Journal articles: '16S rDNA profiling'

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Gu, F., Y. Li, C. Zhou, D. T. W. Wong, C. M. Ho, F. Qi, and W. Shi. "Bacterial 16S rRNA/rDNA Profiling in the Liquid Phase of Human Saliva." Open Dentistry Journal 3, no. 1 (April 28, 2009): 80–84. http://dx.doi.org/10.2174/1874210600903010080.

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Human saliva can be separated by centrifugation into cell pellet and cell-free supernatant, which are called cellular phase and liquid phase in this study. While it is well documented that the cellular phase of saliva contains hundreds of oral bacteria species, little is known whether the liquid phase of saliva contains any information related to oral microbiota. In this study, we analyzed the bacterial nucleic acid contents of the liquid phase of saliva. Using primers universal to most eubacterial 16S rDNA, we detected large amounts of bacterial 16S rRNA and rDNA in the cell-free phase of saliva. Random sequencing analysis of forty PCR amplicons from the cell-free phase of saliva led to 15 operational taxonomic unit (OTU) groups. Furthermore, using denaturing gradient gel electrophoresis (DGGE), we compared 16S rRNA/rDNA profiles derived from liquid phases and cellular phases of saliva samples, and found positive correlations (Pearson Correlation=0.822,P<0.001) between these sample groups. These findings indicate that the liquid phase of saliva contains numerous bacterial 16S rRNA/rDNA molecules that have correlations with bacteria existing in the cellular phase.

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Wood, Jacqueline, Karen P. Scott, Gorazd Avguštin, C. James Newbold, and Harry J. Flint. "Estimation of the Relative Abundance of DifferentBacteroides and Prevotella Ribotypes in Gut Samples by Restriction Enzyme Profiling of PCR-Amplified 16S rRNA Gene Sequences." Applied and Environmental Microbiology 64, no. 10 (October 1, 1998): 3683–89. http://dx.doi.org/10.1128/aem.64.10.3683-3689.1998.

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ABSTRACT We describe an approach for determining the genetic composition ofBacteroides and Prevotellapopulations in gut contents based on selective amplification of 16S rRNA gene sequences (rDNA) followed by cleavage of the amplified material with restriction enzymes. The relative contributions of different ribotypes to total Bacteroides andPrevotella 16S rDNA are estimated after end labelling of one of the PCR primers, and the contribution ofBacteroides and Prevotellasequences to total eubacterial 16S rDNA is estimated by measuring the binding of oligonucleotide probes to amplified DNA.Bacteroides and Prevotella 16S rDNA accounted for between 12 and 62% of total eubacterial 16S rDNA in samples of ruminal contents from six sheep and a cow. Ribotypes 4, 5, 6, and 7, which include most cultivated rumen Prevotellastrains, together accounted for between 20 and 86% of the total amplified Bacteroides andPrevotella rDNA in these samples. The most abundantBacteroides or Prevotella ribotype in four animals, however, was ribotype 8, for which there is only one known cultured isolate, while ribotypes 1 and 2, which include many colonic Bacteroides spp., were the most abundant in two animals. This indicates that some abundantBacteroides and Prevotella groups in the rumen are underrepresented among cultured rumenPrevotella isolates. The approach described here provides a rapid, convenient, and widely applicable method for comparing the genotypic composition of bacterial populations in gut samples.

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Galovic, Vladislava, Milan Drekic, Sreten Vasic, Sinisa Andrasev, Sasa Pekec, Dejan Stojanovic, and Verica Vasic. "Mitochondrial 16s rDNA profiling and phylogenetic analysis suggest genetic diversity of ash weevil (Stereonichus fraxini De Geer) in Serbia." Genetika 51, no. 2 (2019): 675–86. http://dx.doi.org/10.2298/gensr1902675g.

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This study contributes to knowledge of ash weevil (Stereonychus fraxini De Geer) molecular taxonomy, phylogeny and genetic diversity. Adult and larvae stages of insect were collected from several locations covering northern and central part of Serbia cojoined with homologous sequences with respect to their different geographic origin and hypothesis of their evolutionary relationships. Due to its slow rates of evolution the gene region that covers mitochondrial 16S rDNA, was choice for sequence profiling and phylogenetic reconstruction of ash weevil in correspondence with sequences of related tribes Cionini. Phylogenetic analyses demonstrating clear separation of the native weevil populations and the Cionini tribes. Even though bioinformatic tools confirm that all native specimens belong to species Stereonychus fraxini, different profile of the mitochondrial 16S rDNA in the clade of Serbian specimens indicate intraspecific genomic rearrangement in one specimen detached it to northern geographic position. Those particular specimens invade also different Fraxinus species. Genetic distinctness of other imagos from this particular individual proved by indels and point mutations found in their sequences. By screening the mitochondrial 16S rDNA, molecular evidence suggests the existence of the specimen with rearranged genome that indicate genetic variability in native populations of ash weevil species.

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Adesetan, Titilayo O., Moses O. Efuntoye, and Olubukola O. Babalola. "Genotypic Profiling of Bacillus cereus Recovered from Some Retail Foods in Ogun State, Nigeria, and Their Phylogenetic Relationship." International Journal of Microbiology 2020 (September 14, 2020): 1–9. http://dx.doi.org/10.1155/2020/3750948.

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Identifying Bacillus cereus with conventional methods is neither specific nor rapid because of the close relatedness of the B. cereus group, hence the need for molecular methods. Genotypic profiling of B. cereus isolates from food was obtained by Random Amplified Polymorphic DNA-polymerase chain reaction (RAPD-PCR) using OPR13 primer. A dendrogram was drawn with the Numerical Taxonomy System of Statistic (NTSYS) software. Thirty of the isolates were subjected to molecular identification by 16S rDNA sequencing. The thirty sequences were deposited in GenBank for accession number. Phylogenetic relationship of the 16S rDNA sequence obtained was carried out with the Multiple Alignment using Fast Fourier Transform (MAFFT) software version 7.0. The evolutionary tree was drawn using the Molecular Evolutionary Genetics Analysis (MEGA 6) software. The dendrogram generated for the RAPD profile showed that all the strains are closely related, with a similarity coefficient of 70%. The isolates were confirmed with 16S rDNA sequencing as B. cereus. The thirty sequences deposited in GenBank were given accession numbers: KX574760–KX574769, KX610811–KX610820, MT757957-MT757963, and MT772282-MT772284. By comparing the phylogenetic relationship, eleven of the strains did not cluster with the reference strains from the GenBank but form distinct clades, which means they are likely to be of different ancestors. Conventional methods rarely differentiate bacteria of the same species into clade, neither can it describe their ancestral lineage. Therefore, it is important to employ molecular methods in identifying bacteria to give detailed information about them.

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Raju, Sajan C., Sonja Lagström, Pekka Ellonen, Willem M. de Vos, Johan G. Eriksson, Elisabete Weiderpass, and Trine B. Rounge. "Reproducibility and repeatability of six high-throughput 16S rDNA sequencing protocols for microbiota profiling." Journal of Microbiological Methods 147 (April 2018): 76–86. http://dx.doi.org/10.1016/j.mimet.2018.03.003.

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Dickinson, Danielle N., Myron T. La Duc, William E. Haskins, Igor Gornushkin, James D. Winefordner, David H. Powell, and Kasthuri Venkateswaran. "Species Differentiation of a Diverse Suite of Bacillus Spores by Mass Spectrometry-Based Protein Profiling." Applied and Environmental Microbiology 70, no. 1 (January 2004): 475–82. http://dx.doi.org/10.1128/aem.70.1.475-482.2004.

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ABSTRACT In this study, we demonstrate the versatility of matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOFMS) protein profiling for the species differentiation of a diverse suite of Bacillus spores. MALDI-TOFMS protein profiles of 11 different strains of Bacillus spores, encompassing nine different species, were evaluated. Bacillus species selected for MALDI-TOFMS analysis represented the spore-forming bacterial diversity of typical class 100K clean room spacecraft assembly facilities. A one-step sample treatment and MALDI-TOFMS preparation were used to minimize the sample preparation time. A library of MALDI-TOFMS spectra was created from these nine Bacillus species, the most diverse protein profiling study of the genus reported to date. Linear correlation analysis was used to successfully differentiate the MALDI-TOFMS protein profiles from all strains evaluated in this study. The MALDI-TOFMS protein profiles were compared with 16S rDNA sequences for their bacterial systematics and molecular phylogenetic affiliations. The MALDI-TOFMS profiles were found to be complementary to the 16S rDNA analysis. Proteomic studies of Bacillus subtilis 168 were pursued to identify proteins represented by the biomarker peaks in the MALDI-TOFMS spectrum. Four small, acid-soluble proteins (A, B, C, and D), one DNA binding protein, hypothetical protein ymf J, and four proteins associated with the spore coat and spore coat formation (coat JB, coat F, coat T, and spoIVA) were identified. The ability to visualize higher-molecular-mass coat proteins (10 to 25 kDa) as well as smaller proteins (<10 kDa) with MALDI-TOFMS profiling is critical for the complete and effective species differentiation of the Bacillus genus.

