Routine DNA analysis based on 12S rRNA gene sequencing as a tool in the management of captive primates

A non-destructive method of collecting samples for DNA analysis of the Chinese giant salamander is described and validated. DNA was extracted from the skin secretion and shedding using a Chelex-based method, and partial 12S rRNA gene sequences were amplified and sequenced. Sequences from skin secretion and shedding were cross-checked against the reported sequences from liver and were found to be identical. This method provides a non-destructive way of carrying out larger studies of the genetics of rare amphibians and may be of general use for genetic-based field studies of amphibians.

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FIRST ANALYSIS FROM UK IBD TWIN BIOBANK; 16S RRNA GENE SEQUENCING IDENTIFIES REDUCED DIVERSITY IN ACTIVE IBD AND TAXA ASSOCIATED WITH ACTIVE DISEASE PHENOTYPE TO LEVEL OF SPECIES

10.26226/morressier.59a6b347d462b80290b54eec ◽

2017 ◽

Author(s):

Marcus Harbord

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Gene Sequencing ◽

16S Rrna Gene Sequencing ◽

Rrna Gene ◽

Disease Phenotype ◽

Rrna Gene Sequencing ◽

Active Disease

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Screening and Molecular Characterization of Cellulase Producing Actinobacteria from Litchi Orchard

Current Chemical Biology ◽

10.2174/2212796812666180718114432 ◽

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.

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A preliminary study on the potential of Nanopore MinION and Illumina MiSeq 16S rRNA gene sequencing to characterize building-dust microbiomes

Scientific Reports ◽

10.1038/s41598-020-59771-0 ◽

2020 ◽

Vol 10 (1) ◽

Cited By ~ 5

Author(s):

Anders B. Nygaard ◽

Hege S. Tunsjø ◽

Roger Meisal ◽

Colin Charnock

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Gene Sequencing ◽

16S Rrna Gene Sequencing ◽

Illumina Miseq ◽

Rrna Gene ◽

Preliminary Study ◽

Rrna Gene Sequencing ◽

Building Dust

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

Microbiome ◽

10.1186/s40168-021-01012-1 ◽

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.

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First Molecular Confirmation of Equine Ocular Setaria digitata in China

Veterinary Sciences ◽

10.3390/vetsci8040055 ◽

2021 ◽

Vol 8 (4) ◽

pp. 55

Author(s):

Feng Yu ◽

Bo Liu ◽

Shulei Chen ◽

Ziwen Yi ◽

Xianyong Liu ◽

...

Keyword(s):

Phylogenetic Analysis ◽

Light Microscopy ◽

Corneal Opacity ◽

12S Rrna ◽

Rrna Gene ◽

Asian Countries ◽

Setaria Digitata ◽

12S Rrna Gene ◽

The Right ◽

Ocular Discharge

A 5-year-old Mongolian mare (Equus caballus Linnaeus, 1758) was observed to have corneal opacity and excessive ocular discharge. An ophthalmic examination revealed a moving thread-like cylindrical worm in the anterior chamber of the right eye. The parasite was successfully removed surgically. The worm was observed under light microscopy and confirmed as Setaria digitata by 12S rRNA gene amplification and sequencing. Phylogenetic analysis demonstrated similarity with Setaria digitata in the National Center for Biotechnology Information (NCBI) GenBank database isolated from other Asian countries. This report is the first confirmed case of equine ocular setariasis by molecular diagnosis in China, which may indicate its presence in livestock and promote research on its epidemiology.

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

Bulletin of the National Research Centre ◽

10.1186/s42269-021-00528-8 ◽

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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