Urinary Microbiota Research Articles

Search for Microorganisms in Men with Urologic Chronic Pelvic Pain Syndrome: A Culture-Independent Analysis in the MAPP Research Network

We used next-generation, state-of-the-art, culture independent methodology to survey urine microbiota of males with urologic chronic pelvic pain syndrome and control participants enrolled in the MAPP Network to investigate a possible microbial etiology. Male patients with urologic chronic pelvic pain syndrome and matched controls were asked to provide initial, midstream and post-prostatic massage urine specimens. Specimens were analyzed with Ibis T-5000 Universal Biosensor technology to provide comprehensive identification of bacterial and select fungal species. Differences between urologic chronic pelvic pain syndrome and control study participants for the presence of species or species variation in a higher taxonomic grouping (genus) were evaluated using permutational multivariate analysis of variance and logistic regression. Initial and midstream urine specimens were obtained from 110 (post-prostatic massage urine in 67) participants with urologic chronic pelvic pain syndrome and 115 (post-prostatic massage urine in 62) controls. Overall 78, 73 and 54 species (42, 39 and 27 genera) were detected in initial, midstream and post-prostatic massage urine specimens, respectively. Mean (SD) initial, midstream and post-prostatic massage urine species count per person was 1.62 (1.28), 1.38 (1.36) and 1.33 (1.24) for cases, and 1.75 (1.32), 1.23 (1.15) and 1.56 (0.97) for controls, respectively. Overall species and genus composition differed significantly between participants with urologic chronic pelvic pain syndrome and controls in initial stream urine (p=0.002 species level, p=0.004 genus level), with Burkholderia cenocepacia overrepresented in urologic chronic pelvic pain syndrome. No significant differences were observed at any level in midstream or post-prostatic massage urine samples. Assessment of baseline culture-independent microbiological data from male subjects enrolled in the MAPP Network has identified overrepresentation of B. cenocepacia in urologic chronic pelvic pain syndrome. Future studies are planned to further evaluate microbiota associations with variable and changing urologic chronic pelvic pain syndrome symptom patterns.

Urinary microbiota in patients with prostate cancer and benign prostatic hyperplasia.

IntroductionInflammation is associated with promotion of the initiation of various malignancies, partly due to bacterial infection-induced microenvironmental changes. However, the exact association between microbiota in urine, seminal fluid and the expressed prostatic secretions and benign prostatic hypertrophy and prostate cancer is not clear.Material and methodsIn the present study, we investigated the type of microbiota in the expressed prostatic secretions (EPS) of patients with prostate cancer and benign prostatic hyperplasia (BPH) by the polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) method using universal bacterial primers. In order to understand the possible association between various bacteria and prostate cancer, quantitative real-time PCR assay was performed to quantify the amount of strains of bacteria in urine, EPS and seminal fluid.ResultsThe prostate cancer group had a significantly increased number of Bacteroidetes bacteria, Alphaproteobacteria, Firmicutes bacteria, Lachnospiraceae, Propionicimonas, Sphingomonas, and Ochrobactrum, and a decrease in Eubacterium and Defluviicoccus compared to the BPH group. The number of Escherichia coli in the prostate cancer group was significantly decreased in urine and increased in the EPS and seminal fluid, while the number of Enterococcus was significantly increased in the seminal fluid with little change in urine and EPS.ConclusionsBased on these results, we suggest that there are significant changes in the microbial population in EPS, urine and seminal fluid of subjects with prostate cancer and BPH, indicating a possible role for these bacteria in these two conditions.

Interplay between bladder microbiota and urinary antimicrobial peptides: mechanisms for human urinary tract infection risk and symptom severity.

