Assessing diversity of the female urine microbiota by high throughput sequencing of 16S rDNA amplicons

<p>Abstract</p> <p>Background</p> <p>Urine within the urinary tract is commonly regarded as "sterile" in cultivation terms. Here, we present a comprehensive in-depth study of bacterial 16S rDNA sequences associated with urine from healthy females by means of culture-independent high-throughput sequencing techniques.</p> <p>Results</p> <p>Sequencing of the V1V2 and V6 regions of the 16S ribosomal RNA gene using the 454 GS FLX system was performed to characterize the possible bacterial composition in 8 culture-negative (<100,000 CFU/ml) healthy female urine specimens. Sequences were compared to 16S rRNA databases and showed significant diversity, with the predominant genera detected being <it>Lactobacillus</it>, <it>Prevotella </it>and <it>Gardnerella</it>. The bacterial profiles in the female urine samples studied were complex; considerable variation between individuals was observed and a common microbial signature was not evident. Notably, a significant amount of sequences belonging to bacteria with a known pathogenic potential was observed. The number of operational taxonomic units (OTUs) for individual samples varied substantially and was in the range of 20 - 500.</p> <p>Conclusions</p> <p>Normal female urine displays a noticeable and variable bacterial 16S rDNA sequence richness, which includes fastidious and anaerobic bacteria previously shown to be associated with female urogenital pathology.</p

Prevalence of sexually transmitted infections among pregnant women with known HIV status in northern Tanzania

<p>Abstract</p> <p>Objectives</p> <p>To determine the prevalence of sexually transmitted infections (STIs) and other reproductive tract infections (RTIs) among pregnant women in Moshi, Tanzania and to compare the occurrence of STIs/RTIs among human immunodeficiency virus (HIV)-infected and uninfected women.</p> <p>Methods</p> <p>Pregnant women in their 3^rdtrimester (N = 2654) were recruited from two primary health care clinics between June 2002 and March 2004. They were interviewed, examined and genital and blood samples were collected for diagnosis of STIs/RTIs and HIV.</p> <p>Results</p> <p>The prevalence of HIV, active syphilis and herpes simplex virus – type 2 (HSV-2) were 6.9%, 0.9% and 33.6%, respectively, while 0.5% were positive for <it>N gonorrhoeae</it>, 5.0% for <it>T vaginalis </it>and 20.9% for bacterial vaginosis. Genital tract infections were more prevalent in HIV-seropositive than seronegative women, statistically significant for syphilis (3.3% vs 0.7%), HSV-2 (43.2% vs 32.0%), genital ulcers (4.4% vs 1.4%) and bacterial vaginosis (37.2% vs 19.6%). In comparison with published data, a declining trend for curable STIs/RTIs (syphilis, trichomoniasis and bacterial vaginosis) was noted.</p> <p>Conclusion</p> <p>Rates of STIs and RTIs are still high among pregnant women in Moshi. Where resources allow, routine screening and treatment of STIs/RTIs in the antenatal care setting should be offered. Higher STIs/RTIs in HIV-seropositive women supports the expansion of HIV-counseling and testing services to all centers offering antenatal care. After identification, STIs/RTIs need to be aggressively addressed in HIV-seropositive women, both at antenatal and antiretroviral therapy care clinics.</p

HIV Genetic Diversity in Cameroon: Possible Public Health Importance

To monitor the evolving molecular epidemiology and genetic diversity of HIV in a country where many distinct strains cocirculate, we performed genetic analyses on sequences from 75 HIV-1-infected Cameroonians: 74 were group M and 1 was group O. Of the group M sequences, 74 were classified into the following env gp41 subtypes or recombinant forms: CRF02 (n = 54), CRF09 (n = 2), CRF13 (n = 2), A (n = 5), CRF11 (n = 4), CRF06 (n = 1), G (n = 2), F2 (n = 2), and E (n = 1, CRF01), and 1 was a JG recombinant. Comparison of phylogenies for 70 matched gp41 and protease sequences showed inconsistent classifications for 18 (26%) strains. Our data show that recombination is rampant in Cameroon with recombinant viruses continuing to recombine, adding to the complexity of circulating HIV strains. This expanding genetic diversity raises public health concerns for the ability of diagnostic assays to detect these unique HIV mosaic variants and for the development of broadly effective HIV vaccines.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/63150/1/aid.2006.22.812.pd

CMV quantitative PCR in the diagnosis of CMV disease in patients with HIV-infection – a retrospective autopsy based study

