scholarly journals First Experience of a Multicenter External Quality Assessment of Molecular 16S rRNA Gene Detection in Bone and Joint Infections

2014 ◽  
Vol 53 (2) ◽  
pp. 419-424 ◽  
Author(s):  
Chloé Plouzeau ◽  
Pascale Bémer ◽  
Anne Sophie Valentin ◽  
Geneviève Héry-Arnaud ◽  
Didier Tandé ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Joint Infection ◽  
Rrna Gene ◽  
Gene Detection ◽  
External Quality ◽  
Multicenter Studies ◽  
Joint Infections ◽  
Significant Difference ◽  
Bead Mill

The objective of this study was to assess the performance of seven French laboratories for 16S rRNA gene detection by real-time PCR in the diagnosis of bone and joint infection (BJI) to validate a large multicenter study. External quality control (QC) was required owing to the differences in extraction procedures and the molecular equipment used in the different laboratories. Three proficiency sets were organized, including four bacterial DNA extracts and four bead mill-pretreated osteoarticular specimens. Extraction volumes, 16S rRNA gene primers, and sequencing interpretation rules were standardized. In order to assess each laboratory's ability to achieve the best results, scores were assigned, and each QC series was classified as optimal, acceptable, or to be improved. A total of 168 QCs were sent, and 160 responses were analyzed. The expected results were obtained for 93.8%, with the same proportion for extracts (75/80) and clinical specimens (75/80). For the specimens, there was no significant difference between manual and automated extraction. This QC demonstrated the ability to achieve good and homogeneous results using the same 16S rRNA gene PCR with different equipment and validates the possibility of high-quality multicenter studies using molecular diagnosis for BJI.

scholarly journals Role of Universal 16S rRNA Gene PCR and Sequencing in Diagnosis of Prosthetic Joint Infection

2011 ◽  
Vol 50 (3) ◽  
pp. 583-589 ◽  
Author(s):  
M. Marin ◽  
J. M. Garcia-Lechuz ◽  
P. Alonso ◽  
M. Villanueva ◽  
L. Alcala ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Prosthetic Joint Infection ◽  
Joint Infection ◽  
Rrna Gene ◽  
Prosthetic Joint

scholarly journals Anaplasma Infection in Ticks in Southeastern Region of Iran

2020 ◽  
Author(s):  
Reza Ranjbar ◽  
Mehdi Anjomruz ◽  
Ahmad Ali Enayati ◽  
Mehdi Khoobdel ◽  
Atiyeh Rafinejad ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Cold Chain ◽  
Rrna Gene ◽  
Infection Rates ◽  
Male And Female ◽  
Significant Difference ◽  
High Infection ◽  
Hyalomma Marginatum ◽  
Southeastern Region

Background: Anaplasmosis and Ehrlichiosis are the most important tick-borne diseases. This study was conducted in three cities of Kerman Province in Iran to investigate the circulation of the bacteria in ticks collected from sheep. Methods: Ticks were collected from animals using Srkj forceps and transferred to the Entomology lab in cold chain. After specimen’s identification, they kept at -70 ºC. Tick DNA was extracted using Bioneers DNA extraction kits followed by Nested PCR technique to amplify ribosomal 16S rRNA gene to detect Anaplasma infection in ticks. Results: 472 sheep were examined from which 349 ticks were collected and identified in laboratory using valid keys. Tick specimens belonged to two genera and four species; Hyalomma marginatum (62.47%) was the most frequent and Hylomma asiaticum (5.73%) showed the least abundance. The infestation rate to different tick species was different in three regions of Kerman Province. Observation revealed that 24 specimens (58.3%) were positive for Anaplasma. There is a significant difference between male and female infection rate. However, there is no significant difference between these variables in each of these cities. Conclusion: This study shows high infection rates to Anaplasma in hard ticks. It is essential for health and veterinary authorities and farmers to use appropriate strategies to control ticks to reduce the infestation.

scholarly journals Detection of Prosthetic Hip Infection at Revision Arthroplasty by Immunofluorescence Microscopy and PCR Amplification of the Bacterial 16S rRNA Gene

1999 ◽  
Vol 37 (10) ◽  
pp. 3281-3290 ◽  
Author(s):  
Michael M. Tunney ◽  
Sheila Patrick ◽  
Martin D. Curran ◽  
Gordon Ramage ◽  
Donna Hanna ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Immunofluorescence Microscopy ◽  
Joint Infection ◽  
Pcr Amplification ◽  
Rrna Gene ◽  
Bacterial Dna ◽  
Prosthetic Joint ◽  
Culture Positive ◽  
Bacterial 16S Rrna Gene

