The lower genital tract microbiota in relation to cytokine-, SLPI- and endotoxin levels: application of checkerboard DNA-DNA hybridization (CDH).

In the present study the lower genital tract microbiota in asymptomatic fertile women (n=34) was identified and quantified by culturing vaginal secretions. Also, vaginal and cervical samples were analyzed by a semiquantitative checkerboard DNA-DNA hybridization technique (CDH) based on genomic probes prepared from 13 bacterial species (Bacteroides ureolyticus, Escherichia coli, Fusobacterium nucleatum, Gardnerella vaginalis, Mobiluncus curtisii ss curtisii, Prevotella bivia, Prevotella disiens, Prevotella melaninogenica, Atopobium vaginae, Lactobacillus iners, Staphylococcus aureus ss aureus, Streptococcus anginosus, and Streptococcus agalactiae). The bacterial species found by either culture or CDH were correlated with proinflammatory cytokines (IL-1 alpha, IL-1 beta, IL-6, IL-8), secretory leukocyte protease inhibitor (SLPI), and endotoxin in the cervicovaginal samples. Grading the women into healthy, intermediate, or bacterial vaginosis (BV) as based on Gram staining of vaginal smears, the viable counts of lactobacilli (L. gasseri) and of streptococci-staphylococci combined were highest in the intermediate group. In BV, particularly the high concentrations of Actinomyces urogenitalis, Atopobium vaginae, and Peptoniphilus harei were noted (>or=10(11) per ml). The total viable counts correlated with both cervical IL-1 alpha and IL-1 beta. A strong negative correlation was observed between L. iners and total viable counts, G. vaginalis, or cervical IL-1 alpha, while it correlated positively with SLPI. Analysis of vaginal and cervical samples from 26 out of the 34 women by CDH showed that anaerobic bacteria were more frequently detected by CDH compared to culture. By this method, A. vaginae correlated with G. vaginalis, and L. iners with S. aureus. With regard to cytokines, B. ureolyticus correlated with both cervical and vaginal IL-1 alpha as well as with cervical IL-8, while F. nucleatum, S. agalactiae, S. anginosus, or S. aureus correlated with vaginal IL-1 alpha. Furthermore, all Gram-negative bacteria taken together, as measured by CDH, correlated with vaginal endotoxin and inversely with vaginal SLPI. The significance of the results is discussed. In summary, mapping of the identity and quantity of vaginal bacterial species and their association with locally produced host innate immune factors will help in defining various types of abnormal vaginal microbiota, developing new ways of assessing the risk of ascending subclinical infections, and in treating them. CDH appears to be a suitable tool for future analyses of large numbers of clinical samples with an extended number of bacterial probes.

Transfer of members of the genus Falcivibrio to the genus Mobiluncus, and emended description of the genus Mobiluncus.

It has long been thought that the genera Mobiluncus and Falcivibrio contain the same organisms. Using a polyphasic approach, it was found that Mobiluncus curtisii and Mobiluncus mulieris were the same as Falcivibrio vaginalis and Falcivibrio grandis, respectively. As the genus name Mobiluncus takes precedence, it is proposed that F. vaginalis and F. grandis be transferred to the genus Mobiluncus. In agreement with previous studies, results from phenotypic tests did not support the separation of M. curtisii strains into its two subspecies, M. curtisii subsp. curtisii and M. curtisii subsp. holmesii. Phenotypic complexity within M. curtisii dictates that the species should be treated as a complex until more in-depth analyses of the species have been performed. Phylogenetic analyses, based on 16S rRNA gene sequences, demonstrated that the genus Mobiluncus was associated with Varibaculum cambriense and the two subspecies of Actinomyces neuii, and that A. neuii is only distantly related to Actinomyces sensu stricto.

Luteococcus sanguinis sp. nov., isolated from human blood.