Resident bacterial communities (microbiota) and host antimicrobial peptides (AMPs) are both essential components of normal host innate immune responses that limit infection and pathogen induced inflammation. However, their interdependence has not been investigated in the context of urinary tract infection (UTI) susceptibility. Here, we explored the interrelationship between the urinary microbiota and host AMP responses as mechanisms for UTI risk. Using prospectively collected day of surgery (DOS) urine specimens from female pelvic floor surgery participants, we report that the relative abundance and/or frequency of specific urinary microbiota distinguished between participants who did or did not develop a post-operative UTI. Furthermore, UTI risk significantly correlated with both specific urinary microbiota and β-defensin AMP levels. Finally, urinary AMP hydrophobicity and protease activity were greater in participants who developed UTI, and correlated positively with both UTI risk and pelvic floor symptoms. These data demonstrate an interdependency between the urinary microbiota, AMP responses and symptoms, and identify a potential mechanism for UTI risk. Assessment of bacterial microbiota and host innate immune AMP responses in parallel may identify increased risk of UTI in certain populations.

MP16-08 SEARCH FOR MICROORGANISMS IN MEN WITH UROLOGIC CHRONIC PELVIC PAIN SYNDROME: A CULTURE-INDEPENDENT ANALYSIS OF CASES AND CONTROLS ENROLLED IN THE TRANS-MAPP EPIDEMIOLOGY/PHENOTYPING (EP) STUDY

You have accessJournal of UrologyInfections/Inflammation of the Genitourinary Tract: Prostate & Genitalia1 Apr 2014MP16-08 SEARCH FOR MICROORGANISMS IN MEN WITH UROLOGIC CHRONIC PELVIC PAIN SYNDROME: A CULTURE-INDEPENDENT ANALYSIS OF CASES AND CONTROLS ENROLLED IN THE TRANS-MAPP EPIDEMIOLOGY/PHENOTYPING (EP) STUDY J. Curtis Nickel, Alisa Stephens, Jun Chen, Richard Landis, Christopher Mullins, Garth Ehrlich, and The MAPP Research Network J. Curtis NickelJ. Curtis Nickel More articles by this author , Alisa StephensAlisa Stephens More articles by this author , Jun ChenJun Chen More articles by this author , Richard LandisRichard Landis More articles by this author , Christopher MullinsChristopher Mullins More articles by this author , Garth EhrlichGarth Ehrlich More articles by this author , and The MAPP Research Network More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2014.02.581AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES We hypothesized that differences in lower urinary tract microbial populations between male UCPPS patients and normal subjects may be implicated in the etiology of male UCPPS. We used novel state-of-the-art culture-independent methodology to compare the microbiota of the lower urinary tract in men with UCPPS and controls enrolled in the NIH trans-MAPP EP study. METHODS Each participant (cases and non-UCPPS controls) were asked to provide initial (VB1), midstream (VB2) and post-prostatic massage (VB3) urine specimen. Specimens were analyzed with Ibis T-5000 Universal Biosensor technology which employs a polymerase chain reaction – electron spray ionization – time-of-flight – mass spectoscopic- based system to provide comprehensive identification of all bacterial species. Differences between UCPPS patients and controls in the presence of species or the number of different species within a higher level group were tested using PERMANOVA, and Fisher’s Exact test. All tests were conducted at the species, genus,and gram-stain level. RESULTS 417 male subjects (191 cases; 226 controls) were enrolled in the trans-MAPP-EP study. Baseline VB1 and VB2 urine specimens were obtained from 110 UCPPS cases and 115 matched controls (subjects with no UCPPS symptoms). VB3 samples were provided by 67 CP/CPPS and 62 controls. A total of 78, 73 and 54 species (42, 39 and 27 genera) were detected in VB1, VB2 and VB3 respectively. Mean (SD) VB1, VB2 and VB3 species count per person was 1.62 (1.28), 1.38 (1.36) and 1.33(1.24) for cases and 1.75(1.32), 1.23(1.15) and 1.56 (0.97) for controls respectively. Overall species and genus composition significantly differed between UCPPS patients and controls in VB1 only (p=0.008 species level, p=0.011 genus level, p=0.079 Gram-stain level) with overall differences driven by B. cenocepacia (RR=3.34, p=0.009), P. acnes (RR 0.38, p=0.01), and S. capitis/capare (RR=0.17, p=0.008). These differences (and similar trends at genus level) remained significant after adjustment for multiple comparisons. No significant differences were observed at the Gram-stain level in VB1 or at any level in VB2 or VB3. CONCLUSIONS While our results make it difficult to implicate the minor differences in lower urinary tract microbiota (VB1 only) in patients vs controls for a role in the etiology and pathogenesis of UCPPS in males, further work is required to determine if microbial composition or fluctuations influence symptom patterns over time. © 2014FiguresReferencesRelatedDetails Volume 191Issue 4SApril 2014Page: e156 Advertisement Copyright & Permissions© 2014MetricsAuthor Information J. Curtis Nickel More articles by this author Alisa Stephens More articles by this author Jun Chen More articles by this author Richard Landis More articles by this author Christopher Mullins More articles by this author Garth Ehrlich More articles by this author The MAPP Research Network More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Human Microbiota Characterization in the Course of Renal Transplantation