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Rogers, G. B., C. A. Hart, J. R. Mason, M. Hughes, M. J. Walshaw, and K. D. Bruce. "Bacterial Diversity in Cases of Lung Infection in Cystic Fibrosis Patients: 16S Ribosomal DNA (rDNA) Length Heterogeneity PCR and 16S rDNA Terminal Restriction Fragment Length Polymorphism Profiling." Journal of Clinical Microbiology 41, no. 8 (August 1, 2003): 3548–58. http://dx.doi.org/10.1128/jcm.41.8.3548-3558.2003.

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Apajalahti, Juha H. A., Anu Kettunen, Michael R. Bedford, and William E. Holben. "Percent G+C Profiling Accurately Reveals Diet-Related Differences in the Gastrointestinal Microbial Community of Broiler Chickens." Applied and Environmental Microbiology 67, no. 12 (December 1, 2001): 5656–67. http://dx.doi.org/10.1128/aem.67.12.5656-5667.2001.

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ABSTRACT Broiler chickens from eight commercial farms in Southern Finland were analyzed for the structure of their gastrointestinal microbial community by a nonselective DNA-based method, percent G+C-based profiling. The bacteriological impact of the feed source and in-farm whole-wheat amendment of the diet was assessed by percent G+C profiling. Also, a phylogenetic 16S rRNA gene (rDNA)-based study was carried out to aid in interpretation of the percent G+C profiles. This survey showed that most of the 16S rDNA sequences found could not be assigned to any previously known bacterial genus or they represented an unknown species of one of the taxonomically heterogeneous genera, such as Ruminococcus or Clostridium. The data from bacterial community profiling were analyzed byt-test, multiple linear regression, and principal-component statistical approaches. The percent G+C profiling method with appropriate statistical analyses detected microbial community differences smaller than 10% within each 5% increment of the percent G+C profiles. Diet turned out to be the strongest determinant of the cecal bacterial community structure. Both the source of feed and local feed amendment changed the bacteriological profile significantly, whereas profiles of individual farms with identical feed regimens hardly differed from each other. This suggests that the management of typical Finnish farms is relatively uniform or that hygiene on the farm, in fact, has little impact on the structure of the cecal bacterial community. Therefore, feed compounders should have a significant role in the modulation of gut microflora and consequently in prevention of gastrointestinal disorders in farm animals.

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Wood, J., KP Scott, G. Avguštin, CJ Newbold, F. McIntosh, and HJ Flint. "A 16S rDNA-based molecular profiling approach for studying relative changes in ruminal bacterial populations." Reproduction Nutrition Development 37, Suppl. 1 (1997): 30–31. http://dx.doi.org/10.1051/rnd:19970710.

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Scheldeman, Patsy, Marina Rodríguez-Díaz, Johan Goris, Annelies Pil, Elke De Clerck, Lieve Herman, Paul De Vos, Niall A. Logan, and Marc Heyndrickx. "Bacillus farraginis sp. nov., Bacillus fortis sp. nov. and Bacillus fordii sp. nov., isolated at dairy farms." International Journal of Systematic and Evolutionary Microbiology 54, no. 4 (July 1, 2004): 1355–64. http://dx.doi.org/10.1099/ijs.0.63095-0.

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Forty-eight bacterial strains were isolated at dairy farms from raw milk, the milking apparatus, green fodder or feed concentrate after a heat treatment of 30 min at 100 °C. In this way, spore-forming bacteria with a very high intrinsic heat resistance were selected for. The aerobic spore-forming isolates were subjected to a polyphasic taxonomical study, including repetitive element sequence-based PCR typing, whole-cell protein profiling, 16S rDNA sequence analysis, DNA–DNA hybridizations, DNA base composition, fatty acid analysis, and morphological and biochemical characteristics. A comparison of the REP- and (GTG)5-PCR and whole-cell protein SDS-PAGE profiles resulted in three clusters of similar strains. Analysis of the 16S rDNA sequences and DNA–DNA relatedness data showed that these clusters represented three novel species. The highest 16S rDNA similarity to a recognized species found for the three groups was around 94 % with Bacillus lentus and Bacillus sporothermodurans. Further phenotypic characterization supported the proposal of three novel species in the genus Bacillus, Bacillus farraginis, Bacillus fortis and Bacillus fordii. The respective type strains are R-6540T (=LMG 22081T=DSM 16013T), R-6514T (=LMG 22079T=DSM 16012T) and R-7190T (=LMG 22080T=DSM 16014T); their G+C DNA base contents are 43·7, 44·3 and 41·9 mol%, respectively. Although in variable amounts, a predominance of the branched fatty acids iso-C15 : 0 and anteiso-C15 : 0 was observed in all three novel species.

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GUNAY-ESIYOK, OZLEM, NEFISE AKCELIK, and MUSTAFA AKCELIK. "Identification of Genomic Heterogeneity among Lactococcus lactis Strains by Plasmid Profiling, PFGE and 16S rDNA Sequence Analysis." Polish Journal of Microbiology 63, no. 2 (2014): 157–66. http://dx.doi.org/10.33073/pjm-2014-021.

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Lactococcus lactis strains are used commonly as starters, which contribute to desirable flavour and texture properties known as strain-specific, in dairy industry. Genomic heterogeneity of 30 L. lactis strains originating from Turkey and characterized phenotypically were investigated in this study. Plasmid profiling, PFGE and 16S rDNA sequence analyses were performed to determine the genetic variability of strains. High degree of heterogeneity was detected among the L. lactis strains. Plasmid profiles of strains showed that compared to the plasmid free control strains, namely; L. lactis subsp. lactis IL1403 and L. lactis subsp. cremoris MG1614, all tested strains carried one to ten plasmids with molecular size ranging from 1.5 to 41.5kb. The fingerprints of strains obtained by PFGE from digestion with ApaI, SmaI and I-CeuI restriction endonucleases of chromosomal DNA's were compared with each other. All strains out of four were grouped into a large cluster A with at least 44% similarity level. The other four strains formed a minor duster B, distinctively different from major cluster A. PFGE results were confirmed by 16S rDNA sequence analysis and strains included in cluster B were identified as members of different species. These results suggested that morphologic and biochemical methods should be verified by reliable molecular approaches for the purpose of strain typing. Also, PFGE was found suitable to determine genomic differentiations among inter- and intra species.

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Khan, Shams Tabrez, Yoko Horiba, Masamitsu Yamamoto, and Akira Hiraishi. "Members of the Family Comamonadaceae as Primary Poly(3-Hydroxybutyrate-co-3-Hydroxyvalerate)-Degrading Denitrifiers in Activated Sludge as Revealed by a Polyphasic Approach." Applied and Environmental Microbiology 68, no. 7 (July 2002): 3206–14. http://dx.doi.org/10.1128/aem.68.7.3206-3214.2002.

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ABSTRACT The distribution and phylogenetic affiliations of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV)-degrading denitrifying bacteria in activated sludge were studied by a polyphasic approach including culture-independent biomarker and molecular analyses as well as cultivation methods. A total of 23 strains of PHBV-degrading denitrifiers were isolated from activated sludges from different sewage treatment plants. 16S ribosomal DNA (rDNA) sequence comparisons showed that 20 of the isolates were identified as members of the family Comamonadaceae, a major group of β-Proteobacteria. When the sludges from different plants were acclimated with PHBV under denitrifying conditions in laboratory scale reactors, the nitrate removal rate increased linearly during the first 4 weeks and reached 20 mg NO3 −-N h−1 g of dry sludge−1 at the steady state. The bacterial-community change in the laboratory scale sludges during the acclimation was monitored by rRNA-targeted fluorescence in situ hybridization and quinone profiling. Both approaches showed that the population of β-Proteobacteria in the laboratory sludges increased sharply during acclimation regardless of their origins. 16S rDNA clone libraries were constructed from two different acclimated sludges, and a total of 37 clones from the libraries were phylogenetically analyzed. Most of the 16S rDNA clones were grouped with members of the family Comamonadaceae. The results of our polyphasic approach indicate that β-Proteobacteria, especially members of the family Comamonadaceae, are primary PHBV-degrading denitrifiers in activated sludge. Our data provide useful information for the development of a new nitrogen removal system with solid biopolymer as an electron donor.

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Sliwinski, Marek K., and Robert M. Goodman. "Spatial Heterogeneity of Crenarchaeal Assemblages within Mesophilic Soil Ecosystems as Revealed by PCR-Single-Stranded Conformation Polymorphism Profiling." Applied and Environmental Microbiology 70, no. 3 (March 2004): 1811–20. http://dx.doi.org/10.1128/aem.70.3.1811-1820.2004.

