scholarly journals Infective Arthritis: Bacterial 23S rRNA Gene Sequencing as a Supplementary Diagnostic Method

2008 ◽  
Vol 2 (1) ◽  
pp. 85-88 ◽  
Author(s):  
Claus Moser ◽  
Keld Andresen ◽  
Anne Kjerulf ◽  
Suheil Salamon ◽  
Michael Kemp ◽  
...  
Keyword(s):  
Diagnostic Method ◽  
Gene Sequencing ◽  
23S Rrna ◽  
Rrna Gene ◽  
23S Rrna Gene ◽  
Rrna Gene Sequencing ◽  
Infective Arthritis

scholarly journals Investigation of Linezolid Resistance in Staphylococci and Enterococci

2016 ◽  
Vol 54 (5) ◽  
pp. 1289-1294 ◽  
Author(s):  
Christopher D. Doern ◽  
Jason Y. Park ◽  
Michael Gallegos ◽  
Debbie Alspaugh ◽  
Carey-Ann D. Burnham
Keyword(s):  
Susceptibility Testing ◽  
Gene Sequencing ◽  
Disk Diffusion ◽  
23S Rrna ◽  
Poor Correlation ◽  
Rrna Gene ◽  
Broth Microdilution ◽  
Linezolid Resistance ◽  
23S Rrna Gene ◽  
Rrna Gene Sequencing

The objective of this study was to investigate an apparent increase in linezolid-nonsusceptible staphylococci and enterococci following a laboratory change in antimicrobial susceptibility testing from disk diffusion to an automated susceptibility testing system. Isolates with nonsusceptible results (n= 27) from Vitek2 were subjected to a battery of confirmatory testing which included disk diffusion, Microscan broth microdilution, Clinical and Laboratory Standards Institute (CLSI) reference broth microdilution, gradient diffusion (Etest), 23S rRNA gene sequencing, andcfrPCR. Our results show that there is poor correlation between methods and that only 70 to 75% of isolates were confirmed as linezolid resistant with alternative phenotypic testing methods (disk diffusion, Microscan broth microdilution, CLSI broth microdilution, and Etest). 23S rRNA gene sequencing identified mutations previously associated with linezolid resistance in 16 (59.3%) isolates, and thecfrgene was detected in 3 (11.1%) isolates. Mutations located at positions 2576 and 2534 of the 23S rRNA gene were most common. In addition, two previously undescribed variants (at positions 2083 and 2345 of the 23S rRNA gene) were also identified and may contribute to linezolid resistance.

scholarly journals Characterization of Lactic Bacteria Isolated from Raw Milk and Their Antibacterial Activity against Bacteria as the Cause of Clinical Bovine Mastitis

2021 ◽  
Vol 2021 ◽  
pp. 1-8
Author(s):  
Nada K. Alharbi ◽  
Albandary Nasser Alsaloom
Keyword(s):  
Bovine Mastitis ◽  
16S Rrna Gene Sequencing ◽  
Raw Milk ◽  
Antibacterial Activities ◽  
Antagonistic Activity ◽  
23S Rrna ◽  
Rrna Gene ◽  
Technological Properties ◽  
23S Rrna Gene ◽  
Rrna Gene Sequencing

The objectives of this study were the selection of lactic acid bacteria (LAB) isolated from raw milk and studying their technological properties and antibacterial activities against bacteria as the cause of cattle mastitis. Biochemical and molecular identification using 16S–23S rRNA gene spacer analysis and 16S rRNA gene sequencing highlighted the presence of three species: Lactiplantibacillus plantarum, Lactococcus lactis, and Levilactobacillus brevis. The enzymatic characterization followed by the determination of technofunctional properties showed that LAB strains did not exhibit any hemolytic effect and were able to produce protease and lipase enzymes. Isolates showed very high antagonistic activity against Gram-positive and Gram-negative bacteria by producing H2O2, bacteriocin(s), and organic acid(s). APIZYM micromethod demonstrated that all selected strains are capable of producing valine arylamidase, cystine arylamidase, N-acetyl-β-glucosaminidase, and ᾳ-mannosidase. The antibiotic susceptibility assay showed that all selected strains were sensible to the majority of tested antibiotics. Based on these results, it can be concluded that the technological properties of the selected LAB allow considering their industrial use in order to formulate bioactive functional foods or drug(s).

scholarly journals Diversity of Domain V of 23S rRNA Gene Sequence in Different Enterococcus Species

