scholarly journals Characterization of captive and wild 13-lined ground squirrel cecal microbiotas using Illumina-based sequencing

2022 ◽  
Vol 4 (1) ◽  
Author(s):  
Edna Chiang ◽  
Courtney L. Deblois ◽  
Hannah V. Carey ◽  
Garret Suen
Keyword(s):  
16S Rrna ◽  
Beta Diversity ◽  
Current Knowledge ◽  
Winter Season ◽  
Ground Squirrels ◽  
Rrna Gene ◽  
Active Season ◽  
Operational Taxonomic Units ◽  
The 16S Rrna Gene

Abstract Background Hibernating animals experience extreme changes in diet that make them useful systems for understanding host-microbial symbioses. However, most of our current knowledge about the hibernator gut microbiota is derived from studies using captive animals. Given that there are substantial differences between captive and wild environments, conclusions drawn from studies with captive hibernators may not reflect the gut microbiota’s role in the physiology of wild animals. To address this, we used Illumina-based sequencing of the 16S rRNA gene to compare the bacterial cecal microbiotas of captive and wild 13-lined ground squirrels (TLGS) in the summer. As the first study to use Illumina-based technology to compare the microbiotas of an obligate rodent hibernator across the year, we also reported changes in captive TLGS microbiotas in summer, winter, and spring. Results Wild TLGS microbiotas had greater richness and phylogenetic diversity with less variation in beta diversity when compared to captive microbiotas. Taxa identified as core operational taxonomic units (OTUs) and found to significantly contribute to differences in beta diversity were primarily in the families Lachnospiraceae and Ruminococcaceae. Captive TLGS microbiotas shared phyla and core OTUs across the year, but active season (summer and spring) microbiotas had different alpha and beta diversities than winter season microbiotas. Conclusions This is the first study to compare the microbiotas of captive and wild rodent hibernators. Our findings suggest that data from captive and wild ground squirrels should be interpreted separately due to their distinct microbiotas. Additionally, as the first study to compare seasonal microbiotas of obligate rodent hibernators using Illumina-based 16S rRNA sequencing, we reported changes in captive TLGS microbiotas that are consistent with previous work. Taken together, this study provides foundational information for improving the reproducibility and experimental design of future hibernation microbiota studies.

scholarly journals Characterization of captive and wild 13-lined ground squirrel cecal microbiotas using Illumina-based sequencing

2021 ◽  
Author(s):  
Edna Chiang ◽  
Courtney L. Deblois ◽  
Hannah V. Carey ◽  
Garret Suen
Keyword(s):  
16S Rrna ◽  
Beta Diversity ◽  
Current Knowledge ◽  
Winter Season ◽  
Ground Squirrels ◽  
Rrna Gene ◽  
Active Season ◽  
Operational Taxonomic Units ◽  
The 16S Rrna Gene

Abstract Background Hibernating animals experience extreme changes in diet that make them useful systems for understanding host-microbial symbioses. However, most of our current knowledge about the hibernator gut microbiota is derived from studies using captive animals. Given that there are substantial differences between captive and wild environments, conclusions drawn from studies with captive hibernators may not reflect the gut microbiota’s role in the physiology of wild animals. To address this, we used Illumina-based sequencing of the 16S rRNA gene to compare the bacterial cecal microbiotas of captive and wild 13-lined ground squirrels (TLGS) in the summer. As the first study to use Illumina-based technology to sequence the microbiotas of an obligate rodent hibernator, we also reported changes in captive TLGS microbiotas across the year (summer, winter, and spring). Results Wild TLGS microbiotas had greater richness and phylogenetic diversity with less variation in beta diversity when compared to captive microbiotas. Taxa identified as core operational taxonomic units (OTUs) and found to significantly contribute to differences in beta diversity were primarily in the families Lachnospiraceae and Ruminococcaceae. Captive TLGS microbiotas shared phyla and core OTUs across the year, but active season (summer and spring) microbiotas had different alpha and beta diversities than winter season microbiotas. Conclusions This is the first study to compare the microbiotas of captive and wild rodent hibernators. Our findings suggest that data from captive and wild ground squirrels should be interpreted separately due to their distinct microbiotas. Additionally, as the first study to investigate the microbiotas of obligate rodent hibernators using Illumina-based 16S rRNA sequencing, we reported seasonal changes in captive TLGS microbiotas that are consistent with previous work. Taken together, this study provides foundational information for improving the reproducibility and experimental design of future hibernation microbiota studies.

