Immune activation and suppression by group B streptococcus in a murine model of urinary tract infection - PubMed (original) (raw)
Immune activation and suppression by group B streptococcus in a murine model of urinary tract infection
Kimberly A Kline et al. Infect Immun. 2011 Sep.
Abstract
Group B streptococcus (GBS) is a common commensal of the gastrointestinal and vaginal mucosa and a leading cause of serious infections in newborns, the elderly, and immunocompromised populations. GBS also causes infections of the urinary tract. However, little is known about host responses to GBS urinary tract infection (UTI) or GBS virulence factors that participate in UTI. Here we describe a novel murine model of GBS UTI that may explain some features of GBS urinary tract association in the human host. We observed high titers and heightened histological signs of inflammation and leukocyte recruitment in the GBS-infected kidney. However, extensive inflammation and leukocyte recruitment were not observed in the bladder, suggesting that GBS may suppress bladder inflammation during cystitis. Acute GBS infection induced the localized expression of proinflammatory cytokines interleukin-1α (IL-1α), macrophage inflammatory protein-1α (MIP-1α), MIP-1β, and IL-9, as well as IL-10, more commonly considered an anti-inflammatory cytokine. Using isogenic GBS strains with different capsule structures, we show that capsular sialic acid residues contribute to GBS urinary tract pathogenesis, while high levels of sialic acid O-acetylation attenuate GBS pathogenesis in the setting of UTI, particularly in direct competition experiments. In vitro studies demonstrated that GBS sialic acids participate in the suppression of murine polymorphonuclear leukocyte (PMN) bactericidal activities, in addition to reducing levels of IL-1α, tumor necrosis factor alpha, IL-1β, MIP-1α, and KC produced by PMNs. These studies define several basic molecular and cellular events characterizing GBS UTI in an animal model, showing that GBS participates simultaneously in the activation and suppression of host immune responses in the urinary tract.
Figures
Fig. 1.
GBS preferentially colonizes the murine kidney. Young C3H/HeN female mice (7 to 8 weeks old) were transurethrally inoculated with 107 CFU GBS or UPEC in 50 μl, and the CFU in the bladder (A) or kidney (B) was enumerated at the indicated time points. The numbers of GBS or UPEC CFU were directly compared at 24 hpi (C). Results represent a compilation of 1 to 2 experiments (A and B) and a compilation of 4 experiments (C), using at least 5 mice per time point. The limit of detection of the assay is indicated by a dashed line. Statistical significance was determined by a two-tailed Mann-Whitney U test: *, P < 0.05; **, P < 0.01; n.s., not significant. Statistical significance shown in panel B denotes comparison to the numbers of bladder CFU at the same time point.
Fig. 2.
Cytokine/chemokine induction in the bladder and serum of GBS-infected mice. At 24 hpi, the bladders from C3H/HeN mice infected transurethrally with GBS or mock infected with PBS were removed and homogenized (A) or the serum was collected (B), and cytokine expression was assessed. The data are a composite of 2 separate experiments consisting of 4 to 5 mice each and are calculated as fold change compared to the results for PBS mock-infected animals. Dark gray bars and asterisks indicate values significantly different from those for mice mock infected with PBS (P < 0.05), as determined by a 1-sample t test; light gray bars, P < 0.10; dotted lines, 2-fold induction cutoff.
Fig. 3.
Robust inflammation in the kidney of GBS-infected animals. Hematoxylin-eosin staining of GBS-infected C3H/HeN murine kidneys (A to G) or bladders (H to K) at 3 hpi (A B, E, F, and H) or 24 hpi (C, G, I, and J). PBS mock-infected kidney (D) and bladder (K) at 24 hpi are shown. Representative images are shown at magnifications of ×10 (A to D, K), ×40 (I and J), ×60 (F to H), and ×100 (E). (E to G) Boxed areas from panels A to C, respectively.
Fig. 4.
Unmodified capsular sialic acids of GBS suppress oxidative burst and bactericidal activity of murine neutrophils. (A) Biochemical similarity of the surfaces of the bladder and GBS is shown by high-pressure liquid chromatography analysis of sialic acids from uninfected murine bladder and GBS strains used in this study. The latter have been previously published and are shown here for comparison. The most common sialic acid in mammals is _N_-acetylneuraminic acid (Neu5Ac). O-acetylation of Neu5Ac occurs at the carbon 7 and 9 positions of the sialic acid molecule (Neu5,7Ac2 and Neu5,9Ac2, respectively). (B) GBS was incubated with murine peritoneal PMNs at an MOI of 10 to 20:1 bacteria/PMNs, and bacterial survival was assessed after 30 min. Data shown are the compilation of 4 separate experiments. Kinetic measurement (C) and maximal luminescence measurement (D) of in vitro oxidative burst of murine peritoneal PMNs were assessed after incubation with GBS-hiOAc or GBS-loOAc. The MOI was 20:1 bacteria/PMNs. Similar results were obtained at an MOI of 200:1. Significant differences were determined by the paired t test: *, P < 0.05; **, P < 0.01.
Fig. 5.
GBS capsular sialic acid modification alters PMN cytokine production. Murine PMN culture supernatants were collected after 60 min of incubation with live GBS or PBS (A) or 180 min of incubation with heat-killed GBS or PBS (B) and assayed for cytokine production. Data are represented as fold change compared to the results for PBS-treated PMN at the same time point (n = 3). Significant differences between GBS-hiOAc or GBS-loOAc infection were measured by the unpaired t test: *, P < 0.05; **, P < 0.01. Each panel and statistical evaluation includes data from at least 3 independent experiments.
Fig. 6.
GBS capsular sialic acid O-acetylation limits virulence in the bladder. C3H/HeN mice were infected with isogenic GBS bearing surface structural alterations of the capsular polysaccharide. The numbers of CFU in bladder (A) and kidney (B) were enumerated at 24 hpi (n = 3 to 5 experiments with at least 5 mice per group). The limit of detection of the assay is indicated by a dashed line, and statistical analysis was performed as described in the legend to Fig. 1. (C) Competition experiments were performed by coinoculation of 107 each of GBS-hiOAC and GBS-loOAc containing different antibiotic resistance markers, followed by bacterial enumeration at 24 hpi.
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