Gut colonization with methanobrevibacter smithii is associated with childhood weight development

OBJECTIVE: To prospectively investigate the presence and counts of archaea in feces of 472 children in association with weight development from 6 to 10 years of age. METHODS: Within the KOALA Birth Cohort Study, a single fecal sample from each child was analyzed by quantitative polymerase chain reaction to quantify archaea (Methanobrevibacter smithii, Methanosphera stadtmanae). Anthropometric outcomes (overweight [body mass index {BMI} >/= 85th percentile], age- and sex-standardized BMI, weight, and height z-scores) were repeatedly measured at ages (mean +/- SD) of 6.2 +/- 0.5, 6.8 +/- 0.5, 7.8 +/- 0.5, and 8.8 +/- 0.5 years. Generalized estimating equation was used for statistical analysis while controlling for confounders. RESULTS: Methanobrevibacter smithii colonization was associated with an increased risk of overweight (adjusted odds ratio [OR] = 2.69; 95% confidence interval [CI] 0.96-7.54) from 6 to 10 years of age. Children with high levels (>7 log10 copies/g feces) of this archaeon were at highest risk for overweight (OR = 3.27; 95% CI 1.09-9.83). Moreover, M. smithii colonization was associated with higher weight z-scores (adj. beta 0.18; 95% CI 0.00-0.36), but not with height. For BMI z-scores, the interaction (P = 0.008) between M. smithii and age was statistically significant, implying children colonized with M. smithii had increasing BMI z-scores with age. CONCLUSIONS: Presence and higher counts of M. smithii in the gut of children are associated with higher weight z-scores, higher BMI z-scores, and overweight

Fecal microbial composition of ulcerative colitis and Crohn's disease patients in remission and subsequent exacerbation

Contains fulltext : 138027.pdf (publisher's version ) (Open Access)BACKGROUND: Limited studies have examined the intestinal microbiota composition in relation to changes in disease course of IBD over time. We aimed to study prospectively the fecal microbiota in IBD patients developing an exacerbation during follow-up. DESIGN: Fecal samples from 10 Crohn's disease (CD) and 9 ulcerative colitis (UC) patients during remission and subsequent exacerbation were included. Active disease was determined by colonoscopy and/or fecal calprotectine levels. Exclusion criteria were pregnancy, antibiotic use, enema use and/or medication changes between consecutive samples. The microbial composition was assessed by 16S rDNA pyrosequencing. RESULTS: After quality control, 6,194-11,030 sequences per sample were available for analysis. Patient-specific shifts in bacterial composition and diversity were observed during exacerbation compared to remission, but overarching shifts within UC or CD were not observed. Changes in the bacterial community composition between remission and exacerbation as assessed by Bray-Curtis dissimilarity, were significantly larger in CD versus UC patients (0.59 vs. 0.42, respectively; p = 0.025). Thiopurine use was found to be a significant cause of clustering as shown by Principal Coordinate Analysis and was associated with decreases in bacterial richness (Choa1 501.2 vs. 847.6 in non-users; p<0.001) and diversity (Shannon index: 5.13 vs. 6.78, respectively; p<0.01). CONCLUSION: Shifts in microbial composition in IBD patients with changing disease activity over time seem to be patient-specific, and are more pronounced in CD than in UC patients. Furthermore, thiopurine use was found to be associated with the microbial composition and diversity, and should be considered when studying the intestinal microbiota in relation to disease course

Rapid plasmid replicon typing by real time PCR melting curve analysis

BACKGROUND: Genes encoding Extended Spectrum Beta Lactamases are usually located on transferable plasmids. Each plasmid contains its own replication mechanism. Carattoli et al. developed an extended PCR-based replicon typing method to characterize and identify the replicons of the major plasmid incompatibility groups in Enterobacteriaceae. Based on this method, we designed a rapid approach with amplicon detection by real-time melting curve analysis. This method appeared to be fast, sensitive, less laborious, less prone to contamination and applicable in a routine laboratory environment. RESULTS: We successfully integrated the post-PCR analysis of the replicon typing into a closed system, which leads to a 10-fold increase in sensitivity compared to agarose gel visualization. Moreover, the use of crude lysates and SYBR-green saves a considerable amount of hands-on time without decreasing the sensitivity or specificity. CONCLUSIONS: This real-time melting curve replicon typing method appears to be fast, sensitive, less laborious, less prone to contamination and applicable in a routine laboratory environment

