Background. Elucidation of a pathogen's antimicrobial susceptibility requires subculture after the organism is first isolated. This takes several days, requiring patients to be treated with broad-spectrum antibiotics. This approach contributes to the development of bacterial resistance. Methods. Microtiter wells were coated with a polyclonal antibody targeting the pathogen of interest. Bacterial suspensions were added in the presence/absence of selected antibiotics. After washing, captured bacteria were detected. Findings. Group B streptococcus (GBS), Enterococcus faecalis, and Neisseria gonorrhoeae were each detected at 105 bacteria/mL following a 20-minute incubation period. Susceptibility to select antibiotics was discernable following a 6-hour incubation period (GBS and Enterococcus). Sensitivity was increased to 10−2 bacteria/mL for GBS, 10−1 bacteria/mL for E. faecalis, and 101 bacteria/mL for N. gonorrhoeae following 18–24-hour culture. Conclusion. This novel assay allows for the highly sensitive and specific identification of a pathogen and simultaneous determination of its antimicrobial susceptibility in a reduced time.
Objective. To determine the validity of a novel Group B Streptococcus (GBS) diagnostic assay for the detection of GBS in antepartum patients. Study Design. Women were screened for GBS colonization at 35 to 37 weeks of gestation. Three vaginal-rectal swabs were collected per patient; two were processed by traditional culture (commercial laboratory versus in-house culture), and the third was processed by an immunoblot-based test, in which a sample is placed over an antibody-coated nitrocellulose membrane, and after a six-hour culture, bound GBS is detected with a secondary antibody. Results. 356 patients were evaluated. Commercial processing revealed a GBS prevalence rate of 85/356 (23.6%). In-house culture provided a prevalence rate of 105/356 (29.5%). When the accelerated GBS test result was compared to the in-house GBS culture, it demonstrated a sensitivity of 97.1% and a specificity of 88.4%. Interobserver reliability for the novel GBS test was 88.2%. Conclusions. The accelerated GBS test provides a high level of validity for the detection of GBS colonization in antepartum patients within 6.5 hours and demonstrates a substantial agreement between observers.
Objective. To assess the effect of universal screening and administration of intrapartum antibiotic prophylaxis to prevent early-onset neonatal GBS sepsis at a private tertiary care hospital since issuance of the 2002 CDC guidelines for preventing perinatal GBS disease. Methods. Retrospective analysis of women delivering between January 1, 2003 and December 31, 2004 at a private tertiary care hospital in Houston, Texas. The percentage of women screened, GBS positive women receiving intrapartum antibiotic prophylaxis, and infants developing early-onset GBS sepsis were determined. Results. 2,108 women delivered 2,135 infants with 1,874 (89%) screened for GBS. Of those screened, 1,322 (71%) tested negative and 552 (29%) tested positive for GBS. In this analysis of 2,135 infants, 3 (0.94 cases/1,000 live births) were diagnosed with invasive GBS sepsis. Conclusion. High rates of screening of pregnant women for GBS colonization and use of intrapartum antibiotic prophylaxis for GBS carriers can be achieved in a private tertiary care hospital setting. “Synopsis: High screening rates for group B streptococcus in a private tertiary care hospital reduce the incidence of maternal and early onset neonatal GBS infection.”
Bacterial vaginosis (BV), a condition affecting millions of women each year, is primarily caused by the gram-variable organism Gardnerella vaginalis. A number of organisms associated with BV cases have been reported to develop multidrug resistance, leading to the need for alternative therapies. Previously, we reported the antimicrobial peptide subtilosin has proven antimicrobial activity against G. vaginalis, but not against the tested healthy vaginal microbiota of lactobacilli. After conducting tissue sensitivity assays using an ectocervical tissue model, we determined that human cells remained viable after prolonged exposures to partially-purified subtilosin, indicating the compound is safe for human use. Subtilosin was shown to eliminate the motility and forward progression of human spermatozoa in a dose-dependent manner, and can therefore be considered a general spermicidal agent. These results suggest subtilosin would be a valuable component in topical personal care products aimed at contraception and BV prophylaxis and treatment.
