In this work, we describe the first Leishmania-infecting leishbunyavirus—the first virus other than Leishmania RNA virus (LRV) found in trypanosomatid parasites. Its host is Leishmania martiniquensis, a human pathogen causing infections with a wide range of manifestations from asymptomatic to severe visceral disease. This virus (LmarLBV1) possesses many characteristic features of leishbunyaviruses, such as tripartite organization of its RNA genome, with ORFs encoding RNA-dependent RNA polymerase, surface glycoprotein, and nucleoprotein on L, M, and S segments, respectively. Our phylogenetic analyses suggest that LmarLBV1 originated from leishbunyaviruses of monoxenous trypanosomatids and, probably, is a result of genomic re-assortment. The LmarLBV1 facilitates parasites’ infectivity in vitro in primary murine macrophages model. The discovery of a virus in L. martiniquensis poses the question of whether it influences pathogenicity of this parasite in vivo, similarly to the LRV in other Leishmania species.

Leishmania spp. are important pathogens causing a vector-borne disease with a broad range of clinical manifestations from self-healing ulcers to the life-threatening visceral forms. Presence of Leishmania RNA virus (LRV) confers survival advantage to these parasites by suppressing anti-leishmanial immunity in the vertebrate host. The two viral species, LRV1 and LRV2 infect species of the subgenera Viannia and Leishmania, respectively. In this work we investigated co-phylogenetic patterns of leishmaniae and their viruses on a small scale (LRV2 in L. major) and demonstrated their predominant coevolution, occasionally broken by intraspecific host switches. Our analysis of the two viral genes, encoding the capsid and RNA-dependent RNA polymerase (RDRP), revealed them to be under the pressure of purifying selection, which was considerably stronger for the former gene across the whole tree. The selective pressure also differs between the LRV clades and correlates with the frequency of interspecific host switches. In addition, using experimental (capsid) and predicted (RDRP) models we demonstrated that the evolutionary variability across the structure is strikingly different in these two viral proteins.

Trypanosoma cruzi dysregulates the gene expression profile of primary human cardiomyocytes (PHCM) during the early phase of infection through a mechanism which remains to be elucidated. The role that small non-coding RNAs (sncRNA) including PIWI-interacting RNA (piRNA) play in regulating gene expression during the early phase of infection is unknown. To understand how T. cruzi dysregulate gene expression in the heart, we challenged PHCM with T. cruzi trypomastigotes and analyzed sncRNA, especially piRNA, by RNA-sequencing. The parasite induced significant differential expression of host piRNAs, which can target and regulate the genes which are important during the early infection phase. An average of 21,595,866 (88.40%) of clean reads mapped to the human reference genome. The parasite induced 217 unique piRNAs that were significantly differentially expressed (q ≥ 0.8). Of these differentially expressed piRNAs, 6 were known and 211 were novel piRNAs. In silico analysis showed that some of the dysregulated known and novel piRNAs could target and potentially regulate the expression of genes including NFATC2, FOS and TGF-β1, reported to play important roles during T. cruzi infection. Further evaluation of the specific functions of the piRNAs in the regulation of gene expression during the early phase of infection will enhance our understanding of the molecular mechanism of T. cruzi pathogenesis. Our novel findings constitute the first report that T. cruzi can induce differential expression of piRNAs in PHCM, advancing our knowledge about the involvement of piRNAs in an infectious disease model, which can be exploited for biomarker and therapeutic development.

Trypanosoma cruzi, the causative agent of Chagas Disease, is an obligate intracellular parasite that has caused severe mortality and morbidity historically in Central and South America. Due to international travel, globalization, and migration, this neglected tropical disease is now present in most industrialized countries. About 30% of those who develop chronic T. cruzi infection will exhibit various pathologies, including cardiomyopathies, neurological and/or gastrointestinal diseases. We and others have implicated gene dysregulation as necessary in facilitating T. cruzi infection. However, the role of small noncoding RNAs during infection is still being investigated. Hence, the molecular mechanisms that contribute to chronic chagas cardiomyopathies remains to be elucidated. In this study, we aim to delineate dysregulation of a class of small non‐coding RNAs, piwi‐interacting RNAs (piRNAs), during the early phase of T. cruzi infection in primary human cardiomyocytes (PHCM). We hypothesize that dysregulated piRNAs target genes that may play important roles in infection. After parasite challenge, we extracted PHCM RNA for subsequent RNA sequencing. We then utilized bioinformatic tools in order to analyze differential expression of piRNAs, to predict mRNA targeting, and to map piRNAs to biological pathways. We found an average of 21,595,866 of clean reads (88.4%) mapped to the human reference genome. During the course of infection, T. cruzi induced differential expression of 217 unique piRNAs. In silico analysis predicted that both dysregulated known and novel piRNAs could target several genes of interest, including NFATC2, FOS, and TGF‐b1, and related genes known to interact with these targets. These novel findings are the first to implicate that Trypanosoma cruzi induces differential expression of piRNAs in PHCM, which advances our knowledge of piRNAs in infectious disease pathogenesis.