Viability of Yersinia pestis subcultures in agar‐stabs

Since its identification as the causative agent of plague in 1894, thousands of Yersinia pestisstrains have been isolated and stored. Here, we report the ability of Y. pestis to survive up to 47 years in agar stabs, in rubber-stoppered tubes, under refrigeration (+4 to +10°C), although overall subculturerecovery rates were poor and inversely relatedto the length of time stored. Genetic characterization of virulence gene presence among these subcultureswas suggestive of significant variation in the genomic stability of Y. pestis subculturesstored under these conditions. Specifically, we found variation in the presence of plasmid and chromosomal virulence markers (genes pla, lcrV, caf1 and irp2) among multiple subculturesof Y. pestis strains in the 'Collection of Yersinia pestis' (Fiocruz-CYP) maintained by the SRP of FIOCRUZ-PE in Brazil. This variation, together with all of the inherent temporal, geographic and other genetic variation represented by all of the recoverable strains in this historical collection was preserved in new frozen culture stocks stored at -70°C as a result of this study. These frozen culture stocks represent a valuable resource for future comparative studies of Y. pestis.We report the ability of Yersinia pestisto survive up to 47 years in agar stabs, in rubber-stoppered tubes, under refrigeration (+4 to +10°C), although overall subculturerecovery rates were poor and inversely relatedto the length of time stored. Genetic characterization of virulence gene presence among these subcultureswas suggestive of significant variation in the genomic stability of Y. pestis subculturesstored under these conditions. This variation, together with all of the inherent temporal, geographic and other genetic variation represented by all of the recoverable strains in the historical 'Collection of Yersinia pestis' (Fiocruz-CYP) maintained by the SRP of FIOCRUZ-PE in Brazil was preserved in new frozen culture stocks stored at -70°C as a result of this study. These frozen culture stocks represent a valuable resource for future comparative studies of Y. pestis.