Urogenital Mucosa Research Articles

Vaccination utilizing the FimCH complex as a strategy to prevent Escherichia coli urinary tract infections.

Eseherichia coli are the main causative agents of uncomplicated urinary tract infection (UTI) and account for >85% of cases of acute cystitis and pyelonephritis, >60% of recurrent cystitis, and at least 35% of recurrent pyelonephritis [1]. The reservoir for uropathogenic E. coli is fecal flora, from which bacteria spread to the urogenital mucosa, ascend into the bladder, and adhere to bladder epithelium. Once established in the bladder, the bacteria multiply and establish a local infection (cystitis) and may further ascend to involve the ureters and kidneys (pyelonephritis). While many factors contribute to the acquisition and progression of E. coli UTI, colonization of the urogenital tract by pathogenic bacteria appears to be a prerequisite for disease [2]. One promising strategy for the prevention of UTI is the development of a vaccine that targets the mechanism(s) by which uropathogens adhere to bladder epithelial cells.

Experimental gonococcal genital tract infection and opacity protein expression in estradiol-treated mice.

The development of effective prophylactic agents against gonorrhea and the study of adaptation by Neisseria gonorrhoeae to the urogenital mucosa are hindered by the lack of a well-established animal model of gonococcal genital tract infection. Here, a murine model of long-term gonococcal genital tract infection is described. Female BALB/c mice were treated with 17-beta-estradiol and inoculated intravaginally with wild-type gonococcal strain FA1090 or MS11. N. gonorrhoeae was recovered from vaginal swabs for an average of 12 to 13 days following inoculation with 10(6) CFU of either strain. Inflammation occurred in over 80% of infected mice, and diplococci were associated with epithelial cells and neutrophils in stained vaginal smears. Ascended infection occurred in 17 to 20% of mice inoculated with strain FA1090. An outbred mouse strain (SLC:ddY) previously reported to be naturally susceptible to N. gonorrhoeae was also tested; however, as with BALB/c mice, estradiol was required for prolonged infection. Although piliation was not maintained during experimental murine infection, 46 to 100% of vaginal isolates from four of eight BALB/c mice and three of four SLC:ddY mice expressed one or more opacity (Opa) proteins within 4 days after inoculation with an Opa-negative variant of strain FA1090. The observed selection for and/or induction of gonococcal Opa protein expression during murine infection appears to parallel events that occur during experimental urethritis in volunteers.

HLA-B27 modulates the survival of Salmonella enteritidis in transfected L cells, possibly by impaired nitric oxide production.

Reactive arthritis is triggered by certain microbes that cause primary infections mainly on the gastrointestinal or urogenital mucosa. The disease is strongly associated with HLA-B27. Long persistence of causative microbes or their structures in the body has been thought to have an important role in the pathogenesis of reactive arthritis. This suggests that the elimination of the microbes causing reactive arthritis is ineffective or disturbed in HLA-B27-positive individuals developing this complication. We examined the role of the HLA-B27 antigen in microbe-host interaction in vitro by monitoring the invasion and intracellular survival of Salmonella enteritidis in mouse fibroblasts transfected with HLA-B27, HLA-B7, or beta2-microglobulin only. S. enteritidis invaded into all the three transfectants with the same efficiency. However, at 6 and 10 days after incubation, there were more living intracellular Salmonella organisms in HLA-B27 transfectants than in the other transfected cell lines (P < 0.05), suggesting that the bactericidal effect is impaired in these cells. Impaired NO production in HLA-B27-transfected cells was indicated as a possible mechanism, since the amount of nitrite in the supernatants of the Salmonella-infected HLA-B27-transfected cells was smaller than that in the supernatants of the Salmonella-infected HLA-B7- or beta2-microglobulin-transfected cells (P < 0.001). The inhibition of NO synthesis by N-monomethyl-L-arginine resulted in impaired elimination of Salmonella also in HLA-B7and beta2-microglobulin-transfected cells. The inverse correlation between intracellular survival of Salmonella and the amount of nitrite detected in culture supernatants supports the hypothesis that the L-arginine-dependent NO pathway plays an important role in the murine fibroblast response against Salmonella. We suggest that a major histocompatibility complex class I antigen, HLA-B27, may contribute to the intracellular persistence of Salmonella by a mechanism which involves the NO pathway.