Recent studies demonstrate that the human microbiota, the collection of microorganisms growing on and in individuals, have numerous bidirectional interactions with the host, influencing immunity, resistance to infection, inflammation and metabolism. Little has been done to study the potential associations between microbiota composition and transplant outcome. Here, we investigated the longitudinal changes in the blood, urinary, oral and rectal microbiota of renal allograft recipients before and at 1 and 6 months after transplantation. The results showed major changes in microbiota composition as a result of the transplant episode and associated medications, and these changes persisted over time. The high interindividual variation as well as differences in response to transplantation suggested that it is unlikely that the same specific microbiota members can serve as universal diagnostic markers. Rather, longitudinal changes in each individual's microbiota have the potential to be indicative of health or disease. Use of sensitive nucleic acid-based testing showed that urine, irrespective of disease states, more often harbors a diverse microbiota than appreciated by conventional culture techniques. These results lay the groundwork to construct more comprehensive future investigations to identify microbiota characteristics that can serve as diagnostic markers for transplant health and to guide intervention strategies to improve transplant outcome.

Urine Is Not Sterile: Use of Enhanced Urine Culture Techniques To Detect Resident Bacterial Flora in the Adult Female Bladder

Our previous study showed that bacterial genomes can be identified using 16S rRNA sequencing in urine specimens of both symptomatic and asymptomatic patients who are culture negative according to standard urine culture protocols. In the present study, we used a modified culture protocol that included plating larger volumes of urine, incubation under varied atmospheric conditions, and prolonged incubation times to demonstrate that many of the organisms identified in urine by 16S rRNA gene sequencing are, in fact, cultivable using an expanded quantitative urine culture (EQUC) protocol. Sixty-five urine specimens (from 41 patients with overactive bladder and 24 controls) were examined using both the standard and EQUC culture techniques. Fifty-two of the 65 urine samples (80%) grew bacterial species using EQUC, while the majority of these (48/52 [92%]) were reported as no growth at 10(3) CFU/ml by the clinical microbiology laboratory using the standard urine culture protocol. Thirty-five different genera and 85 different species were identified by EQUC. The most prevalent genera isolated were Lactobacillus (15%), followed by Corynebacterium (14.2%), Streptococcus (11.9%), Actinomyces (6.9%), and Staphylococcus (6.9%). Other genera commonly isolated include Aerococcus, Gardnerella, Bifidobacterium, and Actinobaculum. Our current study demonstrates that urine contains communities of living bacteria that comprise a resident female urine microbiota.

