Bile-induced DNA damage in Salmonella enterica - PubMed (original) (raw)

Bile-induced DNA damage in Salmonella enterica

Ana I Prieto et al. Genetics. 2004 Dec.

Abstract

In the absence of DNA adenine methylase, growth of Salmonella enterica serovar Typhimurium is inhibited by bile. Mutations in any of the mutH, mutL, and mutS genes suppress bile sensitivity in a Dam(-) background, indicating that an active MutHLS system renders Dam(-) mutants bile sensitive. However, inactivation of the MutHLS system does not cause bile sensitivity. An analogy with Escherichia coli, in which the MutHLS system sensitizes Dam(-) mutants to DNA-injuring agents, suggested that bile might cause DNA damage. In support of this hypothesis, we show that bile induces the SOS response in S. enterica and increases the frequency of point mutations and chromosomal rearrangements. Mutations in mutH, mutL, or mutS cause partial relief of virulence attenuation in a Dam(-) background (50- to 100-fold by the oral route and 10-fold intraperitoneally), suggesting that an active MutHLS system reduces the ability of Salmonella Dam(-) mutants to cope with DNA-damaging agents (bile and others) encountered during the infection process. The DNA-damaging ability of bile under laboratory conditions raises the possibility that the phenomenon may be relevant in vivo, since high bile concentrations are found in the gallbladder, the niche for chronic Salmonella infections.

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Figures

Figure 1.—

Plate tests of SOS induction by sodium deoxycholate. Each plate was seeded with >107 cells (which form a lawn after growth). A small amount of sodium deoxycholate powder was placed in the center of the plate. Plates were incubated overnight at 37°. Left, strain SV4933 (_recA+/_pGE108); right, strain SV4870 (_recA1/_pGE108).

Figure 2.—

SOS induction by ox bile extract. All the strains carried the cea::lacZ fusion of pGE108. The strains shown are, from left to right, SV4933 (wild type/pGE108), SV4870 (RecA−/pGE108), and SV4930 (Dam−/pGE108). Open bars show β-galactosidase activity of the cea::lacZ fusion in LB. Solid bars show β-galactosidase activity of the cea::lacZ fusion after exposure to 10% ox bile extract. Because Dam− mutants are sensitive to 10% ox bile extract, strain SV4930 was preadapted by growth in the presence of LB containing 2% ox bile extract. The data shown are means and standard deviations of more than five independent experiments.

Figure 3.—

Effect of ox bile extract on the frequency of duplication segregation. Open bars show the background level of duplication segregation for each merodiploid strain. Solid bars show the percentage of segregants after exposure to 15% ox bile extract. Because of the high numerical dispersion typical of these experiments, the median is shown instead of the average, and error bars are not included (see C

amacho

and C

asades

2001). Each median has been calculated from more than eight independent experiments.

Figure 4.—

Effect of ox bile extract on the frequency of reversion in strains SV1408 (leuA414), SV2045 (hisG46), and TR947 (hisC3072). Open bars show the frequency of spontaneous reversion. Shaded bars show the frequency of reversion after exposure to 10% ox bile extract. Solid bars show the frequency of reversion after exposure to 15% ox bile extract. Stippled bars show the frequency of reversion in a LexA(Ind−) background after exposure to 15% ox bile extract. In the latter set of experiments the strains used were SV5006 [lexA(Ind−) _leuA414_], SV5007 [lexA(Ind−) _hisG46_], and SV5008 [lexA(Ind−) _hisC3072_]. The data shown are means and standard deviations of 20 independent experiments.

Figure 5.—

Graphical representation of competitive index analysis of Mut−, Dam−, and Mut− Dam− strains, after oral or intraperitoneal infections of BALB/c mice. The mixed infections performed were as follows: (a) MutS−/wild type, oral infection; (b) MutL−/wild type, oral; (c) MutH−/wild type, oral; (d) MutS− Dam−/Dam−, oral; (e) MutL− Dam−/Dam−, oral; (f) MutH− Dam−/Dam−, oral; and (g) MutL− Dam−/Dam−, intraperitoneal. (h and i) Internal controls. In these infections the wild type was co-inoculated with strain SV4676. The latter carries a trg::Mu_d_J insertion, previously shown to be neutral for virulence (S

egura

et al. 2004). (h) Oral infection; (i) intraperitoneal infection. The indexes represented are the means from more than three infections. Error bars represent the standard deviation.

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Bile-induced DNA damage in Salmonella enterica - PubMed (original) (raw)