BackgroundThe haemoflagellate Trypanosoma lewisi is a kinetoplastid parasite which, as it has been recently reported to cause human disease, deserves increased attention. Characteristic features of all kinetoplastid flagellates are a uniquely structured mitochondrial DNA or kinetoplast, comprised of a network of catenated DNA circles, and RNA editing of mitochondrial transcripts. The aim of this study was to describe the kinetoplast DNA of T. lewisi.Methods/ResultsIn this study, purified kinetoplast DNA from T. lewisi was sequenced using high-throughput sequencing in combination with sequencing of PCR amplicons. This allowed the assembly of the T. lewisi kinetoplast maxicircle DNA, which is a homologue of the mitochondrial genome in other eukaryotes. The assembly of 23,745 bp comprises the non-coding and coding regions. Comparative analysis of the maxicircle sequence of T. lewisi with Trypanosoma cruzi, Trypanosoma rangeli, Trypanosoma brucei and Leishmania tarentolae revealed that it shares 78 %, 77 %, 74 % and 66 % sequence identity with these parasites, respectively. The high GC content in at least 9 maxicircle genes of T. lewisi (ATPase6; NADH dehydrogenase subunits ND3, ND7, ND8 and ND9; G-rich regions GR3 and GR4; cytochrome oxidase subunit COIII and ribosomal protein RPS12) implies that their products may be extensively edited. A detailed analysis of the non-coding region revealed that it contains numerous repeat motifs and palindromes.ConclusionsWe have sequenced and comprehensively annotated the kinetoplast maxicircle of T. lewisi. Our analysis reveals that T. lewisi is closely related to T. cruzi and T. brucei, and may share similar RNA editing patterns with them rather than with L. tarentolae. These findings provide novel insight into the biological features of this emerging human pathogen.Electronic supplementary materialThe online version of this article (doi:10.1186/s13071-015-1281-8) contains supplementary material, which is available to authorized users.

Viral infections generally cause disease symptoms by interfering with the microRNA (miRNA)-mediated regulation of gene expression of host plants. In tomato leaves, the accumulation levels of eleven miRNAs and ten target mRNAs were enhanced by different degrees upon Cucumber mosaic virus (CMV)-Fny and Tomato aspermy virus (TAV)-Bj infections. The ability of CMV-Fny to interfere with miRNA pathway was dramatically suppressed in the addition of the benign satellite (sat) RNA variant (satYn12), but was slightly affected when CMV-Fny was co-inoculated with the aggressive satRNA variant (satT1). In plants harboring the infection of CMV-FnyΔ2b (a CMV-Fny 2b-deletion mutant), the unaltered miRNAs and target mRNAs levels compared with mock inoculated plants indicated that 2b ORF was essential for perturbation of miRNA metabolism. When the amounts of viral open reading frames (ORFs) in these infections were quantified, we found satYn12 caused a higher reduction of CMV-Fny accumulation levels than satT1. These results indicate the complex mechanism by which satRNAs participate in CMV-tomato interaction, and suggest that the severity of disease symptoms positively correlates to some extent with the perturbation of miRNA pathway in tomato.

MicroRNAs (miRNAs) are post‐transcriptional regulators that are involved in numerous biological processes in both plants and animals. In plants, root and stem tissues play essential roles in their anchorage to soil as well as in nutrient and water uptake and transfer. We have quantified the expression alterations of eleven miRNAs and their target mRNAs in tomato root and stem tissues after infection with Cucumber mosaic virus (CMV)‐Fny, CMV‐FnyΔ2b (a CMV‐Fny 2b‐deletion mutant), CMV‐Fny‐satT1 (CMV‐Fny plus an aggressive satellite (sat) RNA variant satT1) and Tomato aspermy virus (TAV)‐Bj. From 21 days postinoculation onwards, we found the stem tissues were ‘hollow’ in CMV‐Fny‐satT1 infected tomatoes, and a ‘fibrosis’ phenotype was observed in stems of TAV‐Bj infected plants. In addition, this phenotype was associated with the decreasing of tomato lateral root numbers upon the two infections. Consistence with these phenotypic alterations, tomato miRNA/mRNA levels upon different viral infections were changed by different degrees. Based on our results, the interference of different viral infections with tomato miRNA pathways were analysed and the correlations between miRNAs/mRNAs expression alterations and tomato symptoms were discussed.

The economic importance of Solanaceae plant species is well documented, and tomato has become a model for fleshy fruit development and ripening studies. Plant microRNAs (miRNAs) are small endogenous RNAs that are involved in a variety of activities including plant development, signal transduction and protein degradation, as well as response to environment stress and pathogen invasion. Here in this study, we aimed at quantifying the expression alterations of nine miRNAs and target mRNAs in tomato flower and fruit development upon Cucumber mosaic virus (CMV) and Tomato aspermy virus infections. Three different CMV strains CMV-Fny, CMV-FnyΔ2b and CMV-Fny-satT1 were used in our investigation, and the miRNA/mRNA expression alterations were analyzed by real-time quantitative RT-PCR. The results shown the levels of several miRNA/mRNA pairs were increased upon virus infections. However, the increased level of individual miRNA differed for different virus strains, reflecting differences in severity of symptom phenotypes. The altered expression patterns of these miRNA/mRNA pairs and their predicted functions indicate the possible roles in flower and fruit development, and provide experimental data for understanding the miRNA-mediated phenotype alterations in tomato fruit.

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Trypanosoma musculi is a, globally distributed, mouse-specific haemoflagellate, of the family Trypanosomatidae, which shares similar characteristics in morphology with Trypanosoma lewisi. The kinetoplast (mitochondrial) DNA of Trypanosomatidae flagellates is comprised of catenated maxicircles and minicircles. However, genetic information on the T. musculi kinetoplast remains largely unknown. In this study, the T. musculi maxicircle genome was completely assembled, with PacBio and Illumina sequencing, and the size was confirmed at 34 606 bp. It consisted of 2 distinct parts: the coding region and the divergent regions (DRs, DRI and II). In comparison with other trypanosome maxicircles (Trypanosoma brucei, Trypanosoma cruzi and T. lewisi), the T. musculi maxicircle has a syntenic distribution of genes and shares 73.9, 78.0 and 92.7% sequence identity, respectively, over the whole coding region. Moreover, novel insertions in MURF2 (630 bp) and in ND5 (1278 bp) were found, respectively, which are homologous to minicircles. These findings support an evolutionary scenario similar to the one proposed for insertions in Trypanosoma cruzi, the pathogen of American trypanosomiasis. These novel insertions, together with a deletion (281 bp) in ND4, question the role of Complex I in T. musculi. A detailed analysis of DRII indicated that it contains numerous repeat motifs and palindromes, the latter of which are highly conservative and contain A5C elements. The comprehensively annotated kinetoplast maxicircle of T. musculi reveals a high degree of similarity between this parasite and the maxicircle of T. lewisi and suggests that the DRII could be a valuable marker for distinguishing these evolutionarily related species.