Detection of a single identical cytomegalovirus (CMV) strain in recently seroconverted young women

Background

Infection with multiple CMV strains is common in immunocompromised hosts, but its occurrence in normal hosts has not been well-studied.

Methods

We analyzed CMV strains longitudinally in women who acquired CMV while enrolled in a CMV glycoprotein B (gB) vaccine trial. Sequencing of four variable genes was performed in samples collected from seroconversion and up to 34 months thereafter.

Results

199 cultured isolates from 53 women and 65 original fluids from a subset of 19 women were sequenced. 51 women were infected with one strain each without evidence for genetic drift; only two women shed multiple strains. Genetic variability among strains increased with the number of sequenced genetic loci. Nevertheless, 13 of 53 women proved to be infected with an identical CMV strain based on sequencing at all four variable genes. CMV vaccine did not alter the degree of genetic diversity amongst strains.

Conclusions

Primary CMV infection in healthy women nearly always involves shedding of one strain that remains stable over time. Immunization with CMVgB-1 vaccine strain is not selective against specific strains. Although 75% of women harbored their unique strain, or a strain shared with only one other woman, 25% shared a single common strain, suggesting that this predominant strain with a particular combination of genetic loci is advantageous in this large urban area.     

Pathogenic characteristics of Escherichia coli strains in cases of asymptomatic bacteriuria and symptomatic urinary tract infections

The aim of this study was to examine if E. coli isolated from asymptomatic bacteriuria differed in pathogenic features from strains isolated from symptomatic infections of urinary tract. In this study 130 strains of E. coli isolated from women having asymptomatic bacteriuria and 112 strains isolated from patients with symptoms of urinary tract infection were examined. It was shown that E. coli isolated from patients with symptomatic urinary tract infection showed the more frequently ability to cause mannose-resistant haemagglutination of human erythrocytes, resistance to bactericidal activity of serum and haemolytic properties than those isolated from asymptomatic bacteriuria. These strains showed also the higher ability to adhere to Vero cells in tissue culture. Among E. coli strains isolated from persons with asymptomatic bacteriuria the pathogenic features were most frequently found in strains from healthy women and the most rarely in isolated from diabetic women.     

Susceptibility to antimicrobial agents of bacteria isolated from pregnant, postpartum women, and newborns from zones with high irradiation risk]

Susceptibility of 350 strains of staphylococci, isolated from pregnant women, puerperal and newborns and of 170 strains, isolated from obstetrical hospital environment at the region with high risk of radiation after accident was determined. The tested bacterial strains were the following: 56--Staphylococcus aureus, 152--S. epidermidis, 142--S. saprophyticus, isolated from women and newborns; strains isolated from hospital environment included 120 strains of S. saprophyticus and 50 strains E. coli. The tested antimicrobial factors were the following: 22 antibiotics, 2 bioantibiotics of the breast milk and blood (lysozyme, betalysin) not separated from the substrate and tested on test-cultures of M. luteus and B. subtilis, and 5 desinfectants. Analysis was performed by common disk-diffusion method and by contact method elaborated before. High resistance of staphylococci to many antibiotics, bioantibiotics and disinfectants was demonstrated. The resistance is considered to be due to high radiation background because it is much higher than resistance frequency of the similar strains isolated at the safe areas. The resistance to antimicrobial factors was higher among the opportunistic and saprophyte staphylococci along with appearance of pathogenic factors and activation of persistence properties.     