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ABSTRACT Microbial ecologists have discovered novel rRNA genes (rDNA) in mesophilic soil habitats worldwide, including sequences that affiliate phylogenetically within the division Crenarchaeota (domain Archaea). To characterize the spatial distribution of crenarchaeal assemblages in mesophilic soil habitats, we profiled amplified crenarchaeal 16S rDNA sequences from diverse soil ecosystems by using PCR-single-stranded-conformation polymorphism (PCR-SSCP) analysis. PCR-SSCP profiles provide a measure of relative microbial diversity in terms of richness (number of different phylotypes as estimated from the number of unique PCR-SSCP peaks) and evenness (abundance of each phylotype as estimated from the relative area under a peak). Crenarchaeal assemblages inhabiting prairie, forest, turf, and agricultural soils were characterized at six sampling locations in southern and central Wisconsin. Phylotype richness was found to be more stable than evenness among triplicate samples collected within 30 cm at each sampling location. Transformation of the PCR-SSCP data by principal-component analysis, followed by statistical testing (analysis of variance [P < 0.0001] and least-significant-difference analysis [α = 0.5]), supported the conclusion that each location exhibited a unique profile. To further characterize the spatial distribution of crenarchaeal assemblages at one location, additional soil samples (a total of 30) were collected from agricultural field plots at the Hancock Agricultural Research Station. PCR-SSCP revealed a patchy spatial distribution of crenarchaeal assemblages within and between these plots. This mosaic of crenarchaeal assemblages was characterized by differences in phylotype evenness that could not be correlated with horizontal distance (15 to 30 m) or with depth (0 to 20 cm below the surface). Crenarchaeal 16S rDNA clone libraries were produced and screened for unique SSCP peaks. Clones representing the dominant phylotypes at each location were identified, sequenced, and found to group phylogenetically with sequences in crenarchaeal clade C1b.

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Merrill, Lori, Jennifer Richardson, Cheryl R. Kuske, and John Dunbar. "Fluorescent Heteroduplex Assay for Monitoring Bacillus anthracis and Close Relatives in Environmental Samples." Applied and Environmental Microbiology 69, no. 6 (June 2003): 3317–26. http://dx.doi.org/10.1128/aem.69.6.3317-3326.2003.

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ABSTRACT A fluorescent heteroduplex method was developed to assess the presence of 16S rRNA gene (rDNA) sequences from Bacillus anthracis and close relatives in PCR-amplified 16S rDNA sequence mixtures from environmental samples. The method uses a single-stranded, fluorescent DNA probe, 464 nucleotides in length, derived from a B. anthracis 16S rRNA gene. The probe contains a unique, engineered deletion such that all probe-target duplexes are heteroduplexes with an unpaired G at position 343 (ΔG343). Heteroduplex profiles of sequences ≥85% similar to the probe were produced using an ABI 377 sequencer in less than 3 h. The method divides strains of the Bacillus cereus-Bacillus thuringiensis-B. anthracis group into two subgroups. Each subgroup is defined by a specific 16S rRNA gene sequence type. Sequence type A, containing one mismatch with the probe, occurs in B. anthracis and a small number of closely related clonal lineages represented mostly by food-borne pathogenic isolates of B. cereus and B. thuringiensis. Sequence type B, containing two mismatches with the probe, is found in the majority of B. cereus and B. thuringiensis strains examined to date. Sequence types A and B, when hybridized to the probe, generate two easily differentiated heteroduplexes. Thus, from heteroduplex profiles, the presence of B. cereus-B. thuringiensis-B. anthracis subgroups in environmental samples can be inferred unambiguously. The results show that fluorescent heteroduplex analysis is an effective profiling technique for detection and differentiation of sequences representing small phylogenetic or functional groups in environmental samples.

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Röling, Wilfred F. M., Boris M. van Breukelen, Martin Braster, Bin Lin, and Henk W. van Verseveld. "Relationships between Microbial Community Structure and Hydrochemistry in a Landfill Leachate-Polluted Aquifer." Applied and Environmental Microbiology 67, no. 10 (October 1, 2001): 4619–29. http://dx.doi.org/10.1128/aem.67.10.4619-4629.2001.

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ABSTRACT Knowledge about the relationship between microbial community structure and hydrogeochemistry (e.g., pollution, redox and degradation processes) in landfill leachate-polluted aquifers is required to develop tools for predicting and monitoring natural attenuation. In this study analyses of pollutant and redox chemistry were conducted in parallel with culture-independent profiling of microbial communities present in a well-defined aquifer (Banisveld, The Netherlands). Degradation of organic contaminants occurred under iron-reducing conditions in the plume of pollution, while upstream of the landfill and above the plume denitrification was the dominant redox process. Beneath the plume iron reduction occurred. Numerical comparison of 16S ribosomal DNA (rDNA)-based denaturing gradient gel electrophoresis (DGGE) profiles of Bacteria andArchaea in 29 groundwater samples revealed a clear difference between the microbial community structures inside and outside the contaminant plume. A similar relationship was not evident in sediment samples. DGGE data were supported by sequencing cloned 16S rDNA. Upstream of the landfill members of the β subclass of the class Proteobacteria(β-proteobacteria) dominated. This group was not encountered beneath the landfill, where gram-positive bacteria dominated. Further downstream the contribution of gram-positive bacteria to the clone library decreased, while the contribution of δ-proteobacteria strongly increased and β-proteobacteria reappeared. The β-proteobacteria (Acidovorax,Rhodoferax) differed considerably from those found upstream (Gallionella, Azoarcus). Direct comparisons of cloned 16S rDNA with bands in DGGE profiles revealed that the data from each analysis were comparable. A relationship was observed between the dominant redox processes and the bacteria identified. In the iron-reducing plume members of the familyGeobacteraceae made a strong contribution to the microbial communities. Because the only known aromatic hydrocarbon-degrading, iron-reducing bacteria areGeobacter spp., their occurrence in landfill leachate-contaminated aquifers deserves more detailed consideration.

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Horz, Hans-Peter, Merlin Tchawa Yimga, and Werner Liesack. "Detection of Methanotroph Diversity on Roots of Submerged Rice Plants by Molecular Retrieval ofpmoA, mmoX, mxaF, and 16S rRNA and Ribosomal DNA, Including pmoA-Based Terminal Restriction Fragment Length Polymorphism Profiling." Applied and Environmental Microbiology 67, no. 9 (September 1, 2001): 4177–85. http://dx.doi.org/10.1128/aem.67.9.4177-4185.2001.

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ABSTRACT The diversity of methanotrophic bacteria associated with roots of submerged rice plants was assessed using cultivation-independent techniques. The research focused mainly on the retrieval ofpmoA, which encodes the α subunit of the particulate methane monooxygenase. A novel methanotroph-specific community-profiling method was established using the terminal restriction fragment length polymorphism (T-RFLP) technique. The T-RFLP profiles clearly revealed a more complex root-associated methanotrophic community than did banding patterns obtained bypmoA-based denaturing gradient gel electrophoresis. The comparison of pmoA-based T-RFLP profiles obtained from rice roots and bulk soil of flooded rice microcosms suggested that there was a substantially higher abundance of type I methanotrophs on rice roots than in the bulk soil. These were affiliated to the generaMethylomonas, Methylobacter,Methylococcus, and to a novel type I methanotroph sublineage. By contrast, type II methanotrophs of theMethylocystis-Methylosinus group could be detected with high relative signal intensity in both soil and root compartments. Phylogenetic treeing analyses and a set of substrate-diagnostic amino acid residues provided evidence that a novelpmoA lineage was detected. This branched distinctly from all currently known methanotrophs. To examine whether the retrieval ofpmoA provided a complete view of root-associated methanotroph diversity, we also assessed the diversity detectable by recovery of genes coding for subunits of soluble methane monooxygenase (mmoX) and methanol dehydrogenase (mxaF). In addition, both 16S rRNA and 16S ribosomal DNA (rDNA) were retrieved using a PCR primer set specific to type I methanotrophs. The overall methanotroph diversity detected by recovery of mmoX,mxaF, and 16S rRNA and 16S rDNA corresponded well to the diversity detectable by retrieval of pmoA.

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Visioli, Giovanna, Anna Maria Sanangelantoni, Teofilo Vamerali, Cristian Dal Cortivo, and Massimo Blandino. "16S rDNA Profiling to Reveal the Influence of Seed-Applied Biostimulants on the Rhizosphere of Young Maize Plants." Molecules 23, no. 6 (June 15, 2018): 1461. http://dx.doi.org/10.3390/molecules23061461.

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Yadav, Shailendra, Arvind Kumar, Manish Gupta, and S. S. Maitra. "Cross-Reactivity of Prokaryotic 16S rDNA-Specific Primers to Eukaryotic DNA: Mistaken Microbial Community Profiling in Environmental Samples." Current Microbiology 75, no. 8 (April 2, 2018): 1038–45. http://dx.doi.org/10.1007/s00284-018-1482-4.

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Sahin, F., P. A. Abbasi, M. L. Lewis Ivey, J. Zhang, and S. A. Miller. "Diversity Among Strains of Xanthomonas campestris pv. vitians from Lettuce." Phytopathology® 93, no. 1 (January 2003): 64–70. http://dx.doi.org/10.1094/phyto.2003.93.1.64.

