scholarly journals Using DNA-based stable isotope probing to reveal novel propionate- and acetate-oxidizing bacteria in propionate-fed mesophilic anaerobic chemostats

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Hui-Zhong Wang ◽  
Xiao-Meng Lv ◽  
Yue Yi ◽  
Dan Zheng ◽  
Min Gou ◽  
...  
Keyword(s):  
Stable Isotope ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
High Throughput Sequencing ◽  
Anaerobic Fermentation ◽  
Stable Isotope Probing ◽  
Rrna Gene ◽  
Metabolic Characteristics ◽  
Rate Limiting Step ◽  
Propionate Degradation

AbstractPropionate is one of the most important intermediates of anaerobic fermentation. Its oxidation performed by syntrophic propionate-oxidizing bacteria coupled with hydrogenotrophic methanogens is considered to be a rate-limiting step for methane production. However, the current understanding of SPOB is limited due to the difficulty of pure culture isolation. In the present study, two anaerobic chemostats fed with propionate as the sole carbon source were operated at different dilution rates (0.05 d−1 and 0.15 d−1). The propionate- and acetate-oxidizing bacteria in the two methanogenic chemostats were investigated combining DNA-stable isotope probing (DNA-SIP) and 16S rRNA gene high-throughput sequencing. The results of DNA-SIP with 13C-propionate/acetate suggested that, Smithella, Syntrophobacter, Cryptanaerobacter, and unclassified Rhodospirillaceae may be putative propionate-oxidizing bacteria; unclassified Spirochaetaceae, unclassified Synergistaceae, unclassified Elusimicrobia, Mesotoga, and Gracilibacter may contribute to acetate oxidation; unclassified Syntrophaceae and Syntrophomonas may be butyrate oxidizers. By DNA-SIP, unclassified OTUs with 16S rRNA gene abundance higher than 62% of total Bacteria in the PL chemostat and 38% in the PH chemostat were revealed to be related to the degradation of propionate. These results suggest that a variety of uncultured bacteria contribute to propionate degradation during anaerobic digestion. The functions and metabolic characteristics of these bacteria require further investigation.

scholarly journals Identification of a Toluene-Degrading Bacterium from a Soil Sample through H218O DNA Stable Isotope Probing

2011 ◽  
Vol 77 (17) ◽  
pp. 5995-5999 ◽  
Author(s):  
Angela Woods ◽  
Maribeth Watwood ◽  
Egbert Schwartz
Keyword(s):  
Stable Isotope ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Stable Isotope Probing ◽  
Rrna Gene ◽  
Soil Dna ◽  
Content Type ◽  
Degrading Bacterium ◽  
Rhodococcus Jostii ◽  
Oxygen Atoms

ABSTRACTDNA stable isotope probing (DNA-SIP) with H218O was used to identify a toluene-degrading bacterium in soil amended with 48 ppm toluene. After quantification of toluene degradation rates in soil, DNA was extracted from soil incubated with H218O, H216O, H216O and 48 ppm toluene, or H218O and 48 ppm toluene. A single DNA band formed along a cesium chloride gradient after isopycnic centrifugation of extracts from soils incubated with H216O. With extracts from soils to which only H218O was added, two distinct DNA bands formed, while three bands formed when DNA extracted from soil incubated with both H218O and toluene was analyzed. We suggest that this third band formed because toluene does not contain any oxygen atoms and toluene-degrading organisms had to transfer oxygen atoms from H218O into metabolic intermediates to form nucleic acidsde novo. We extracted the third DNA band and amplified a large fraction of the bacterial 16S rRNA gene. Direct sequencing of the PCR product obtained from the labeled DNA, as well as cloned 16S rRNA amplicons, identified a known toluene degrader,Rhodococcus jostiiRHA1. A toluene-degrading bacterial strain was subsequently isolated from soil and shown to beRhodococcus jostiiRHA1. Finally, quantitative real-time PCR analysis showed that the abundance of the 16S rRNA gene ofRhodococcus jostiiRHA1 increased in soil after toluene exposure but not in soils from which toluene was withheld. This study indicates that H218O DNA-SIP can be a useful method for identifying pollutant-degrading bacteria in soil.

scholarly journals Putative glycogen-accumulating organisms belonging to the Alphaproteobacteria identified through rRNA-based stable isotope probing

Microbiology ◽  
2006 ◽  
Vol 152 (2) ◽  
pp. 419-429 ◽  
Author(s):  
Rikke Louise Meyer ◽  
Aaron Marc Saunders ◽  
Linda Louise Blackall
Keyword(s):  
Stable Isotope ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Phosphorus Removal ◽  
Stable Isotope Probing ◽  
Metabolic Phenotype ◽  
Rrna Gene ◽  
Biological Phosphorus Removal ◽  
16S Rrna Gene Analysis

