System of Yersinia pseudotuberculosis MLVA14-Typing Based on Multiple-Locus Variable-Number Tandem Repeat Analysis

Yersinia pseudotuberculosis bacterium is the etiologic agent of pseudotuberculosis, a natural-focal disease reported in various regions of the world. Strains of pseudotuberculosis microbe are characterized by significant genetic diversity and differ in pathogenicity and ability to cause group and sporadic diseases in humans. Since 2022, 1,090 cases of pseudotuberculosis infection have been identified in the Russian Federation; however, the incidence remains underestimated. Current advances in molecular-genetic technologies facilitate the development of highly effective methods for detecting and differentiating Y. pseudotuberculosis strains.
The aim of this study was to develop an effective MLVA14 typing system for intraspecific genetic differentiation of Y. pseudotuberculosis, based on multilocus VNTR analysis.
Materials and methods. This study involved molecular-genetic and phylogenetic assessment, as well as in silico O-genotyping of 94 Y. pseudotuberculosis strains based on their nucleotide sequences deposited in the NCBI RefSeq database.
Results and discussion. The serological affiliation, pathogenicity factors, and genetic group assignments have been determined. Phylogenetic relationships of the Y. pseudotuberculosis strains used in this study have been characterized, fourteen not previously described VNTR loci with distinct properties and discriminatory abilities identified. Flanking primers have been designed for the identified molecular targets using the Primer-BLAST web service. Applying in silico, MLVA typing of strains was performed and a cladogram of relationships was constructed, demonstrating that the developed MLVA14 method discriminates closely related Y. pseudotuberculosis strains into the same groups as whole-genome SNP analysis, but using a smaller number of molecular targets. The developed MLVA14 typing method is highly effective for intraspecific genetic differentiation of Y. pseudotuberculosis strains.

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