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Diversity in host range, pathogenicity, phenotypic characteristics, repetitive extragenic palindromic polymerase chain reaction (rep-PCR) profiles, and sequence of the 16S-23S rDNA spacer region was examined among 44 Xanthomonas strains isolated from lettuce. Forty-two of the strains were divided into two groups, designated A and B. Seventy percent were Group A, and most of the remaining strains including a reference strain (LMG 938) were Group B. Group A strains induced both local and systemic symptoms, whereas Group B strains caused only distinct necrotic spots. Two strains, including the X. campestris pv. vitians type strain, were distinct from the Group A and B strains and were not pathogenic on lettuce. Analysis of fatty acid profiles, serotype, carbon substrate utilization patterns, and protein fingerprints confirmed this grouping. The Group A and B strains also formed two unique clusters (I and II) by rep-PCR profiling that corresponded to the two groups. Direct sequencing of a PCR-amplified DNA fragment (680 bp) from the 16S-23S rDNA spacer region of four representative strains, however, did not differentiate these groups. Serology and rep-PCR fingerprinting can be used to diagnose and identify X. campestris pv. vitians strains, while the other analyses evaluated are useful for strain characterization.

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Ramachandran, Umesh, Nathan Wrana, Nazim Cicek, Richard Sparling, and David B. Levin. "Isolation and characterization of a hydrogen- and ethanol-producing Clostridium sp. strain URNW." Canadian Journal of Microbiology 57, no. 3 (March 2011): 236–43. http://dx.doi.org/10.1139/w11-005.

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Identification, characterization, and end-product synthesis patterns were analyzed in a newly identified mesophilic, anaerobic Clostridium sp. strain URNW, capable of producing hydrogen (H2) and ethanol. Metabolic profiling was used to characterize putative end-product synthesis pathways of the Clostridium sp. strain URNW, which was found to grow on cellobiose; on hexose sugars, such as glucose, sucrose, and mannose; and on sugar alcohols, like mannitol and sorbitol. When grown in batch cultures on 2 g cellobiose·L–1, Clostridium sp. strain URNW showed a cell generation time of 1.5 h, and the major end-products were H2, formate, carbon dioxide (CO2), lactate, butyrate, acetate, pyruvate, and ethanol. The total volumetric H2production was 14.2 mmol·(L culture)–1and the total production of ethanol was 0.4 mmol·(L culture)–1. The maximum yield of H2was 1.3 mol·(mol glucose equivalent)–1at a carbon recovery of 94%. The specific production rates of H2, CO2, and ethanol were 0.45, 0.13, and 0.003 mol·h–1·(g dry cell mass)–1, respectively. BLAST analyses of 16S rDNA and chaperonin 60 (cpn60) sequences from Clostridium sp. strain URNW revealed a 98% nucleotide sequence identity with the 16S rDNA and cpn60 sequences from Clostridium intestinale ATCC 49213. Phylogenetic analyses placed Clostridium sp. strain URNW within the butyrate-synthesizing clostridia.

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Rudi, Knut, Olav M. Skulberg, Randi Skulberg, and Kjetill S. Jakobsen. "Application of Sequence-Specific Labeled 16S rRNA Gene Oligonucleotide Probes for Genetic Profiling of Cyanobacterial Abundance and Diversity by Array Hybridization." Applied and Environmental Microbiology 66, no. 9 (September 1, 2000): 4004–11. http://dx.doi.org/10.1128/aem.66.9.4004-4011.2000.

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ABSTRACT DNA sequence information for the small-subunit rRNA gene (16S rDNA) obtained from cyanobacterial cultures was used to investigate the presence of cyanobacteria and their abundance in natural habitats. Eight planktonic communities developing in lakes characterized by relatively low algal biomass (mesotrophic) and in lakes with correspondingly high biomass (eutrophic) were selected for the study. The organismal compositions of the water samples were analyzed genetically, using multiplex sequence-specific labeling of oligonucleotide probes targeted to 16S rDNA and subsequent hybridization of the labeled probes to their respective complements spotted onto a solid support (DNA array). Ten probes were established to determine the relative abundances of the discernible cyanobacteria encountered in the selected lakes. The probes were generally specific for their targets, as determined through analyses of clone cultures. Reproducible abundance profiles were established for the lakes investigated in the subsequent analyses of natural cyanobacterial communities. The results from the genetic analyses were then compared with information obtained from standard hydrobiological and hydrochemical analyses. Qualitatively, there were relatively good correlations among the groups of organisms (Nostoc,Microcystis, and Planktothrix species) found in the different lakes. The levels of correlation were lower for the quantitative data. This may, however, be due to differences in sample processing technique. The conclusions from these comparisons are that the genetic abundance profiles may provide a foundation for separating and quantifying genetically distinct groups of cyanobacteria in their natural habitats.

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Highlander, Sarah K. "High throughput sequencing methods for microbiome profiling: application to food animal systems." Animal Health Research Reviews 13, no. 1 (June 2012): 40–53. http://dx.doi.org/10.1017/s1466252312000126.

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AbstractAnalysis of microbial communities using high throughput sequencing methods began in the mid 2000s permitting the production of 1000s to 10,000s of sequence reads per sample and megabases of data per sequence run. This then unprecedented depth of sequencing allowed, for the first time, the discovery of the ‘rare biosphere’ in environmental samples. The technology was quickly applied to studies in several human subjects. Perhaps these early studies served as a reminder that though the microbes that inhabit mammals are known to outnumber host cells by an order of magnitude or more, most of these are unknown members of our second genome, or microbiome (as coined by Joshua Lederberg), because of our inability to culture them. High throughput methods for microbial 16S ribosomal RNA gene and whole genome shotgun (WGS) sequencing have now begun to reveal the composition and identity of archaeal, bacterial and viral communities at many sites, in and on the human body. Surveys of the microbiota of food production animals have been published in the past few years and future studies should benefit from protocols and tools developed from large-scale human microbiome studies. Nevertheless, production animal-related resources, such as improved host genome assemblies and increased numbers and diversity of host-specific microbial reference genome sequences, will be needed to permit meaningful and robust analysis of 16S rDNA and WGS sequence data.

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Griffiths, Steve, Krista Melville, Marcia Cook, Sherry Vincent, Maurice Pierre, and Carole Lanteigne. "Profiling of Bacterial Species Associated with Haddock Larviculture by PCR Amplification of 16S rDNA and Denaturing Gradient Gel Electrophoresis." Journal of Aquatic Animal Health 13, no. 4 (December 2001): 355–63. http://dx.doi.org/10.1577/1548-8667(2001)013<0355:pobsaw>2.0.co;2.

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Mataragas, Marios, Valentina Alessandria, Ilario Ferrocino, Kalliopi Rantsiou, and Luca Cocolin. "A bioinformatics pipeline integrating predictive metagenomics profiling for the analysis of 16S rDNA/rRNA sequencing data originated from foods." Food Microbiology 76 (December 2018): 279–86. http://dx.doi.org/10.1016/j.fm.2018.05.009.

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Guest, Claire, Rob Harris, Karen S. Sfanos, Eva Shrestha, Alan W. Partin, Bruce Trock, Leslie Mangold, et al. "Feasibility of integrating canine olfaction with chemical and microbial profiling of urine to detect lethal prostate cancer." PLOS ONE 16, no. 2 (February 17, 2021): e0245530. http://dx.doi.org/10.1371/journal.pone.0245530.

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Prostate cancer is the second leading cause of cancer death in men in the developed world. A more sensitive and specific detection strategy for lethal prostate cancer beyond serum prostate specific antigen (PSA) population screening is urgently needed. Diagnosis by canine olfaction, using dogs trained to detect cancer by smell, has been shown to be both specific and sensitive. While dogs themselves are impractical as scalable diagnostic sensors, machine olfaction for cancer detection is testable. However, studies bridging the divide between clinical diagnostic techniques, artificial intelligence, and molecular analysis remains difficult due to the significant divide between these disciplines. We tested the clinical feasibility of a cross-disciplinary, integrative approach to early prostate cancer biosensing in urine using trained canine olfaction, volatile organic compound (VOC) analysis by gas chromatography-mass spectroscopy (GC-MS) artificial neural network (ANN)-assisted examination, and microbial profiling in a double-blinded pilot study. Two dogs were trained to detect Gleason 9 prostate cancer in urine collected from biopsy-confirmed patients. Biopsy-negative controls were used to assess canine specificity as prostate cancer biodetectors. Urine samples were simultaneously analyzed for their VOC content in headspace via GC-MS and urinary microbiota content via 16S rDNA Illumina sequencing. In addition, the dogs’ diagnoses were used to train an ANN to detect significant peaks in the GC-MS data. The canine olfaction system was 71% sensitive and between 70–76% specific at detecting Gleason 9 prostate cancer. We have also confirmed VOC differences by GC-MS and microbiota differences by 16S rDNA sequencing between cancer positive and biopsy-negative controls. Furthermore, the trained ANN identified regions of interest in the GC-MS data, informed by the canine diagnoses. Methodology and feasibility are established to inform larger-scale studies using canine olfaction, urinary VOCs, and urinary microbiota profiling to develop machine olfaction diagnostic tools. Scalable multi-disciplinary tools may then be compared to PSA screening for earlier, non-invasive, more specific and sensitive detection of clinically aggressive prostate cancers in urine samples.