2000 ◽  
Vol 38 (11) ◽  
pp. 3991-3993 ◽  
Author(s):  
Sotirios Tsiodras ◽  
Howard S. Gold ◽  
Eoin P. G. Coakley ◽  
Christine Wennersten ◽  
Robert C. Moellering ◽  
...  
Keyword(s):  
16S Rrna Gene Sequencing ◽  
23S Rrna ◽  
Rrna Gene ◽  
Dna Encoding ◽  
Enterococcus Casseliflavus ◽  
23S Rrna Gene ◽  
Rrna Gene Sequencing ◽  
Biochemical Testing ◽  
Enterococcus Gallinarum ◽  
Domain V

The highly conserved central loop of domain V of 23S RNA (nucleotides 2042 to 2628; Escherichia coli numbering) is implicated in peptidyltransferase activity and represents one of the target sites for macrolide, lincosamide, and streptogramin B antibiotics. DNA encoding domain V (590 bp) of several species ofEnterococcus was amplified by PCR. Twenty enterococcal isolates were tested, including Enterococcus faecium (six isolates), Enterococcus faecalis, Enterococcus avium, Enterococcus durans, Enterococcus gallinarum, Enterococcus casseliflavus (two isolates of each), and Enterococcus raffinosus, Enterococcus mundtii, Enterococcus malodoratus, andEnterococcus hirae (one isolate of each). For all isolates, species identification by biochemical testing was corroborated by 16S rRNA gene sequencing. The sequence of domain V of the 23S rRNA gene from E. faecium and E. faecalis differed from those of all other enterococci. The domain V sequences of E. durans and E. hirae were identical. This was also true for E. gallinarum and E. casseliflavus. E. avium differed from E. casseliflavus by 23 bases, from E. durans by 16 bases, and from E. malodoratus by 2 bases. E. avium differed fromE. raffinosus by one base. Despite the fact that domain V is considered to be highly conserved, substantial differences were identified between several enterococcal species.

Screening and Molecular Characterization of Cellulase Producing Actinobacteria from Litchi Orchard

2019 ◽  
Vol 13 (1) ◽  
pp. 90-101
Author(s):  
Sanju Kumari ◽  
Utkarshini Sharma ◽  
Rohit Krishna ◽  
Kanak Sinha ◽  
Santosh Kumar
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Molecular Characterization ◽  
Biochemical Characterization ◽  
Evolutionary Relationship ◽  
Gene Sequencing ◽  
Cellulase Activity ◽  
16S Rrna Gene Sequencing ◽  
Rrna Gene ◽  
Rrna Gene Sequencing

Background: Cellulolysis is of considerable economic importance in laundry detergents, textile and pulp and paper industries and in fermentation of biomass into biofuels. Objective: The aim was to screen cellulase producing actinobacteria from the fruit orchard because of its requirement in several chemical reactions. Methods: Strains of actinobacteria were isolated on Sabouraud’s agar medium. Similarities in cultural and biochemical characterization by growing the strains on ISP medium and dissimilarities among them perpetuated to recognise nine groups of actinobacteria. Cellulase activity was measured by the diameter of clear zone around colonies on CMC agar and the amount of reducing sugar liberated from carboxymethyl cellulose in the supernatant of the CMC broth. Further, 16S rRNA gene sequencing and molecular characterization were placed before NCBI for obtaining recognition with accession numbers. Results: Prominent clear zones on spraying Congo Red were found around the cultures of strains of three groups SK703, SK706, SK708 on CMC agar plates. The enzyme assay for carboxymethylcellulase displayed extra cellulase activity in broth: 0.14, 0.82 and 0.66 µmol mL-1 min-1, respectively at optimum conditions of 35°C, pH 7.3 and 96 h of incubation. However, the specific cellulase activities per 1 mg of protein did not differ that way. It was 1.55, 1.71 and 1.83 μmol mL-1 min-1. The growing mycelia possessed short compact chains of 10-20 conidia on aerial branches. These morphological and biochemical characteristics, followed by their verification by Bergey’s Manual, categorically allowed the strains to be placed under actinobacteria. Further, 16S rRNA gene sequencing, molecular characterization and their evolutionary relationship through phylogenetics also confirmed the putative cellulase producing isolates of SK706 and SK708 subgroups to be the strains of Streptomyces. These strains on getting NCBI recognition were christened as Streptomyces glaucescens strain SK91L (KF527284) and Streptomyces rochei strain SK78L (KF515951), respectively. Conclusion: Conclusive evidence on the basis of different parameters established the presence of cellulase producing actinobacteria in the litchi orchard which can convert cellulose into fermentable sugar.

