Comparative analysis of two 16S rRNA gene-based PCR primer sets provides insight into the diversity distribution patterns of anammox bacteria in different environments

2015 ◽  
Vol 99 (19) ◽  
pp. 8163-8176 ◽  
Author(s):  
Shuailong Wang ◽  
Yiguo Hong ◽  
Jiapeng Wu ◽  
Xiang-Rong Xu ◽  
Liying Bin ◽  
...  
Keyword(s):  
Comparative Analysis ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Distribution Patterns ◽  
Anammox Bacteria ◽  
Rrna Gene ◽  
Primer Sets ◽  
Insight Into ◽  
Pcr Primer

scholarly journals Molecular Evidence for the Broad Distribution of Anaerobic Ammonium-Oxidizing Bacteria in Freshwater and Marine Sediments

2006 ◽  
Vol 72 (10) ◽  
pp. 6829-6832 ◽  
Author(s):  
C. Ryan Penton ◽  
Allan H. Devol ◽  
James M. Tiedje
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Marine Sediments ◽  
Permafrost Soil ◽  
Anammox Bacteria ◽  
Molecular Evidence ◽  
Rrna Gene ◽  
Gene Sequences ◽  
16S Rrna Gene Sequences ◽  
Primer Sets

ABSTRACT Previously available primer sets for detecting anaerobic ammonium-oxidizing (anammox) bacteria are inefficient, resulting in a very limited database of such sequences, which limits knowledge of their ecology. To overcome this limitation, we designed a new primer set that was 100% specific in the recovery of ∼700-bp 16S rRNA gene sequences with >96% homology to the “Candidatus Scalindua” group of anammox bacteria, and we detected this group at all sites studied, including a variety of freshwater and marine sediments and permafrost soil. A second primer set was designed that exhibited greater efficiency than previous primers in recovering full-length (1,380-bp) sequences related to “Ca. Scalindua,” “Candidatus Brocadia,” and “Candidatus Kuenenia.” This study provides evidence for the widespread distribution of anammox bacteria in that it detected closely related anammox 16S rRNA gene sequences in 11 geographically and biogeochemically diverse freshwater and marine sediments.

scholarly journals Hydrazine Synthase, a Unique Phylomarker with Which To Study the Presence and Biodiversity of Anammox Bacteria

2011 ◽  
Vol 78 (3) ◽  
pp. 752-758 ◽  
Author(s):  
Harry R. Harhangi ◽  
Mathilde Le Roy ◽  
Theo van Alen ◽  
Bao-lan Hu ◽  
Joost Groen ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Environmental Samples ◽  
Anammox Bacteria ◽  
Rrna Gene ◽  
A Subunit ◽  
Primer Sets ◽  
Marine Environmental ◽  
The 16S Rrna Gene ◽  
Wastewater Treatment Systems

ABSTRACTAnaerobic ammonium-oxidizing (anammox) bacteria play an important role in the biogeochemical cycling of nitrogen. They derive their energy for growth from the conversion of ammonium and nitrite into dinitrogen gas in the complete absence of oxygen. Several methods have been used to detect the presence and activity of anammox bacteria in the environment, including 16S rRNA gene-based approaches. The use of the 16S rRNA gene to study biodiversity has the disadvantage that it is not directly related to the physiology of the target organism and that current primers do not completely capture the anammox diversity. Here we report the development of PCR primer sets targeting a subunit of the hydrazine synthase (hzsA), which represents a unique phylogenetic marker for anammox bacteria. The tested primers were able to retrievehzsAgene sequences from anammox enrichment cultures, full-scale anammox wastewater treatment systems, and a variety of freshwater and marine environmental samples, covering all known anammox genera.

scholarly journals Comparative Analysis of Bacterial Diversity in Freshwater Sediment of a Shallow Eutrophic Lake by Molecular and Improved Cultivation-Based Techniques

2005 ◽  
Vol 71 (4) ◽  
pp. 2162-2169 ◽  
Author(s):  
Hideyuki Tamaki ◽  
Yuji Sekiguchi ◽  
Satoshi Hanada ◽  
Kazunori Nakamura ◽  
Nakao Nomura ◽  
...  
Keyword(s):  
Comparative Analysis ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Bacterial Diversity ◽  
Clone Library ◽  
Eutrophic Lake ◽  
Freshwater Sediment ◽  
Gene Clone ◽  
Rrna Gene ◽  
Shallow Eutrophic Lake

ABSTRACT Comparative analysis of bacterial diversity in freshwater sediment collected from a shallow eutrophic lake was performed by using 16S rRNA gene clone library and improved cultivation-based techniques. Our study demonstrated that the use of gellan gum as a gelling reagent instead of agar was more effective at increasing culturability, cultivating a diverse array of novel microbes, and reducing the gaps of the results between molecular and cultivation-based analyses.

