Influence of Salmonella enterica Serovar Typhimurium ssrB on Colonization of Eastern Oysters (Crassostrea virginica) as Revealed by a Promoter Probe Screen

Abstract

cAlthough Salmonella has been isolated from 7.4 to 8.6% of domestic raw oysters, representing a significant risk for food-borne illness, little is known about the factors that influence their initial colonization by Salmonella. This study tested the hypothesis that specific regulatory changes enable a portion of the invading Salmonella population to colonize oysters. An in vivo promoter probe library screen identified 19 unique regions as regulated during colonization. The mutants in the nearest corresponding downstream genes were tested for colonization defects in oysters. Only one mutation, in ssrB, resulted in a significantly reduced ability to colonize oysters compared to that of wild-type Salmonella. Because ssrB regulates Salmonella pathogenicity island 2 (SPI-2)-dependent infections in vertebrate macrophages, the possibility that ssrB mediated colonization of oyster hemocytes in a similar manner was examined. However, no difference in hemocyte colonization was observed. The complementary hypothesis that signal exchange between Salmonella and the oyster's native microbial community aids colonization was also tested. Signals that triggered responses in quorum sensing (QS) reporters were shown to be produced by oyster-associated bacteria and present in oyster tissue. However, no evidence for signal exchange was observed in vivo. The sdiA reporter responded to salinity, suggesting that SdiA may also have a role in environmental sensing. Overall, this study suggests the initial colonization of live oysters by Salmonella is controlled by a limited number of regulators, including ssrB. Market surveys in the United States have isolated Salmonella in 7.4 to 8.6% of raw, market-ready oysters, with local contamination rates as high as 77% within a single sample (1-3). Because oysters are typically consumed raw or lightly cooked, they present a significant risk for exposure to live pathogens (4, 5). Oysters are capable of sustaining high filtration rates and are known to concentrate bacteria, including pathogens, present in the environment (6-8). Colonization is rapid; detectable contamination occurs in as little as 15 min following exposure to water containing Salmonella (9). Despite the clear evidence that Salmonella colonizes oysters, the mechanisms mediating Salmonella colonization of, and persistence inside, shellfish remain unclear.Oysters are typically colonized by a diverse microbial community, with 90% of the inhabiting bacteria being associated with the digestive tract. The composition of oyster microbiota is derived, but clearly differs, from the surrounding seawater and becomes progressively dissimilar from the seawater from the stomach to the lower intestine (10). This divergence in community structure indicates selection for certain species and suggests that the ability to resist shedding is responsible for the enhanced persistence. Experimental contamination of oysters by Salmonella typically results in a rapid initial colonization at a density 1-to 2-log-fold below that of the inoculum, followed by a ...

Improved Microbial Safety of Direct Ozone-Depurated Shellstock Eastern Oysters (Crassostrea virginica) by Superchilled Storage

“…However, little is currently known about how Salmonella interacts with marine invertebrates. According to Cox et al (2016) , the ssrB regulated effector could contribute to the fitness of Salmonella in oysters through regulatory mechanisms which are not currently understood. ssrB is required for intracellular survival in vertebrate macrophages.…”

Computational Identification of the Plausible Molecular Vaccine Candidates of Multidrug-Resistant Salmonella enterica

“…SsrB is responsible for the survival and replication of Salmonella in the host cell and plays an important role in the transcription of multiple genes of SPI-2. SsrB has been claimed as one of the most important factors for Salmonella's virulence by the fact that, a mutated ssrB, resulted in reduced ability of colonization on comparing with the wild type [70]. Moreover, one alteration in the gene ssrB, preferentially silencing the acquired DNA, can have a high contribution towards low transcription in the virulence factors of Salmonella [71].…”

Polymicrobial Interactions Operative during Pathogen Transmission

“…Human pathogens such as Vibrio vulnificus and V. parahaemolyticus are normal commensals of oysters (108,109). Further, interactions between normal commensals of oysters with S. enterica in vitro can activate quorum-sensing virulence regulons (110), providing a further mechanism for foodborne-associated illness from shellfish.…”