Species diversity and relative abundance of vaginal lactic acid bacteria from women in Uganda and Korea

AIMS: Lactobacilli play an important role in maintaining vaginal health of women. The aim of this study was to compare the species richness and relative abundance of Lactobacillus and other lactic acid bacteria in women of two geographically distant countries, Uganda and Korea. METHODS AND RESULTS: Vaginal samples were obtained from two women populations in Uganda and Korea. The Lactobacillus Rogosa SL agar was used for initial isolation of lactic acid bacteria. After phenotypic analyses, the 16S rRNA gene was amplified by polymerase-chain reaction and analysed by the BLAST program and phylogenetic tree construction. A total of 338 (128 Korean and 210 Ugandan) vaginal lactic acid bacterial strains were isolated, including five genera: Lactobacillus, Leuconostoc, Pediococcus, Streptococcus and Weissella. While Lactobacillus crispatus was common in both populations, Lactobacillus fermentum was common only in Korean women, and Lactobacillus gasseri, Lactobacillus reuteri and Lactobacillus vaginalis only in Ugandan women. Among other lactic acid bacteria, Weissella was more common in Ugandan, and Pediococcus in Korean women. All Weissella strains produced hydrogen peroxide, and all Pediococcus strains inhibited Candida species. CONCLUSION: Although many lactic acid bacteria colonize women, their species distributions may be different in women of geographically separated communities. SIGNIFICANCE AND IMPACT OF THE STUDY: The knowledge of species richness and relative abundance of vaginal lactic acid bacteria, including Lactobacillus, Pediococcus and Weissella, may lead to the design of better probiotic products as bacterial replacement therapy.     

Characterization of microbial communities found in the human vagina by analysis of terminal restriction fragment length polymorphisms of 16S rRNA genes

To define and monitor the structure of microbial communities found in the human vagina, a cultivation-independent approach based on analyses of terminal restriction fragment length polymorphisms (T-RFLP) of 16S rRNA genes was developed and validated. Sixteen bacterial strains commonly found in the human vagina were used to construct model communities that were subsequently used to develop efficient means for the isolation of genomic DNA and an optimal strategy for T-RFLP analyses. The various genera in the model community could best be resolved by digesting amplicons made using bacterial primers 8f and 926r with HaeIII; fewer strains could be resolved using other primer-enzyme combinations, and no combination successfully distinguished certain species of the same genus. To demonstrate the utility of the approach, samples from five women that had been collected over a 2-month period were analyzed. Differences and similarities among the vaginal microbial communities of the women were readily apparent. The T-RFLP data suggest that the communities of three women were dominated by a single phylotype, most likely species of Lactobacillus. In contrast, the communities of two other women included numerically abundant populations that differed from Lactobacillus strains whose 16S rRNA genes had been previously determined. The T-RFLP profiles of samples from all the women were largely invariant over time, indicating that the kinds and abundances of the numerically dominant populations were relatively stable throughout two menstrual cycles. These findings show that T-RFLP of 16S rRNA genes can be used to compare vaginal microbial communities and gain information about the numerically dominant populations that are present.     

Characterization of Microbial Communities Found in the Human Vagina by Analysis of Terminal Restriction Fragment Length Polymorphisms of 16S rRNA Genes

To define and monitor the structure of microbial communities found in the human vagina, a cultivation-independent approach based on analyses of terminal restriction fragment length polymorphisms (T-RFLP) of 16S rRNA genes was developed and validated. Sixteen bacterial strains commonly found in the human vagina were used to construct model communities that were subsequently used to develop efficient means for the isolation of genomic DNA and an optimal strategy for T-RFLP analyses. The various genera in the model community could best be resolved by digesting amplicons made using bacterial primers 8f and 926r with HaeIII; fewer strains could be resolved using other primer-enzyme combinations, and no combination successfully distinguished certain species of the same genus. To demonstrate the utility of the approach, samples from five women that had been collected over a 2-month period were analyzed. Differences and similarities among the vaginal microbial communities of the women were readily apparent. The T-RFLP data suggest that the communities of three women were dominated by a single phylotype, most likely species of Lactobacillus. In contrast, the communities of two other women included numerically abundant populations that differed from Lactobacillus strains whose 16S rRNA genes had been previously determined. The T-RFLP profiles of samples from all the women were largely invariant over time, indicating that the kinds and abundances of the numerically dominant populations were relatively stable throughout two menstrual cycles. These findings show that T-RFLP of 16S rRNA genes can be used to compare vaginal microbial communities and gain information about the numerically dominant populations that are present.     

