Presence of Bacteria in Spontaneous Achilles Tendon Ruptures

2017 ◽  
Vol 45 (9) ◽  
pp. 2061-2067 ◽  
Author(s):  
Christer G. Rolf ◽  
Sai-Chuen Fu ◽  
Chelsea Hopkins ◽  
Ju Luan ◽  
Margaret Ip ◽  
...  
Keyword(s):  
Achilles Tendon ◽  
Acl Reconstruction ◽  
Bacterial Infections ◽  
Bacterial Species ◽  
Hamstring Tendon ◽  
Universal Primers ◽  
Rrna Gene ◽  
Low Grade ◽  
Bacterial Dna ◽  
Normal Tendon

Background: The structural pathology of Achilles tendon (AT) ruptures resembles tendinopathy, but the causes remain unknown. Recently, a number of diseases were found to be attributed to bacterial infections, resulting in low-grade inflammation and progressive matrix disturbance. The authors speculate that spontaneous AT ruptures may also be influenced by the presence of bacteria. Hypothesis: Bacteria are present in ruptured ATs but not in healthy tendons. Study Design: Cross-sectional study; Level of evidence, 3. Methods: Patients with spontaneous AT ruptures and patients undergoing anterior cruciate ligament (ACL) reconstruction were recruited for this study. During AT surgical repair, excised tendinopathic tissue was collected, and healthy tendon samples were obtained as controls from hamstring tendon grafts used in ACL reconstruction. Half of every sample was reserved for DNA extraction and the other half for histology. Polymerase chain reaction (PCR) was conducted using 16S rRNA gene universal primers, and the PCR products were sequenced for the identification of bacterial species. A histological examination was performed to compare tendinopathic changes in the case and control samples. Results: Five of 20 AT rupture samples were positive for the presence of bacterial DNA, while none of the 23 hamstring tendon samples were positive. Sterile operating and experimental conditions and tests on samples, controlling for harvesting and processing procedures, ruled out the chance of postoperative bacterial contamination. The species identified predominantly belonged to the Staphylococcus genus. AT rupture samples exhibited histopathological features characteristic of tendinopathy, and most healthy hamstring tendon samples displayed normal tendon features. There were no apparent differences in histopathology between the bacterial DNA–positive and bacterial DNA–negative AT rupture samples. Conclusion: The authors have demonstrated the presence of bacterial DNA in ruptured AT samples. It may suggest the potential involvement of bacteria in spontaneous AT ruptures.

scholarly journals Two-Step PCR-Based Assay for Identification of Bacterial Etiology of Otitis Media with Effusion in Infected Lebanese Children

1998 ◽  
Vol 36 (5) ◽  
pp. 1185-1188 ◽  
Author(s):  
Ghassan M. Matar ◽  
Nada Sidani ◽  
Michel Fayad ◽  
Usamah Hadi
Keyword(s):  
Otitis Media ◽  
Otitis Media With Effusion ◽  
Bacterial Species ◽  
Cost Effective ◽  
Universal Primers ◽  
Rrna Gene ◽  
Specific Primer ◽  
Specific Primers ◽  
Bacterial Dna ◽  
Species Specific