scholarly journals Much ado about nothing? Off-target amplification can lead to false-positive bacterial brain microbiome detection in healthy and Parkinson’s disease individuals

Microbiome ◽  
2021 ◽  
Vol 9 (1) ◽  
Author(s):  
Janis R. Bedarf ◽  
Naiara Beraza ◽  
Hassan Khazneh ◽  
Ezgi Özkurt ◽  
David Baker ◽  
...  
Keyword(s):  
Parkinson’S Disease ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
False Positive ◽  
Gene Sequencing ◽  
Bacterial Biomass ◽  
16S Rrna Gene Sequencing ◽  
Amplicon Sequencing ◽  
Rrna Gene ◽  
Rrna Gene Sequencing

Abstract Background Recent studies suggested the existence of (poly-)microbial infections in human brains. These have been described either as putative pathogens linked to the neuro-inflammatory changes seen in Parkinson’s disease (PD) and Alzheimer’s disease (AD) or as a “brain microbiome” in the context of healthy patients’ brain samples. Methods Using 16S rRNA gene sequencing, we tested the hypothesis that there is a bacterial brain microbiome. We evaluated brain samples from healthy human subjects and individuals suffering from PD (olfactory bulb and pre-frontal cortex), as well as murine brains. In line with state-of-the-art recommendations, we included several negative and positive controls in our analysis and estimated total bacterial biomass by 16S rRNA gene qPCR. Results Amplicon sequencing did detect bacterial signals in both human and murine samples, but estimated bacterial biomass was extremely low in all samples. Stringent reanalyses implied bacterial signals being explained by a combination of exogenous DNA contamination (54.8%) and false positive amplification of host DNA (34.2%, off-target amplicons). Several seemingly brain-enriched microbes in our dataset turned out to be false-positive signals upon closer examination. We identified off-target amplification as a major confounding factor in low-bacterial/high-host-DNA scenarios. These amplified human or mouse DNA sequences were clustered and falsely assigned to bacterial taxa in the majority of tested amplicon sequencing pipelines. Off-target amplicons seemed to be related to the tissue’s sterility and could also be found in independent brain 16S rRNA gene sequences. Conclusions Taxonomic signals obtained from (extremely) low biomass samples by 16S rRNA gene sequencing must be scrutinized closely to exclude the possibility of off-target amplifications, amplicons that can only appear enriched in biological samples, but are sometimes assigned to bacterial taxa. Sequences must be explicitly matched against any possible background genomes present in large quantities (i.e., the host genome). Using close scrutiny in our approach, we find no evidence supporting the hypothetical presence of either a brain microbiome or a bacterial infection in PD brains.

scholarly journals Molecular pathways to high-level azithromycin resistance in Neisseria gonorrhoeae

2021 ◽  
Author(s):  
J G E Laumen ◽  
S S Manoharan-Basil ◽  
E Verhoeven ◽  
S Abdellati ◽  
I De Baetselier ◽  
...  
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Ribosomal Proteins ◽  
Efflux Pump ◽  
23S Rrna ◽  
Rrna Gene ◽  
23S Rrna Gene ◽  
Evolution Of Resistance ◽  
High Level ◽  
Azithromycin Resistance ◽  
Level Resistance

Abstract Background The prevalence of azithromycin resistance in Neisseria gonorrhoeae is increasing in numerous populations worldwide. Objectives To characterize the genetic pathways leading to high-level azithromycin resistance. Methods A customized morbidostat was used to subject two N. gonorrhoeae reference strains (WHO-F and WHO-X) to dynamically sustained azithromycin pressure. We tracked stepwise evolution of resistance by whole genome sequencing. Results Within 26 days, all cultures evolved high-level azithromycin resistance. Typically, the first step towards resistance was found in transitory mutations in genes rplD, rplV and rpmH (encoding the ribosomal proteins L4, L22 and L34 respectively), followed by mutations in the MtrCDE-encoded efflux pump and the 23S rRNA gene. Low- to high-level resistance was associated with mutations in the ribosomal proteins and MtrCDE efflux pump. However, high-level resistance was consistently associated with mutations in the 23S ribosomal RNA, mainly the well-known A2059G and C2611T mutations, but also at position A2058G. Conclusions This study enabled us to track previously reported mutations and identify novel mutations in ribosomal proteins (L4, L22 and L34) that may play a role in the genesis of azithromycin resistance in N. gonorrhoeae.

scholarly journals Molecular characterization of a ceftriaxone-resistant Neisseria gonorrhoeae strain found in Switzerland: a case report