Colony formation by human fibroblasts: Inhibition by multitransfusion and multiparous antisera

Human isoantisera from various sources were evaluated as potential selective agents for human diploid fibroblasts. Antisera inhibiting colony formation of some fibroblast strains were obtained from among subjects receiving multiple transfusions and also from multiparous women. Of 18 multitransfusion sera tested, 6 inhibited colony formation by some strains of a test panel of fibroblast strains. Thirty sera from multiparous women were tested for colony-inhibiting antibody against fibroblast strains from their offspring; two were positive. Antiserum fractionated by density gradient centrifugation contained colony-inhibiting activity in both the IgG and the IgM fractions. Multiparous antiserum reacted with a smaller proportion of tests strains than any of the multi-transfusion sera, suggesting that it had fewer antibody specificities. From studies of survival of mixtures of cells from an antiserum-sensitive and an antiserum-resistant strain in the presence of antiserum, it appeared that human isoantisera would be effective selective agents for human diploid fibroblasts.Supported by a grant (5R01 GM15883) and a Research Career Development Award (1K3 HD31, 721) from the National Institutes of Health.     

The genetic diversity and phenotypic characterisation of Streptococcus agalactiae isolates from Rio de Janeiro, Brazil

Streptococcus agalactiae isolates are more common among pregnant women, neonates and nonpregnant adults with underlying diseases compared to other demographic groups. In this study, we evaluate the genetic and phenotypic diversity in S. agalactiae strains from Rio de Janeiro (RJ) that were isolated from asymptomatic carriers. We analysed these S. agalactiae strains using pulsed-field gel electrophoresis (PFGE), serotyping and antimicrobial susceptibility testing, as well as by determining the macrolide resistance phenotype, and detecting the presence of the ermA/B, mefA/E and lnuB genes. The serotypes Ia, II, III and V were the most prevalent serotypes observed. The 60 strains analysed were susceptible to penicillin, vancomycin and levofloxacin. Resistance to clindamycin, chloramphenicol, erythromycin, rifampin and tetracycline was observed. Among the erythromycin and/or clindamycin resistant strains, the ermA, ermB and mefA/E genes were detected and the constitutive macrolides, lincosamides and streptogramin B-type resistance was the most prevalent phenotype observed. The lnuB gene was not detected in any of the strains studied. We found 56 PFGE electrophoretic profiles and only 22 of them were allocated in polymorphism patterns. This work presents data on the genetic diversity and prevalent capsular serotypes among RJ isolates. Approximately 85% of these strains came from pregnant women; therefore, these data may be helpful in developing future prophylaxis and treatment strategies for neonatal syndromes in RJ.     

Characterization of Staphylococcus aureus strains involved in human and bovine mastitis

Staphylococcus aureus is one of the main etiological agents of mastitis in different mammalian species. At present, it is unknown whether strains isolated from human mastitis cases share phenotypic properties and genetic background with those obtained from animal mastitis cases. Therefore, the objective of this study was to characterize S. aureus strains isolated from women with lactational mastitis and to compare them with the strains responsible for bovine mastitis and noninfectious strains. All the strains were genotyped by both pulsed field gel electrophoresis and multilocus sequence typing and submitted to a characterization scheme that included diverse assays related to pathogenic potential and antibiotic resistance. Apart from siderophore production, no significant association was observed between the strains from bovine and human mastitis. Statistical differences between human- and bovine-mastitis-associated strains were detected for some traits and virulence determinants, such as the presence of prophages and cna and hlb genes, which were more frequently found within the bovine group. On the contrary, resistance to penicillin was significantly higher among strains isolated from human lactational mastitis, probably related to the common presence of the blaZ gene. A high genetic diversity was found among the strains involved in mastitis in breastfeeding women.     