We developed and evaluated a two-step PCR-based assay with universal primers and genus- or species-specific primers for the detection of the most prevalent bacterial etiologies of otitis media with effusion (OME) in children from Lebanese hospitals. These etiologies included Haemophilus, Streptococcus, and Moraxella (Branhamella)catarrhalis, which were detected in middle-ear effusion (MEE) samples taken from children with OME. A total of 47 MEE samples were aspirated from 36 patients during insertion of a tympanostomy tube performed particularly for OME. The duration of effusion in all patients was ≥2 months. DNA was extracted from MEE samples, and PCR was initially done with DNA extracts by using the universal primers RW01 and DG74, which flank an ∼370-bp fragment found in the 16S rRNA gene of all bacterial species. For the identification of specific bacteria, we used in three separate reaction mixtures the following genus- or species-specific primers: (i) aHaemophilus-specific probe (probe RDR125) as a primer along with DG74, (ii) a Streptococcus-specific primer (primer STR1; designed by us) along with DG74, and (iii) an M. catarrhalis-specific primer pair (primer pair MCA1-MCA2). Thirty-five MEE samples (74.5%) gave the expected 370-bp band, indicating the presence of bacterial DNA in the tested samples. Of the 35 PCR-positive samples tested, 33 (94.3%) were positive forHaemophilus, 3 (8.6%) were positive forStreptococcus, and 10 (28.6%) were positive for M. catarrhalis. Ten samples (28.6%) exhibited a mixed infection and were positive for both Haemophilus and M. catarrhalis. Culture was simultaneously performed for all 47 MEE samples. Ten of the 47 MEE samples (21.3%) exhibited bacterial growth. These 10 were PCR positive for bacterial DNA. The remaining 25 PCR-positive samples were negative by culture, thus showing about 53% discordance between PCR results and those of culture. The PCR assay proved to be more sensitive than culture, more rapid, less cumbersome, and more cost-effective than the available PCR-Southern hybridization-based assays.

scholarly journals Whipple's Disease and “Tropheryma whippelii”

2001 ◽  
Vol 14 (3) ◽  
pp. 561-583 ◽  
Author(s):  
Fabrizio Dutly ◽  
Martin Altwegg
Keyword(s):  
Bacterial Species ◽  
Periodic Acid ◽  
Clinical Manifestations ◽  
The Body ◽  
Rrna Gene ◽  
Low Grade ◽  
Whipple’S Disease ◽  
Major Step ◽  
Whipple's Disease ◽  
Initial Symptoms

SUMMARY Whipple's disease is a rare bacterial infection that may involve any organ system in the body. It occurs primarily in Caucasian males older than 40 years. The gastrointestinal tract is the most frequently involved organ, with manifestations such as abdominal pain, malabsorption syndrome with diarrhea, and weight loss. Other signs include low-grade fever, lymphadenopathy, skin hyperpigmentation, endocarditis, pleuritis, seronegative arthritis, uveitis, spondylodiscitis, and neurological manifestations, and these signs may occur in the absence of gastrointestinal manifestations. Due to the wide variability of manifestations, clinical diagnosis is very difficult and is often made only years or even decades after the initial symptoms have appeared. Trimethoprim-sulfamethoxazole for at least 1 year is usually considered adequate to eradicate the infection. The microbiological diagnosis of this insidious disease is rendered difficult by the virtual lack of culture and serodiagnostic methods. It is usually based on the demonstration of periodic acid-Schiff-positive particles in infected tissues and/or the presence of bacteria with an unusual trilaminar cell wall ultrastructure by electron microscopy. Recently, the Whipple bacteria have been characterized at the molecular level by amplification of their 16S rRNA gene(s). Phylogenetic analysis of these sequences revealed a new bacterial species related to the actinomycete branch which was named “Tropheryma whippelli.” Based on its unique 16S ribosomal DNA (rDNA) sequence, species-specific primers were selected for the detection of the organism in clinical specimens by PCR. This technique is currently used as one of the standard methods for establishing the diagnosis of Whipple's disease. Specific and broad-spectrum PCR amplifications mainly but not exclusively from extraintestinal specimens have significantly improved diagnosis, being more sensitive than histopathologic analysis. However, “T. whippelii” DNA has also been found in persons without clinical and histological evidence of Whipple's disease. It is unclear whether these patients are true asymptomatic carriers or whether differences in virulence exist among strains of “T. whippelii” that might account for the variable clinical manifestations. So far, six different “T. whippelii” subtypes have been found by analysis of their 16S-23S rDNA spacer region. Further studies of the pathogen “T. whippelii” as well as the host immune response are needed to fully understand this fascinating disease. The recent cultivation of the organisms is a promising major step in this direction.