OBJECTIVES: To determine the mechanism of antimicrobial action of lactocin 160, a bacteriocin produced by the healthy vaginal strain of Lactobacillus rhamnosus, using an established model, with Micrococcus luteus ATCC 10420 as a test organism. METHODS: Sensitivity of M. luteus to lactocin 160 was determined by the diffusion assay. Loss of cellular ATP in the lactocin-treated cells was elucidated using a commercially available ATP determination kit (luciferin-luciferase bioluminescence assay). Luminescence intensity as a reflection of ATP quantity was determined using a luminometer. Dissipation of membrane potential (Deltapsi) was studied using fluorophore DiSC3(5) with the fluorescence spectrum sensitive to changes in Deltapsi. RESULTS: Lactocin 160 inhibited growth of M. luteus ATCC 10420 at a concentration of 5 microg/ml. There were no significant changes in the intracellular ATP level of M. luteus upon the addition of 20 microg/ml of lactocin 160. However, the extracellular ATP level increased significantly. This means that the treatment of cells with lactocin 160 resulted in an efflux of ATP from inside the cells. Therefore, a partially purified lactocin 160 preparation (16 microg /ml of the bacteriocin in the sample) killed sensitive cells and dissipated 3.12 +/- 0.36% of Deltapsi. CONCLUSION: Lactocin 160 has a mode of action typical for bacteriocins. It disturbs the cellular membrane (Deltapsi dissipation) and induces ATP efflux, most likely because of the pore formation, which is a common mechanism of action for many bacteriocins.
OBJECTIVE: To determine whether a single dose of Clindesse vaginal cream is comparable in efficacy and safety to Cleocin vaginal cream administered once daily for 7 days in the treatment of bacterial vaginosis. STUDY DESIGN: This multicenter, randomized, single-blind, parallel-group study enrolled 540 patients with BV infections. Treatment consisted of either a single intravaginal dose of Clindesse or 7 daily doses of Cleocin. Efficacy and safety were assessed 21-30 days after the start of treatment. The efficacy endpoints were Investigator Cure, Clinical Cure (a composite of all 4 Amsel's criteria and Investigator Cure), Nugent Cure (Nugent score < 4), and Therapeutic Cure (a composite of Clinical Cure and Nugent Cure). Resolution of individual Amsel's criteria was also evaluated. Treatment-emergent adverse events were monitored throughout the study. RESULTS: There were no significant differences in cure rates between the Clindesse and Cleocin treatment groups in Investigator Cure (P=0.702), Clinical Cure (P=0.945), Nugent Cure (P=0.788), or Therapeutic Cure (P=0.572). Results were also similar for 3 of 4 and 2 of 4 Amsel's criteria and for each individual Amsel's criterion (all P-values >0.200). Ninety-five percent confidence intervals for each endpoint were consistent with equivalence between the 2 products. There was no significant difference between the treatment groups in the incidence of treatment-emergent adverse events (P=0.386). CONCLUSIONS: A single dose of Clindesse vaginal cream is equivalent in safety and efficacy to a 7-dose regimen of Cleocin vaginal cream in the treatment of bacterial vaginosis. This represents a significant advance in the treatment of bacterial vaginosis.