An unusual catalase-positive, Gram-positive, coccus-shaped bacterium that originated from a human blood specimen was subjected to a polyphasic taxonomic study. Cell-wall murein and lipid composition analyses indicated that the unknown isolate was a member of the genus Luteococcus. The results of comparative 16S rRNA gene sequence analysis were consistent with chemotaxonomic findings and showed that the unidentified bacterium represents a hitherto unknown sublineage within the genus Luteococcus that is closely related to, but distinct from, Luteococcus japonicus. On the basis of both phenotypic and phylogenetic evidence, it is proposed that the unknown bacterium from human blood should be classified as Luteococcus sanguinis sp. nov., with the type strain CCUG 33897(T) (=CIP 107216(T)).

Interleukin-18 in cervical mucus and amniotic fluid: relationship to microbial invasion of the amniotic fluid, intra-amniotic inflammation and preterm delivery.

OBJECTIVE: To evaluate the relationship between interleukin (IL)-18 in cervical mucus and amniotic fluid and microbial invasion of amniotic fluid, preterm delivery and intra-amniotic inflammation in women in preterm labour, with preterm prelabour rupture of membranes and at term. DESIGN: A prospective follow up study. SETTING: Sahlgrenska University Hospital, Göteborg, Sweden. SAMPLE: Women with singleton pregnancies (<34 weeks) presenting with preterm labour (n = 87) or preterm prelabour rupture of membranes (n = 47) and women, not in labour, at term (n = 28). METHODS: Amniotic fluid was retrieved transabdominally. Cervical mucus was taken from the uterine cervix of women in preterm labour and at term. IL-18 was analysed with enzyme-linked immunosorbent assay. MAIN OUTCOME MEASURES: IL-18 in relation to microbial invasion of the amniotic fluid, delivery within seven days or <34 weeks of gestation and intra-amniotic inflammation. RESULTS: The levels of IL-18 in cervical mucus and amniotic fluid were higher in women with preterm labour than in those not in labour at term. In the preterm labour group, significant associations were found between elevated IL-18 in amniotic fluid and microbial invasion of the amniotic fluid, as well as between delivery within seven days or <34 weeks of gestation and intra-amniotic inflammation. Delivery was delayed longer in the preterm prelabour rupture of membranes subgroup with IL-18 >or=1.0 ng/mL than in that with IL-18 <1.0 ng/mL. CONCLUSIONS: In the preterm labour group, high IL-18 in amniotic fluid (but not in the cervix) was associated with microbial invasion of the amniotic fluid, intra-amniotic inflammation and prompt delivery. On the other hand, elevated IL-18 in preterm prelabour rupture of the membranes group correlated with a longer interval to delivery.

Microbial invasion and cytokine response in amniotic fluid in a Swedish population of women with preterm prelabor rupture of membranes.

BACKGROUND: Previous studies have shown an association between intra-amniotic microbial invasion and/or inflammation and spontaneous preterm birth. The aim of this study was to investigate the occurrence of intra-amniotic microorganisms and cytokines [interleukin (IL)-6 and IL-8] in a Swedish population, with low incidence of preterm birth, of women with preterm prelabor rupture of membranes and their correlation to preterm birth. METHODS: Amniotic fluid was retrieved transabdominally from 58 patients with preterm prelabor rupture of membranes before 34 weeks of gestation. Polymerase chain reaction (PCR) analyses for Ureaplasma urealyticum and Mycoplasma hominis and culture for aerobic and anaerobic bacteria were performed. IL-6 and IL-8 were analyzed with enzyme-linked immunosorbent assay (ELISA). RESULTS: Microorganisms in amniotic fluid were detected in 13 patients (25%). Patients with bacteria detected in the amniotic fluid had significantly higher levels of IL-6 and IL-8. An amniotic fluid concentration of IL-6 >/= 0.80 ng/ml [relative risk 1.93, 95% confidence interval (CI) 1.13-3.29, sensitivity 63%, specificity 75%] was associated with an increased risk of delivery within 7 days. There was also an association between IL-8 and preterm birth (< 34 weeks). CONCLUSIONS: Intra-amniotic microbial invasion and inflammation in this population of Swedish women with preterm prelabor rupture of membranes were similar to data reported from populations with a higher incidence of preterm delivery. Amniotic IL-6 correlated to the presence of microorganisms and delivery within 7 days and IL-8 to delivery before 34 weeks.