Prevention of mucosal Escherichia coli infection by FimH-adhesin-based systemic vaccination.

Virtually all uropathogenic strains of Escherichia coli, the primary cause of cystitis, assemble adhesive surface organelles called type 1 pili that contain the FimH adhesin. Sera from animals vaccinated with candidate FimH vaccines inhibited uropathogenic E. coli from binding to human bladder cells in vitro. Immunization with FimH reduced in vivo colonization of the bladder mucosa by more than 99 percent in a murine cystitis model, and immunoglobulin G to FimH was detected in urinary samples from protected mice. Furthermore, passive systemic administration of immune sera to FimH also resulted in reduced bladder colonization by uropathogenic E. coli. This approach may represent a means of preventing recurrent and acute infections of the urogenital mucosa.

Characterization of transferrin binding proteins 1 and 2 in invasive type b and nontypeable strains of Haemophilus influenzae.

Haemophilus influenzae has the ability to obtain iron from human transferrin via two bacterial cell surface transferrin binding proteins, Tbp1 and Tbp2. Although a wide array of strains have been shown to express these receptor proteins, two studies have recently identified a series of isolates which appeared to lack the ability to bind transferrin. Included in this group were the members of a cryptic genospecies of nontypeable biotype IV strains which appear to possess a tropism for female urogenital tissues and are major etiologic agents of neonatal and postpartum bacteremia due to H. influenzae. The present study employed oligonucleotide primers specific for genes encoding the Tbp proteins of a type b biotype I strain of H. influenzae to probe the genomic DNAs of isolates from the previous studies. The tbpA and tbpB genes which encode Tbp1 and Tbp2, respectively, were detected in all of the strains tested either by PCR amplification directly or by Southern hybridization analysis. All of the strains displayed a transferrin binding phenotype, and affinity isolation of receptor proteins with transferrin-conjugated Sepharose recovered Tbp1 and/or Tbp2 from 11 of 14 strains, including 2 of the nontypeable biotype IV strains. In addition, all of the strains were capable of growing on human transferrin specifically, indicating that the mechanism of iron assimilation from transferrin is functional and is not siderophore mediated. These results confirm the presence of tbp genes in all of the invasive H. influenzae isolates characterized to date, suggesting that Tbp-mediated iron acquisition is important in disease which initiates from either the respiratory or urogenital mucosa.

Generation of humoral immune responses against herpes simplex virus type 2 in the murine female genital tract

A murine model of genital infection with a thymidine kinase-deficient (tk −) strain of herpes simplex virus type 2 (HSV-2) was utilized to examine the local urogenital antibody response to HSV-2. Vaginal inoculation with HSV-2 tk − protected against a subsequent genital challenge with a lethal dose of virulent HSV-2. After primary vaginal infection, predominantly HSV-specific IgG antibodies were detected in serum and vaginal secretions. HSV-specific IgG antibody-secreting cells (ASC) were detected first and in greatest numbers in the genital lymph nodes (gLN) after primary HSV-2 tk − inoculation. HSV specific IgG or IgA ASC were not detected in the urogenital mucosa after primary HSV-2 vaginal infection. Vaginal HSV-2 challenge of HSV-immune mice resulted in increased HSV-specific serum IgG antibody and vaginal IgA antibody titers. HSV-specific IgG ASC were detected by 4 days postchallenge in gLN and by Days 6 and 7 postchallenge in the spleen and genital mucosa. These results suggest that urogenital humoral responses originate in the gLN following HSV genital infection and that gLN may serve as the primary source of the HSV-specific IgG- and IgA-secreting cells present in the urogenital mucosa after vaginal challenge.

Quantitation of effects of subinhibitory concentrations of trimethoprim on P fimbria expression and in vitro adhesiveness of uropathogenic Escherichia coli.

The ability to adhere to and colonize urogenital mucosa is an important virulence attribute of uropathogenic Escherichia coli. This adherence, which appears to be mediated by P fimbriae, may be affected by antibiotics or other agents that affect fimbrial expression. We describe here an enzyme immunofiltration assay to quantitate fimbriation and the application of that technique to measurement of the effects of sublethal doses of trimethoprim on P fimbrial expression. Effects on P fimbriation correlated with effects on the adherence of treated bacteria to cultured T24 bladder carcinoma epithelial cells; i.e., trimethoprim treatment decreased both P fimbriation and bacterial adherence. It was possible to quantitate effects on P fimbriation when type 1 fimbriae were also present. The enzyme immunofiltration assay may be useful for studies on the role of fimbriae in the pathogenesis of bacterial infections, and it may facilitate identification of antimicrobial agents that interfere with bacterial adherence to mucosal surfaces.

