scholarly journals Characterization of polybacterial clinical samples using a set of group-specific broad-range primers targeting the 16S rRNA gene followed by DNA sequencing and RipSeq analysis

2011 ◽  
Vol 60 (7) ◽  
pp. 927-936 ◽  
Author(s):  
Øyvind Kommedal ◽  
Katrine Lekang ◽  
Nina Langeland ◽  
Harald G. Wiker
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Dna Sequencing ◽  
Clinical Samples ◽  
Rrna Gene ◽  
The 16S Rrna Gene

scholarly journals Molecular Characterization of anEndozoicomonas-Like Organism Causing Infection in the King Scallop (Pecten maximusL.)

2017 ◽  
Vol 84 (3) ◽  
Author(s):  
Irene Cano ◽  
Ronny van Aerle ◽  
Stuart Ross ◽  
David W. Verner-Jeffreys ◽  
Richard K. Paley ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Molecular Characterization ◽  
Mass Mortality ◽  
Rrna Gene ◽  
Gene Sequences ◽  
16S Rrna Gene Sequences ◽  
Content Type ◽  
The 16S Rrna Gene

ABSTRACTOne of the fastest growing fisheries in the UK is the king scallop (Pecten maximusL.), also currently rated as the second most valuable fishery. Mass mortality events in scallops have been reported worldwide, often with the causative agent(s) remaining uncharacterized. In May 2013 and 2014, two mass mortality events affecting king scallops were recorded in the Lyme Bay marine protected area (MPA) in Southwest England. Histopathological examination showed gill epithelial tissues infected with intracellular microcolonies (IMCs) of bacteria resemblingRickettsia-like organisms (RLOs), often with bacteria released in vascular spaces. Large colonies were associated with cellular and tissue disruption of the gills. Ultrastructural examination confirmed the intracellular location of these organisms in affected epithelial cells. The 16S rRNA gene sequences of the putative IMCs obtained from infected king scallop gill samples, collected from both mortality events, were identical and had a 99.4% identity to 16S rRNA gene sequences obtained from “CandidatusEndonucleobacter bathymodioli” and 95% withEndozoicomonasspecies.In situhybridization assays using 16S rRNA gene probes confirmed the presence of the sequenced IMC gene in the gill tissues. Additional DNA sequences of the bacterium were obtained using high-throughput (Illumina) sequencing, and bioinformatic analysis identified over 1,000 genes with high similarity to protein sequences fromEndozoicomonasspp. (ranging from 77 to 87% identity). Specific PCR assays were developed and applied to screen for the presence of IMC 16S rRNA gene sequences in king scallop gill tissues collected at the Lyme Bay MPA during 2015 and 2016. There was 100% prevalence of the IMCs in these gill tissues, and the 16S rRNA gene sequences identified were identical to the sequence found during the previous mortality event.IMPORTANCEMolluscan mass mortalities associated with IMCs have been reported worldwide for many years; however, apart from histological and ultrastructural characterization, characterization of the etiological agents is limited. In the present work, we provide detailed molecular characterization of anEndozoicomonas-like organism (ELO) associated with an important commercial scallop species.

scholarly journals Characterization of a protein binding sequence in the promoter region of the 16S rRNA gene of the spinach chloroplast genome

1991 ◽  
Vol 19 (13) ◽  
pp. 3577-3581 ◽  
Author(s):  
Laurence Baeza ◽  
Alain Bertrand ◽  
Régis Mache ◽  
Silva Lerbs-Mache
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Protein Binding ◽  
Promoter Region ◽  
Rrna Gene ◽  
Spinach Chloroplast ◽  
Protein Binding Sequence ◽  
Binding Sequence ◽  
The 16S Rrna Gene

scholarly journals MOLECULAR CHARACTERIZATION OF GENE 16S rRNA MICRO SYMBIONTS IN SPONGE AT MELAWAI BEACH, EAST KALIMANTAN

2018 ◽  
Author(s):  
Ismail Marzuki ◽  
Alfian Noor ◽  
Nursiah La Nafie
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Molecular Characterization ◽  
Morphological Identification ◽  
Rrna Gene ◽  
Isolation And Purification ◽  
East Kalimantan ◽  
Degree Of Similarity ◽  
The 16S Rrna Gene

