HIV-1 co-receptor expression and epithelial immune cells of the cervix in asymptomatic women attending a genitourinary medicine clinic.

OBJECTIVES: The aim of the study was to qualitatively and semiquantitatively characterize the expression of the principal HIV co-receptors chemokine (C-C motif) receptor 5 (CCR5) and chemokine (C-X-C motif) receptor 4 (CXCR4) on susceptible CD4 T-helper cell, monocyte/macrophage and Langerhans dendritic cell populations within the cervical epithelia of asymptomatic women attending a genitourinary medicine clinic. METHODS: Of 77 asymptomatic women recruited, 35 were excluded: 21 because they were found to have bacterial vaginosis, eight because they were found to have candida and six for other reasons. Cervical cytobrush samples from 11 women with Chlamydia trachomatis infection and 31 women without any detectable genital infection were stained with fluorescently labelled antibodies specific for cell surface CCR5, CXCR4, CD4, CD3, CD1a and CD19 expression, then analysed by flow cytometry. RESULTS: CD4/CD3 T-helper cells (84%), CD1a Langerhans dendritic cells (75%) and CD4/CD14 monocytes/macrophages (59%) were detected in the samples. CCR5 and CXCR4 HIV co-receptor expression was observed on 46-86% of the above subsets. CD1a cells exhibited significantly higher CCR5 and CXCR4 positivity and median fluorescence than CD4 cells and higher CXCR4 positivity and median fluorescence than CD14 cells (P < 0.05 or less). Increased detection of CCR5 over CXCR4 was seen in CD14 cells (P < 0.05). No significant differences in CCR5 or CXCR4 expression were found in samples from asymptomatic women with or without chlamydial infection. CONCLUSIONS: Co-receptor expression confirms the potential for CD1a Langerhans cells, monocytes/macrophages and T-helper cells in the cervix as primary targets for HIV infection. Previously observed selective transmission of CCR5-tropic isolates cannot be accounted for by a lack of CXCR4-expressing CD4 cervical immune cells. We were unable to identify any specific impact of chlamydial infection on co-receptor expression in this study.

Apoptosis of ejaculated human sperm is induced by co-incubation with Chlamydia trachomatis lipopolysaccharide.

BACKGROUND: Previous work has shown that co-incubation of human sperm with Chlamydia trachomatis serovars E and LGV leads to premature sperm death and that this is due primarily to chlamydial lipopolysaccharide (LPS). Here, we investigated the possible involvement of apoptosis in this premature sperm death. METHODS: Highly motile preparations of sperm from normozoospermic patients were co-incubated for 6 h with extracted LPS from C. trachomatis serovars E and LGV. Three different methods were used to determine if LPS-treated sperm underwent apoptosis, including: (i) flow cytometry; (ii) measurement of ADP:ATP ratios; and (iii) measurement of mono- and oligonucleosomal DNA fragments. Caspase activity was also investigated by fluorimetry and by use of a pan-caspase inhibitor and caspase-3 inhibitor. RESULTS: All three methods used for detection indicated that C. trachomatis LPS induced some apoptosis in sperm after 6 h when compared with a staurosporine (apoptosis-positive) control. Moreover, a greater degree of apoptosis was seen with C. trachomatis serovar E than with serovar LGV. It was also shown that C. trachomatis LPS-induced apoptosis of sperm could be blocked with a pan-caspase inhibitor and a caspase-3 inhibitor. Moreover, by using a fluorogenic substrate, apoptosis was shown to be caspase-mediated. CONCLUSIONS: In general it is believed that apoptosis does not occur in C. trachomatis-infected host cells. However, using three different methods, our findings clearly indicate that co-incubation of sperm with C. trachomatis LPS results in cellular death which is in part due to apoptosis and is caspase-mediated. These findings provide an explanation as to how C. trachomatis can mediate premature death in human sperm.

Polymorphisms in matrix metalloproteinase-1, -3, -9, and -12 genes in relation to subarachnoid hemorrhage.

BACKGROUND AND PURPOSE: Intracranial aneurysm, which underlies the vast majority of subarachnoid hemorrhage incidences, has a multifactorial etiology, and the importance of genetic factors is increasingly recognized. Development and rupture of intracranial aneurysms involve degradation and remodeling of the vascular wall matrix in which the matrix metalloproteinases (MMPs) play an important role. The possible impact of MMP gene polymorphisms on susceptibility to intracranial aneurysms is still controversial, with conflicting data from different reported studies. METHODS: In this study we analyzed 5 different functional promoter polymorphisms in the MMP-1, MMP-3, MMP-9, and MMP-12 genes in a sample of 92 patients with aneurysmal subarachnoid hemorrhage and 158 healthy control subjects, all from southern England. RESULTS: No significant difference was detected between the patient and control groups in genotype distribution of any of the polymorphisms studied. CONCLUSIONS: The data do not support the hypothesis that MMP gene variations influence the development of intracranial aneurysms in the population studied.