2021 ◽  
Vol 20 (1) ◽  
Author(s):  
Konrad Egli ◽  
Anna Roditscheff ◽  
Ursula Flückiger ◽  
Martin Risch ◽  
Lorenz Risch ◽  
...  
Keyword(s):  
Neisseria Gonorrhoeae ◽  
23S Rrna ◽  
Rrna Gene ◽  
Limited Information ◽  
Specific Pcr ◽  
Appropriate Treatment ◽  
23S Rrna Gene ◽  
Allele Type ◽  
Ciprofloxacin Resistance

Abstract Background The resistance of Neisseria gonorrhoeae to ceftriaxone is unusual in Switzerland. The underlying genotype responsible for resistance is suspected to be novel. Generally, resistance in Neisseria gonorrhoeae (Ng) involves a comprehensive set of genes with many different mutations leading to resistance to different β-lactams and fluoroquinolones. Case presentation A patient had a positive result from specific PCR for Ng. We routinely culture all clinical specimens with a positive NG-PCR. In this particular case, we isolated a strain with resistance to ceftriaxone in Switzerland. A total of seven different genes (penA, ponA, porinB, mtr, gyrA, parC, 23S rRNA gene) in this strain were partially sequenced for comparison with phenotypic susceptibility testing. Interestingly, two different mutations in the porinB gene were observed, and data on this gene are limited. Information on the identified allele type of the penA gene is very limited as well. Three different mutations of parC and gyrA that correlate with ciprofloxacin resistance were found. The combination of ceftriaxone and ciprofloxacin resistance makes an appropriate treatment difficult to obtain due to multidrug resistance. Conclusion The combined results for all genes show the appearance of new mutations in central Europe either due to worldwide spread or the emergence of new genetic combinations of mutations.

scholarly journals The growth response of rice (Oryza sativa L. var. FARO 44) in vitro after inoculation with bacterial isolates from a typical ferruginous ultisol

2021 ◽  
Vol 45 (1) ◽  
Author(s):  
Musa Saheed Ibrahim ◽  
Beckley Ikhajiagbe
Keyword(s):  
16S Rrna ◽  
Bacillus Cereus ◽  
Proteus Mirabilis ◽  
Growth Response ◽  
Gene Sequencing ◽  
16S Rrna Gene Sequencing ◽  
Rrna Gene ◽  
Rice Seedlings ◽  
Rrna Gene Sequencing

Abstract Background Rice forms a significant portion of food consumed in most household worldwide. Rice production has been hampered by soil factors such as ferruginousity which has limited phosphorus availability; an important mineral component for the growth and yield of rice. The presence of phosphate-solubilizing bacteria (PSB) in soils has been reported to enhance phosphate availability. In view of this, the present study employed three bacteria species (BCAC2, EMBF2 and BCAF1) that were previously isolated and proved P solubilization capacities as inocula to investigate the growth response of rice germinants in an in vitro setup. The bacteria isolates were first identified using 16S rRNA gene sequencing and then applied as inoculum. The inolula were prepared in three concentrations (10, 7.5 and 5.0 ml) following McFarland standard. Viable rice (var. FARO 44) seeds were sown in petri dishes and then inoculated with the three inocula at the different concentrations. The setup was studied for 28 days. Results 16S rRNA gene sequencing identified the isolates as: isolate BCAC2= Bacillus cereus strain GGBSU-1, isolate BCAF1= Proteus mirabilis strain TL14-1 and isolate EMBF2= Klebsiella variicola strain AUH-KAM-9. Significant improvement in rice germination, morphology, physiology and biomass parameters in the bacteria-inoculated setups was observed compared to the control. Germination percentage after 4 days was 100 % in the inoculated rice germinants compared to 65% in the control (NiS). Similarly, inoculation with the test isolates enhanced water-use efficiency by over 40%. The rice seedlings inoculated with Bacillus cereus strain GGBSU-1 (BiS) showed no signs of chlorosis and necrosis throughout the study period as against those inoculated with Proteus mirabilis strain TL14-1 (PiS) and Klebsiella variicola strain AUH-KAM-9 (KiS). Significant increase in chlorophyll-a, chlorophyll-b and alpha amylase was observed in the rice seedlings inoculated with BiS as against the NiS. Conclusion Inoculating rice seeds with Bacillus cereus strain GGBSU-1, Proteus mirabilis strain TL14-1 and Klebsiella variicola strain AUH-KAM-9 in an in vitro media significantly improved growth parameters of the test plant. Bacillus cereus strain GGBSU-1 showed higher efficiency due to a more improved growth properties observed.

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