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 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 Bacterial diversity in the saliva of patients with different oral hygiene indexes

2012 ◽  
Vol 23 (4) ◽  
pp. 409-416 ◽  
Author(s):  
Juliana Vianna Pereira ◽  
Luciana Leomil ◽  
Fabíola Rodrigues-Albuquerque ◽  
José Odair Pereira ◽  
Spartaco Astolfi-Filho
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Bacterial Diversity ◽  
Oral Hygiene ◽  
Dental Plaque ◽  
High Index ◽  
Rrna Gene ◽  
Operational Taxonomic Units ◽  
Gene Libraries ◽  
The 16S Rrna Gene

The objective of the present study was to evaluate the bacterial diversity in the saliva of patients with different oral hygiene indexes using of two 16S rRNA gene libraries. Each library was composed of samples from patients with different averages of the differentiated Silness-Löe biofilm index: the first library (A) with an index between 1.0 and 3.0 (considered a high index) and the second library (B) between 0 and 0.5 (considered a low index). Saliva DNA was extracted and the 16S rRNA gene was amplified and cloned. The obtained sequences were compared with those stored at NCBI and RDP GenBank. The saliva of patients with high index presented five known genera - Streptococcus, Granulicatella, Gemella, Veillonella and Peptostreptococcus - and 33.3% of nonculturable bacteria grouped into 23 operational taxonomic units (OTUs). The saliva of patients with low index differed significantly from the first library (p=0.000) and was composed of 42 OTUs distributed into 11 known genera - Streptococcus, Granulicatella, Gemella, Veillonella, Oribacterium, Haemophilus, Escherichia, Neisseria, Prevotella, Capnocytophaga, Actinomyces - including 24.87% of nonculturable bacteria. It was possible to conclude that there is greater bacterial diversity in the saliva of patients with low dental plaque in relation to patients with high dental plaque.

Ultrastructural and molecular characterization of endosymbionts of the reed beetle genusMacroplea(Chrysomelidae, Donaciinae), and proposal of “CandidatusMacropleicola appendiculatae” and “CandidatusMacropleicola muticae”

2009 ◽  
Vol 55 (11) ◽  
pp. 1250-1260 ◽  
Author(s):  
Gregor Kölsch ◽  
Corinna Matz-Grund ◽  
Bo V. Pedersen
Keyword(s):  
Phylogenetic Analysis ◽  
16S Rrna ◽  
Fluorescent In Situ Hybridization ◽  
Malpighian Tubules ◽  
Rrna Gene ◽  
Beetle Species ◽  
Bacterial Symbionts ◽  
The 16S Rrna Gene

Intracellular bacterial symbionts are known from various insect groups, particularly from those feeding on unbalanced diets, where the bacteria provide essential nutrients to the host. In the case of reed beetles (Coleoptera: Chrysomelidae, Donaciinae), however, the endosymbionts appear to be associated with specialized “glands” that secrete a material used for the beetles’ unusual water-tight cocoon. These glands were discovered over a century ago, but the bacteria they contain have yet to be characterized and placed in a phylogenetic context. Here, we describe the ultrastructure of two endosymbiotic species (“ Candidatus Macropleicola appendiculatae” and “ Candidatus Macropleicola muticae”) that reside in cells of the Malpighian tubules of the reed beetle species Macroplea appendiculata and Macroplea mutica , respectively. Fluorescent in situ hybridization using oligonucleotides targeting the 16S rRNA gene specific to Macroplea symbionts verified the localization of the symbionts in these organs. Phylogenetic analysis of 16S rRNA placed “Candidatus Macropleicola” in a clade of typically endosymbiotic Enterobacteriaceae (γ-proteobacteria). Finally, we discuss the evidence available for the hypothesis that the beetle larvae use a secretion produced by the bacteria for the formation of an underwater cocoon.

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 Molecular characterization of Mycobacterium intracellulare-related strains based on the sequence analysis of hsp65, internal transcribed spacer and 16S rRNA genes

2010 ◽  
Vol 59 (9) ◽  
pp. 1037-1043 ◽  
Author(s):  
Joo-Hee Park ◽  
Tae-Sun Shim ◽  
Seung-Ae Lee ◽  
Hyungki Lee ◽  
In-Kyung Lee ◽  
...  
Keyword(s):  
Sequence Analysis ◽  
16S Rrna ◽  
Internal Transcribed Spacer ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Mycobacterium Intracellulare ◽  
Epidemiological Features ◽  
The 16S Rrna Gene

We investigated the molecular epidemiological features of 94 Mycobacterium intracellulare-related strains, isolated from Korean patients, using sequence analysis targeting 3 independent chronometer molecules, hsp65, the internal transcribed spacer 1 region and the 16S rRNA gene. By collective consideration of these three gene-based approaches, the 94 strains were divided into 5 groups (INT1, INT2, INT3, INT4 and INT5). The frequencies of genotype INT1, 2, 3, 4 and 5 in the 94 isolates were 57.4 % (54), 27.7 % (26), 6.4 % (6), 5.3 % (5) and 3.2 % (3), respectively. When correlations between genotypes and clinical parameters (age, sex, radiological type and the presence of a cavity) were analysed in 78 patients with non-tuberculous mycobacteria pulmonary diseases, no relationships were observed with respect to age, sex and radiological type, but genotype and the presence of a cavity tended to be related (P=0.051).

Sign in / Sign up

Export Citation Format

Share Document