OBJECTIVES: To determine the in vitro resistance of group B streptococcus (GBS) to 12 antibiotics. To determine if there has been any decrease in sensitivity to the penicillins or other antibiotics currently used for GBS chemoprophylaxis in pregnant women. Find suitable alternative antibiotics to penicillin. Find an antibiotic that will have minimal selective pressure for resistance among the endogenous resident vaginal microflora. METHODS: The antibiotic susceptibility profiles of 52 clinical isolates of GBS were evaluated to 12 antibiotics: ampicillin, azithromycin, cefamandole, cefazolin, ceftriaxone, ciprofloxacin, clindamycin, erythromycin, nitrofurantoin, ofloxacin, penicillin and vancomycin. Antibiotic sensitivities were determined using disk diffusion and microdilution methods according to the guidelines of the National Committee for Clinical Laboratory Standards (NCCLS). RESULTS: All isolates were sensitive to vancomycin, ofloxacin, ampicillin, ciprofloxacin, nitrofurantoin and penicillin. However, the following number of clinical isolates exhibited intermediate or decreased sensitivity, nine (17%) to ampicillin, eight (15%) to penicillin, 14 (32%) to ciprofloxacin and one (2%) to nitrofurantoin. Thirty-one percent of the isolates were resistant to azithromycin and ceftriaxone, 19% to clindamycin, 15% to cefazolin and 13% to cefamandole. Eighteen (35%) of the clinical isolates tested were resistant to 6 of the 12 antibiotics tested. CONCLUSIONS: The relatively high rates of resistance for 6 of the 12 antibiotics tested suggest that for women allergic to penicillin and colonized with GBS, antibiotic sensitivities to their isolates should be determined. The antibiotic selected for intrapartum chemoprophylaxis should be guided by the organism's antibiotic sensitivity pattern. Patients with GBS bacteriuria should be treated with nitrofurantoin.
OBJECTIVE: To identify alterations in the cytokine profile and microbial ecosystem of the vagina in association with cervical dysplasia. METHODS: Demographics, lifestyle variables and Papanicolau (Pap) smear results of subjects presenting to the same site for gynecologic complaints, obstetric visits or colposcopy were prospectively recorded. Vaginal smear for Gram stain, aerobic and anaerobic culture, pH, and wet mount and KOH examination for Trichomonas vaginalis, Gardnerella vaginalis and yeast organisms were performed. Vaginal lavage specimens were centrifuged, and the pellets and supernatants were assayed for human papillomavirus (HPV) by polymerase chain reaction and for cytokines interleukin (IL)-1beta IL-6, IL-10 and IL-12 by enzyme-linked immunosorbent assay (ELISA) respectively. Subjects with abnormal Pap smears underwent colposcopy and biopsy as indicated. RESULTS: Of 51 patients, 32 were referred for colposcopy, 12 presented with gynecologic needs, and seven presented for obstetric visits. Median age was 24 years. Demographics did not differ significantly between the dysplasia and control groups except for a trend towards more sexual partners in the dysplasia group. Biopsies were performed in 81% (26/32) of patients presenting for colposcopy and 17 revealed cervical intraepithelial neoplasia. IL-1beta, IL-6, IL-10, and IL-12 levels were elevated in 63% (20/32), 38% (15/39), 4% (2/49), and 0% of samples respectively. Elevated vaginal lavage IL-1beta was associated with a 6.1 odds ratio (95% confidence interval 1.06-35) of cervical dysplasia. Alterations in other variables studied were not associated with cervical dysplasia. CONCLUSIONS: Elevated IL-1beta, possibly representing a complex host inflammatory response to multiple pathogens, was demonstrated in patients with cervical dysplasia.
Objective: To evaluate the in vitro effect of varying concentrations of clindamycin on Lactobacillus spp.
Methods: Concentrations of clindamycin ranging from 1.95–20 000 mg/ml were studied for their effect on the
growth of six strains of Lactobacillus .
Results: Clindamycin concentrations between 1.95–31.25 mg/ml had no statistically significant effect on growth of
lactobacilli (p > 0.05). Concentrations 125 and 250 mg/ml had a bacteriostatic effect. The mean minimum inhibitory
concentration (MIC) for studied Lactobacillus strains was determined as 1000 mg/ml.
Conclusion: High concentrations of clindamycin achieved in the vagina by intravaginal application might be
inhibitory for Lactobacillus .
Objective: To determine whether metronidazole has an adverse effect on the growth of Lactobacillus.
Methods: Hydrogen peroxide- and bacteriocin-producing strains of Lactobacillus were used as test strains.