scholarly journals Sensitive Identification of Bacterial DNA in Clinical Specimens by Broad-Range 16S rRNA Gene Enrichment

2020 ◽  
Vol 58 (12) ◽  
Author(s):  
Sara Rassoulian Barrett ◽  
Noah G. Hoffman ◽  
Christopher Rosenthal ◽  
Andrew Bryan ◽  
Desiree A. Marshall ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Nucleic Acids ◽  
Bacterial Species ◽  
False Negative ◽  
Case Series ◽  
Rrna Gene ◽  
Metagenomic Sequencing ◽  
Bacterial Dna ◽  
Clinical Specimens

ABSTRACT The broad-range detection and identification of bacterial DNA from clinical specimens are a foundational approach in the practice of molecular microbiology. However, there are circumstances under which conventional testing may yield false-negative or otherwise uninterpretable results, including the presence of multiple bacterial templates or degraded nucleic acids. Here, we describe an alternative, next-generation sequencing approach for the broad range detection of bacterial DNA using broad-range 16S rRNA gene hybrid capture (“16S Capture”). The method is able to deconvolute multiple bacterial species present in a specimen, is compatible with highly fragmented templates, and can be readily implemented when the overwhelming majority of nucleic acids in a specimen derive from the human host. We find that this approach is sensitive to detecting as few as 17 Staphylococcus aureus genomes from a background of 100 ng of human DNA, providing 19- to 189-fold greater sensitivity for identifying bacterial sequences than standard shotgun metagenomic sequencing, and is able to successfully recover organisms from across the eubacterial tree of life. Application of 16S Capture to a proof-of-principle case series demonstrated its ability to identify bacterial species that were consistent with histological evidence of infection, even when diagnosis could not be established using conventional broad range bacterial detection assays. 16S Capture provides a novel means for the efficient and sensitive detection of bacteria embedded in human tissues and for specimens containing highly fragmented template DNA.

scholarly journals Isolation of Vibrio alginolyticus and Vibrio splendidus from Aquacultured Carpet Shell Clam (Ruditapes decussatus) Larvae Associated with Mass Mortalities

2005 ◽  
Vol 71 (1) ◽  
pp. 98-104 ◽  
Author(s):  
J. G�mez-Le�n ◽  
L. Villamil ◽  
M. L. Lemos ◽  
B. Novoa ◽  
A. Figueras
Keyword(s):  
Enzymatic Activity ◽  
Pure Culture ◽  
Bacterial Infections ◽  
Soft Tissues ◽  
Bacterial Species ◽  
Larval Mortality ◽  
Vibrio Alginolyticus ◽  
Rrna Gene ◽  
Ruditapes Decussatus ◽  
Vibrio Splendidus

ABSTRACT Two episodes of mortality of cultured carpet shell clams (Ruditapes decussatus) associated with bacterial infections were recorded during 2001 and 2002 in a commercial hatchery located in Spain. Vibrio alginolyticus was isolated as the primary organism from moribund clam larvae that were obtained during the two separate events. Vibrio splendidus biovar II, in addition to V. alginolyticus, was isolated as a result of a mixed Vibrio infection from moribund clam larvae obtained from the second mortality event. The larval mortality rates for these events were 62 and 73%, respectively. Mortality was also detected in spat. To our knowledge, this is the fist time that these bacterial species have been associated with larval and juvenile carpet shell clam mortality. The bacterial strains were identified by morphological and biochemical techniques and also by PCR and sequencing of a conserved region of the 16S rRNA gene. In both cases bacteria isolated in pure culture were inoculated into spat of carpet shell clams by intravalvar injection and by immersion. The mortality was attributed to the inoculated strains, since the bacteria were obtained in pure culture from the soft tissues of experimentally infected clams. V. alginolyticus TA15 and V. splendidus biovar II strain TA2 caused similar histological lesions that affected mainly the mantle, the velum, and the connective tissue of infected organisms. The general enzymatic activity of both live cells and extracellular products (ECPs), as evaluated by the API ZYM system, revealed that whole bacterial cells showed greater enzymatic activity than ECPs and that the activity of most enzymes ceased after heat treatment (100�C for 10 min). Both strain TA15 and strain TA2 produced hydroxamate siderophores, although the activity was greater in strain TA15. ECPs from both bacterial species at high concentrations, as well as viable bacteria, caused significant reductions in hemocyte survival after 4 h of incubation, whereas no significant differences in viability were observed during incubation with heat-killed bacteria.

