Application of Long-Read Sequencing for Genotyping, Epigenetic Profiling and Surveillance of Yersinia pestis Isolates from Natural Foci and Disease Outbreaks in Central Asia

INTRODUCTION: This study explores the application of long-read sequencing technologies for genotyping, epigenetic profiling, and epidemiological monitoring of Yersinia pestis isolates obtained from natural foci in Central Asia and previous zoonotic outbreaks. METHODS: Computational tools for genome assembly and genotyping were developed, enabling high-precision identification of both chromosomal and plasmid sequences, including the small plasmid pCKF. RESULTS: Genotyping based on genetic polymorphisms distinguished the major Y. pestis biovars and lineages, and revealed cluster-specific diversity among Medievalis (2.MED) isolates, including groups of strains associated with different plague foci and disease outbreaks in domestic animals and humans. These specific genomic polymorphisms identified in subclades of 2.MED isolates allow their high-precision identification. The distribution of pCKF-positive strains in the region and the potential involvement of these pathogens in disease outbreaks are discussed. Additionally, comparative epigenomic analysis uncovered strain-specific cytosine methylation patterns at cgGATCG motifs. Further studies involving more sequenced strains are needed to determine whether this cytosine methylation specificity is linked to genome function regulation and adaptation to different hosts and environments. CONCLUSIONS: These findings demonstrate the effectiveness of long-read sequencing technologies in revealing both genetic and epigenetic features of bacterial pathogens, contributing to our understanding of the evolutionary mechanisms underlying the emergence and spread of this especially dangerous infection.