BackgroundStreptococcus agalactiae (group B streptococcus; GBS) is a significant cause of perinatal and neonatal infections worldwide. To detect GBS colonization in pregnant women, the CDC recommends isolation of the bacterium from vaginal and anorectal swab samples by growth in a selective enrichment medium, such as Lim broth (Todd-Hewitt broth supplemented with selective antibiotics), followed by subculture on sheep blood agar. However, this procedure may require 48 h to complete. We compared different sampling and culture techniques for the detection of GBS.MethodsA total of 300 swabs was taken from 100 pregnant women at 35-37 weeks of gestation. For each subject, one rectovaginal, one vaginal and one rectal ESwab were collected. Plating onto Columbia CNA agar (CNA), group B streptococcus differential agar (GBSDA) (Granada Medium) and chromID Strepto B agar (CA), with and without Lim broth enrichment, were compared. The isolates were confirmed as S. agalactiae using the CAMP test on blood agar and by molecular identification with tDNA-PCR or by 16S rRNA gene sequence determination.ResultsThe overall GBS colonization rate was 22%. GBS positivity for rectovaginal sampling (100%) was significantly higher than detection on the basis of vaginal sampling (50%), but not significantly higher than for rectal sampling (82%). Direct plating of the rectovaginal swab on CNA, GBSDA and CA resulted in detection of 59, 91 and 95% of the carriers, respectively, whereas subculturing of Lim broth yielded 77, 95 and 100% positivity, respectively. Lim broth enrichment enabled the detection of only one additional GBS positive subject. There was no significant difference between GBSDA and CA, whereas both were more sensitive than CNA. Direct culture onto GBSDA or CA (91 and 95%) detected more carriers than Lim broth enrichment and subculture onto CNA (77%). One false negative isolate was observed on GBSDA, and three false positives on CA.ConclusionsIn conclusion, rectovaginal sampling increased the number GBS positive women detected, compared to vaginal and/or rectal sampling. Direct plating on CA and/or GBSDA provided rapid detection of GBS that was at least as sensitive and specific as the CDC recommended method of Lim broth subcultured onto non chromogenic agar.

BackgroundNasal carriage of Staphylococcus aureus among hospital personnel is a common cause of hospital acquired infections. Emergence of drug resistant strains especially methicillin resistant S. aureus (MRSA) is a serious problem in hospital environment. Therefore, the aim of this study was to determine the nasal carriage rate of S. aureus and MRSA among Health Care Workers (HCWs) at Al Shifa Hospital, the major hospital in Gaza Strip.MethodsA cross sectional study was conducted on 200 HCWs. Nasal swabs were collected during February — April 2015, and cultured on blood and mannitol salt agar. The isolates were identified as S. aureus based on morphology, coagulase test, DNase test and mannitol salt agar fermentation. Disk diffusion antibiotic susceptibility tests were performed according to the guidelines of the Clinical and Laboratory Standards Institute. MRSA were confirmed by detection of the mecA gene by PCR.ResultsOut of the 200 healthcare workers, 62 (31%) carried S. aureus, of which 51 (82.3%) were MRSA. Therefore, 25.5% of all HCWs were identified as MRSA carriers. MRSA carriage rate was highest among nurses (30.4%) whereas the carriage rate among doctors was (16%). The majority of MRSA carriers were workers of internal medicine department and surgical wards (41.3 and 35% respectively). Out of the 51 MRSA isolates identified by oxacillin disc resistance, 40 were confirmed by PCR targeting the mecA gene. Penicillin showed the highest rate of resistance among MRSA and MSSA isolates (100%).ConclusionsThe high rate of nasal MRSA carriage among healthcare workers found in this study is alarming and highlights the need for adjusted infection control measures to prevent MRSA transmission from HCWs to the vulnerable patient.

BackgroundThe vaginal microflora is important for maintaining vaginal health and preventing infections of the reproductive tract. The rectum has been suggested as the major source for the colonisation of the vaginal econiche.MethodsTo establish whether the rectum can serve as a possible bacterial reservoir for colonisation of the vaginal econiche, we cultured vaginal and rectal specimens from pregnant women at 35-37 weeks of gestation, identified the isolates to the species level with tRNA intergenic length polymorphism analysis (tDNA-PCR) and genotyped the isolates for those subjects from which the same species was isolated simultaneously vaginally and rectally, by RAPD-analysis.One vaginal and one rectal swab were collected from a total of each of 132 pregnant women at 35-37 weeks of gestation. Swabs were cultured on Columbia CNA agar and MRS agar. For each subject 4 colonies were selected for each of both sites, i.e. 8 colonies in total.ResultsAmong the 844 isolates that could be identified by tDNA-PCR, a total of 63 bacterial species were present, 9 (14%) only vaginally, 26 (41%) only rectally, and 28 (44%) in both vagina and rectum. A total of 121 (91.6%) of 132 vaginal samples and 51 (38.6%) of 132 rectal samples were positive for lactobacilli. L. crispatus was the most frequently isolated Lactobacillus species from the vagina (40% of the subjects were positive), followed by L. jensenii (32%), L. gasseri (30%) and L. iners (11%). L. gasseri was the most frequently isolated Lactobacillus species from the rectum (15%), followed by L. jensenii (12%), L. crispatus (11%) and L. iners (2%).A total of 47 pregnant women carried the same species vaginally and rectally. This resulted in 50 vaginal/rectal pairs of the same species, for a total of eight different species. For 34 of the 50 species pairs (68%), isolates with the same genotype were present vaginally and rectally and a high level of genotypic diversity within species per subject was also established.ConclusionIt can be concluded that there is a certain degree of correspondence between the vaginal and rectal microflora, not only with regard to species composition but also with regard to strain identity between vaginal and rectal isolates.These results support the hypothesis that the rectal microflora serves as a reservoir for colonisation of the vaginal econiche.