scholarly journals A multi-view model for relative and absolute microbial abundances

2019 ◽  
Author(s):  
Brian D. Williamson ◽  
James P. Hughes ◽  
Amy D. Willis
Keyword(s):  
Relative Abundance ◽  
Estimation Error ◽  
Bacterial Species ◽  
Critical Role ◽  
Sensitivity Study ◽  
Universal Primers ◽  
Rrna Gene ◽  
Absolute Abundance ◽  
Abundance Data ◽  
The Absolute

AbstractThe absolute abundance of bacterial taxa in human host-associated environments play a critical role in reproductive and gastrointestinal health. However, obtaining the absolute abundance of many bacterial species is typically prohibitively expensive. In contrast, relative abundance data for many species is comparatively cheap and easy to collect (e.g., with universal primers for the 16S rRNA gene). In this paper, we propose a method to jointly model relative abundance data for many taxa and absolute abundance data for a subset of taxa. Our method provides point and interval estimates for the absolute abundance of all taxa. Crucially, our proposal accounts for differences in the efficiency of taxon detection in the relative and absolute abundance data. We show that modeling taxon-specific efficiencies substantially reduces the estimation error for absolute abundance, and controls the coverage of interval estimators. We demonstrate the performance of our proposed method via a simulation study, a sensitivity study where we jackknife the taxa with observed absolute abundances, and a study of women with bacterial vaginosis.

scholarly journals Characterization of Natural Lactic Flora in a Soft Cheese “Camembert of the Tessala” Made From Thermised Milk of Local Breed Cow “Brown of the Atlas”

2020 ◽  
Vol 39 (01) ◽  
Author(s):  
A. , A. , A. Dahou ◽  
A. M.A. Bekada ◽  
M. Medjahed ◽  
H. Tahlaiti ◽  
S. N. Rechidi ◽  
...  
Keyword(s):  
Bacterial Species ◽  
Large Degree ◽  
Taste Quality ◽  
Universal Primers ◽  
Bacterial Dna ◽  
Local Breed ◽  
Isolation And Purification ◽  
Soft Cheese ◽  
Positive Controls

Microbial communities play an important role in the maturation of cheese and determine to a large degree its taste quality. The typicality and the sensory richness of Tessala camembert diversity is preserved by this microflora. In the present study, we tried after isolation and purification to characterize genotypically lactic microflora of this cheese at the end of maturation of the transformation from thermized milk of local breed cow. The bacterial DNA from twenty-two purified lactic culture was established by an amplification of ribosomal DNA 16S by specific universal primers of prokaryotes with strains reference of each isolated bacterial species used as positive controls. The results show the characteristic of the Tessala camembert by the diversity of its original lactic flora dominated by lactococci, enterococci, lactobacilli, pediococci and leuconostocs, preserved by the initial thermization of milk used to maintain the technologically-interested dairy microbial ecosystem.

scholarly journals Identification of a Lactococcus lactis Isolated from a Fresh Local Cheese of the Western Algerian Steppe “J’ben of Naama”

2021 ◽  
Author(s):  
A.A. Dahou ◽  
A.A. Bekada ◽  
A. Homrani
Keyword(s):  
Dna Sequencing ◽  
Mixed Culture ◽  
Lactococcus Lactis ◽  
Ribosomal Dna ◽  
Bacterial Species ◽  
Sheep Breed ◽  
Universal Primers ◽  
Bacterial Dna ◽  
Genotypic Identification ◽  
Lactic Bacteria