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Farnleitner, Andreas H., Franziska Zibuschka, Martina M. Burtscher, Gerhard Lindner, Georg Reischer, and Robert L. Mach. "Eubacterial 16S-rDNA amplicon profiling: a rapid technique for comparison and differentiation of heterotrophic plate count communities from drinking water." International Journal of Food Microbiology 92, no. 3 (May 2004): 333–45. http://dx.doi.org/10.1016/j.ijfoodmicro.2003.08.014.

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Złoch, Michał, Agnieszka Rodzik, Katarzyna Pauter, Małgorzata Szultka-Młyńska, Agnieszka Rogowska, Wojciech Kupczyk, Paweł Pomastowski, and Bogusław Buszewski. "Problems with identifying and distinguishing salivary streptococci: a multi-instrumental approach." Future Microbiology 15, no. 12 (August 2020): 1157–71. http://dx.doi.org/10.2217/fmb-2020-0036.

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Aim: The purpose of this study was to create an alternative protocol for the DNA-based identification of salivary microbiota focused on the distinguishing of Streptococcus species. Materials & methods: Salivary bacteria were identified using 16S rDNA sequencing and proteins and lipids profiling using MALDI-TOF/MS as well as FTIR analysis. Results: Most of the isolates belonged to streptococci - mostly the salivarious group indistinguishable by the molecular technique. In turn, MALDI analysis allowed for their fast and reliable classification. Although FTIR spectroscopy demonstrated the correct species classification, the spectra interpretation was time consuming and complicated. Conclusion: MALDI-TOF/MS demonstrated the biggest effectiveness in the identification and discrimination between the salivary streptococci, which could be easily incorporated in the workflow of routine microbiological laboratories.

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McCaig, Allison E., L. Anne Glover, and James I. Prosser. "Numerical Analysis of Grassland Bacterial Community Structure under Different Land Management Regimens by Using 16S Ribosomal DNA Sequence Data and Denaturing Gradient Gel Electrophoresis Banding Patterns." Applied and Environmental Microbiology 67, no. 10 (October 1, 2001): 4554–59. http://dx.doi.org/10.1128/aem.67.10.4554-4559.2001.

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ABSTRACT Bacterial diversity in unimproved and improved grassland soils was assessed by PCR amplification of bacterial 16S ribosomal DNA (rDNA) from directly extracted soil DNA, followed by sequencing of ∼45 16S rDNA clones from each of three unimproved and three improved grassland samples (A. E. McCaig, L. A. Glover, and J. I. Prosser, Appl. Environ. Microbiol. 65:1721–1730, 1999) or by denaturing gradient gel electrophoresis (DGGE) of total amplification products. Semi-improved grassland soils were analyzed only by DGGE. No differences between communities were detected by calculation of diversity indices and similarity coefficients for clone data (possibly due to poor coverage). Differences were not observed between the diversities of individual unimproved and improved grassland DGGE profiles, although considerable spatial variation was observed among triplicate samples. Semi-improved grassland samples, however, were less diverse than the other grassland samples and had much lower within-group variation. DGGE banding profiles obtained from triplicate samples pooled prior to analysis indicated that there was less evenness in improved soils, suggesting that selection for specific bacterial groups occurred. Analysis of DGGE profiles by canonical variate analysis but not by principal-coordinate analysis, using unweighted data (considering only the presence and absence of bands) and weighted data (considering the relative intensity of each band), demonstrated that there were clear differences between grasslands, and the results were not affected by weighting of data. This study demonstrated that quantitative analysis of data obtained by community profiling methods, such as DGGE, can reveal differences between complex microbial communities.

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Mäkivuokko, Harri, Kirsti Tiihonen, Soile Tynkkynen, Lars Paulin, and Nina Rautonen. "The effect of age and non-steroidal anti-inflammatory drugs on human intestinal microbiota composition." British Journal of Nutrition 103, no. 2 (August 25, 2009): 227–34. http://dx.doi.org/10.1017/s0007114509991553.

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Ageing has been suggested to cause changes in the intestinal microbial community. In the present study, the microbiota of a previously well-defined group of elderly subjects aged between 70 and 85 years, both non-steroidal anti-inflammatory drugs (NSAID) users (n9) and non-users (n9), were further compared with young adults (n14) with a mean age of 28 years, by two DNA-based techniques: percentage guanine+cytosine (%G+C) profiling and 16S rDNA sequencing. Remarkable changes in microbiota were described with both methods: compared with young adults a significant reduction in overall numbers of microbes in both elderly groups was measured. Moreover, the total number of microbes in elderly NSAID users was higher than in elderly without NSAID. In 16S rDNA sequencing, shifts in all major microbial phyla, such as lower numbers of Firmicutes and an increase in numbers of Bacteroidetes in the elderly were monitored. On the genus level an interesting link between reductions in the proportion of known butyrate producers belonging toClostridiumcluster XIVa, such asRoseburiaandRuminococcus, could be demonstrated in the elderly. Moreover, in the Actinobacteria group, lower numbers ofCollinsellaspp. were evident in the elderly subjects with NSAID compared both with young adults and the elderly without NSAID, suggesting that the use of NSAID along with age may also influence the composition of intestinal microbiota. Furthermore, relatively high numbers ofLactobacillusappeared only in the elderly subjects without NSAID. In general, the lowered numbers of microbial members in the major phyla, Firmicutes, together with changes in the epithelial layer functions can have a significant effect on the colon health of the elderly.

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Castro, Juan C., J. Dylan Maddox, Hicler N. Rodríguez, Richard B. Orbe, Gad E. Grandez, Kevin A. Feldheim, Marianela Cobos, et al. "Metagenomic 16S rDNA amplicon data on bacterial diversity profiling and its predicted metabolic functions of varillales in Allpahuayo-Mishana National Reserve." Data in Brief 30 (June 2020): 105625. http://dx.doi.org/10.1016/j.dib.2020.105625.

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Gu, Ganyu, Hsin-Bai Yin, Andrea Ottesen, Samantha Bolten, Jitendra Patel, Steve Rideout, and Xiangwu Nou. "Microbiomes in Ground Water and Alternative Irrigation Water, and Spinach Microbiomes Impacted by Irrigation with Different Types of Water." Phytobiomes Journal 3, no. 2 (January 2019): 137–47. http://dx.doi.org/10.1094/pbiomes-09-18-0037-r.

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Irrigation water, particularly if applied overhead, could be an important source of bacterial contamination to fresh produce. The colonization, survival, and proliferation of exogenous bacterial pathogens can be strongly influenced by the produce microbiota. In this study, spinach grown in an organic field was irrigated with ground water and potential alternative irrigation water including reclaimed wastewater, and urban runoff water, over a period of 2 weeks. Water and spinach samples were collected before and after irrigation for bacterial plate count, qPCR, and community profiling using 16S rDNA high-throughput sequencing analyses. The average bacterial population densities on spinach (6.50 ± 0.04 log CFU/g, 7.40 ± 0.10 log 16S copies/g) were significantly higher than those in irrigation water (3.61 ± 0.12 log CFU/ml, 4.94 ± 0.13 log 16S copies/ml). The composition and relative abundance of spinach microbiomes varied with different types of irrigation waters; however, the most abundant microbial taxa on spinach were not significantly affected by the irrigation with different types of water. Shigella, Salmonella, Listeria, Campylobacter spp., and pathogenic Escherichia coli were not detected in this study. This study provides information on the microbial ecology of diverse bacterial communities on spinach surface after irrigation by different types of water, which can benefit future studies on the interaction of microbes on produce, and the prevention of foodborne pathogens and plant disease.

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Liew, Winnie-Pui-Pui, Sabran Mohd-Redzwan, and Leslie Than. "Gut Microbiota Profiling of Aflatoxin B1-Induced Rats Treated with Lactobacillus casei Shirota." Toxins 11, no. 1 (January 17, 2019): 49. http://dx.doi.org/10.3390/toxins11010049.