Biochemical typing as a method of differentiating group B streptococcus]

In epidemiological studies on the group B streptococcus the serological typing is used. The paper present the results of a study on usefulness of biochemical typing for differentiation of the group B streptococcus. For that purpose, 210 strains descended from colonized infants and pregnant women were put to typing with both of mentioned methods. We showed that each of the method distinguishes similar number of biotypes and serotypes. However, ought to be marked that significant number of strains (93.8%) belonged to the three out of eight biochemical types. Similar results were achieved in serological typing, three of the most numerous serotypes contained 81.4% strains. Analysis of the relationship between serological and biochemical types did not reveal statistical association because the strains belonged to various serotypes. Obtained results show that both methods of typing--biochemical and serological, have similar value in differentiation of the strains. The method of biochemical typing is quite simple and can be used in laboratory conditions.     

A numerical taxonomic study of anaerobic gram-negative bacilli classified as Bacteroides ureolyticus isolated from patients with non-gonococcal urethritis

A numerical taxonomic study of 64 strains of anaerobic Gram-negative bacilli isolated from men with non-gonococcal urethritis, two unclassified laboratory strains of 'corroding bacilli', and 12 other strains of anaerobic Gram-negative bacilli, including nine received as anaerobic curved rods and three as 'Bacteroides corrodens' (B. ureolyticus), isolated from women with bacterial vaginosis, was undertaken. Seventeen reference anaerobic strains belonging to the genera Bacteroides, Fusobacterium, Mobiluncus, Mitsuokella and Wolinella were included. Morphological, biochemical and physiological characteristics were examined in 103 tests. The resemblance between the 95 strains was calculated using the SSM, SJ and DP coefficients for cluster analyses based on the UPGMA method. All three approaches gave similar groupings, and the estimated average probability of test error was 2.46%. The strains fell into 10 phenons. The unclassified strains from men and three from women with lower genital-tract infections, and the laboratory strains of 'corroding bacilli' clustered in one phenon with the reference strains of B. ureolyticus, indicating that they correspond to B. ureolyticus. The other unclassified strains of anaerobic curved rods clustered as a distinct phenon. They correspond to species of the newly described genus Mobiluncus. The taxonomic data and the compilation of diagnostic tables serve as a useful guide for the laboratory identification of clinical isolates regarded as B. ureolyticus.     

Interleukin-18 and gestosis: correlation with Helicobacter pylori seropositivity

The aim of this brief communication was to determine the correlation between pre-eclampsia (PE), Helicobacter pylori pathogenic strains seropositivity, and interleukin-18 (IL-18) levels. To this purpose 25 pre-eclamptic women and 25 healthy parturient women of similar age were evaluated for: IL-18 levels, by ELISAH. pylori seropositivity, by anti-IgG ELISAAnti-Cag-A antibodies using a commercial immunoblot assay.We report similar values of IL-18 in our pre-eclamptic patients and in healthy parturient women (respectively 350 +/- 150 vs. 399 +/- 132 pg ml(-1); p = 0.23). However the seropositivity for H. pylori was 84 and 32% (p < 0.001), and anti-Cag-A antibodies were present respectively in 80 and 28% of the two populations. On the basis of our data we hypothesize that H. pylori infection from Cag-A strains can be involved in some cases of PE and that the microorganism could modulate IL-18 release. In fact, differences on IL-18 production have been described by different authors between pre-eclamptic and healthy pregnant women, independently from infective pathology.     