Toxin production, adherence and protein expression by clinical Aeromonas spp. isolates in broth and human pooled ileostomy fluid.

The physiological behaviour of clinical Aeromonas spp. isolates was compared following culture in a conventional broth and human pooled ileostomy fluid (PIF). Protein expression was markedly affected by the growth medium, with an overall reduction in whole cell proteins in bacteria grown in ileostomy fluid. In addition, novel outer membrane proteins were produced in PIF but not in broth. The majority of A. hydrophila and A. sobria isolates produced toxin in both broth and PIF, whereas no cytotoxin positive A. caviae were found. Toxin titres were at least two doubling dilutions higher in 40% and 21% of A. hydrophila and A. sobria isolates, respectively, following culture in brain heart infusion broth compared with PIF. Bacterial adherence to Vero and A-549 cells was significantly more common in A. hydrophila (53%) and A. sobria (64%) than in A. caviae (15%) (P < 0.01). We observed increased adherence by 6 aeromonas strains previously classified as adherence-positive, but not by 6 non-adherers, in PIF compared with brain heart infusion broth. The influence of growth medium on the expression of potential virulence determinants by Aeromonas spp. provides a rationale for the use of human ileostomy fluid in future in vitro studies, in order to simulate the nutrient conditions found in vivo.

In-vitro activity of imipenem, in comparison with cefuroxime and ciprofloxacin, against coagulase-negative staphylococci in broth and peritoneal dialysis fluid.

The activity of cefuroxime, ciprofloxacin and imipenem against 50 coagulase-negative staphylococci (CNS) strains isolated from patients with peritonitis while on continuous ambulatory peritoneal dialysis (CAPD) was examined. Strains were cultured in both used peritoneal dialysis fluid (PUD) and broth (ISB) in an atmosphere of air with 5% CO2. MICs of cefuroxime, ciprofloxacin and imipenem differed significantly between the two media for 40%, 26% and 42% of strains, respectively. In three-quarters of these cases the MICs were higher in PUD compared to ISB. The ability of each antibiotic to kill six strains of CNS adherent to silicone rubber was also examined. Once again, differences in killing were noted between the two media. At least two-thirds of the strains remained viable in the presence of concentrations of antibiotic equal to the MBC of each antibiotic and at least one-third survived antibiotic concentrations equivalent to those attained during treatment. Assessment of activity in PUD may provide a more realistic test for antibiotics to be used in the treatment of CAPD peritonitis.

Comparison of the in vitro activities of fenticonazole, other imidazoles, metronidazole, and tetracycline against organisms associated with bacterial vaginosis and skin infections.

The in vitro antibacterial activity of the antifungal compound fenticonazole was compared with those of clotrimazole, miconazole, tetracycline, and metronidazole against 177 strains of bacterial species associated with either bacterial vaginosis (BV) or skin infections by agar dilution MIC determinations. BV-associated Bacteroides isolates of the Bacteroides melaninogenicus-B. oralis group, Gardnerella vaginalis, Mobiluncus spp., and anaerobic, gram-positive cocci were highly susceptible to fenticonazole, clotrimazole, and miconazole; but Bacteroides spp. not associated with BV, Bacteroides ureolyticus and the Bacteroides fragilis group, were resistant. All Bacteroides strains were susceptible to metronidazole, but the susceptibility of G. vaginalis and Mobiluncus spp. varied. Among the skin bacteria, Staphylococcus aureus, coryneforms, and streptococci were highly susceptible to the imidazoles; but Staphylococcus epidermidis strains were generally resistant. This antibacterial activity may give fenticonazole a useful role in the topical treatment of vaginal discharge and in mycotic skin infections that are superinfected with bacteria.

Activity of pefloxacin and thirteen other antimicrobial agents in vitro against isolates from hospital and genitourinary infections.

The in-vitro activity of the quinolone derivative pefloxacin was compared with that of three other quinolones, five beta-lactam antibiotics and three aminoglycosides against 367 isolates from hospital patients and from out-patients with genitourinary infections. MIC90 of pefloxacin and norfloxacin for each strain was the same; that of ciprofloxacin was a little lower. All strains except Escherichia coli were resistant to nalidixic acid. Pefloxacin was highly active against Staphylococcus aureus (39 strains; MIC90 1.0 mg/l) and most strains of coagulase-negative staphylococci (56; 4 mg/l), Esch. coli (50; 0.25 mg/l), other enterobacteria (33; 1.0 mg/l) and Pseudomonas aeruginosa 6; 0.25 mg/l). With Bacteroides spp. (total 78; 64 mg/l), the fragilis group (23) and the fusobacteria (19) were resistant, but the melaninogenicus-oralis group (31; range 0.06- greater than 64 mg/l) and B. ureolyticus (22; 0.125- greater than 64 mg/l) gave variable results. Amongst genitourinary isolates, Neisseria gonorrhoeae (15) and Haemophilus ducreyi (34) were sensitive (less than 0.06 mg/l) but Gardnerella vaginalis (25) and Mobiluncus spp. (11) were resistant (32 mg/l). Pefloxacin was more active than ceftazidime, cefotaxime, ceftizoxime, latamoxef and piperacillin against S. aureus and coagulase-negative staphylococci and than gentamicin, tobramycin and amikacin against coagulase-negative staphylococci. No enterobacteria or pseudomonads were resistant to pefloxacin or other quinolones, whereas some were resistant to beta-lactams and aminoglycosides.

