Downloaded from genesdev.cshlp.org on October 7, 2021 - Published by Cold Spring Harbor Laboratory Press REVIEW Survival in a cruel world: how Vibrio cholerae and Salmonella respond to an unwilling host James Slauch, 1 Ronald Taylor, 2 and Stanley Maloy 1,3 1Department of Microbiology, University of Illinois, Urbana, Illinois 61801 USA; 2Department of Microbiology, Dartmouth Medical School, Hanover, New Hampshire 03755-3842 USA The stressful life of an enteric pathogen tolerate the low pH in the stomach, flagella and chemo- taxis proteins that allow it to migrate to a suitable niche, Enteric pathogens are exposed to a variety of harsh con- adhesins that allow it to colonize the intestine, and tox- ditions: extremes of temperature and pH, strong deter- ins or invasins that allow it to elicit disease. Extracellu- gents and digestive enzymes, competition with an estab- lar pathogens like Vibrio cholerae remain in the lumen lished community of microorganisms, and the host im- of the intestine and produce toxins that cause the disease mune response. To survive, pathogens must express a symptoms. Intracellular pathogens like Salmonella pro- variety of gene products that elude these onslaughts. Of- duce invasins that allow it to penetrate mucosal cells. ten these conditions act as signals to turn on or off the Specific gene products must be turned on to adapt to specific genes needed for adaptation to distinct microen- vironments in the host. each of these conditions, and simultaneously certain other gene products must be turned off. This finely tuned The invading bacterium must be able to distinguish ability to turn "virulence genes" on and off in response and adapt rapidly to differences in growth conditions be- to different conditions in the host is essential for patho- tween the outside environment and the host, and within genesis. different microenvironments in the host. Different mi- croenvironments may have distinct temperatures, oxida- tion-reduction potentials, pH, and concentrations of in- What is a virulence gene? organic and organic nutrients. For example, before inges- tion intestinal pathogens are often present in water at a Virulence factors are those properties of an organism low temperature, low osmotic strength, near-neutral pH, that allow a pathogen to infect a host and cause a disease. and with a low concentration of organic nutrients. In Virulence depends upon numerous cell properties: both contrast, once ingested by a mammalian host the patho- housekeeping functions required for growth and replica- gen must adapt to a higher temperature and higher os- tion in the host, as well as functions specifically in- motic strength. It is transiently exposed to low pH in the volved in pathogenesis. Thus, in a broad sense, any bac- stomach, then within several hours after ingestion it terial property that is required for entry, growth, or sur- passes to the intestine where it encounters a higher pH, vival of a bacterium in the host may be considered a high concentrations of bile salts, anaerobiosis, and abun- virulence factor. Virulence factors include properties of dant organic nutrients. In addition, essentially all the the bacterial cell surface that prevent killing by comple- iron in the host is tightly bound to proteins such as lac- ment, adhesins that allow the bacterial cell to adhere to toferrin and transferrin, and it is actively sequestered by host cells, the ability to adapt to the acidic conditions in the host during infection. The ability to acquire iron un- the host, and the ability to synthesize nutrients that are der these conditions is essential for growth of most limiting in the host. For example, the aroA gene product, pathogens. If it survives these obstacles, the pathogen which is required for aromatic amino acid biosynthesis, must then make intimate contact with appropriate host is essential for virulence of Salmonella presumably be- cells upon arriving in the lumen of the small bowel to cause aromatic amino acids are limiting in the intracel- stably colonize the intestine. lular environment (Stocker 1993), an insight that led to To survive this journey the pathogen must rapidly ex- the development of a live, attenuated vaccine for Salmo- press a variety of gene products: iron chelators that allow nella typhi. it to scavenge the limiting iron, proteins that allow it to Genes that encode virulence factors are often called virulence genes. When virulence genes are lost by muta- tion or segregation there is a considerable decrease in 3Corresponding author. virulence. It has been estimated that 5%-10% of the E-MAIL [email protected]; FAX (217) 244-6697. genes in both V. cholerae and Salmonella typhimurium GENES & DEVELOPMENT 11:1761-1774 9 1997by Cold Spring Harbor Laboratory Press ISSN 