Molecular characterization studies have been conducted 16S rRNA gene micro symbiont of sponge origin Melawai Beach, Balikpapan in East Kalimantan. Objective analysis of histo- morphological research, isolation-purification, molecular characterization of micro-symbiont genes in order to search symbiont bacteria that can live in extreme environments contaminated hydrocarbon waste. The research method that morphological identification, isolation-purification and molecular characterization of the 16S rRNA gene with Chain Reaction Polymerization method. The results of histo-morphological analysis concluded sponge samples with species of Callyspongia sp. Isolation and purification mikro symbionts of sponge obtained 2 (two) isolates. Characteristics of Isolates 1; spherical shape, colonize and creamy, while isolates 2; jagged shape, oval and white colonies. Molecular characterization of the 16S rRNA gene by PCR, Bacillus subtilis strain BAB-684 identification for isolates one is the number of nucleotide pairs reached 899 bp and the degree of similarity in GenBank reached 89% homologous, while the second is a Bacillus flexus strain PHCDB20 isolates the number reached 950 bp nucleotide pairs with the degree of similarity in GenBank reached 99% homologous

scholarly journals Astacopsidrilus hibernicus sp. nov. (Phreodrilidae, Oligochaeta, Annelida) from Irish peatlands

Zoosymposia ◽  
2020 ◽  
Vol 17 (1) ◽  
pp. 34-44
Author(s):  
RÜDIGER M. SCHMELZ ◽  
MÅRTEN J. KLINTH ◽  
RACHEL WISDOM ◽  
THOMAS BOLGER
Keyword(s):  
United Kingdom ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Dna Sequencing ◽  
Dorsal Side ◽  
Rrna Gene ◽  
The United Kingdom ◽  
The 16S Rrna Gene ◽  
Epidermal Gland ◽  
Whole Mounts

The discovery of a large and flourishing population of Phreodrilidae in terrestrial peatlands in northwest Ireland was surprising on two counts: these oligochaete worms are usually aquatic and most of the species occur in the Southern Hemisphere. The phreodrilids were discovered in a project that targeted Enchytraeidae, therefore methods adapted to the investigation of enchytraeids could be applied, including the study of living animals and properly fixed whole mounts. DNA sequencing was also performed. All worms identified here belong to one species, new to science, and placed in the genus Astacopsidrilus, because of the ventral position of the spermathecal pores and the opening of the female funnels inside the spermathecal vestibule. Astacopsidrilus hibernicus sp. nov. is mainly distinguished by thick segmental cushions of epidermal gland cells on the dorsal side of the posterior body half. Male sexual organs and spermathecae are comparatively small and without the often-observed bizarre modifications common in species of this family. DNA sequencing yielded a fragment of the 16S rRNA gene. This is the first description of a phreodrilid species from Europe; the few previous recordings of this family in Ireland and the United Kingdom had been left unidentified.

scholarly journals DNA sequencing reveals limited heterogeneity in the 16S rRNA gene from the rrnB operon among five Mycoplasma hominis isolates

1998 ◽  
Vol 48 (3) ◽  
pp. 1067-1071 ◽  
Author(s):  
T. Mygind ◽  
S. Birkelund ◽  
G. Christiansen
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Dna Sequencing ◽  
Mycoplasma Hominis ◽  
Rrna Gene ◽  
The 16S Rrna Gene

scholarly journals Ochrobactrum pseudintermedium sp. nov., a novel member of the family Brucellaceae, isolated from human clinical samples

2007 ◽  
Vol 57 (5) ◽  
pp. 1007-1013 ◽  
Author(s):  
Corinne Teyssier ◽  
Hélène Marchandin ◽  
Hélène Jean-Pierre ◽  
Agnès Masnou ◽  
Ghislaine Dusart ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Gene Sequence ◽  
Sequence Similarity ◽  
Clinical Samples ◽  
Rrna Gene ◽  
The Family ◽  
Ochrobactrum Intermedium ◽  
Physiological And Biochemical ◽  
The 16S Rrna Gene

Three novel Gram-negative, non-fermenting aerobic bacilli were isolated from human clinical samples. They shared more than 99.8 % of the 16S rRNA gene nucleotide positions. The strains were related to Ochrobactrum intermedium with about 97.48 % 16S rRNA gene sequence similarity. In 16S rRNA gene-, dnaK- and rpoB-based phylogenies, the strains were grouped in a lineage that was distinct from other Ochrobactrum species in the family Brucellaceae. Fatty acid composition, polar lipids, quinone system, DNA–DNA relatedness, genome organization, and physiological and biochemical data differentiated these isolates from recognized species of the genus Ochrobactrum. The three clinical strains therefore represent a novel species within the genus Ochrobactrum, for which the name Ochrobactrum pseudintermedium sp. nov., is proposed. The type strain is ADV31T (=CIP 109116T=DSM 17490T). The DNA G+C content of strain ADV31T was 54.5 mol%.

scholarly journals Critical Relevance of Stochastic Effects on Low-Bacterial-Biomass 16S rRNA Gene Analysis

mBio ◽  
2020 ◽  
Vol 11 (3) ◽  
Author(s):  
John R. Erb-Downward ◽  
Nicole R. Falkowski ◽  
Jennifer C. D’Souza ◽  
Lisa M. McCloskey ◽  
Roderick A. McDonald ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Bacterial Biomass ◽  
Critical Examination ◽  
Rrna Gene ◽  
Stochastic Noise ◽  
Oral Rinse ◽  
Healthy Humans ◽  
The 16S Rrna Gene