In this study the detection rates of bacterial infection of hip prostheses by culture and nonculture methods were compared for 120 patients with total hip revision surgery. By use of strict anaerobic bacteriological practice during the processing of samples and without enrichment, the incidence of infection by culture of material dislodged from retrieved prostheses after ultrasonication (sonicate) was 22%. Bacteria were observed by immunofluorescence microscopy in 63% of sonicate samples with a monoclonal antibody specific forPropionibacterium acnes and polyclonal antiserum specific for Staphylococcus spp. The bacteria were present either as single cells or in aggregates of up to 300 bacterial cells. These aggregates were not observed without sonication to dislodge the biofilm. Bacteria were observed in all of the culture-positive samples, and in some cases in which only one type of bacterium was identified by culture, both coccoid and coryneform bacteria were observed by immunofluorescence microscopy. Bacteria from skin-flake contamination were readily distinguishable from infecting bacteria by immunofluorescence microscopy. Examination of skin scrapings did not reveal large aggregates of bacteria but did reveal skin cells. These were not observed in the sonicates. Bacterial DNA was detected in 72% of sonicate samples by PCR amplification of a region of the bacterial 16S rRNA gene with universal primers. All of the culture-positive samples were also positive for bacterial DNA. Evidence of high-level infiltration either of neutrophils or of lymphocytes or macrophages into associated tissue was observed in 73% of patients. Our results indicate that the incidence of prosthetic joint infection is grossly underestimated by current culture detection methods. It is therefore imperative that current clinical practice with regard to the detection and subsequent treatment of prosthetic joint infection be reassessed in the light of these results.

Organotrophic and Mixotrofic Sulfur Oxidation in an Acidic Salt Flat in Northern Chile

2015 ◽  
Vol 1130 ◽  
pp. 63-66 ◽  
Author(s):  
Lorena Escudero ◽  
Jonathan Bijman ◽  
Guajardo M. Mariela ◽  
Juan José Pueyo Mur ◽  
Guillermo Chong ◽  
...  
Keyword(s):  
Microbial Community ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Microbial Community Composition ◽  
Iron Concentration ◽  
Rrna Gene ◽  
Salt Flat ◽  
Significant Difference ◽  
Acidic Salt ◽  
The 16S Rrna Gene

To understand the microbial community inhabiting in an acidic salt flat the phylogenetic diversity and the geochemistry of this system was compared to acid mine drainage (AMD) systems. The microbial community structure was assessed by DNA extraction/PCR/DGGE and secuencing for the 16S rRNA gene and the geochemistry was analyzed using several approaches. Prediction of metagenome functional content was performed from the 16S rRNA gene survey using the bioinformatics software package Phylogenetic Investigation of Communities by Reconstruction of Unobserved States (PICRUSt). The geochemical results revealed a much lower iron concentration in the salt flat than in AMD systems (39 and 21804 mg L-1, respectively) and a significant difference in chloride levels. Sequences inferred to be from potential sulfur metabolizing organisms constituted up to 70% of the microbial community in the acidic salt flat meanwhile predominat iron-metabolizing acidophile populations were reported in AMD systems. Interestingly, the microbial assemblage in the acidic salt flat was dominated by mixotrophic and organotrophic sulfur oxidizers as well as by photoautotrophic acidophiles. Our results suggests that the salt concentration in Salar de Gorbea (average Cl-= 40 gL-1) is in the limit for the occurrence of chemolithotrophic oxidation of sulfur compounds. In addition, the investigation allows concluding that salinity rather than extremes of pH is the major environmental determinant of microbial community composition.

scholarly journals The impact of persistent bacterial bronchitis on the pulmonary microbiome of children

2017 ◽  
Author(s):  
Leah Cuthbertson ◽  
Vanessa Craven ◽  
Lynne Bingle ◽  
William O.C.M. Cookson ◽  
Mark L. Everard ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Bacterial Diversity ◽  
Community Composition ◽  
Bacterial Communities ◽  
Microbial Community Analysis ◽  
Rrna Gene ◽  
Healthy Children ◽  
Significant Difference ◽  
The Impact

AbstractPersistent bacterial bronchitis is a leading cause of chronic wet cough in young children. This study aimed to characterise the respiratory bacterial microbiota of healthy children and to assess the impact of the changes associated with the development of persistent bacterial bronchitis.Blind, protected brushings were obtained from 20 healthy controls and 24 children with persistent bacterial bronchitis, with an additional directed sample obtained from persistent bacterial bronchitis patients. DNA was extracted, quantified using a 16S rRNA gene quantitative PCR assay prior to microbial community analysis by 16S rRNA gene sequencing.No significant difference in bacterial diversity or community composition (R2 = 0.01, P = 0.36) was observed between paired blind and non-blind brushes, showing that blind brushings are a valid means of accessing the airway microbiota. This has important implications for collecting lower respiratory samples from healthy children. A significant decrease in bacterial diversity (P < 0.001) and change in community composition (R2 = 0.08, P = 0.004) was observed between controls and patients. Bacterial communities within patients with PBB were dominated by Proteobacteria, and indicator species analysis showed that Haemophilus and Neisseria were significantly associated with the patient group. In 15 (52.9%) cases the dominant organism by sequencing was not identified by standard routine clinical culture.The bacteria present in the lungs of patients with persistent bacterial bronchitis were less diverse in terms of richness and evenness. The results validate the clinical diagnosis, and suggest that more attention to bacterial communities in children with chronic cough may lead to more rapid recognition of this condition with earlier treatment and reduction in disease burden.

