNAs (piRNAs) is activated by viral infection. Finally, endogenous microRNAs (miRNA) may also regulate host immune responses. Here, we analyzed the small non-coding RNA response to Vesicular stomatitis virus (VSV) infection in the sandfly Lutzoymia longipalpis. We detected abundant production of virus-derived siRNAs after VSV infection in adult sandflies. However, there was no production of virus-derived piRNAs and only mild changes in the expression of vector miRNAs in response to infection. We also observed abundant production of virus-derived siRNAs against two other viruses in Lutzomyia Lulo cells. Together, our results suggest that the siRNA but not the piRNA pathway mediates an antiviral response in sandflies. In agreement with this hypothesis, pre-treatment of cells with dsRNA against VSV was able to inhibit viral replication while knock-down of the central siRNA component, Argonaute-2, led to increased virus levels. Our work begins to elucidate the role of RNAi mechanisms in the interaction between L. longipalpis and viruses and should also open the way for studies with other sandfly-borne pathogens.

Arboviruses and the Challenge to Establish Systemic and Persistent Infections in Competent Mosquito Vectors: The Interaction With the RNAi Mechanism

“…Since their involvement in antiviral defense was not clearly demonstrated (Varjak et al, 2018b), the function of vpiRNAs during viral infection remains to be elucidated. Their production may be a secondary effect of the expansion of PIWI genes in aedine and culicine mosquitoes since canonical vpiRNAs (with ping-pong signature) do not appear during arbovirus infections of anopheline mosquitoes, sandflies or midges that (presumably) have a more standard set of PIWI genes (Myles et al, 2009; Schnettler et al, 2013b; Ferreira et al, 2018; Dietrich et al, 2017b). The production of vpiRNAs in aedine and culicine mosquitoes occurs with specific patterns according to the type of virus (alphavirus, flavivirus, or bunyavirus) (Varjak et al, 2018b).…”

“…Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis reveals the identification of immune genes among different predicted cellular targets (Liu et al, 2015, 2016; Xing et al, 2016). However, in some studies, no significant changes in miRNA abundance are observed following arbovirus infection (Miesen et al, 2016b; Ferreira et al, 2018).…”

MicroRNAs: Biological Regulators in Pathogen–Host Interactions

“…Regarding the invertebrate host, phlebotomine sandflies are well recognized as vectors for species of Leishmania and a wide range of pathogens, such as bacteria and viruses, with the regulation of miRNAs favoring pathogen survival and transmission [94][95][96]. Yang and Wu discriminated miRNAs from other noncoding RNAs in Lutzomya longipalpis, a vector of leishmaniasis in the Americas and also suggest the role of miRNA targeting genes involved in Leishmania infection, such as llo-miR-9388-5p and llo-miR-3871-5p [97] in Phlebotomus perniciosus, a major vector of Leishmania infantum in Europe [98].…”

The insect reservoir of biodiversity for viruses and for antiviral mechanisms

“…It involves the RNaseIII enzyme Dicer-2, which senses replicating viral RNAs and processes them into 21 nucleotide (nt)-long siRNAs (reviewed in Paro et al 2015, Aguiar et al 2016). These siRNAs originate from both the genome and the antigenome of the virus, as observed during infection by the rhabdovirus Vesicular Stomatitis virus (VSV) in the model organism Drosophila melanogaster and the sandfly Lutzomyia longipalpis, which is one natural vector for this arthropod-borne virus ( Figure 2) (Mueller et al 2010, Marques et al 2013, Ferreira et al 2018. In Aedes mosquitoes, another RNAi pathway called piRNA pathway, which involves 24-29nt long Piwi-interacting (pi) RNAs, is also activated upon viral infections (Morazzani et al 2012, Aguiar et al 2015, Dietrich et al 2017.…”

“…siRNAs originate from both the viral genome and antigenome and can be detected along the entire extension of the virus sequence. Adapted fromFerreira et al 2018 andMueller et al 2010. An Acad Bras Cienc (2019) 91(Suppl. 3) e20190122 5 | 7…”