0890-9369/97 $5.00 1761 Downloaded from genesdev.cshlp.org on October 7, 2021 - Published by Cold Spring Harbor Laboratory Press Slauch et al. encode virulence factors. Many virulence genes are lo- virulence indirectly. Mutations that affect chromosome cated on mobile genetic elements or in localized regions structure may fall into this category. DNA supercoiling of the chromosome called "pathogenicity islands" has been reported to be a common feature in the regula- {Groisman and Ochman 1996; Hacker et al. 1997). Typi- tion of many virulence genes (e.g., see Galan and Curtiss cally, pathogenicity islands are present in the genome of 1990; Parsot and Mekalanos 1992). This is not surprising pathogenic strains but absent from closely related non- because changes in DNA supercoiling affect many im- pathogenic strains. The G + C content of these regions is portant processes, including DNA replication, recombi- often very different from that of the rest of the genome, nation, and transcription. The extent of supercoiling suggesting that they were inherited by horizontal gene may be modulated by a variety of environmental factors transfer. They are often large regions that include mul- such as temperature, anaerobiosis, and osmolarity con- tiple virulence genes. For example, the SPI I pathogenesis ditions, which are likely to be experienced by pathogenic island in S. typhimurium is -40 kb and has a G + C content bacteria during infection (for review, see Dorman 1991, of 40%-47% relative to an average G + C content of 52%. 1996). Thus, it is possible that the level of supercoiling may be involved in coordinating expression of a variety Regulation of virulence genes of pleiotropic virulence genes. However, there remain many unanswered questions about the extent and Not surprisingly, expression of virulence genes is regu- mechanism of this regulation. One problem is that cer- lated by a diversity of environmental signals (Mekalanos tain promoters are sensitive to supercoiling only when 1992). Furthermore, virulence genes are commonly regu- maintained in the correct DNA context, and transcrip- lated by multiple conditions, allowing different genes to tion of adjacent genes can also affect the extent of local respond to several different environmental signals indi- supercoiling. Thus, potential differences between local vidually or only when multiple signals occur at the same and global supercoiling may make it difficult to interpret time (Tables 1 and 2). results obtained when genes are cloned onto plasmids to Many virulence genes are regulated by global regula- study the effect of supercoiling. A second problem is that tory systems, that is, regulatory systems that control ex- supercoiling increases or decreases the expression of pression of multiple genes simultaneously. For example, many genes that have no known role in virulence; esti- a common signal for inducing virulence gene expression mates suggest that -50% of all genes are affected by su- in V. cholerae and Salmonella is lack of iron availability, percoiling. Bacterial histone-like proteins may also affect which stimulates derepression or activation of transcrip- virulence indirectly as a result of changes in chromo- tion by the global regulatory protein Fur. some structure (e.g., see Higgins et al. 1990). Even if su- Two-component signal transduction systems play a percoiling and histone-like proteins play a direct role in major role in regulation of virulence genes (Dziejman the global regulation of virulence genes, most virulence and Mekalanos 1995). These systems typically have a genes clearly are regulated by other systems as well. membrane-associated sensor module that responds to an extracellular signal and transmits the signal to an intra- cellular response regulator module, which carries out the Identifying virulence genes adaptive response, usually by directly binding DNA and regulating gene expression (Parkinson 1995). Most often A variety of approaches have been used to identify viru- the sensor and receiver modules are distinct proteins. lence genes (Maloy et al. 1996). The brute force approach However, a variety of alternatives exist as well. For ex- is to isolate a large collection of null mutants and screen ample, the toxR gene from V. cholerae encodes a single them individually for loss of virulence in tissue culture integral membrane protein that may function as both a cells or animal models (Fields et al. 1986). Signature tag sensor and transcriptional activator (DiRita 1995). mutagenesis is an alternative approach for identifying Because virulence relies on so many different genes mutants that fail to survive in the host. This approach and regulatory signals, mutations that compromise the involves mutagenesis with transposons carrying unique ability