Background Patients with advanced HIV infection at the time of diagnosis and patients not responding to antiretroviral therapy are at risk of cytomegalovirus (CMV) disease. Earlier studies of patients with HIV infection have demonstrated that the diagnosis is often first made post-mortem. In recent years new molecular biological tests have become available for diagnosis of CMV disease. Although clinical evaluation of tests for diagnosis of CMV disease in HIV-infected individuals is suboptimal without autopsy, no results from such studies have been published. The aim of this study was to explore the diagnostic utility of CMV quantitative polymerase chain reaction (PCR) in plasma from HIV and CMV seropositive patients who died during the period 1991–2002 and in whom autopsy was performed. Methods Autopsy was performed in all cases, as part of routine evaluation of HIV-infected cases followed at Ullevaal University Hospital. Of 125 patients included, 53 had CMV disease, 37 of whom were first diagnosed at autopsy. CMV disease was diagnosed either by ophthalmoscopic findings typical of CMV retinitis, biopsy or autopsy. One or two plasma samples taken prior to the first diagnosis of CMV disease (alive or at autopsy) or death without CMV disease were analysed by CMV quantitative PCR. Sensitivity, specificity, positive and negative predictive values were calculated for different CMV viral load cut-offs and according to detection of viraemia in one versus two samples. Results Twenty-seven of 53 patients with CMV disease (51%) and 10 of 72 patients without CMV disease (14%) had detectable viraemia in at least one sample. Sensitivity and negative predictive value (NPV) of the test, maximised with a cut-off at the test's limit of detection of CMV viraemia (400 copies/mL), were 47% and 70%, respectively. With cut-off at 10 000 copies/mL, specificity and positive predictive value (PPV) were 100%. With a requirement for CMV viraemia in two samples, specificity and PPV were 100% in patients with CMV viraemia above the limit of detection. Conclusion Our results indicate that quantitative CMV PCR is best used to rule in, rather than to rule out CMV disease in HIV-infected individuals at high risk

Alterations of microbiota in urine from women with interstitial cystitis

Background Interstitial Cystitis (IC) is a chronic inflammatory condition of the bladder with unknown etiology. The aim of this study was to characterize the microbial community present in the urine from IC female patients by 454 high throughput sequencing of the 16S variable regions V1V2 and V6. The taxonomical composition, richness and diversity of the IC microbiota were determined and compared to the microbial profile of asymptomatic healthy female (HF) urine. Results The composition and distribution of bacterial sequences differed between the urine microbiota of IC patients and HFs. Reduced sequence richness and diversity were found in IC patient urine, and a significant difference in the community structure of IC urine in relation to HF urine was observed. More than 90% of the IC sequence reads were identified as belonging to the bacterial genus Lactobacillus, a marked increase compared to 60% in HF urine. Conclusion The 16S rDNA sequence data demonstrates a shift in the composition of the bacterial community in IC urine. The reduced microbial diversity and richness is accompanied by a higher abundance of the bacterial genus Lactobacillus, compared to HF urine. This study demonstrates that high throughput sequencing analysis of urine microbiota in IC patients is a powerful tool towards a better understanding of this enigmatic disease

Alterations of microbiota in urine from women with interstitial cystitis

Abstract Background Interstitial Cystitis (IC) is a chronic inflammatory condition of the bladder with unknown etiology. The aim of this study was to characterize the microbial community present in the urine from IC female patients by 454 high throughput sequencing of the 16S variable regions V1V2 and V6. The taxonomical composition, richness and diversity of the IC microbiota were determined and compared to the microbial profile of asymptomatic healthy female (HF) urine. Results The composition and distribution of bacterial sequences differed between the urine microbiota of IC patients and HFs. Reduced sequence richness and diversity were found in IC patient urine, and a significant difference in the community structure of IC urine in relation to HF urine was observed. More than 90% of the IC sequence reads were identified as belonging to the bacterial genus Lactobacillus, a marked increase compared to 60% in HF urine. Conclusion The 16S rDNA sequence data demonstrates a shift in the composition of the bacterial community in IC urine. The reduced microbial diversity and richness is accompanied by a higher abundance of the bacterial genus Lactobacillus, compared to HF urine. This study demonstrates that high throughput sequencing analysis of urine microbiota in IC patients is a powerful tool towards a better understanding of this enigmatic disease.</p

Bacteria, biofilm and honey: A study of the effects of honey on &apos;planktonic&apos; and biofilm-embedded chronic wound bacteria

Abstract Chronically infected wounds are a costly source of suffering. An important factor in the failure of a sore to heal is the presence of multiple species of bacteria, living cooperatively in highly organized biofilms. The biofilm protects the bacteria from antibiotic therapy and the patient&apos;s immune response. Honey has been used as a wound treatment for millennia. The components responsible for its antibacterial properties are now being elucidated. The study aimed to determine the effects of different concentrations of &apos;Medihoney TM &apos; therapeutic honey and Norwegian Forest Honey 1) on the real-time growth of typical chronic wound bacteria; 2) on biofilm formation; and 3) on the same bacteria already embedded in biofilm. Reference strains of MRSE, MRSA, ESBL Klebsiella pneumoniae and Pseudomonas aeruginosa were incubated with dilution series of the honeys in microtitre plates for 20 h. Growth of the bacteria was assessed by measuring optical density every 10 min. Growth curves, biofilm formation and minimum bactericidal concentrations are presented. Both honeys were bactericidal against all the strains of bacteria. Biofilm was penetrated by biocidal substances in honey. Reintroduction of honey as a conventional wound treatment may help improve individual wound care, prevent invasive infections, eliminate colonization, interrupt outbreaks and thereby preserve current antibiotic stocks