Leptospira transcriptome sequencing using long-read technology reveals unannotated transcripts and potential polyadenylation of RNA molecules

Leptospirosis, caused by the spirochete bacteria Leptospira, is an emerging zoonosis that causes life-threatening disease in humans and animals. However, a comprehensive understanding of leptospiral RNA profiles is limited. In this study, we sequenced and analyzed the transcriptome of multiple Leptospira strains using Oxford Nanopore Technologies' direct cDNA and direct RNA sequencing methods. We identified new operons, RNA molecules, and evidence of potential posttranscriptional polyadenylation in Leptospira transcriptomes. Some RNA molecules that had not been previously annotated could be potential sRNA or noncoding RNA molecules that play a role in gene expression regulation in Leptospira. Interestingly, the majority of the new RNA molecules identified in this study were not detected in the nonpathogenic Leptospira, suggesting potential virulence-related functions for these molecules. Overall, our study highlights the utility of Oxford Nanopore Technologies's sequencing in studying prokaryotic transcriptome profiles and offers a tool to improve our understanding of prokaryotic RNA landscapes and polyadenylation. Nonetheless, the findings from our study also warrant that the presence of homopolymers of adenine bases in the transcripts may interfere with the interpretation of bacterial transcriptome profiles. Carefully designed experiments are needed to explore the role of the features described in this study in Leptospira virulence and pathogenesis.