Analysis of the DNA region mediating increased thermotolerance at 58°C in Cronobacter sp. and other enterobacterial strains

Abstract

Cronobacter spp. are opportunistic pathogens associated with serious infections in neonates. The increased stress tolerance, including thermoresistance, of some Cronobacter strains can promote their survival in production facilities and thus raise the possibility of contamination of dried infant milk formula, which has been identified as a potential source of infection. In this study, we characterized a DNA region which is present in some Cronobacter strains and which contributes to their prolonged survival at 58°C. The 18 kbp long region containing 22 open reading frames was sequenced in Cronobacter sakazakii ATCC 29544. The major feature of the region contained a cluster of conserved genes, most of them having significant homologies with bacterial proteins involved in some type of stress response, including heat, oxidation and acid stress. The same thermoresistance DNA region was detected in strains belonging to the genera Cronobacter, Enterobacter, Citrobacter and Escherichia and its presence positively correlated with increased thermotolerance.

Contribution of the thermotolerance genomic island to increased thermal tolerance in Cronobacter strains

“…T possessed the highest thermotolerance of all the strains, which is in agreement with our previously reported results (Gajdosova et al 2011). Strains with the longer version of the island (C. sakazakii ATCC 29544 T , C. malonaticus LMG23826 T and NTU507) were 2-10 times more thermotolerant than strain C. sakazakii NTU701 containing the sequence shortened to the thrBCD and thrOP genes.…”

“…By transformation of an E. coli rpoS mutant with the thermotolerance island plasmids we observed increased thermotolerance in transformed strains but the cell survival was lower than that of the same constructs in wild-type E. coli (data not shown). This experiment confirmed our previous observations that genes from the thermotolerance island are expressed in both exponential and stationary phases but exponentially grown bacteria are not as tolerant as stationary bacteria (Gajdosova et al 2011;Orieskova et al 2013). Subsequently, a recombinant plasmid containing rpoS from E. coli under the control of the arabinose promoter was constructed and used to complement the C. sakazakii NTU696 strain.…”

“…The island from the C. sakazakii ATCC 29544 T type strain, which is 18 kbp long and harbours thrB-Q genes ( Fig. 1, Table S3), was described in our previous studies (Gajdosova et al 2011;Orieskova et al 2013). Recently, genomes of fourteen Cronobacter strains covering seven species were published ) and a comparison with C. sakazakii ATCC 29544…”

“…A protein marker unique for thermotolerant Cronobacter strains was later identified by mass spectrometry (Williams et al 2005). In our previous work, we found that this thermotolerance marker is encoded by a gene localised on a genomic island which contained a cluster of conserved genes encoding several stress response proteins (Gajdosova et al 2011). We observed that an isogenic mutant lacking the thermotolerance island was defective in survival at increased temperatures both in exponential and stationary growth phase (Orieskova et al 2013).…”

Cronobacter spp. – opportunistic food-borne pathogens. A review of their virulence and environmental-adaptive traits

“…during processing and reconstitution of PIF (Al-Nabulsi et al, 2011). Recently, an 18 kbp region containing 22 ORFs that are upregulated under heat adaptation conditions has been reported (Gajdosova et al, 2011). The major feature of the region is a cluster of conserved genes -most of them having significant homologies with known bacterial proteins involved in some type of stress response, including heat, oxidation and acid stress.…”