Background: Small non-coding RNAs play critical regulatory roles in post-transcription. However, their characteristics in Trichomonas vaginalis, the causative agent of human sexually transmitted trichomoniasis, still remain to be determined. Methods: Small RNA transcriptomes from Trichomonas trophozoites were deep sequenced using the Illumina NextSeq 500 system and comprehensively analyzed to identify Trichomonas microRNAs (miRNAs) and transfer RNA (tRNA)-derived small RNAs (tsRNAs). The tsRNA candidates were confirmed by stem-loop quantitative reverse transcription-PCR, and motifs to guide the cleavage of tsRNAs were predicted using the GLAM2 algorithm. Results: The miRNAs were found to be present in T. vaginalis but at an extremely low abundance (0.0046%). Three categories of endogenous Trichomonas tsRNAs were identified, namely 5′tritsRNAs, mid-tritsRNAs and 3′tritsRNAs, with the 5′tritsRNAs constituting the dominant category (67.63%) of tsRNAs. Interestingly, the cleavage site analysis verified both conventional classes of tRNA-derived fragments (tRFs) and tRNA-halves in tritsRNAs, indicating the expression of tRNA-halves in the non-stress condition. A total of 25 tritsRNAs were experimentally confirmed, accounting for 78.1% of all tested candidates. Three motifs were predicted to guide the production of tritsRNAs. The results prove the expression of tRFs and tRNA-halves in the T. vaginalis transcriptome. Conclusions: This is the first report of genome-wide investigation of small RNAs, particularly tsRNAs and miRNAs, from Trichomonas parasites. Our findings demonstrate the expression profile of tsRNAs in T. vaginalis, while miRNA was barely detected. These results may promote further research aimed at gaining a better understanding of the evolution of small non-coding RNA in T. vaginalis and their functions in the pathogenesis of trichomoniasis. Graphical

Plasmodium vivax remains the predominant species at the China–Myanmar border, imposing a major challenge to the recent gains in regional malaria elimination. To closely supervise the emerging of drug resistance in this area, we surveyed the variations in genes potentially correlated with drug resistance in P. vivax parasite and the possible drug selection with time. A total of 235 P. vivax samples were collected from patients suffering uncomplicated malaria at Yingjiang, Tengchong, and Longling counties, and Nabang port in China, Yunnan province, and Laiza sub-township in Myanmar, from 2008 to 2017. Five potential drug resistance genes were amplified utilizing nested-PCR and analyzed, including pvdhfr, pvdhps, pvmdr1, pvcrt-o, and pvk12. The Pearson’s Chi-squared test or Fisher’s exact test were applied to determine the statistical frequency differences of mutations between categorical data. The pvdhfr F57I/L, S58R, T61M and S117T/N presented in 40.6%, 56.7%, 40.1%, and 56.0% of the sequenced P. vivax isolates, and these mutations significantly decreased with years. The haplotype formed by these quadruple mutations predominated in Yingjiang, Tengchong, Longling and Nabang. While a mutation H99S/R (56.6%) dominated in Laiza and increased with time. In pvdhps, the A383G prevailed in 69.2% of the samples, which remained the most prevalent haplotype. However, a significant decrease of its occurrence was also noticed over the time. The S382A/C and A553G existed in 8.4% and 30.8% of the isolates, respectively. In pvmdr1, the mutation Y976F occurred at a low frequency in 5/232 (2.2%), while T958M was fixed and F1076L was approaching fixed (72.4%). The K10 insertion was detected at an occurrence of 33.2% in pvcrt-o, whereas there was no significant difference among the sites or over the time. No mutation was identified in pvk12. Mutations related with resistance to antifolate drugs are prevalent in this area, while their frequencies decrease significantly with time, suggestive of increased susceptibility of P. vivax parasite to antifolate drugs. Resistance to chloroquine (CQ) is possibly emerging. However, since the molecular mechanisms underneath CQ resistance is yet to be better understood, close supervision of clinical drug efficiency and continuous function investigation is urgently needed to alarm drug resistance.