Background: The purpose of the study is to evaluate the diversity of lactococci, lactic bacteria, recovered from “J’ben”, a local cheese made from the milk of the Rembi sheep breed, a product of exploitation in the Algerian steppe regions of Naama.Methods: The bacterial species were isolated from samples of the recovered cheese exploitation and analyzed using genotypic methods. The isolation of bacterial DNA from purified Lactococcus cultures has been established by an amplification of ribosomal DNA 16s using the specific universal primers of prokaryotes. Result: The 16s DNA sequencing of all isolates, for genotypic identification, confirmed the predominance of Lactococcus lactis. This lactic dominance determines the quality and distinctiveness of this cheese in the region. The results obtained from acidification and proteolysis kinetics met the technological requirements and good functionality, from the strains used individually and in mixed culture to the cheese processing.

scholarly journals Identification and Localization of Extraradicular Biofilm-Forming Bacteria Associated with Refractory Endodontic Pathogens

2005 ◽  
Vol 71 (12) ◽  
pp. 8738-8743 ◽  
Author(s):  
Nobuo Noguchi ◽  
Yuichiro Noiri ◽  
Masahiro Narimatsu ◽  
Shigeyuki Ebisu
Keyword(s):  
Biofilm Formation ◽  
Bacterial Species ◽  
Fusobacterium Nucleatum ◽  
Immunohistochemical Localization ◽  
Rrna Gene ◽  
Root Tips ◽  
Bacterial Dna ◽  
Gene Assay ◽  
Periapical Periodontitis ◽  
Root Surfaces

ABSTRACT Bacterial biofilms have been found to develop on root surfaces outside the apical foramen and be associated with refractory periapical periodontitis. However, it is unknown which bacterial species form extraradicular biofilms. The present study aimed to investigate the identity and localization of bacteria in human extraradicular biofilms. Twenty extraradicular biofilms, used to identify bacteria using a PCR-based 16S rRNA gene assay, and seven root-tips, used to observe immunohistochemical localization of three selected bacterial species, were taken from 27 patients with refractory periapical periodontitis. Bacterial DNA was detected from 14 of the 20 samples, and 113 bacterial species were isolated. Fusobacterium nucleatum (14 of 14), Porphyromonas gingivalis (12 of 14), and Tannellera forsythensis (8 of 14) were frequently detected. Unidentified and uncultured bacterial DNA was also detected in 11 of the 14 samples in which DNA was detected. In the biofilms, P. gingivalis was immunohistochemically detected in all parts of the extraradicular biofilms. Positive reactions to anti-F. nucleatum and anti-T. forsythensis sera were found at specific portions of the biofilm. These findings suggested that P. gingivalis, T. forsythensis, and F. nucleatum were associated with extraradicular biofilm formation and refractory periapical periodontitis.

DIVERSITY OF CULTURABLE BACTERIAL MICROBIOTA OF THE Eisenia foetida DIGESTIVE TRACT

2018 ◽  
Vol 41 (3) ◽  
pp. 255-264 ◽  
Author(s):  
J. Abraham Pérez-Pérez ◽  
David Espinosa-Victoria ◽  
Hilda V. Silva-Rojas ◽  
Lucía López-Reyes
Keyword(s):  
Bacterial Diversity ◽  
Digestive Tract ◽  
Bacterial Species ◽  
Brain Heart Infusion ◽  
Rrna Gene ◽  
Eisenia Foetida ◽  
Bacterial Isolates ◽  
Bacterial Microbiota ◽  
Decomposition Of Organic Matter ◽  
Earthworm Gut

