Geert Claeys

BACKGROUND: Patients with severe asthma are at increased risk of exacerbations and lower respiratory tract infections (LRTI). Severe asthma is heterogeneous, encompassing eosinophilic and non-eosinophilic (mainly neutrophilic) phenotypes. Patients with neutropilic airway diseases may benefit from macrolides. METHODS: We performed a randomised double-blind placebo-controlled trial in subjects with exacerbation-prone severe asthma. Subjects received low-dose azithromycin (n=55) or placebo (n=54) as add-on treatment to combination therapy of inhaled corticosteroids and long-acting β2 agonists for 6 months. The primary outcome was the rate of severe exacerbations and LRTI requiring treatment with antibiotics during the 26-week treatment phase. Secondary efficacy outcomes included lung function and scores on the Asthma Control Questionnaire (ACQ) and Asthma Quality of Life Questionnaire (AQLQ). RESULTS: The rate of primary endpoints (PEPs) during 6 months was not significantly different between the two treatment groups: 0.75 PEPs (95% CI 0.55 to 1.01) per subject in the azithromycin group versus 0.81 PEPs (95% CI 0.61 to 1.09) in the placebo group (p=0.682). In a predefined subgroup analysis according to the inflammatory phenotype, azithromycin was associated with a significantly lower PEP rate than placebo in subjects with non-eosinophilic severe asthma (blood eosinophilia ≤200/µl): 0.44 PEPs (95% CI 0.25 to 0.78) versus 1.03 PEPs (95% CI 0.72 to 1.48) (p=0.013). Azithromycin significantly improved the AQLQ score but there were no significant between-group differences in the ACQ score or lung function. Azithromycin was well tolerated, but was associated with increased oropharyngeal carriage of macrolide-resistant streptococci. CONCLUSIONS: Azithromycin did not reduce the rate of severe exacerbations and LRTI in patients with severe asthma. However, the significant reduction in the PEP rate in azithromycin-treated patients with non-eosinophilic severe asthma warrants further study. CLINICALTRIALS.GOV NUMBER: NCT00760838.

BACKGROUND: Despite their antimicrobial potential, vaginal lactobacilli often fail to retain dominance, resulting in overgrowth of the vagina by other bacteria, as observed with bacterial vaginosis. It remains elusive however to what extent interindividual differences in vaginal Lactobacillus community composition determine the stability of this microflora. In a prospective cohort of pregnant women we studied the stability of the normal vaginal microflora (assessed on Gram stain) as a function of the presence of the vaginal Lactobacillus index species (determined through culture and molecular analysis with tRFLP). RESULTS: From 100 consecutive Caucasian women vaginal swabs were obtained at mean gestational ages of 8.6 (SD 1.4), 21.2 (SD 1.3), and 32.4 (SD 1.7) weeks, respectively. Based on Gram stain, 77 women had normal or Lactobacillus-dominated vaginal microflora (VMF) during the first trimester, of which 18 had grade Ia (L. crispatus cell morphotypes) VMF (23.4%), 16 grade Iab (L. crispatus and other Lactobacillus cell morphotypes) VMF (20.8%), and 43 grade Ib (non-L. crispatus cell morphotypes) VMF (55.8%). Thirteen women with normal VMF at baseline, converted in the second or third trimester (16.9%) to abnormal VMF defined as VMF dominated by non-Lactobacillus bacteria. Compared to grade Ia and grade Iab VMF, grade Ib VMF were 10 times (RR = 9.49, 95% CI 1.30 - 69.40) more likely to convert from normal to abnormal VMF (p = 0.009). This was explained by the observation that normal VMF comprising L. gasseri/iners incurred a ten-fold increased risk of conversion to abnormal VMF relative to non-L. gasseri/iners VMF (RR 10.41, 95% CI 1.39-78.12, p = 0.008), whereas normal VMF comprising L. crispatus had a five-fold decreased risk of conversion to abnormal VMF relative to non-L. crispatus VMF (RR 0.20, 95% CI 0.05-0.89, p = 0.04). CONCLUSION: The presence of different Lactobacillus species with the normal vaginal microflora is a major determinant to the stability of this microflora in pregnancy: L. crispatus promotes the stability of the normal vaginal microflora while L. gasseri and/or L. iners predispose to some extent to the occurrence of abnormal vaginal microflora.

BACKGROUND: The pathogenesis of bacterial vaginosis remains largely elusive, although some microorganisms, including Gardnerella vaginalis, are suspected of playing a role in the etiology of this disorder. Recently culture-independent analysis of microbial ecosystems has proven its efficacy in characterizing the diversity of bacterial populations. Here, we report on the results obtained by combining culture and PCR-based methods to characterize the normal and disturbed vaginal microflora. RESULTS: A total of 150 vaginal swab samples from healthy women (115 pregnant and 35 non-pregnant) were categorized on the basis of Gram stain of direct smear as grade I (n = 112), grade II (n = 26), grade III (n = 9) or grade IV (n = 3). The composition of the vaginal microbial community of eight of these vaginal swabs (three grade I, two grade II and three grade III), all from non-pregnant women, were studied by culture and by cloning of the 16S rRNA genes obtained after direct amplification. Forty-six cultured isolates were identified by tDNA-PCR, 854 cloned 16S rRNA gene fragments were analysed of which 156 by sequencing, yielding a total of 38 species, including 9 presumptively novel species with at least five species that have not been isolated previously from vaginal samples. Interestingly, cloning revealed that Atopobium vaginae was abundant in four out of the five non-grade I specimens. Finally, species specific PCR for A. vaginae and Gardnerella vaginalis pointed to a statistically significant co-occurrence of both species in the bacterial vaginosis samples. CONCLUSIONS: Although historically the literature regarding bacterial vaginosis has largely focused on G. vaginalis in particular, several findings of this study--like the abundance of A. vaginae in disturbed vaginal microflora and the presence of several novel species--indicate that much is to be learned about the composition of the vaginal microflora and its relation to the etiology of BV.

A total of 53 field and reference strains, including the type strains of the seven named species (nomenspecies) and belonging to the 18 described genomic species (DNA groups) of the genus Acinetobacter, were studied by amplified ribosomal DNA restriction analysis (ARDRA). Restriction analysis with the enzymes AluI, CfoI, MboI, RsaI, and MspI of the enzymatically amplified 16S rRNA genes allowed us to identify all species except the genomic species 4 (Acinetobacter haemolyticus) and 7 (A. johnsonii), 5 (A. junii) and 17, and 10 and 11, which clustered pairwise in three respective groups. Further analysis with the enzyme HaeIII, HinfI, NciI, ScrFI, or TaqI did not allow us to differentiate the species within these three clusters. However, use of a few additional simple phenotypic tests (hemolysis, growth at 37 degrees C, production of acid from glucose, and gelatin hydrolysis) can be used to differentiate between the species within these clusters. ARDRA proved to be a rapid and reliable method for the identification of most of the Acinetobacter genomic species, including the closely related DNA groups 1 (A. calcoaceticus), 2 (A. baumannii), 3, and 13. The results of this study suggest that ARDRA can be used for the identification of Acinetobacter species and as such may help to elucidate the ecology and clinical significance of the different species of this genus. Since ARDRA uses universal 16S rRNA gene primers, it is expected to be applicable to the identification of most bacterial species. Furthermore, ARDRA is less prone to contamination problems than PCR for detection, since the use of cultured organisms results in a large initial quantity of target DNA.