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Aflatoxin B1 (AFB1) is a ubiquitous carcinogenic food contaminant. Gut microbiota is of vital importance for the host’s health, regrettably, limited studies have reported the effects of xenobiotic toxins towards gut microbiota. Thus, the present study aims to investigate the interactions between AFB1 and the gut microbiota. Besides, an AFB1-binding microorganism, Lactobacillus casei Shirota (Lcs) was tested on its ability to ameliorate the changes on gut microbiota induced by AFB1. The fecal contents of three groups of rats included an untreated control group, an AFB1 group, as well as an Lcs + AFB1 group, were analyzed. Using the MiSeq platform, the PCR products of 16S rDNA gene extracted from the feces were subjected to next-generation sequencing. The alpha diversity index (Shannon) showed that the richness of communities increased significantly in the Lcs + AFB1 group compared to the control and AFB1 groups. Meanwhile, beta diversity indices demonstrated that AFB1 group significantly deviated from the control and Lcs + AFB1 groups. AFB1-exposed rats were especially high in Alloprevotella spp. abundance. Such alteration in the bacterial composition might give an insight on the interactions of AFB1 towards gut microbiota and how Lcs plays its role in detoxification of AFB1.

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Peters, Sabine, Stefanie Koschinsky, Frank Schwieger, and Christoph C. Tebbe. "Succession of Microbial Communities during Hot Composting as Detected by PCR–Single-Strand-Conformation Polymorphism-Based Genetic Profiles of Small-Subunit rRNA Genes." Applied and Environmental Microbiology 66, no. 3 (March 1, 2000): 930–36. http://dx.doi.org/10.1128/aem.66.3.930-936.2000.

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ABSTRACT A cultivation-independent technique for genetic profiling of PCR-amplified small-subunit rRNA genes (SSU rDNA) was chosen to characterize the diversity and succession of microbial communities during composting of an organic agricultural substrate. PCR amplifications were performed with DNA directly extracted from compost samples and with primers targeting either (i) the V4–V5 region of eubacterial 16S rRNA genes, (ii) the V3 region in the 16S rRNA genes of actinomycetes, or (iii) the V8–V9 region of fungal 18S rRNA genes. Homologous PCR products were converted to single-stranded DNA molecules by exonuclease digestion and were subsequently electrophoretically separated by their single-strand-conformation polymorphism (SSCP). Genetic profiles obtained by this technique showed a succession and increasing diversity of microbial populations with all primers. A total of 19 single products were isolated from the profiles by PCR reamplification and cloning. DNA sequencing of these molecular isolates showed similarities in the range of 92.3 to 100% to known gram-positive bacteria with a low or high G+C DNA content and to the SSU rDNA of γ-Proteobacteria. The amplified 18S rRNA gene sequences were related to the respective gene regions of Candida krusei and Candida tropicalis. Specific molecular isolates could be attributed to different composting stages. The diversity of cultivated bacteria isolated from samples taken at the end of the composting process was low. A total of 290 isolates were related to only 6 different species. Two or three of these species were also detectable in the SSCP community profiles. Our study indicates that community SSCP profiles can be highly useful for the monitoring of bacterial diversity and community successions in a biotechnologically relevant process.

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De Santis, Stefania, Marina Liso, Mirco Vacca, Giulio Verna, Elisabetta Cavalcanti, Sergio Coletta, Francesco Maria Calabrese, et al. "Dysbiosis Triggers ACF Development in Genetically Predisposed Subjects." Cancers 13, no. 2 (January 14, 2021): 283. http://dx.doi.org/10.3390/cancers13020283.

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Background: Colorectal cancer (CRC) is the third most common cancer worldwide, characterized by a multifactorial etiology including genetics, lifestyle, and environmental factors including microbiota composition. To address the role of microbial modulation in CRC, we used our recently established mouse model (the Winnie-APCMin/+) combining inflammation and genetics. Methods: Gut microbiota profiling was performed on 8-week-old Winnie-APCMin/+ mice and their littermates by 16S rDNA gene amplicon sequencing. Moreover, to study the impact of dysbiosis induced by the mother’s genetics in ACF development, the large intestines of APCMin/+ mice born from wild type mice were investigated by histological analysis at 8 weeks. Results: ACF development in 8-week-old Winnie-APCMin/+ mice was triggered by dysbiosis. Specifically, the onset of ACF in genetically predisposed mice may result from dysbiotic signatures in the gastrointestinal tract of the breeders. Additionally, fecal transplant from Winnie donors to APCMin/+ hosts leads to an increased rate of ACF development. Conclusions: The characterization of microbiota profiling supporting CRC development in genetically predisposed mice could help to design therapeutic strategies to prevent dysbiosis. The application of these strategies in mothers during pregnancy and lactation could also reduce the CRC risk in the offspring.

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Urakawa, Hidetoshi, Tsutomu Yoshida, Masahiko Nishimura, and Kouichi Ohwada. "Characterization of depth-related population variation in microbial communities of a coastal marine sediment using 16S rDNA-based approaches and quinone profiling." Environmental Microbiology 2, no. 5 (October 2000): 542–54. http://dx.doi.org/10.1046/j.1462-2920.2000.00137.x.

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García-López, Rodrigo, Fernanda Cornejo-Granados, Alonso A. Lopez-Zavala, Andrés Cota-Huízar, Rogerio R. Sotelo-Mundo, Bruno Gómez-Gil, and Adrian Ochoa-Leyva. "OTUs and ASVs Produce Comparable Taxonomic and Diversity from Shrimp Microbiota 16S Profiles Using Tailored Abundance Filters." Genes 12, no. 4 (April 13, 2021): 564. http://dx.doi.org/10.3390/genes12040564.

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The interplay between shrimp immune system, its environment, and microbiota contributes to the organism’s homeostasis and optimal production. The metagenomic composition is typically studied using 16S rDNA profiling by clustering amplicon sequences into operational taxonomic units (OTUs) and, more recently, amplicon sequence variants (ASVs). Establish the compatibility of the taxonomy, α, and β diversity described by both methods is necessary to compare past and future shrimp microbiota studies. Here, we used identical sequences to survey the V3 16S hypervariable-region using 97% and 99% OTUs and ASVs to assess the hepatopancreas and intestine microbiota of L. vannamei from two ponds under standardized rearing conditions. We found that applying filters to retain clusters >0.1% of the total abundance per sample enabled a consistent taxonomy comparison while preserving >94% of the total reads. The three sets turned comparable at the family level, whereas the 97% identity OTU set produced divergent genus and species profiles. Interestingly, the detection of organ and pond variations was robust to the clustering method’s choice, producing comparable α and β-diversity profiles. For comparisons on shrimp microbiota between past and future studies, we strongly recommend that ASVs be compared at the family level to 97% identity OTUs or use 99% identity OTUs, both using tailored frequency filters.

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LOUI, CINDY, GRIGOR GRIGORYAN, HAOHAO HUANG, LEE W. RILEY, and SANGWEI LU. "Bacterial Communities Associated with Retail Alfalfa Sprouts." Journal of Food Protection 71, no. 1 (January 1, 2008): 200–204. http://dx.doi.org/10.4315/0362-028x-71.1.200.

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Fresh produce, including salad, is increasingly implicated in foodborne outbreaks. Although studies have been carried out to detect specific human pathogens from fresh produce, the total bacterial community associated with fresh produce is poorly understood. In this study, we characterized the bacterial community associated with alfalfa sprouts, using a culture-independent method. Four retail-purchased alfalfa sprout samples were obtained from different producers, and the bacterial community associated with each sample was determined by 16S rDNA profiling. Our results indicate that alfalfa sprouts sampled in our study shared significant similarities in their bacterial communities. Proteobacteria was the dominant phylum detected from all alfalfa sprout samples, with Enterobacteriaceae, Oxalobacteraceae, Moraxellaceae, and Sphingomonadaceae as the most frequently detected families. These results indicate that growth conditions of alfalfa sprouts should be taken into consideration to prevent the proliferation of pathogenic proteobacteria such as Escherichia coli O157 and Salmonella.

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Zeng, Su-Ling, Shang-Zhen Li, Ping-Ting Xiao, Yuan-Yuan Cai, Chu Chu, Bai-Zhong Chen, Ping Li, Jing Li, and E.-Hu Liu. "Citrus polymethoxyflavones attenuate metabolic syndrome by regulating gut microbiome and amino acid metabolism." Science Advances 6, no. 1 (January 2020): eaax6208. http://dx.doi.org/10.1126/sciadv.aax6208.

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Metabolic syndrome (MetS) is intricately linked to dysregulation of gut microbiota and host metabolomes. Here, we first find that a purified citrus polymethoxyflavone-rich extract (PMFE) potently ameliorates high-fat diet (HFD)–induced MetS, alleviates gut dysbiosis, and regulates branched-chain amino acid (BCAA) metabolism using 16S rDNA amplicon sequencing and metabolomic profiling. The metabolic protective effects of PMFE are gut microbiota dependent, as demonstrated by antibiotic treatment and fecal microbiome transplantation (FMT). The modulation of gut microbiota altered BCAA levels in the host serum and feces, which were significantly associated with metabolic features and actively responsive to therapeutic interventions with PMFE. Notably, PMFE greatly enriched the commensal bacterium Bacteroides ovatus, and gavage with B. ovatus reduced BCAA concentrations and alleviated MetS in HFD mice. PMFE may be used as a prebiotic agent to attenuate MetS, and target-specific microbial species may have unique therapeutic promise for metabolic diseases.