ABSTRACT The bacterial microbiome of human body sites, previously considered sterile, remains highly controversial because it can be challenging to isolate signal from noise when low-biomass samples are being analyzed. We tested the hypothesis that stochastic sequencing noise, separable from reagent contamination, is generated during sequencing on the Illumina MiSeq platform when DNA input is below a critical threshold. We first purified DNA from serial dilutions of Pseudomonas aeruginosa and from negative controls using three DNA purification kits, quantified input using droplet digital PCR, and then sequenced the 16S rRNA gene in four technical replicates. This process identified reproducible contaminant signal that was separable from an irreproducible stochastic noise, which occurred as bacterial biomass of samples decreased. This approach was then applied to authentic respiratory samples from healthy individuals (n = 22) that ranged from high to ultralow bacterial biomass. Using oral rinse, bronchoalveolar lavage (BAL) fluid, and exhaled breath condensate (EBC) samples and matched controls, we were able to demonstrate (i) that stochastic noise dominates sequencing in real-world low-bacterial-biomass samples that contain fewer than 104 copies of the 16S rRNA gene per sample, (ii) that critical examination of the community composition of technical replicates can be used to separate signal from noise, and (iii) that EBC is an irreproducible sampling modality for sampling the microbiome of the lower airways. We anticipate that these results combined with suggested methods for identifying and dealing with noisy communities will facilitate increased reproducibility while simultaneously permitting characterization of potentially important low-biomass communities. IMPORTANCE DNA contamination from external sources (reagents, environment, operator, etc.) has long been assumed to be the main cause of spurious signals that appear under low-bacterial-biomass conditions. Here, we demonstrate that contamination can be separated from another, random signal generated during low-biomass-sample sequencing. This stochastic noise is not reproduced between technical replicates; however, results for any one replicate taken alone could look like a microbial community different from the controls. Using this information, we investigated respiratory samples from healthy humans and determined the narrow range of bacterial biomass where samples transition from producing reproducible microbial sequences to ones dominated by noise. We present a rigorous approach to studies involving low-bacterial-biomass samples to detect this source of noise and provide a framework for deciding if a sample is likely to be dominated by noise. We anticipate that this work will facilitate increased reproducibility in the characterization of potentially important low-biomass communities.

scholarly journals Negativicoccus succinicivorans gen. nov., sp. nov., isolated from human clinical samples, emended description of the family Veillonellaceae and description of Negativicutes classis nov., Selenomonadales ord. nov. and Acidaminococcaceae fam. nov. in the bacterial phylum Firmicutes

2010 ◽  
Vol 60 (6) ◽  
pp. 1271-1279 ◽  
Author(s):  
Hélène Marchandin ◽  
Corinne Teyssier ◽  
Josiane Campos ◽  
Hélène Jean-Pierre ◽  
Frédéric Roger ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Clinical Samples ◽  
Rrna Gene ◽  
The Novel ◽  
Gene Sequences ◽  
16S Rrna Gene Sequences ◽  
Gram Negative ◽  
The Family ◽  
The 16S Rrna Gene

Three strains of a hitherto unknown, Gram-negative, tiny, anaerobic coccus were collected from human clinical samples originating from skin and soft tissues. The three isolates displayed at least 99.9 % identity in their 16S rRNA gene sequences and more than 99.8 % identity in their dnaK gene sequences. The isolates were affiliated to the family Veillonellaceae, the coccobacillus Dialister micraerophilus being the most closely related species, but there was no more than 91.1 % identity in the 16S rRNA gene sequence between this species and the three isolates. Phylogeny based on the 16S rRNA gene confirmed that the three strains represent a novel and robust lineage within the current family Veillonellaceae. A similar genomic structure was demonstrated for the three isolates by PFGE-based analysis. Morphology and metabolic end products, as well as genotypic and phylogenetic data supported the proposal of the novel genus Negativicoccus gen. nov., with the novel species Negativicoccus succinicivorans sp. nov. [type strain ADV 07/08/06-B-1388T (=AIP 149.07T=CIP 109806T=DSM 21255T=CCUG 56017T) as type species]. Phylogenetic analyses based on the 16S rRNA gene sequences of members of the phylum Firmicutes and other phyla indicated that the family Veillonellaceae forms a robust lineage clearly separated from those of the classes ‘Bacilli’, ‘Clostridia’, Thermolithobacteria and ‘Erysipelotrichi’ in the phylum Firmicutes. Therefore, we propose that this family is a class-level taxon in the phylum Firmicutes, for which the name Negativicutes classis nov. is proposed, based on the Gram-negative type of cell wall of its members, with the type order Selenomonadales ord. nov. In this order, a novel family, Acidaminococcaceae fam. nov., is proposed and description of the family Veillonellaceae is emended.

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