Differential gene expression patterns of Yersinia pestis and Yersinia pseudotuberculosis during infection and biofilm formation in the flea digestive tract

Yersinia pestis, the etiologic agent of plague, emerged as a flea-borne pathogen only within the last 6,000 years. Just five simple genetic changes in the Yersinia pseudotuberculosis progenitor, which served to eliminate toxicity to fleas and to enhance survival and biofilm formation in the flea digestive tract, were key to the transition to the arthropod-borne transmission route. To gain a deeper understanding of the genetic basis for the development of a transmissible biofilm infection in the flea foregut, we evaluated additional gene differences and performed in vivo transcriptional profiling of Y. pestis, Y. pseudotuberculosis wild-type (unable to form biofilm in the flea foregut), and a Y. pseudotuberculosis mutant strain (able to produce foregut-blocking biofilm in fleas) recovered from fleas 1 day and 14 days after an infectious bloodmeal. Surprisingly, the Y. pseudotuberculosis mutations that increased c-di-GMP levels and enabled biofilm development in the flea did not change expression levels of the hms genes responsible for the synthesis and export of the extracellular polysaccharide matrix required for mature biofilm formation. The Y. pseudotuberculosis mutant uniquely expressed much higher levels of one of the Yersinia Type VI secretion systems (T6SS-4) in the flea, and this locus was required for flea blockage by Y. pseudotuberculosis, but not by Y. pestis. Significant differences between the two species in expression of several metabolism genes, the Psa fimbrial genes, quorum sensing related genes, transcriptional regulators, and stress response genes were evident during flea infection. The results provide insights into how Y. pestis has adapted to life in its flea vector and point to evolutionary changes in the regulation of biofilm development pathways in these two closely related species Yersinia strain comparisons