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Sebat, Jonathan L., Frederick S. Colwell, and Ronald L. Crawford. "Metagenomic Profiling: Microarray Analysis of an Environmental Genomic Library." Applied and Environmental Microbiology 69, no. 8 (August 2003): 4927–34. http://dx.doi.org/10.1128/aem.69.8.4927-4934.2003.

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ABSTRACT Genomic libraries derived from environmental DNA (metagenomic libraries) are useful for characterizing uncultured microorganisms. However, conventional library-screening techniques permit characterization of relatively few environmental clones. Here we describe a novel approach for characterization of a metagenomic library by hybridizing the library with DNA from a set of groundwater isolates, reference strains, and communities. A cosmid library derived from a microcosm of groundwater microorganisms was used to construct a microarray (COSMO) containing ∼1-kb PCR products amplified from the inserts of 672 cosmids plus a set of 16S ribosomal DNA controls. COSMO was hybridized with Cy5-labeled genomic DNA from each bacterial strain, and the results were compared with the results for a common Cy3-labeled reference DNA sample consisting of a composite of genomic DNA from multiple species. The accuracy of the results was confirmed by the preferential hybridization of each strain to its corresponding rDNA probe. Cosmid clones were identified that hybridized specifically to each of 10 microcosm isolates, and other clones produced positive results with multiple related species, which is indicative of conserved genes. Many clones did not hybridize to any microcosm isolate; however, some of these clones hybridized to community genomic DNA, suggesting that they were derived from microbes that we failed to isolate in pure culture. Based on identification of genes by end sequencing of 17 such clones, DNA could be assigned to functions that have potential ecological importance, including hydrogen oxidation, nitrate reduction, and transposition. Metagenomic profiling offers an effective approach for rapidly characterizing many clones and identifying the clones corresponding to unidentified species of microorganisms.

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Forbes, Jessica D., Gary Van Domselaar, Michael Sargent, Chris Green, Susan Springthorpe, Denis O. Krause, and Charles N. Bernstein. "Microbiome profiling of drinking water in relation to incidence of inflammatory bowel disease." Canadian Journal of Microbiology 62, no. 9 (September 2016): 781–93. http://dx.doi.org/10.1139/cjm-2016-0219.

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The etiology of inflammatory bowel disease (IBD) is unknown; current research is focused on determining environmental factors. One consideration is drinking water: water systems harbour considerable microbial diversity, with bacterial concentrations estimated at 106–108 cells/L. Perhaps differences in microbial ecology of water sources may impact differential incidence rates of IBD. Regions of Manitoba were geographically mapped according to incidence rates of IBD and identified as high (HIA) or low (LIA) incidence areas. Bulk water, filter material, and pipe wall samples were collected from public buildings in different jurisdictions and their population structure analyzed using 16S rDNA sequencing. At the phylum level, Proteobacteria were observed significantly less frequently (P = 0.02) in HIA versus LIA. The abundance of Proteobacteria was also found to vary according to water treatment distribution networks. Gammaproteobacteria was the most abundant class of bacteria and was observed more frequently (P = 0.006) in LIA. At the genus level, microbes found to associate with HIA include Bradyrhizobium (P = 0.02) and Pseudomonas (P = 0.02). Particular microbes were found to associate with LIA or HIA, based on sample location and (or) type. This work lays out a basis for further studies exploring water as a potential environmental source for IBD triggers.

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Shibulal, Biji, Saif N. Al-Bahry, Yahya M. Al-Wahaibi, Abdulkadir E. Elshafie, Ali S. Al-Bemani, and Sanket J. Joshi. "Analysis of Bacterial Diversity in Different Heavy Oil Wells of a Reservoir in South Oman with Alkaline pH." Scientifica 2018 (2018): 1–10. http://dx.doi.org/10.1155/2018/9230143.

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The identification of potential hydrocarbon utilizing bacteria is an essential requirement in microbial enhanced oil recovery (MEOR). Molecular approaches like proteomic and genomic characterization of the isolates are replacing the traditional method of identification with systemic classification. Genotypic profiling of the isolates includes fingerprint or pattern-based technique and sequence-based technique. Understanding community structure and dynamics is essential for studying diversity profiles and is challenging in the case of microbial analysis. The present study aims to understand the bacterial community composition from different heavy oil contaminated soil samples collected from geographically related oil well areas in Oman and to identify spore-forming hydrocarbon utilizing cultivable bacteria. V4 region of 16S rDNA gene was the target for Ion PGM™. A total of 825081 raw sequences were obtained from Ion torrent from all the 10 soil samples. The species richness and evenness were found to be moderate in all the samples with four main phyla, Firmicutes, Bacteroidetes, Proteobacteria, and Actinobacteria, the most abundant being Firmicutes.Bacillussp. ubiquitously dominated in all samples followed byPaenibacillus, which was followed byBrevibacillus,Planococcus, andFlavobacterium. Principal Coordinate Analysis (PCoA) and UPGMA dendrogram clustered the 10 soil samples into four main groups. Weighted UniFrac significance test determined that there was significant difference in the communities present in soil samples examined. It can be concluded that the microbial community was different in all the 10 soil samples withBacillusandPaenibacillussp. as predominating genus. The 16S rDNA sequencing of cultivable spore-forming bacteria identified the hydrocarbon utilizing bacteria asBacillusandPaenibacillussp. and the nucleotide sequences were submitted to NCBI GenBank under accession numbers KP119097–KP119115.BacillusandPaenibacillussp., which were relatively abundant in the oil fields, can be recommended to be chosen as candidates for hydrocarbon utilization study.

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Röling, W. F. M., B. M. van Breukelen, M. Braster, M. T. Goeltom, J. Groen, and H. W. van Verseveld. "Analysis of Microbial Communities in a Landfill Leachate Polluted Aquifer using a New Method for Anaerobic Physiological Profiling and 16S rDNA Based Fingerprinting." Microbial Ecology 40, no. 3 (August 2000): 177–88. http://dx.doi.org/10.1007/s002480000033.

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Sakai, Kazuko, Marco A. De Velasco, Yurie Kura, and Kazuto Nishio. "Transcriptome Profiling and Metagenomic Analysis Help to Elucidate Interactions in an Inflammation-Associated Cancer Mouse Model." Cancers 13, no. 15 (July 22, 2021): 3683. http://dx.doi.org/10.3390/cancers13153683.

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Colitis is a risk factor for colorectal cancer (CRC) and can change the dynamics of gut microbiota, leading to dysbiosis and contributing to carcinogenesis. The functional interactions between colitis-associated CRC and microbiota remain unknown. In this study, colitis and CRC were induced in BALB/c mice by the administration of dextran sodium sulfate (DSS) and/or azoxymethane (AOM). Whole transcriptome profiling of normal colon was then performed, and gene set enrichment analysis (GSEA) revealed enriched fatty acid metabolism, oxidative phosphorylation, and PI3K-Akt-mTOR signaling in the tissues from DSS/AOM mice. Additionally, immunohistochemical staining showed increased expression levels of phosphorylated S6 ribosomal protein, a downstream target of the PI3K-Akt-mTOR pathway in the inflamed mucosa of DSS/AOM mice. Fecal microbes were characterized using 16S rDNA gene sequencing. Redundancy analysis demonstrated a significant dissimilarity between the DSS/AOM group and the others. Functional analysis inferred from microbial composition showed enrichments of the sphingolipid signal and lipoarabinomannan biosynthetic pathways. This study provides additional insights into alterations associated with DSS/AOM-induced colitis and associates PI3K-Akt-mTOR, sphingolipid-signaling and lipoarabinomannan biosynthetic pathways in mouse DSS/AOM-induced colitis.

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Shevchenko, Margarita, Stanislav Sukhikh, Olga Babich, Svetlana Noskova, Svetlana Ivanova, Valery Lisun, Viktoriya Skripskaya, Andrey Lomtev, and Maria Zimina. "First Insight into the Diversity and Antibacterial Potential of Psychrophilic and Psychotrophic Microbial Communities of Abandoned Amber Quarry." Microorganisms 9, no. 7 (July 16, 2021): 1521. http://dx.doi.org/10.3390/microorganisms9071521.

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Natural habitats, including extreme ones, are potential sources of new antimicrobial compound producers, such as bacteriocins and enzymes, capable of degrading the matrix polysaccharides of bacterial biofilms. This study aimed to investigate biodiversity and evaluate the antibacterial potential of psychrophilic and psychrotrophic microbial communities of the flooded Walter amber quarry (Kaliningrad region, Russia). As a result of 16S rDNA high-throughput profiling, 127 genera of bacteria belonging to 12 phyla of bacteria were found in sediment samples: Acidobacteria sp., Actinobacteria sp., Armatimonadetes sp., Bacteroidetes sp., Chloroflexi sp., Cyanobacteria sp., Firmicutes sp., Gemmatimonadetes sp., Planctomycetes sp., Proteobacteria sp., Tenericutes sp., and Verrucomicrobia sp. The dominant bacteria groups were the families Ruminococcaceae and Lachnospiraceae, belonging to the order Clostridiales phylum Firmicutes. Analysis of enrichment cultures obtained from sediments showed the presence of antibacterial and cellulolytic activity. It seems likely that the bacteria of the studied communities are producers of antimicrobial compounds and have the potential for biotechnological use.