Application of different genotyping methods for Pseudomonas aeruginosa in a setting of endemicity in an intensive care unit

Medical Microbiology, University Hospital Maastricht, Maastricht, The Netherlands. Colonization with Pseudomonas aeruginosa was studied by taking serial swab specimens from the oropharynges and anuses and tracheal and gastric aspirates from patients in an intensive care unit during a 10-month period in a setting of endemicity. Nineteen (10%) of the 192 patients included in the study were colonized on admission, while another 30 (16%) patients acquired P. aeruginosa while in the hospital. Typing of 353 isolates was performed by random amplified polymorphic DNA (RAPD) analysis, and 56 strains were selected for further typing by RAPD analysis, pulsed-field gel electrophoresis (PFGE), and amplified fragment length polymorphism (AFLP) analysis. By these methods, 42, 44, and 44 genotypes were found, respectively. Computer-aided cluster analysis indicated that similar groups of related isolates were obtained by each method. By taking admission periods into account, analysis of the typing results suggested cross-acquisition of P. aeruginosa for five patient pairs. The small number of transfers and the large number of genotypes found indicate that most P. aeruginosa strains were derived from the patients themselves. The numbers of observed typing patterns and band differences between related isolates were counted for each typing method. AFLP analysis with primers without a selective base proved to be the most discriminatory method, followed by PFGE, AFLP analysis (with one selective base), and RAPD analysis. On the basis of a comparison with established strain differentiation criteria for PFGE, the criteria for differentiation of P. aeruginosa by AFLP analysis are presented

Fecal Microbial Composition of Ulcerative Colitis and Crohn's Disease Patients in Remission and Subsequent Exacerbation

BACKGROUND: Limited studies have examined the intestinal microbiota composition in relation to changes in disease course of IBD over time. We aimed to study prospectively the fecal microbiota in IBD patients developing an exacerbation during follow-up. DESIGN: Fecal samples from 10 Crohn's disease (CD) and 9 ulcerative colitis (UC) patients during remission and subsequent exacerbation were included. Active disease was determined by colonoscopy and/or fecal calprotectine levels. Exclusion criteria were pregnancy, antibiotic use, enema use and/or medication changes between consecutive samples. The microbial composition was assessed by 16S rDNA pyrosequencing. RESULTS: After quality control, 6,194-11,030 sequences per sample were available for analysis. Patient-specific shifts in bacterial composition and diversity were observed during exacerbation compared to remission, but overarching shifts within UC or CD were not observed. Changes in the bacterial community composition between remission and exacerbation as assessed by Bray-Curtis dissimilarity, were significantly larger in CD versus UC patients (0.59 vs. 0.42, respectively; p = 0.025). Thiopurine use was found to be a significant cause of clustering as shown by Principal Coordinate Analysis and was associated with decreases in bacterial richness (Choa1 501.2 vs. 847.6 in non-users; p<0.001) and diversity (Shannon index: 5.13 vs. 6.78, respectively; p<0.01). CONCLUSION: Shifts in microbial composition in IBD patients with changing disease activity over time seem to be patient-specific, and are more pronounced in CD than in UC patients. Furthermore, thiopurine use was found to be associated with the microbial composition and diversity, and should be considered when studying the intestinal microbiota in relation to disease course

Composition and diversity of the duodenal mucosa-associated microbiome in children with untreated coeliac disease