Deterioration of enhanced biological phosphorus removal (EBPR) has been linked to the proliferation of glycogen-accumulating organisms (GAOs), but few organisms possessing the GAO metabolic phenotype have been identified. An unidentified GAO was highly enriched in a laboratory-scale bioreactor and attempts to identify this organism using conventional 16S rRNA gene cloning had failed. Therefore, rRNA-based stable isotope probing followed by full-cycle rRNA analysis was used to specifically identify the putative GAOs based on their characteristic metabolic phenotype. The study obtained sequences from a group of Alphaproteobacteria not previously shown to possess the GAO phenotype, but 90 % identical by 16S rRNA gene analysis to a phylogenetic clade containing cloned sequences from putative GAOs and the isolate Defluvicoccus vanus. Fluorescence in situ hybridization (FISH) probes (DF988 and DF1020) were designed to target the new group and post-FISH chemical staining demonstrated anaerobic–aerobic cycling of polyhydroxyalkanoates, as per the GAO phenotype. The successful use of probes DF988 and DF1020 required the use of unlabelled helper probes which increased probe signal intensity up to 6·6-fold, thus highlighting the utility of helper probes in FISH. The new group constituted 33 % of all Bacteria in the lab-scale bioreactor from which they were identified and were also abundant (51 and 55 % of Bacteria) in two other similar bioreactors in which phosphorus removal had deteriorated. Unlike the previously identified Defluvicoccus-related organisms, the group identified in this study were also found in two full-scale treatment plants performing EBPR, suggesting that this group may be industrially relevant.

scholarly journals Wild specimens of sand fly phlebotomine Lutzomyia evansi, vector of leishmaniasis, show high abundance of Methylobacterium and natural carriage of Wolbachia and Cardinium types in the midgut microbiome

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Rafael J. Vivero ◽  
Marcela Villegas-Plazas ◽  
Gloria E. Cadavid-Restrepo ◽  
Claudia Ximena Moreno Herrera ◽  
Sandra I. Uribe ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
High Throughput Sequencing ◽  
Sand Fly ◽  
Human Populations ◽  
Rrna Gene ◽  
Remarkable Feature ◽  
Core Microbiome ◽  
Bacterial Phyla ◽  
Wide Range

AbstractPhlebotomine sand flies are remarkable vectors of several etiologic agents (virus, bacterial, trypanosomatid Leishmania), posing a heavy health burden for human populations mainly located at developing countries. Their intestinal microbiota is involved in a wide range of biological and physiological processes, and could exclude or facilitate such transmission of pathogens. In this study, we investigated the Eubacterial microbiome from digestive tracts of Lu. evansi adults structure using 16S rRNA gene sequence amplicon high throughput sequencing (Illumina MiSeq) obtained from digestive tracts of Lu. evansi adults. The samples were collected at two locations with high incidence of the disease in humans: peri-urban and forest ecosystems from the department of Sucre, Colombia. 289,068 quality-filtered reads of V4 region of 16S rRNA gene were obtained and clustered into 1,762 operational taxonomic units (OTUs) with 97% similarity. Regarding eubacterial diversity, 14 bacterial phyla and 2 new candidate phyla were found to be consistently associated with the gut microbiome content. Proteobacteria, Firmicutes, and Bacteroidetes were the most abundant phyla in all the samples and the core microbiome was particularly dominated by Methylobacterium genus. Methylobacterium species, are known to have mutualistic relationships with some plants and are involved in shaping the microbial community in the phyllosphere. As a remarkable feature, OTUs classified as Wolbachia spp. were found abundant on peri-urban ecosystem samples, in adult male (OTUs n = 776) and unfed female (OTUs n = 324). Furthermore, our results provide evidence of OTUs classified as Cardinium endosymbiont in relative abundance, notably higher with respect to Wolbachia. The variation in insect gut microbiota may be determined by the environment as also for the type of feeding. Our findings increase the richness of the microbiota associated with Lu. evansi. In this study, OTUs of Methylobacterium found in Lu. evansi was higher in engorged females, suggesting that there are interactions between microbes from plant sources, blood nutrients and the parasites they transmit during the blood intake.

scholarly journals Soehngenia longivitae sp. nov., a Fermenting Bacterium Isolated from a Petroleum Reservoir in Azerbaijan, and Emended Description of the Genus Soehngenia

2020 ◽  
Vol 8 (12) ◽  
pp. 1967
Author(s):  
Tamara N. Nazina ◽  
Salimat K. Bidzhieva ◽  
Denis S. Grouzdev ◽  
Diyana S. Sokolova ◽  
Tatyana P. Tourova ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
High Throughput Sequencing ◽  
Biochemical Characterization ◽  
Sequence Similarity ◽  
Phylogenomic Analysis ◽  
Rrna Gene ◽  
Petroleum Reservoir ◽  
Ph Range ◽  
The 16S Rrna Gene

