Intralumenal occluding fibrosis of the oviduct is a sequela of infection with C. muridarum in this model. These observations support the use of the murine model to study pathogenesis of chlamydial upper genital tract infection.
The mouse chlamydial pathogen Chlamydia muridarum has been used as a model organism for the study of human Chlamydia trachomatis urogenital and respiratory tract infections. To date, two commonly used C. muridarum isolates have been used interchangeably and are essentially taken to be identical. Herein, we present data that indicate that this is not the case. The C. muridarum Weiss isolate and C. muridarum Nigg isolate varied significantly in their virulences in vivo and possessed different growth characteristics in vitro. Distinct differences were observed in intravaginal 50% infectious doses and in challenge infections, with the Weiss isolate displaying greater virulence. Respiratory infection by the intranasal route also indicated a greater virulence of the Weiss isolate. In vitro, morphometric analysis revealed that the Weiss isolate produced consistently smaller inclusions in human cervical adenocarcinoma cells (HeLa 229) and smaller plaques in monolayers of mouse fibroblasts (L929) than did the Nigg isolate. In addition, the Weiss isolate possessed significantly higher replicative yields in vitro than did the Nigg isolate. In plaque-purified isolates derived from our stocks of these two strains, total genomic sequencing identified several unique nonsynonymous single nucleotide polymorphisms and insertion/deletion mutations when our Weiss (n ؍ 4) and Nigg (n ؍ 5) isolates were compared with the published Nigg sequence. In addition, the two isolates shared 11 mutations compared to the published Nigg sequence. These results prove that there is genotypic and virulence diversity among C. muridarum isolates. These findings can be exploited to determine factors related to chlamydial virulence and immunity.
The central hypothesis of this study was that matrix metalloproteinases (MMPs) would be enhanced following murine chlamydial infection and that their expression would vary in mouse strains that differ in their susceptibility to chronic chlamydia-induced disease. To address this hypothesis, female C3H/HeN and C57BL/6 mice were infected intravaginally with Chlamydia muridarum. Uterine and oviduct tissues were assessed for transcription of MMP genes and their tissue inhibitors. An increased activity of MMP genes relative to preinfection tissues was observed in the C3H/HeN mice when compared to C57BL/6 mice. Using gelatin zymography, we detected constitutive MMP-2 activity in both strains of mice but an increase in MMP-9. Casein zymography indicated the presence of two elastase-like activities consistent with MMP-12 and possibly MMP-7. Western blotting and antigen capture enzyme-linked immunoassay also confirmed an increase in MMP-9 but constitutive MMP-2 expression subsequent to the infection in both strains of mice. In C57BL/6 mice, MMP-9 was present in monomer and dimer form throughout the 56-day monitoring period. C3H/HeN mice produced dimeric MMP-9, but increases in the monomer form were also observed through day 14. Post-translational modification of MMP-9 between the two strains also differed. Immunohistochemistry revealed neutrophils as a prominent source for MMP-9 in both strains of mice. We conclude that differences in the relative expression and activity of MMPs, particularly MMP-9, occur in mice differing in their susceptibility to the development of chronic chlamydial disease. These differences may account for disparate outcomes with regard to chronic sequelae of the disease.
It was previously reported that female mice resolve a primary Chlamydia trachomatis urogenital infection independent of inducible nitric oxide synthase (iNOS). We now report that although iNOS-deficient (NOS2 ؊/؊ ) mice resolve culture-apparent infection in a fashion similar to that of normal control (NOS2 ؉/؉ ) mice, they sustain significantly increased rates of disease, as assessed by hydrosalpinx formation. PCR amplification of ompA followed by Southern blot detection of amplicands revealed the presence of chlamydial DNA in the lower genital tracts of both NOS2؊/؊ and NOS2 ؉/؉ mice at >120 days postinfection and in upper genital tract tissues at >120 days postinfection. However, only NOS2 ؊/؊ mice shed low numbers of viable chlamydiae from the lower genital tract after immunosuppressive treatment at 120 days postinfection. When cultured primary murine lung fibroblasts were activated in the presence of gamma interferon (IFN-␥), inhibition of chlamydial growth occurred in both NOS2؉/؉ and NOS2 ؊/؊ cells, but the inhibition was reversible after removal of the cytokine in the NOS2 ؊/؊ primary cell culture only. The iNOS-independent inhibition was microbistatic but was independent of 2,3-indoleamine dioxygenase activity. We conclude that chlamydial DNA and antigens persist in mice subsequent to culture-apparent resolution. In addition, IFN-␥ induces in vivo inhibition of chlamydial growth through microbistatic mechanisms in the absence of iNOS activity, but in the presence of iNOS activity, IFN-␥ is microbicidal and effects eradication.
We hypothesized that the plasmid of urogenital isolates of Chlamydia trachomatis would modulate infectivity and virulence in a mouse model. To test this hypothesis, we infected female mice in the respiratory or urogenital tract with graded doses of a human urogenital isolate of C. trachomatis, serovar F, possessing the cognate plasmid. For comparison, we inoculated mice with a plasmid‐free serovar F isolate. Following urogenital inoculation, the plasmid‐free isolate displayed significantly reduced infectivity compared with the wild‐type strain with the latter yielding a 17‐fold lower infectious dose to yield 50% infection. When inoculated via the respiratory tract, the plasmid‐free isolate exhibited reduced infectivity and virulence (as measured by weight change) when compared to the wild‐type isolate. Further, differences in infectivity, but not in virulence were observed in a C. trachomatis, serovar E isolate with a deletion within the plasmid coding sequence 1 when compared to a serovar E isolate with no mutations in the plasmid. We conclude that plasmid loss reduces virulence and infectivity in this mouse model. These findings further support a role for the chlamydial plasmid in infectivity and virulence in vivo.
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