Concentrations of metronidazole used ranged from 128 to 7000 μg/ml. Susceptibility to metronidazole was conducted
by the broth microdilution method recommended by the National Committee for Clinical Laboratory Standards.
Results: Growth of Lactobacillus was partially inhibited at concentrations between 1000 and 4000 μg/ml
(p = 0.014). Concentrations ≥ 5000 μg/ml completely inhibited growth of Lactobacillus. Concentrations between
128 and 256 μg/ml stimulated growth of Lactobacillus (p = 0.025 and 0.005, respectively). Concentrations of
metronidazole between 64 and 128 μg/ml or ≥ 512 μg/ml did not have an inhibitory or a stimulatory effect on the growth of Lactobacillus compared to the control.
Conclusions: High concentration of metronidazole, i.e. between 1000 and 4000 μg/ml, partially inhibited the
growth of Lactobacillus. Concentrations ≥ 5000 μg/ml completely suppressed the growth of Lactobacillus.
Concentrations between ≥ 128 and ≤ 256 μg/ml stimulated the growth of Lactobacillus. Further investigation to
determine the ideal concentration of metronidazole is needed in order to use the antimicrobial agent effectively in
the treatment of bacterial vaginosis.
Objective: To isolate bacteriocin from a vaginal strain of Lactobacillus acidophilus.
Methods: L. acidophilus 160 was grown on two media. The first was MRS broth for 18 hours; the cells were
harvested, washed, and placed into a chemically defined medium. The second medium resembled vaginal fluid
minus protein. Bacteriocin was precipitated from both media using ammonium sulfate. The growth-inhibiting
activity of bacteriocin was determined by a bioassay using nine different isolates of Gardnerella vaginalis.
Results: MRS broth is not a suitable medium for extracting bacteriocin, because it binds with Tween 80.
Bacteriocin was isolated, without contaminating constituents, from chemically defined medium and identified as a
single band by electrophoresis. Bacteriocin has a molecular weight of 3.8 kDa. All nine isolates of Gardnerella were inhibited by the bacteriocin isolated from L. acidophilus 160.
Conclusions: Bacteriocin produced by L. acidophilus 160 was isolated from the chemically defined medium
(starvation medium) in a partially pure form. L. acidophilus 160 bacteriocin inhibited growth of all nine isolates of Gardnerella vaginalis.
Objective: We sought to determine if preterm labor is associated with the degree of maternal hypoxia
in pregnant women with community-acquired pneumonia but no other maternal diseases.
Methods: We retrospectively reviewed the medical records of all antepartum patients admitted
with a diagnosis of community-acquired pneumonia to an inner-city university hospital between
1983 and 1987. Included in this review were only the patients with radiologically confirmed diagnose
of pneumonia and documented arterial blood gases on room air at the time of admission, but no
other maternal diseases.
Results: A total of 22 cases were identified. There was no maternal mortality, but there were 2
patients (9%) who developed respiratory failure requiring mechanical ventilation. Bacteremia with
Streptococcus pneumoniae was documented in 1 patient (5%). Preterm labor complicated 5 cases
(23%) and led to preterm delivery in 3 patients (14%). Terbutaline tocolysis was instituted in 3
patients, but was discontinued in 1 patient who was allowed to deliver because of her worsening
condition. Preterm labor was associated with the WBC count on admission, usually > 18,000/mm3,
but no statistically significant correlation with the severity of maternal hypoxia was noted. Five
patients (23%) were incorrectly diagnosed at the time of admission, 4 with an initial diagnosis of
pyelonephritis and 1 with an initial diagnosis of cholecystitis.
Conclusions: Community-acquired pneumonia in the antepartum period is responsible for significant
maternal and fetal complications even in the absence of other maternal diseases. Preterm
labor and delivery remain frequent, and tocolysis should be used cautiously. At the time of admission,
the diagnosis may be difficult. The degree of maternal hypoxia on admission does not correlate
with the presence of preterm labor.