The human urinary microbiome; bacterial DNA in voided urine of asymptomatic adults

The urinary microbiome of healthy individuals and the way it alters with ageing have not been characterized and may influence disease processes. Conventional microbiological methods have limited scope to capture the full spectrum of urinary bacterial species. We studied the urinary microbiota from a population of healthy individuals, ranging from 26 to 90 years of age, by amplification of the 16S rRNA gene, with resulting amplicons analyzed by 454 pyrosequencing. Mid-stream urine (MSU) was collected by the “clean-catch” method. Quantitative PCR of 16S rRNA genes in urine samples, allowed relative enumeration of the bacterial loads. Analysis of the samples indicates that females had a more heterogeneous mix of bacterial genera compared to the male samples and generally had representative members of the phyla Actinobacteria and Bacteroidetes. Analysis of the data leads us to conclude that a “core” urinary microbiome could potentially exist, when samples are grouped by age with fluctuation in abundance between age groups. The study also revealed age-specific genera Jonquetella, Parvimonas, Proteiniphilum, and Saccharofermentans. In conclusion, conventional microbiological methods are inadequate to fully identify around two-thirds of the bacteria identified in this study. Whilst this proof-of-principle study has limitations due to the sample size, the discoveries evident in this sample data are strongly suggestive that a larger study on the urinary microbiome should be encouraged and that the identification of specific genera at particular ages may be relevant to pathogenesis of clinical conditions.

Alterations of microbiota in urine from women with interstitial cystitis

BackgroundInterstitial 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.ResultsThe 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.ConclusionThe 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.

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

BackgroundUrine 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.ResultsSequencing 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 Lactobacillus, Prevotella and Gardnerella. 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.ConclusionsNormal 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.

Species interactions in mixed-community crystalline biofilms on urinary catheters

Previous experimental investigations of the crystalline biofilms that colonize and block urinary catheters have focussed on their formation by pure cultures of Proteus mirabilis. In the urine of patients undergoing long-term catheterization, P. mirabilis is commonly found in mixed communities with other urinary tract pathogens. Little is known about the effect that the other species have on the rate at which P. mirabilis encrusts catheters. In the present study, a set of data on the nature of the bacterial communities on 106 catheter biofilms has been analysed and it was found that while species such as Providencia stuartii and Klebsiella pneumoniae were commonly associated with P. mirabilis, when Escherichia coli, Morganella morganii or Enterobacter cloacae were present, P. mirabilis was rarely or never found. The hypothesis that the absence of P. mirabilis from some biofilm communities could be due to its active exclusion by other species has also been examined. Experiments in laboratory models showed that co-infection of P. mirabilis with M. morganii, K. pneumoniae or E. coli had no effect on the ability of P. mirabilis to encrust and block catheters. Co-infection with Ent. cloacae or Pseudomonas aeruginosa, however, significantly increased the time that catheters took to block (P <0.05). The growth of Ent. cloacae, M. morganii, K. pneumoniae or E. coli in the model for 72 h prior to superinfection with P. mirabilis significantly delayed catheter blockage. In the case of Ent. cloacae, for example, the mean time to blockage was extended from 28.7 h to 60.7 h (P < or =0.01). In all cases, however, P. mirabilis was able to generate alkaline urine, colonize the biofilms, induce crystal formation and block the catheters. The results suggest that although there is a degree of antagonism between P. mirabilis and some of the other urinary tract organisms, the effects are temporary and whatever the pre-existing urinary microbiota, infection with P. mirabilis is thus likely to lead to catheter encrustation and blockage.

NEWER MACROLIDES

In clinical microbiology laboratories, advancements in the methods used for routine organism identification have facilitated more accurate species level resolution. This, along with increasing knowledge regarding “new” pathogens, has provided new insights into biology and has revealed clinical associations not previously known. Aerococcus urinae, Alloscardovia omnicolens, and Actinotignum schaalii are Gram-positive bacteria associated with urinary tract infections but can also be members of the urinary tract microbiota. In addition, invasive infections have been reported for the three bacteria, usually with a urinary tract source. Importantly, the use of routine biochemical testing methods for identification of these bacteria can result in misidentification or the incorrect determination that they are not clinically significant. Of these bacteria, only A. urinae has antimicrobial susceptibility testing criteria and interpretative breakpoints. Microbiologists and clinicians need to be aware of these urinary tract pathogens and how to correctly identify them.