BACKGROUND: Intestinal microbiome may play a role in the pathogenesis of coeliac disease (CD). Studies comparing intestinal microbiome in children with and without CD are contradictory. AIM: To compare the composition and diversity of the duodenal mucosa-associated microbiome in children with untreated CD and control children without CD and to identify specific gut bacteria associated with CD at diagnosis. METHODS: Total microbiome profile in small bowel biopsies of 42 children (21 with untreated CD and 21 age-matched controls) were analyzed by means of IS-pro, a 16S-23S interspacer (IS) region-based profiling method. RESULTS: Both groups showed a similar mucosa-associated microbiome pattern and diversity, with high concentrations of the genera Streptococcus, Lactobacillus, and Clostridium. CONCLUSION: Mucosa-associated duodenal microbiome composition and diversity did not differ between children with untreated CD and control children. Duodenal mucosa-associated bacteria do not seem to play an important role in the pathogenesis of CD

Real-time PCR detection of ruminant DNA

To control the spread of bovine spongiform encephalopathy, several DNA methods have been described for the detection of the species origin of meat and bone meal. Most of these methods are based on the amplification of a mitochondrial DNA segment. We have developed a semiquantitative method based on real-time PCR for detection of ruminant DNA, targeting an 88-bp segment of the ruminant short interspersed nuclear element Bov-A2. This method is specific for ruminants and is able to detect as little as 10 fg of bovine DNA. Autoclaving decreased the amount of detectable DNA, but positive signals were observed in feeding stuff containing 10␋ovine material if this had not been rendered in accordance with the regulations, i.e., heated at 134°C for 3 instead of 20 min

Uniparental maternal disomy 6 in a renal transplant patient.

Uniparental maternal disomy 6 in a renal transplant patient. van den Berg-Loonen EM, Savelkoul P, van Hooff H, van Eede P, Riesewijk A, Geraedts J. Tissue Typing Laboratory, University Hospital Maastricht, The Netherlands. HLA analysis of the family of a renal transplant patient revealed an extremely rare condition. On repeated typings the only demonstrable HLA antigens shown in the propositus were from the maternal haplotype, HLA-A11,-B46,-CW1,-DR14,-DQ1. No paternal antigens could be demonstrated either by serologic or by DNA-typing methods. A paternity investigation was carried out to exclude the possibility of the legal father not being the biological father. The results of this investigation showed a paternity index I = > 20000 and a fatherhood probability W = > 99.995%. Karyotyping of the patient showed two normal chromosomes 6 and no other chromosomal abnormalities. Maternal isodisomy was demonstrated from the analysis of polymorphic DNA markers, involving the short as well as the long arm of chromosome 6. These data are consistent with this patient having the first uniparental maternal disomy 6 reported (inheritance of two identical chromosome 6 haplotypes from the mother and none from the father)

No human transmission of Mycobacterium malmoense in a perfect storm setting

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Prevalence and risk factors for carriage of ESBL-producing Enterobacteriaceae in Amsterdam

OBJECTIVES: The objectives of this study were to determine the prevalence of carriage of ESBL-producing Enterobacteriaceae (ESBL-E) in a representative sample of the general adult Dutch community, to identify risk factors and to gain understanding of the epidemiology of these resistant strains. METHODS: Adults enrolled in five general practices in Amsterdam were approached by postal mail and asked to fill in a questionnaire and to collect a faecal sample. Samples were analysed for the presence of ESBL-E. ESBL genes were characterized by PCR and sequencing. Strains were typed using MLST and amplified fragment length polymorphism (AFLP) and plasmids were identified by PCR-based replicon typing. Risk factors for carriage were investigated by multivariate analysis. RESULTS: ESBL-E were found in 145/1695 (8.6%) samples; 91% were Escherichia coli. Most ESBL genes were of the CTX-M group (blaCTX-M-1 and blaCTX-M-15). MLST ST131 was predominant and mainly associated with CTX-M-15-producing E. coli. One isolate with reduced susceptibility to ertapenem produced OXA-48. In multivariate analyses, use of antimicrobial agents, use of antacids and travel to Africa, Asia and Northern America were associated with carriage of ESBL-E, in particular strains with blaCTX-M-14/15. CONCLUSIONS: This study showed a high prevalence of ESBL-E carriage in the general Dutch community. Also, outside hospitals, the use of antibiotics was a risk factor; interestingly, use of antacids increased the risk of carriage. A major risk factor in the general population was travel to countries outside Europe, in particular to Asia, Africa and Northern America