Differential Regulation ofSalmonellaTyphimurium Genes Involved in O-Antigen Capsule Production and Their Role in Persistence Within Tomato Fruit

Abstract

Enteric pathogens, including non-typhoidal Salmonella spp. and enterovirulent Escherichia coli, are capable of persisting and multiplying within plants. Yet, little is still known about the mechanisms of these interactions. This study identified the Salmonella yihT gene (involved in synthesis of the O-antigen capsule) as contributing to persistence in immature tomato fruit. Deletion of yihT reduced competitive fitness of S. enterica sv. Typhimurium in green (but not ripe, regardless of color) tomato fruit by approximately 3 logs. The yihT recombinase-based in vivo expression technology (RIVET) reporter was strongly activated in unripe tomato fruit, and fitness of the mutant inversely correlated with the level of the yihT gene expression. Expression of yihT in mature tomato fruit was low, and yihT did not affect competitive fitness within mature fruit. To better understand the molecular basis of the phenotype, behaviors of the yihT RIVET reporter and the yihT mutant were tested in tomato fruit defective in ethylene signaling. These experiments suggest a role for functional ethylene-mediated signaling in the persistence of Salmonella spp. within tomato fruit. Furthermore, jasmonic acid and its precursors strongly reduced expression of yihT.Over the past decade, at least a dozen major outbreaks of salmonellosis have been traced to the consumption of fresh fruit and vegetables (Batz et al. 2011;Mandrell 2009). By some estimates, the burden of the non-typhoidal Salmonella infections contracted as a result of raw produce consumption is comparable with the burden of salmonellosis caused by foods of animal origin (Batz et al. 2011). In the aftermath (and, likely, as a consequence) of each produce-linked outbreak of gastroenteritis, the commodity price drops dramatically ). Initially, it was hypothesized that these outbreaks are a result of poor sanitation during various production stages. However, produce-linked outbreaks still occur even after significant science-driven improvements in pre-and postharvest handling of fresh produce. It is now well accepted that, under suitable conditions, enteric pathogens are capable of colonizing and persisting within various plant tissues in the field (Danyluk et al. 2008;Greene et al. 2008;Uesugi et al. 2007), possibly as a part of their normal lifecycle (Brandl et al. 2013). Furthermore, growth within plants quickly selects for variants and spontaneous mutants of both Salmonella enterica and enterovirulent Escherichia coli that are more fit for habitation in plant tissues (Parker et al. 2012;Zaragoza et al. 2012). Despite the apparent importance of the plant-associated lifestyle in the ecology of enterics, relatively little is still known about mechanisms of interactions between enterics and plants, and even less is known about the genetics of Salmonella-tomato interactions.Colonization of tomato surfaces and vegetative and reproductive tissues depends on specific Salmonella genes and is a function of the tomato genotype and the physiological state of the plant and specific tissues...

Salmonella Persistence in Tomatoes Requires a Distinct Set of Metabolic Functions Identified by Transposon Insertion Sequencing

“…The capacity of S. enterica (and other enteric pathogens) to readily multiply within plant tissues led to the hypothesis that persistence on plants is a part of the Salmonella life cycle, serving as reservoirs prior to reinfection of the preferred animal hosts (2)(3)(4)(5). Several studies have dissected the molecular basis of plant-Salmonella interactions (6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16). However, the entire complement of genetic functions required for plant colonization by Salmonella is not yet known.…”

“…It involves major changes in the bacterial transcriptome and requires genes involved in amino acid biosynthesis and transport, cellulose production, fimbriae, regulators, and surface structures (6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17). Some of the same mechanisms were involved in the colonization of both vegetative and reproductive tissues on different plant species.…”

“…Colonization of the tomato pericarp involves differential regulation of at least 50 genes (9), some of which were also required for proliferation, including the gene cysB, encoding a cysteine metabolism regulator (9). Surface structures, like the O-antigen capsule and curli fimbriae, were also required for the colonization of tomatoes, as indicated by reduced fitness of the corresponding Salmonella mutants (11,17). The O-antigen capsule also plays a role in the colonization of alfalfa sprouts by Salmonella spp., as do curli fimbriae, suggesting that these structures may be broadly required for the colonization and/or persistence within plants (7,14).…”

Acquisition of Iron Is Required for Growth of Salmonella spp. in Tomato Fruit

“…A genome-wide screen using Salmonella enterica serovar Typhimurium identified a number of metabolic and regulatory genes induced during growth in tomatoes but did not demonstrate genes required for bacterial survival in this environment (20). Further work in this area showed that the O-antigen portion of bacterial lipopolysaccharide is importance for Salmonella survival in tomatoes (21), as it is in animal hosts.…”

Salmonella and Tomatoes

“…The FabH gene was upregulated most strongly in immature tomatoes, probably in response to changes in the concentrations of linoleic acid. Deletion of SirA and MotA (involved in motility) modestly increased fitness, and deletion of YihT (involved in synthesis of the capsule) contributed to fitness in green but not ripe tomatoes, but interestingly, known Salmonella genes associated with motility and virulence in animals such as hila, flhDC, and fliF did not contribute to the fitness of the bacteria in the fruit (Marvasi et al, 2013; Noel et al, 2010). …”