A methanogenic enrichment growing on a medium with methanol was obtained from a petroleum reservoir (Republic of Azerbaijan) and stored for 33 years without transfers to fresh medium. High-throughput sequencing of the V4 region of the 16S rRNA gene revealed members of the genera Desulfovibrio, Soehngenia, Thermovirga, Petrimonas, Methanosarcina, and Methanomethylovorans. A novel gram-positive, rod-shaped, anaerobic fermentative bacterium, strain 1933PT, was isolated from this enrichment and characterized. The strain grew at 13–55 °C (optimum 35 °C), with 0–3.0% (w/v) NaCl (optimum 0–2.0%) and in the pH range of 6.7–8.0 (optimum pH 7.0). The 16S rRNA gene sequence similarity, the average nucleotide identity (ANI) and in silico DNA–DNA hybridization (dDDH) values between strain 1933PT and the type strain of the most closely related species Soehngenia saccharolytica DSM 12858T were 98.5%, 70.5%, and 22.6%, respectively, and were below the threshold accepted for species demarcation. Genome-based phylogenomic analysis and physiological and biochemical characterization of the strain 1933PT (VKM B-3382T = KCTC 15984T) confirmed its affiliation to a novel species of the genus Soehngenia, for which the name Soehngenia longivitae sp. nov. is proposed. Genome analysis suggests that the new strain has potential in the degradation of proteinaceous components.

scholarly journals Toolbox Approaches Using Molecular Markers and 16S rRNA Gene Amplicon Data Sets for Identification of Fecal Pollution in Surface Water

2015 ◽  
Vol 81 (20) ◽  
pp. 7067-7077 ◽  
Author(s):  
W. Ahmed ◽  
C. Staley ◽  
M. J. Sadowsky ◽  
P. Gyawali ◽  
J. P. S. Sidhu ◽  
...  
Keyword(s):  
Molecular Markers ◽  
Bacterial Community ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Water Samples ◽  
High Throughput Sequencing ◽  
Community Analysis ◽  
Fecal Pollution ◽  
Rrna Gene ◽  
Potential Sources

ABSTRACTIn this study, host-associated molecular markers and bacterial 16S rRNA gene community analysis using high-throughput sequencing were used to identify the sources of fecal pollution in environmental waters in Brisbane, Australia. A total of 92 fecal and composite wastewater samples were collected from different host groups (cat, cattle, dog, horse, human, and kangaroo), and 18 water samples were collected from six sites (BR1 to BR6) along the Brisbane River in Queensland, Australia. Bacterial communities in the fecal, wastewater, and river water samples were sequenced. Water samples were also tested for the presence of bird-associated (GFD), cattle-associated (CowM3), horse-associated, and human-associated (HF183) molecular markers, to provide multiple lines of evidence regarding the possible presence of fecal pollution associated with specific hosts. Among the 18 water samples tested, 83%, 33%, 17%, and 17% were real-time PCR positive for the GFD, HF183, CowM3, and horse markers, respectively. Among the potential sources of fecal pollution in water samples from the river, DNA sequencing tended to show relatively small contributions from wastewater treatment plants (up to 13% of sequence reads). Contributions from other animal sources were rarely detected and were very small (<3% of sequence reads). Source contributions determined via sequence analysis versus detection of molecular markers showed variable agreement. A lack of relationships among fecal indicator bacteria, host-associated molecular markers, and 16S rRNA gene community analysis data was also observed. Nonetheless, we show that bacterial community and host-associated molecular marker analyses can be combined to identify potential sources of fecal pollution in an urban river. This study is a proof of concept, and based on the results, we recommend using bacterial community analysis (where possible) along with PCR detection or quantification of host-associated molecular markers to provide information on the sources of fecal pollution in waterways.

scholarly journals Bacterial Diversity and Community in Regional Water Microbiota between Different Towns in World’s Longevity Township Jiaoling, China

Diversity ◽  
2021 ◽  
Vol 13 (8) ◽  
pp. 361
Author(s):  
Lei Wu ◽  
Xinqiang Xie ◽  
Jumei Zhang ◽  
Yu Ding ◽  
Qingping Wu
Keyword(s):  
16S Rrna ◽  
High Throughput Sequencing ◽  
The Other ◽  
Rrna Gene ◽  
Metabolic Characteristics ◽  
Kegg Pathways ◽  
Healthy Longevity ◽  
And Function ◽  
Regional Water

Healthy longevity is associated with many factors, however, the potential correlation between longevity and microbiota remains elusive. To address this, we explored environmental microbiota from one of the world’s longevity townships in China. We used 16S rRNA gene high-throughput sequencing to analyze the composition and function of water microbiota. The composition and diversity of water microbiota significantly differed between the towns. Lactobacillus, Streptococcus, Bacteroides, Faecalibacterium, and Stenotrophomonas were only dominant in Xinpu, a town with an exceptionally high centenarian population. Several biomarkers were identified, including Flavobacterium, Acinetobacter, Paracoccus, Lactobacillales, Psychrobacter, Bacteroides, Ruminococcaceae, and Faecalibacterium, and these shown to be responsible for the significant differences between towns. The main species contributing to the differences between towns were Cyanobacteria, Cupriavidus and Ralstonia. Based on KEGG pathways showed that the predicted metabolic characteristics of the water microbiota in Xinpu towns were significantly different to those of the other towns. The results revealed significant differences in the composition and diversity of water microbiota in the longevity township. These findings provide a foundation for further research on the role of water microbiota in healthy longevity.

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