Characterization of vaginal lactobacilli in women after kidney transplantation

Limited number of publications described vaginal microflora after kidney transplantation. Our PubMed search revealed only 18 publications including words “vaginal bacteria &; kidney transplant” in the period of 1978–2011. The aim of this study was to characterize lactobacilli isolated from vaginal swabs of women after kidney transplantation, compared with healthy women. Eighteen renal transplant recipients (mean age 36.1) and 20 healthy women (mean age 36.0) were evaluated. Lactobacilli were cultured on MRS and Columbia blood agars. Biochemical identification with API 50 CHL (bioMerieux, Marcy L’Etoile, France) and multiplex PCR according to Song et al. was performed. Lactobacilli were tested for production of H₂O₂. Minimal inhibitory concentrations (MICs) of selected antimicrobial agents were determined with E-tests (bioMerieux, Marcy L’Etoile, France) and interpreted with CLSI and EUCAST criteria. No bacterial vaginosis was found among studied women. Two strains of group I were identified as Lactobacillus delbrueckii; 18 strains as Lactobacillus gasseri and 15 strains as Lactobacillus crispatus. Only 3 strains from group II were not identified by species-specific mPCR. Group IV was represented with 2 unidentified strains. Vaginal lactobacilli isolated from healthy women represented more homogenous group compared with heterogenous renal transplant recipients. Biochemical identification of lactobacilli by API 50 CHL kits was concordant with mPCR results only in 7 cases (17.5%), all 7 strains were identified as L. crispatus. Majority (93%) of lactobacilli were H₂O₂ producers. All isolated lactobacilli (100%) demonstrated high resistance to metronidazole (MIC > 256 μg/ml). Only 2 strains resistant to vancomycin (MICs: 32 and 256 μg/ml respectively), in the study and control group, and one to moxifloxacin (MIC = 32 μg/ml), were found. Resistance to metronidazole and vancomycin was concordant in CLSI and EUCAST (2010) criteria. Although significant differences between lactobacilli isolated from vaginas of kidney transplant and healthy women were not demonstrated, we demonstrated strains resistant to metronidazole, vancomycin and moxifloxacin in groups of examined women. Our study was performed on a small group of kidney transplant recipients and further more detailed molecular studies on a larger group of patients are required to confirm our results.     

Identification and plasmid profiles of Lactobacillus species from the vagina of 100 healthy women

Abstract It was hypothesized that Lactobacillus acidophilus might not be the dominant aerobic or microaerophilic Lactobacillus species in the human vagina, and that the dominant isolates are likely to express a variety of properties associated with colonization. To examine the question, vaginal swabs were collected from 100 healthy premenopausal women and cultured to detect the dominant aerobic or microaerophilic isolates of Lactobacillus . Only eight species were detected, with Lactobacillus jensenii being the dominant colonizers in the highest number (35) of women. Twelve different plasmid profiles were found amongst the isolates, but there was no association between plasmids and vaginal colonization or hydrogen peroxide production. Of the strains tested, few were resistant to the spermicidal agent nonoxynol-9. The data identify species within which strains may possess properties associated with maintenance of a healthy vaginal ecology     

Antibiotic sensitivity of Enterobacteriaceae isolated from women vagina and uterine cervix

Based on performed investigation in the group of 200 women treated for recurrent and chronic vaginitis and cervicitis, the characteristic of isolated microorganism was done. There were found series of drug-resistant bacteria in the vagina and uterine cervical canal in women with recurrent vaginitis and chronic cervicitis. In 42.6% of patients with diagnosed chronic cervicitis (Cervicitis chronica n = 50) Escherichia coli strains were isolated, 5% of which produced extended spectrum beta-lactamases (ES beta L). In women with recurrent vaginitis (Colpitis recidivans n = 150) Escherichia coli strains where isolated in 45.6%. Among them 4.5% produced ES beta L. Expression of beta-lactamases with broadened substrate spectrum was done in double-disc-test. CONCLUSIONS: 1. Study of antibiotic sensitivity of isolated bacteria should be a diagnostic standard in bacterial infections of uterine cervix and vagina. 2. The choice of appropriate antibiotics should take into consideration the drug-resistance mechanisms of isolated bacteria. 3. When drug-resistant bacteria are isolated, combined therapy should be applied.     

Genotypes and antibiotic resistances of Campylobacter jejuni and Campylobacter coli isolates from domestic and travel-associated human cases

Multilocus sequence typing (MLST) extended with flaB typing of 425 Campylobacter jejuni isolates and 42 Campylobacter coli isolates revealed quite a low overlap between human isolates from travel-associated and domestic cases in Switzerland. Men were more frequently affected by Campylobacter than women, but strains from women and, overall, from travel-associated cases showed mutations conferring quinolone resistance more frequently than strains from men and domestic cases, respectively.     