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Montero, Olimpio. "Lipid Profiling of Synechococcus sp. PCC7002 and Two Related Strains by HPLC Coupled to ESI-(Ion Trap)-MS/MS." Zeitschrift für Naturforschung C 66, no. 3-4 (April 1, 2011): 149–58. http://dx.doi.org/10.1515/znc-2011-3-409.

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The lipid profiles of Synechococcus sp. PCC7002 and two related 16S rDNA (99% identity) strains were established by a new method of high-performance liquid chromatography coupled to electrospray-mass spectrometry (HPLC-MS). Lipids were analysed in the positive and negative ionization mode, and fragmentation patterns are reported. No differences in the lipid profi le between the three strains could be observed, but the relative content of some species differed. Major lipid species were found to be 1-octadecatrienoyl- 2-hexadecanoyl-3-(6’-sulfo-α-D-quinovosyl)-sn-glycerol [SQDG (18:3/16:0)] and 1-octadecatrienoyl-2-hexadecenoyl-3-β-D-monogalactosyl-sn-glycerol [MGDG (18:3/16:1)]. Ten species of SQDG, six species of PG (phosphatidyl-glycerol), seven species of MGDG, and two species of DGDG (digalactosyl-diacyl-glycerol) were detected. A PG species (m/z 761) containing hydroxylinolenic acid or oxophytodienoic acid acyl ester (C18H32O3), and SQDG species containing C17:1 and C17:3 fatty acyl esters are reported for the first time in cyanobacteria. The method also allowed the separation of two pairs of closely related isobaric MGDG species (m/z 770 and m/z 772 in positive ionization)

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Wyatt, Morgan A., Jonghyun Lee, Yasodha Ahilan, and Nathan A. Magarvey. "Bioinformatic evaluation of the secondary metabolism of antistaphylococcal environmental bacterial isolates." Canadian Journal of Microbiology 59, no. 7 (July 2013): 465–71. http://dx.doi.org/10.1139/cjm-2013-0016.

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The increasing occurrence of drug-resistant Staphylococcus aureus is exacerbated with a declining rate of antibiotic discovery, particularly those with new mechanisms of action. The decline in antibiotic discovery from traditional sources, such as soil actinobacteria, necessitates examination of lesser studied microbes. Here, we present a strategy to select for organisms that may have a propensity to result in new antistaphylococcal agents by using S. aureus as a bait organism, and selecting organisms that have a natural lytic activity towards it. We have isolated over 80 environmental isolates and typed these organisms using 16S rDNA sequence comparison and deployed bioinformatics to assess the secondary metabolic potential of the isolated antistaphylococcal bacteria using genomic sequences. Bioinformatic analysis highlights the enriched and unique suite of potential antibiotic polyketides and nonribosomal peptides and lantibiotic gene clusters from these organisms. Profiling organic microbial extracts further showed that many of the organisms from the 10 staphylolytic genera secrete agents with antistaphylococcal activity and may serve as new sources for future antistaphylococcal drug discovery.

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Jarabo-Lorenzo, Adriana, Encarna Velázquez, Ricardo Pérez-Galdona, Maria C. Vega-Hernández, Eustoquio Martínez-Molina, Pedro F. Mateos, Pablo Vinuesa, Esperanza Martínez-Romero, and Milagros León-Barrios. "Restriction Fragment Length Polymorphism Analysis of 16S rDNA and Low Molecular Weight RNA Profiling of Rhizobial Isolates from Shrubby Legumes Endemic to the Canary Islands." Systematic and Applied Microbiology 23, no. 3 (October 2000): 418–25. http://dx.doi.org/10.1016/s0723-2020(00)80073-9.

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Lu, Shi-En, and Dennis C. Gross. "Drippy Pod of White Lupine: A New Bacterial Disease Caused by a Pathovar of Brenneria quercina." Plant Disease 94, no. 12 (December 2010): 1431–40. http://dx.doi.org/10.1094/pdis-05-10-0365.

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Drippy pod is a unique bacterial disease of Mediterranean white lupine (Lupinus albus) that first appeared in commercial fields in Eastern Washington State in the mid-1980s. The disease is most noticeable in the field as water-soaked lesions on lupine pods that produce an abundance of whitish-colored ooze with a sticky and foamy consistency. As the disease progresses, yellowing of lupine plants occurs with ooze characteristically dripping down the infected pods and stems and solidifying. A gram-negative rod-shaped bacterium with facultative anaerobic growth was repeatedly isolated from infected lupine tissues, and subsequently confirmed by Koch's postulates to infect lupines. Physiological and biochemical tests, including the API 20E and 50CHE strip assays, showed a highly uniform phenotype for the lupine strains that was distinctive for the genus Brenneria and most closely resembled the oak pathogen Brenneria quercina. Furthermore, sequence analyses of the 16S rDNA gene and the 16S-23S intergenic region of lupine strains revealed the highest similarity (>97%) to the corresponding regions of B. quercina and less similarity to the next closest species, B. salicis. Fatty acid profiling demonstrated that lupine strains were qualitatively similar in composition to Brenneria spp., and supported placement of the drippy pod bacterium in the species B. quercina. Oak strains of B. quercina, however, did not incite drippy pod disease on lupine. Consequently, the lupine strains that cause bacterial drippy pod disease were classified as B. quercina pv. lupinicola pv. nov.

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Thayanukul, Parinda, Futoshi Kurisu, Ikuro Kasuga, and Hiroaki Furumai. "Characterization of bacterial isolates from water reclamation systems on the basis of substrate utilization patterns and regrowth potential in reclaimed water." Water Science and Technology 68, no. 7 (October 1, 2013): 1556–65. http://dx.doi.org/10.2166/wst.2013.395.

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Microbial regrowth causes problems during water reuse. Comprehensive understanding of the microorganisms that can regrow in reclaimed water and their substrate requirements are necessary. In this study, potential regrowth organisms were isolated from seven water reclamation plants in Japan. Based on 16S rDNA analysis, the isolates were grouped into 34 operational taxonomic units, belonging to Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria, Actinobacteria, Bacteroidetes, and Firmicutes. Substrate utilization profiling using Biolog microplate™ classified the isolates into four groups. Bacteria in Cluster 1 (e.g., Methylobacterium sp. and Acinetobacter sp.) mainly utilized polymers, esters, amides, and alcohol. Isolates in Cluster 2 (e.g., Flavobacterium sp. and Microbacterium sp.) preferred to utilize polymers, carbohydrates, and esters. Isolates in Cluster 3 (e.g., Pseudomonas sp. and Acidovorax sp.) mainly utilized esters, carboxylic acids, and amino acids. Isolates in Cluster 4 (e.g., Enterobacter sp. and Rhodococcus sp.) utilized carbohydrates, esters, and amino acids. All isolates grew in reclaimed water treated by sand filtration, whereas some isolates could not grow in reclaimed water treated by coagulation and ozonation. Most bacteria in the same Biolog clusters exhibited similar growth characteristics in water samples. The potential of bacteria to regrow in reclaimed water likely depended on substrate requirement.

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Hertz, Frederik Boetius, Andries E. Budding, Malieka van der Lugt-Degen, Paul H. Savelkoul, Anders Løbner-Olesen, and Niels Frimodt-Møller. "Effects of Antibiotics on the Intestinal Microbiota of Mice." Antibiotics 9, no. 4 (April 17, 2020): 191. http://dx.doi.org/10.3390/antibiotics9040191.

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Studies on human and mouse gastrointestinal microbiota have correlated the composition of the microbiota to a variety of diseases, as well as proved it vital to prevent colonization with resistant bacteria, a phenomenon known as colonization resistance. Antibiotics dramatically modify the gut community and there are examples of how antibiotic usage lead to colonization with resistant bacteria [e.g., dicloxacillin usage selecting for ESBL-producing E. coli carriage], as shown by Hertz et al. Here, we investigated the impact of five antibiotics [cefotaxime, cefuroxime, dicloxacillin, clindamycin, and ciprofloxacin] on the intestinal microbiota in mice. Five different antibiotics were each given to groups of five mice. The intestinal microbiotas were profiled by use of the IS-pro analysis; a 16S–23S rDNA interspace [IS]-region-based profiling method. For the mice receiving dicloxacillin and clindamycin, we observed dramatic shifts in dominating phyla from day 1 to day 5. Of note, diversity increased, but overall bacterial load decreased. For ciprofloxacin, cefotaxime, and cefuroxime there were few overall changes. We speculate that antibiotics with efficacy against the abundant anaerobes in the gut, particularly Bacteroidetes, can in fact be selected for resistant bacteria, disregarding the spectrum of activity.

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Journal articles on the topic '16S rDNA profiling'

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