Infection of British deer and farm animals with epizootic haemorrhagic disease of deer virus

Epizootic haemorrhagic disease of deer virus, an orbivirus related to bluetongue virus, was inoculated intravenously into red, fallow, roe and muntjac deer, and into sheep, goats, pigs and cattle. Low level viraemias were recorded in each species apart from goats and pigs but clinical disease was not observed. Virus was not isolated from swabs taken of the nasal, urogenital or rectal mucosae. Control animals in direct contact with inoculated animals did not develop a detectable viraemia or serum antibody. In viraemic animals the virus was associated with the cell fraction of the blood. Beyond 7 days after inoculation the chance of virus isolation was enhanced if the cells were washed and lysed prior to inoculation of the cell cultures. It is suggested that cattle may play a role in the epizootiology of epizootic haemorrhagic disease of deer in North America.

The adherence of pilate and non-pilate strains of neisseria gonorrhoeae to human and guinea-pig epithelial tissues

Strains of Neisseria gonorrhoeae adhered to pieces of human endocervix and appeared to be embedded in the surface mucus. Although a pilate strain adhered better than a non-pilate strain, the difference was small and pilation did not appear to be exclusively responsible for adherence. The pilate strain showed better adherence to pieces of human ectocervix and fallopian tube, but both strains were similarly adsorbed to human bronchus and guinea-pig uterus, cervix, male urethra and bladder, although to different degrees for different tissues. Since gonococci adhered to all tissues examined, their ability to infect human endocervix and fallopian tube and their failure to infect human ectocervix or guinea-pig urogenital tract mucosae are determined by factors other than a capacity for primary adherence to the tissue.

Studies on Organ Specificity

Summary The sera of rabbits immunized with pooled rabbit pancreas incorporated into Freund adjuvants contain antibodies that react with certain other rabbit pancreas extracts. They fail to react in vitro with the antibody-producing animal's own pancreas extract. No gross or microscopic evidence of damage to the pancreas was observed. The antibodies were therefore designated isoantibodies rather than auto-antibodies. These rabbit pancreas isoantibodies could be demonstrated by complement fixation, precipitation, and passive cutaneous anaphylaxis. The isoantibodies of the sera could be removed by absorption with some individual pancreas extracts but not with others. Absorption of the anti-rabbit pancreas rabbit sera with rabbit serum, erythrocytes, gastrointestinal and urogenital tract mucosae or other rabbit organs did not remove the rabbit pancreas isoantibodies. This finding was true even if the absorptions were carried out with organs from a rabbit whose pancreas extract provided a positive reaction. Absorption of these antisera with human A, B or O red blood cells or sheep red blood cells did not inhibit the homologous reaction with rabbit pancreas extract. The existence of at least three different rabbit pancreas isoantigens could be shown by gel diffusion precipitation experiments within the heterogenous strain of rabbits used in the study. Similar pancreas isoantigens have not been found so far in other experimental animals such as guinea pigs or dogs, even though these species contain the pancreas-specific antigens demonstrable by heteroimmunization of rabbits. The anti-rabbit pancreas rabbit sera did not cross-react with pancreas extracts of other species. “Natural antibodies” of the type described by Kidd and Friedewald could be domonstrated in these antisera both before and after immunization with rabbit pancreas extract, although an occasional increase in titer was observed during immunization. These antibodies could be differentiated from the pancreas isoantibodies by specific absorption and heat inactivation properties as well as by gel precipitation. The electrophoretic mobilities of the rabbit pancreas isoantigens were compared to that of human serum albumin in agar gel with the following results: one isoantigen had an electrophoretic mobility slightly greater than albumin, one isoantigen had an electrophoretic mobility similar to that of albumin, whereas the other had a lower electrophoretic mobility than human serum albumin. Immunoelectrophoresis studies showed the rabbit pancreas isoantibodies to be in the γ-globulin region.