In-vitro and in-vivo activity of metronidazole against Gardnerella vaginalis, Bacteroides spp. and Mobiluncus spp. in bacterial vaginosis.

An open, randomized, culture-controlled clinical study was designed to compare the efficacy of a single 2 g dose of metronidazole (Elyzol) with standard 7-day therapy in the treatment of bacterial vaginosis (BV). Forty-one of 47 (87%) patients given the single dose and 30 of 33 (91%) given the 7-day treatment were found to be cured seven days after treatment. At final assessment, 24 of 34 (71%) patients given the single dose and 22 of 28 (79%) given the 7-day treatment remained cured. The two regimes were equally efficaceous in eradicating Gardnerella vaginalis, Bacteroides spp. and Mobiluncus spp. (anaerobic curved rods) from vaginal specimens from patients with BV. The in-vitro activity of metronidazole and its hydroxy metabolite was determined for 11 strains of Gard. vaginalis, 17 strains of Bacteroides spp. and 14 strains of Mobiluncus spp. which had been isolated from patients prior to treatment. The MIC of metronidazole against Gard. vaginalis varied between 2 and greater than or equal to 128 mg/l (median MIC 32 mg/l), but the hydroxy metabolite showed a markedly increased activity against eight of the strains tested (median MIC 4 mg/l). The MIC of metronidazole against the Mobiluncus spp. varied between 0.5 and greater than or equal to 128 mg/l (median MIC 16 mg/l) and the hydroxy metabolite showed little increased activity (median MIC also 16 mg/l). The Bacteroides organisms were highly susceptible to metronidazole and to the hydroxy metabolite, each having a median MIC of 1 mg/l.

In vitro susceptibility of Gardnerella vaginalis and Bacteroides organisms, associated with nonspecific vaginitis, to sulfonamide preparations.

Recent reports suggest that anaerobic Bacteroides organisms are frequently found with Gardnerella vaginalis in nonspecific vaginitis. Specimens taken from 96 women with vaginal discharge were tested simultaneously for these organisms. G. vaginalis was found in 73% of the specimens, Bacteroides was found in 53%, and both organisms were found in 47%. Sulfonamides have been widely used in the successful treatment of vaginitis. Paradoxically, G. vaginalis is reported to be resistant, and it has been suggested that it could be the vehicle of the drugs which effects the cure. Little is known of the susceptibility of vaginal anaerobes to the sulfonamides. G. vaginalis and Bacteroides isolates were therefore tested in vitro against the individual excipients of sulfonamide tablets, and minimal inhibitory concentration tests were also performed against the three active drugs. The excipients had no effect on G. vaginalis, but Bacteroides strains were susceptible to the urea component. All strains of both organisms were susceptible to at least two of the three sulfonamides at high concentrations.

Balanoposthitis associated with Gardnerella vaginalis infection in men.

Fourteen of 194 (7.2%) consecutive unselected men had positive culture results from genital swabs for Gardnerella vaginalis. A higher yield of isolates was obtained from preputial (93%) than from urethral swabs (64%). Of the 14 men, two had no detectable genital abnormality, eight non-gonococcal urethritis, and nine balanoposthitis. The urethral isolation rates for G vaginalis in men with and without non-gonococcal urethritis were not significantly different, but preputial isolation rates were significantly higher (p less than 0.001) in men with balanoposthitis than in those without. The prevalence rate for G vaginalis in men with non-candidal balanoposthitis was 31%. In a second study, concomitant Bacteroides species were isolated in preputial swabs from nine of 12 (75%) men with G vaginalis-associated balanoposthitis and may play a role in its pathogenesis.

Induction of a cell wall variant of the gonococcus by human amylase.

Human amylase has been shown to be an effective and powerful inhibitor of the gonococcus in vitro. Its action appears to be on the bacterial cell wall. When tested in osmotically stabilised liquid media the organism was not inhibited; however the organisms which grew in such cultures appeared to be cell wall variants, which were less sensitive to penicillin than the parent strains. Studies on these variants suggest that they are "transitional-phase variants." Since cervical mucus has much in common with an alkaline osmotically stabilised liquid medium and also contains a high concentration of amylase, it seems possible that these variants may occur in vivo.

Detection and identification of gonococcal L-Forms using a direct immunofluorescence test.

A fluorescent antibody test was used to identify L-Forms of N. gonorrhoeae induced in vitro. It was possible to differentiate the Large Bodies of the L-Forms from parent gonococci and the fluorescent reaction remained specific in the presence of tissue culture cells. A possible method to identify L-Forms of gonococci from patients presenting with postgonococcal urethritis is described.