Although the heterogeneity of Mycobacterium tuberculosis populations and the existence of mixed infections are now generally accepted, systematic studies on their relative importance are rare. In the present study, 10 individual colonies of each M. tuberculosis isolate (primary isolate) from 97 tuberculosis patients in a primarily human immunodeficiency virus-negative population were screened for heterogeneity and detectable mixed infections by spoligotyping, IS6110-based restriction fragment length polymorphism analysis, and mycobacterial interspersed repetitive unit-variable number of tandem repeat typing. The MICs of antituberculosis drugs for colonies with divergent fingerprints were determined. Infections with different bacterial subpopulations were detected in the samples from eight patients (8.2%), and the frequency of detectable mixed infections in the study population was estimated to be 2.1%. Genotypic variations were found to be independent of the drug susceptibilities, and the various molecular markers evolved independently in most cases. The predominant strains and the primary isolates always had concordant drug susceptibility and MIC testing results. These findings have implications on the interpretation of molecular epidemiology results for patient follow-up and in transmission studies.Tuberculosis (TB) patients have often been assumed to be infected with a single Mycobacterium tuberculosis strain, and infection with one strain is thought to confer immunity to additional M. tuberculosis infections. Therefore, a recurrence of disease after treatment is often considered to be caused by the same strain that caused the original infection. However, cases of reinfection by a second M. tuberculosis strain and occasional infection with more than one strain have been documented (2,5,12,22,31), and the occurrence of mixed infections has now become generally accepted. Infection with multiple strains of M. tuberculosis can seriously confuse interpretation of drug susceptibility testing (DST) results and the detection of epidemiological links (2, 44). However, the frequency of mixed infections and the appreciation of their impact on the phenotypic and the genotypic properties of the primary isolates largely remain unknown, essentially because of the lack of systematic studies on the heterogeneity within M. tuberculosis populations from an individual patient.Almost 30 years ago, Mankiewicz and Liivak (19) used phage typing to analyze the heterogeneity among individual colonies obtained from cultures of specimens isolated from 233 Eskimo patients, leading to the conclusion that 14.1% of the patients tested were simultaneously infected with more than one M. tuberculosis strain. The development of DNA fingerprinting tools has, meanwhile, considerably improved the capacity to distinguish M. tuberculosis strains. Among these tools, IS6110-based restriction fragment length polymorphism (IS6110-RFLP) analysis is considered the standard technique for comparison of M. tuberculosis isolates (32, 33). Other techniques, su...

BackgroundGroup B streptococci (GBS), or Streptococcus agalactiae, are the leading bacterial cause of meningitis and bacterial sepsis in newborns. Here we compared different culture media for GBS detection and we compared the occurrence of different genotypes and serotypes of GBS isolates from the vagina and rectum.MethodsStreptococcus agalactiae was cultured separately from both rectum and vagina, for a total of 150 pregnant women, i) directly onto Columbia CNA agar, or indirectly onto ii) Granada agar resp. iii) Columbia CNA agar, after overnight incubation in Lim broth.ResultsThirty six women (24%) were colonized by GBS. Of these, 19 harbored GBS in both rectum and vagina, 9 only in the vagina and 8 exclusively in the rectum. The combination of Lim broth and subculture on Granada agar was the only culture method that detected all GBS positive women. Using RAPD-analysis, a total of 66 genotypes could be established among the 118 isolates from 32 women for which fingerprinting was carried out. Up to 4 different genotypes in total (rectal + vaginal) were found for 4 women, one woman carried 3 different genotypes vaginally and 14 women carried two 2 different genotypes vaginally. Only two subjects were found to carry strains with the same genotype, although the serotype of both of these strains was different.Eighteen of the 19 subjects with GBS at both sites had at least one vaginal and one rectal isolate with the same genotype.We report the presence of two to four different genotypes in 22 (61%) of the 36 GBS positive women and the presence of identical genotypes in both sites for all women but one.ConclusionThe combination of Lim broth and subculture on Granada medium provide high sensitivity for GBS detection from vaginal and rectal swabs from pregnant women. We established a higher genotypic diversity per individual than other studies, with up to four different genotypes among a maximum of 6 isolates per individual picked. Still, 18 of the 19 women with GBS from both rectum and vagina had at least one isolate from each sampling site with the same genotype.