Microorganisms of thePleuropneumonia group (family ofMycoplasmataceae) in man

Summary Sixty one strains of pleuropneumonia-like organisms (P.P.L.O.) have been isolated in man: 25 from the discharge of patients with urogenital disorders, 13 from women who were pregnant or who had borne a child 6 weeks previously. In women the frequency of positive cultures increased with the disappearance of Lactobacilli. Twenty three strains were cultivated from the buccal mucosa of 27 healthy subjects. Classification by means of cultural appearance and biochemical behaviour allowed of differentiation into 4 groups, three of which were in conformity with three species ofMycoplasma described byEdward andFreundt. The only difference as compared with this latter classification is that with our methodM. fermentans andM. salivarius regularl gave clearcut hemolysis, which was not found byEdward. Two urogenital strains which grew well on not specially enriched media could not be identified.     

Presence of various Acinetobacter species in urinary tract infections

Species classification of 39 strains of Acinetobacter isolated from urinary tract infections (UTI) was performed by API 20NE tests. On the whole 24 biotypes were differentiated which formed 4 species and one biotype had no defined species classification. Acinetobacter baumani was most numerously represented. Seventy seven percent of strains were isolated from urine samples from women. About 90% of Acinetobacter strains were isolated as single pathogens. Sensitivity of these strains to 23 antibiotics and chemotherapeutics using ATB-UR and disc diffusion technique was evaluated. Of all cephalosporins tested ceftazidime was the most active. All strains tested were sensitive to imepenem. The high percentage of strains was sensitive to quinolones of IIIrd generation. The high percentage of strains was resistant to nitrofurantoin.     

Improved understanding of the bacterial vaginal microbiota of women before and after probiotic instillation

The vaginal bacterial microbiota of 19 premenopausal women was examined by PCR-denaturing gradient gel electrophoresis (DGGE) and sequencing of the V2-V3 region of the 16S rRNA gene. Ten of the women were studied further to investigate the effect and persistence of vaginally inserted capsules containing viable lactobacilli. PCR-DGGE indicated that most subjects had a microbiota represented by one to three dominant DNA fragments. Analysis of these fragments revealed that 79% of the women possessed sequences with high levels of similarity to Lactobacillus species sequences. Sequences homologous to Lactobacillus iners sequences were the most common and were detected in 42% of the women tested. Alteration of the vaginal microbiota could be detected by PCR-DGGE in several women after the instillation of lactobacilli. Additionally, randomly amplified polymorphic DNA analysis of lactobacilli isolated from selective media demonstrated that the exogenous strains could be detected for up to 21 days in some subjects. This study demonstrates that non-culture-based techniques, such as PCR-DGGE, are useful adjuncts for studies of the vaginal microbiota.     

Role of Staphylococcus saprophyticus in urinary infections]

A proportion of S. saprophyticus in other coagulase-negative staphylococcal isolates from the urine of patients with urinary infections and healthy individuals has been investigated. Certain diagnostic aspect of the urinary infections with S. saprophyticus have also been considered. Hundred four coagulase-negative staphylococcal strains isolated from patients in S?upsk and Gdańsk area and 72 strains of the coagulase-negative staphylococci isolated from the urine of healthy women have been divided into 9 species, according to Kloos and Schleifers' classification. Bacteriologic tests have shown that S. saprophyticus produced 20.4% of the urinary tract infections in S?upsk area holding the second place after S. haemolyticus (27.3%). This species was the most infrequent in the urine of patients in Gdańsk area (3.3%). Its sensitivity to antibiotics did not differ from that of other coagulase-negative staphylococci. In contrast to the majority of other strains, S. saprophyticus has not been isolated from the urinary tract of healthy women and has been encountered most frequently in low bacteriuria. Test of resistance to novobiocin which is considered as a simplified identification method of this species proved to be not very precise as other species have also been resistant to this antibiotic.