Loss of cAMP/CRP regulation confers extreme high hydrostatic pressure resistance in Escherichia coli O157:H7

Application of high hydrostatic pressure (HHP) constitutes a valuable non-thermal pasteurization process in modern food conservation. Triggered by our interest in the rapid adaptive evolution towards HHP resistance in the food-borne pathogen E. coli O157:H7 (strain ATCC 43888) that was demonstrated earlier, we used genetic screening to identify specific loci in which a loss-of-function mutation would be sufficient to markedly increase HHP survival. As such, individual loss of RssB (anti RpoS-factor), CRP (catabolite response protein) and CyaA (adenylate cyclase) were each found to confer significant HHP resistance in the 300 MPa range (i.e. > 1,000-fold), and this phenotype invariably coincided with increased resistance against heat as well. In contrast to loss of RssB, however, loss of CRP or CyaA also conferred significantly increased resistance to 600 MPa (i.e. > 10,000-fold), suggesting cAMP/CRP homeostasis to affect extreme HHP resistance independently of increased RpoS activity. Surprisingly, none of the rapidly emerging HHP-resistant mutants of ATCC 43888 that were isolated previously did incur any mutations in rssB, crp or cyaA, indicating that a number of other loci can guide the rapid emergence of HHP resistance in E. coli O157:H7 as well. The inability of spontaneous rssB, crp or cyaA mutants to emerge during selective enrichment under HHP selection likely stems from their decreased competitive fitness during growth. Overall, this study is the first to shed light on the possible genetic strategies supporting the acquisition of HHP resistance in E. coli O157:H7.