Parasite specific 7SL-derived small RNA is an effective target for diagnosis of active trypanosomiasis infection.

Human and animal African trypanosomiasis (HAT & AAT, respectively) remain a significant health and economic issue across much of sub-Saharan Africa. Effective control of AAT and potential eradication of HAT requires affordable, sensitive and specific diagnostic tests that can be used in the field. Small RNAs in the blood or serum are attractive disease biomarkers due to their stability, accessibility and available technologies for detection. Using RNAseq, we have identified a trypanosome specific small RNA to be present at high levels in the serum of infected cattle. The small RNA is derived from the non-coding 7SL RNA of the peptide signal recognition particle and is detected in the serum of infected cattle at significantly higher levels than in the parasite, suggesting active processing and secretion. We show effective detection of the small RNA in the serum of infected cattle using a custom RT-qPCR assay. Strikingly, the RNA can be detected before microscopy detection of parasitaemia in the blood, and it can also be detected during remission periods of infection when no parasitaemia is detectable by microscopy. However, RNA levels drop following treatment with trypanocides, demonstrating accurate prediction of active infection. While the small RNA sequence is conserved between different species of trypanosome, nucleotide differences within the sequence allow generation of highly specific assays that can distinguish between infections with Trypanosoma brucei, Trypanosoma congolense and Trypanosoma vivax. Finally, we demonstrate effective detection of the small RNA directly from serum, without the need for pre-processing, with a single step RT-qPCR assay. Our findings identify a species-specific trypanosome small RNA that can be detected at high levels in the serum of cattle with active parasite infections. This provides the basis for the development of a cheap, non-invasive and highly effective diagnostic test for trypanosomiasis.

Induction of the cellular microRNA, Hs_154, by West Nile virus contributes to virus-mediated apoptosis through repression of antiapoptotic factors.

MicroRNAs (miRNAs) are a class of noncoding small RNAs that regulate multiple cellular processes, as well as the replication and pathogenesis of many DNA viruses and some RNA viruses. Examination of cellular miRNA profiles in West Nile virus (WNV)-infected HEK293 and SK-N-MC cells revealed increased expression of multiple miRNA species. One of these miRNAs, Hs_154, was significantly induced not only in WNV-infected neuronal cells in culture but also in the central nervous system tissues of infected mice and, upon transfection, caused a significant reduction in viral replication. Analysis of mRNA transcripts enriched through immunoprecipitation of the RNA-induced silencing complex identified several transcripts that contain seed sequence matches to Hs_154 in their 3' untranslated regions (UTRs). Two of these targets, the CCCTC-binding factor (CTCF) and the epidermal growth factor receptor (EGFR)-coamplified and overexpressed protein (ECOP/VOPP1) proteins display reduced expression in WNV-infected cells, and the 3' UTRs of these transcripts were sufficient to cause downregulation of expression in infected cells or in cells transfected with Hs_154, findings consistent with miRNA targeting of these transcripts. CTCF and ECOP have been shown to be associated with cell survival, implicating miRNA-directed repression of these targets in WNV-induced cell death. Consistent with this hypothesis, expression of these genes in WNV-infected cells results in a reduction in the number of cells undergoing apoptosis. These observations suggest that induction of Hs_154 expression after WNV infection modulates the apoptotic response to WNV and that cellular miRNA expression can be quickly altered during WNV infection to control aspects of the host response.