Characterization of Gardnerella vaginalis membrane vesicles reveals a role in inducing cytotoxicity in vaginal epithelial cells.

Bacterial vaginosis (BV) is a common polymicrobial infection affecting women in the reproductive age and is associated with adverse obstetric and gynaecological outcomes. Gardnerella vaginalis is the most virulent anaerobic bacterial species predominantly associated with BV. However, a clear understanding of the mechanisms by which it contributes to the pathogenesis and persistence of BV is lacking. In this report, we demonstrate for the first time, the isolation of membrane vesicles (MVs) from G. vaginalis ATCC 14019. These MVs are approximately 120-260 nm in diameter. Proteomic characterization of the MVs by LC-MS/MS led to the identification of 417 proteins, including proteins involved in cellular metabolism as well as molecular chaperones and certain virulence factors. Immunoblot analysis of the MVs confirmed the presence of vaginolysin, the most well-characterized virulence factor of G. vaginalis. The exposure of the vaginal epithelial cells, VK2/E6E7 to the G. vaginalis MVs resulted in the internalization of the MVs. The MVs induced cytotoxicity and an increase in the levels of the pro-inflammatory cytokine, IL-8 in VK2 cells as well lysis of erythrocytes. The results of the study indicate that G. vaginalis MVs may be involved in the delivery of cytotoxic proteins and other virulence factors to the host cells and could thereby contribute towards enhancing the cellular damage associated with pathogenesis of BV.

Electron microscopy studies on Gardnerella vaginalis grown in conventional and biofilm systems.

The cell-wall characteristics of Gardnerella vaginalis grown in conventional and biofilm systems were studied by electron microscopy. The gram-positive nature of the cell wall was confirmed. Novel cell-wall particles which appeared to be associated with cell division were also identified, particularly in organisms of biofilm origin.

Ultrastructural characteristics of Gardnerella vaginalis infection in the heterosexual couple.

Adhesion and penetration of Gardnerella vaginalis into the male urethral and female vaginal epithelial cells were evaluated in a prospective cohort study of infected females and their sexual partners. Vaginal secretions of 10 women with culture proven G. vaginalis infection and semen samples of their asymptomatic husbands were analyzed with conventional optical and electron microscopy. G. vaginalis was isolated in 50% of the male sexual partners of women harboring the microorganism. G. vaginalis adheres to the plasmatic membrane and penetrates into the cytoplasm of both vaginal and urethral epithelial cells. The ability of G. vaginalis to colonize the male lower genital tract may have clinical relevance with respect to the role of the male partner in the reinfection of women.

[A new zoonosis--investigation of Gardnerella vaginalis disease of fox. II. Identification of pathogenic bacteria].

16 strains isolated from aborted foetus and vaginal exeretions of foxes were used for examination of cell morphology including fine structure of the cell wall and septa, test for homolysis, biochemical reactions and analysis of G+C content of the bacterial DNA. The cells are pleomorphic bacilli and coccobacilli, Gram stain reaction appear positive to variable, catalse and oxidase-negative, with a fermentative type of glucose metabolism, giving acetic and lactic acid as the major end products of fermentation. It's differentiated from the human strains of G. vaginalis by its less fastidious in growth requirements and being facultatively anaerobic to aerobic. We propose the name: a new subspecies--Gardnerella vaginalis subsp. fox, the type strain is U80.

[Gardnerella vaginalis: transport, microscopy, testing resistance]. / Gardnerella vaginalis: Transport, Mikroskopie, Resistenztestung.

UNLABELLED: G. vaginalis is an important pathogen in the aetiology of bacterial vaginosis. Therefore, we investigated the influence of transport systems in isolation, a scoring system for Gram stains, and susceptibility to antimicrobial agents. The comparison between a simple (Transwab) and a sophisticated (Port-A-Cul) system showed no differences with regard to for instance Enterococcus faecalis or Escherichia coli; however, isolation of G. vaginalis, a fastidious microorganism, was significantly higher (alpha < 0.0001) in Port-A-Cul. There was a strong correlation (97.5%) using the scoring system indicating bacterial vaginosis and isolation of G. vaginalis. The minimal inhibitory concentrations (MIC) of metronidazole for 60 strains of G. vaginalis were higher than 32 mg/l, some strains showing heteroresistance. This phenomenon may be an explanation for treatment failures. Clindamycin and erythromycin were much more active, with MIC's between 0.016 and 0.19 mg/l, in-vitro development of resistance being slower for clindamycin than for erythromycin. CONCLUSIONS: (I) for isolation of G. vaginalis, a sophisticated transport system is mandatory; (II) a scoring system for Gram staining is helpful in diagnosis of bacterial vaginosis; (III) in patients with metronidazole treatment failures, clindamycin should be used.

Characterization of Gardnerella vaginalis and G. vaginalis-like organisms from the reproductive tract of the mare.