scholarly journals Prokaryotic diversity and potentially pathogenic bacteria in vended foods and environmental samples

2021 ◽  
Vol 71 (1) ◽  
Author(s):  
Susan W. Muriuki ◽  
Michael S. Rengan ◽  
Nancy L. M. Budambula
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Environmental Samples ◽  
Pathogenic Bacteria ◽  
Risk Group ◽  
Foodborne Diseases ◽  
Rrna Gene ◽  
Prokaryotic Diversity ◽  
Hygiene Measures ◽  
Significant Difference

Abstract Purpose Ready-to-eat fast food vending outlets provide a cheap and readily available food. Foodborne diseases have been previously reported in Embu, Kenya, but data on the prokaryotic metagenome in vended foods is scanty. This study aimed to determine the prokaryotic diversity in fruits, vegetable salad, African sausage, chips (potato fries), fried fish, roasted beef (meat), smokies, samosa, soil, and water collected from food vendors and the surrounding environment in Embu Town and Kangaru Market. Methods The study used 454 pyrosequencing, Illumina high-throughput sequencing of 16S rRNA gene in the analysis of total community DNA extracted from samples using the phenol-chloroform method. The 16S rRNA gene variable region (V4-V7) of the extracted DNA was amplified and library construction performed. Sequence analysis was done using QIIME2. Hierarchical clustering of samples, diversity indices, rarefaction curves, and Venn diagrams were generated using the R programming language in R software version 3.6.3. Results Bacterial operational taxonomic units (OUTs) were distributed in Proteobacteria (52.81%), Firmicutes (31.16%), and Lentisphaerae (0.001%). The OTUs among archaea were Candidatus Nitrososphaera (63.56%) and Nitrososphaera spp. (8.77%). Brucella spp. and Bacillus cereus associated with foodborne diseases were detected. Potential pathogens, Rickettsia spp. in risk group 2 and Brucella spp. in risk group 3, were detected. Uncultured Candidatus Koribacter and Candidatus Solibacter were also detected in the food samples. There was a significant difference in the microbial community structure among the sample types (P<0.1). Conclusion The results demonstrated the presence of some prokaryotes that are associated with food spoilage or foodborne diseases in vended foods and environmental samples. This study also detected uncultured prokaryotes. The presence of potential pathogens calls for stringent hygiene measures in food vending operations.

scholarly journals Stool metabolome-microbiota evaluation among children and adolescents with obesity, overweight, and normal-weight using 1H NMR and 16S rRNA gene profiling

PLoS ONE ◽  
2021 ◽  
Vol 16 (3) ◽  
pp. e0247378
Author(s):  
José Diógenes Jaimes ◽  
Andrea Slavíčková ◽  
Jakub Hurych ◽  
Ondřej Cinek ◽  
Ben Nichols ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Alpha Diversity ◽  
Study Groups ◽  
Normal Weight ◽  
Gene Profiling ◽  
Energy Harvest ◽  
Rrna Gene ◽  
H Nmr ◽  
Significant Difference

Characterization of metabolites and microbiota composition from human stool provides powerful insight into the molecular phenotypic difference between subjects with normal weight and those with overweight/obesity. The aim of this study was to identify potential metabolic and bacterial signatures from stool that distinguish the overweight/obesity state in children/adolescents. Using 1H NMR spectral analysis and 16S rRNA gene profiling, the fecal metabolic profile and bacterial composition from 52 children aged 7 to 16 was evaluated. The children were classified into three groups (16 with normal-weight, 17 with overweight, 19 with obesity). The metabolomic analysis identified four metabolites that were significantly different (p < 0.05) among the study groups based on one-way ANOVA testing: arabinose, butyrate, galactose, and trimethylamine. Significantly different (p < 0.01) genus-level taxa based on edgeR differential abundance tests were genus Escherichia and Tyzzerella subgroup 3. No significant difference in alpha-diversity was detected among the three study groups, and no significant correlations were found between the significant taxa and metabolites. The findings support the hypothesis of increased energy harvest in obesity by human gut bacteria through the growing observation of increased fecal butyrate in children with overweight/obesity, as well as an increase of certain monosaccharides in the stool. Also supported is the increase of trimethylamine as an indicator of an unhealthy state.

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