Bacteria are an unavoidable component of the natural earthworm diet; thus, bacterial diversity in the earthworm gut is directly linked to decomposition of organic matter and development of the surrounding plants. The aim of this research was to isolate and to identify biochemically and molecularly the culturable bacterial microbiota of the digestive tract of Eisenia foetida. Earthworms were sourced from Instituto de Reconversión Productiva y Bioenergética (IRBIO) and Colegio de Postgraduados (COLPOS), México. Bacterial isolation was carried out on plates of Brain Heart Infusion (BHI) culture medium. Fifty six and 44 bacterial isolates were obtained from IRBIO and COLPOS, respectively. The population was composed of 44 Gram-negative and 56 Gram-positive isolates. Over 50 % of the bacterial isolates were rod-shaped cells. The 16S rRNA gene was sequenced and nine genera were identified in worms from IRBIO (Bacillus, Paenibacillus, Solibacillus, Staphylococcus, Arthrobacter, Pantoea, Stenotrophomonas, Acinetobacter and Aeromonas) and six in worms from COLPOS (Bacillus, Paenibacillus, Stenotrophomonas, Staphylococcus, Acinetobacter and Aeromonas). Bacillus was the predominant genus, with eight and six species in the oligochaetes from IRBIO and COLPOS, respectively. The most represented bacteria in the worms from both sites were Bacillus sp. and B. subtilis. The predominance of Bacillus was probably due to spore formation, a reproductive strategy that ensures survival and dispersion in the soil and oligochaetes digestive tract. The gut of E. foetida not only harbored bacterial species of agronomic importance but also species potentially pathogenic for humans (Staphylococcus warneri, Pantoea agglomerans and Stentrophomonas sp.). The larger bacterial diversity in worms from IRBIO could be due to their feeding on cattle manure, which is a rich source of bacteria.

scholarly journals Amplicon-sequencing of raw milk microbiota: impact of DNA extraction and library-PCR

2021 ◽  
Author(s):  
Annemarie Siebert ◽  
Katharina Hofmann ◽  
Lena Staib ◽  
Etienne V. Doll ◽  
Siegfried Scherer ◽  
...  
Keyword(s):  
Sample Preparation ◽  
Dna Extraction ◽  
Raw Milk ◽  
Amplicon Sequencing ◽  
Rrna Gene ◽  
Bacterial Dna ◽  
Microbiome Analysis ◽  
Eukaryotic Dna ◽  
Selective Lysis ◽  
The Impact

Abstract The highly complex raw milk matrix challenges the sample preparation for amplicon-sequencing due to low bacterial counts and high amounts of eukaryotic DNA originating from the cow. In this study, we optimized the extraction of bacterial DNA from raw milk for microbiome analysis and evaluated the impact of cycle numbers in the library-PCR. The selective lysis of eukaryotic cells by proteinase K and digestion of released DNA before bacterial lysis resulted in a high reduction of mostly eukaryotic DNA and increased the proportion of bacterial DNA. Comparative microbiome analysis showed that a combined enzymatic and mechanical lysis procedure using the DNeasy® PowerFood® Microbial Kit with a modified protocol was best suitable to achieve high DNA quantities after library-PCR and broad coverage of detected bacterial biodiversity. Increasing cycle numbers during library-PCR systematically altered results for species and beta-diversity with a tendency to overrepresentation or underrepresentation of particular taxa. To limit PCR bias, high cycle numbers should thus be avoided. An optimized DNA extraction yielding sufficient bacterial DNA and enabling higher PCR efficiency is fundamental for successful library preparation. We suggest that a protocol using ethylenediaminetetraacetic acid (EDTA) to resolve casein micelles, selective lysis of somatic cells, extraction of bacterial DNA with a combination of mechanical and enzymatic lysis, and restriction of PCR cycles for analysis of raw milk microbiomes is optimal even for samples with low bacterial numbers. Key points • Sample preparation for high-throughput 16S rRNA gene sequencing of raw milk microbiota. • Reduction of eukaryotic DNA by enzymatic digestion. • Shift of detected microbiome caused by high cycle numbers in library-PCR.

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