Gardnerella vaginalis has been isolated from women with bacterial vaginosis, from the genital tracts of asymptomatic women, and from several other infected body sites in humans. However, until recently, it has not been isolated from any other animal species. Between June 1988 and October 1989, 31 isolates identified as G. vaginalis and 70 isolates identified as G. vaginalis-like organisms have been recovered from the genital tracts of 93 mares from Michigan and Ohio. Identification was based on biochemical reactions, hemolysis on media containing blood from various animal sources, and susceptibility to select antimicrobial agents. This report details the characterization of G. vaginalis and G. vaginalis-like organism isolates obtained from the reproductive tracts of these mares and compares the equine isolates with human isolates.

[Cytology of bacterial vaginosis]. / Zur Zytologie der bakteriellen Vaginose.

Bacterial vaginosis seems to be one of the most frequent cause of vaginal diseases which may be ascending and separates a not aesthetical discharge. A pH greater than 4.5 is found, the presence of clue cells, and a positive sniff test referring to Mobiluncus. The main microorganisms causing bacterial vaginosis are Gardnerella vaginalis and Mobiluncus sp. These are anaerobic ones. Cytology is able to give a quick and sure diagnosis.

[The ultrastructure of anaerobes isolated from the urogenital tract of women]. / Ul'trastruktura anaérobov, vydelennykh iz urogenital'nogo trakta zhenshchin.

Electron microscopic (ultrathin sections) examination of culture strains of vaginal Gardnerella and bacteroids isolated in aerobic conditions from urogenital tract of women suffering from Gardnerella infection was carried out. Structural analysis of the bacteria is presented, i.e. their shape, size, morphology, and features of multiplication.

Gardnerella vaginalis has a gram-positive cell-wall ultrastructure and lacks classical cell-wall lipopolysaccharide.

Gardnerella vaginalis has a very thin cell wall with a characteristic gram-negative staining pattern and an apparent lamellar structure when viewed at an oblique angle by electronmicroscopy. Examination at right angles to the cell-wall plane and by freeze-etching showed absence of an outer membrane or any other lamellar structure. Cell-wall extracts made by methods specific for lipopolysaccharide (LPS) gave negative reactions by silver staining and for endotoxin in the limulus amoebocyte lysate assay. 2-Keto-3-deoxy-D-manno-2-octonoic acid (KDO), heptose and hydroxy fatty acids specific for LPS were not detected in the extracts. G. vaginalis cell walls are unequivocally gram-positive in their ultrastructural characteristics and chemical composition.

Electron microscopy of adhesive interactions between Gardnerella vaginalis and vaginal epithelial cells, McCoy cells and human red blood cells.

Exfoliated vaginal epithelial cells with attached bacteria, termed 'clue cells', which were procured from a patient with non-specific vaginitis, were stained with ruthenium red and examined by transmission electron microscopy. The attached bacteria appeared to adhere by means of an outer fibrillar coat. An epithelial tissue culture cell line (McCoy) and human red blood cells to which strains of Gardnerella vaginalis attached were similarly examined. The adherence of G. vaginalis to the epithelial cell line appeared to be mediated by an outer fibrillar coat while adherence to red cells appeared to be mediated by fimbriae. Transmission electron microscopy was performed on the Gardnerella strains used. Thin sections of tissue-culture-adherent strains revealed a dense outer fibrillar coat whereas the surface of the haemagglutinating strains showed fine fimbriae. Negative staining of haemagglutinating strains demonstrated fimbriae on a minority of organisms.

Pili on Gardnerella vaginalis studied by electronmicroscopy.

Fourteen recently isolated strains and two laboratory strains of Gardnerella vaginalis were examined by electronmicroscopy for the presence of pili. All strains isolated recently from both men and women were heavily pilated. In contrast only a few pili were seen on organisms of the two laboratory strains, with many of the organisms having no pili. The importance of multiple subculture in this loss was supported by the observation that the degree of pilation of one freshly isolated strain decreased on repeated subculture. Other findings suggested that this was probably due to gradual loss of pili and not to selection of organisms that were non-pilate originally.

Demonstration by electron microscopy of pili on Gardnerella vaginalis.

Eight strains of Gardnerella vaginalis were examined by electron microscopy for the presence of pili. Narrow pili ranging from 3.0 to 7.5 nm in diameter were seen on bacteria from five of the strains studied.

Scanning electron microscopic examination of Bacteroides fragilis and Gardnerella vaginalis after exposure to concentration gradients of metronidazole and tinidazole.

Agar diffusion tests with metronidazole and tinidazole were performed with one strain each of Bacteroides fragilis and Gardnerella vaginalis (Haemophilus vaginalis, Corynebacterium vaginalis). Their cell morphology was studied 'in situ' on agar surfaces by means of scanning electron microscopy (SEM). A sharp growth end-point was found for B. fragilis, whereas with G. vaginalis there was a gradual decrease in the number and size of the colonies close to the agar well. The SEM study also revealed elongation of the B. fragilis cells when exposed to high concentrations of either drug. No such elongation of G. vaginalis was observed.