scholarly journals Anaerobic microbial LCFA degradation in bioreactors

2008 ◽  
Vol 57 (3) ◽  
pp. 439-444 ◽  
Author(s):  
D. Z. Sousa ◽  
M. A. Pereira ◽  
J. I. Alves ◽  
H. Smidt ◽  
A. J. M Stams ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Rrna Gene ◽  
Cloning And Sequencing ◽  
16S Rrna Gene Analysis ◽  
Sulfate Reducing ◽  
Bacterial Consortia ◽  
Enrichment Cultures ◽  
The Family ◽  
Insight Into

This paper reviews recent results obtained on long-chain fatty acids (LCFA) anaerobic degradation. Two LCFA were used as model substrates: oleate, a mono-unsaturated LCFA, and palmitate, a saturated LCFA, both abundant in LCFA-rich wastewaters. 16S rRNA gene analysis of sludge samples submitted to continuous oleate- and palmitate-feeding followed by batch degradation of the accumulated LCFA demonstrated that bacterial communities were dominated by members of the Clostridiaceae and Syntrophomonadaceae families. Archaeal populations were mainly comprised of hydrogen-consuming microorganisms belonging to the genus Methanobacterium, and acetate-utilizers from the genera Methanosaeta and Methanosarcina. Enrichment cultures growing on oleate and palmitate, in the absence or presence of sulfate, gave more insight into the major players involved in the degradation of unsaturated and saturated LCFA. Syntrophomonas-related species were identified as predominant microorganisms in all the enrichment cultures. Microorganisms clustering within the family Syntrophobacteraceae were identified in the methanogenic and sulfate-reducing enrichments growing on palmitate. Distinct bacterial consortia were developed in oleate and palmitate enrichments, and observed differences might be related to the different degrees of saturation of these two LCFA. A new obligately syntrophic bacterium, Syntrophomonas zehnderi, was isolated from an oleate-degrading culture and its presence in oleate-degrading sludges detected by 16S rRNA gene cloning and sequencing.

A PCR-based specific assay reveals a population of bacteria within the Chloroflexi associated with the reductive dehalogenation of polychlorinated biphenyls

Microbiology ◽  
2005 ◽  
Vol 151 (6) ◽  
pp. 2039-2046 ◽  
Author(s):  
Joy E. M. Watts ◽  
Sonja K. Fagervold ◽  
Harold D. May ◽  
Kevin R. Sowers
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Polychlorinated Biphenyls ◽  
Reductive Dehalogenation ◽  
Rrna Gene ◽  
Gene Sequences ◽  
16S Rrna Gene Sequences ◽  
Specific Pcr ◽  
Pure Cultures ◽  
Primer Sets

Polychlorinated biphenyls (PCBs) accumulate and persist in sediments posing a risk to human health and the environment. Highly chlorinated PCBs are reductively dechlorinated in anaerobic sediments and two bacteria, designated o-17 and DF-1, from a novel phylogenetic group that reductively dechlorinate PCBs have recently been identified. However, there is a paucity of knowledge about the distribution, diversity and ecology of PCB-dechlorinating bacteria due to difficulty in obtaining pure cultures and the lack of detection by universal PCR 16S rRNA gene primer sets in sediments. A specific PCR primer was developed and optimized for detection of o-17/DF-1 and other closely related bacteria in the environment. Using this primer set it was determined that bacteria of this group were enriched in sediment microcosms from Baltimore Harbour concurrent with active dechlorination of 2,2′,3,4,4′,5′-hexachlorobiphenyl. Additional 16S rRNA gene sequences that had high levels of similarity to described PCB dechlorinators were detected in sediments from the Elizabeth River tributary of Chesapeake Bay, which had confirmed PCB-dechlorinating activities. Phylogenetic comparison of these detected 16S rRNA gene sequences revealed a relatively diverse group of organisms within the dehalogenating Chloroflexi that are distinct from the Dehalococcoides spp. Results from this study indicate that reductive PCB dechlorination activity may be catalysed by a previously undescribed group of micro-organisms that appear to be prevalent in PCB-impacted sites.

scholarly journals Diversity and Distribution of Planctomycetes and Related Bacteria in the Suboxic Zone of the Black Sea

2006 ◽  
Vol 72 (4) ◽  
pp. 3079-3083 ◽  
Author(s):  
John Kirkpatrick ◽  
Brian Oakley ◽  
Clara Fuchsman ◽  
Sujatha Srinivasan ◽  
James T. Staley ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Black Sea ◽  
Gene Sequence ◽  
The Black Sea ◽  
Anammox Bacteria ◽  
Rrna Gene ◽  
Sequence Information ◽  
Rrna Gene Sequence ◽  
Suboxic Zone

ABSTRACT Samples from six depths of the Black Sea's suboxic zone were analyzed for 16S rRNA gene sequence information. A gradient in phylotype diversity was found. The distributions of known anaerobic ammonium oxidation (anammox) bacteria, many unknown Planctomycetes, and other phylotypes were examined in relation to the local nutrient and redox conditions.

scholarly journals Impact of DNA purification method and primer selection on 16S rRNA gene metabarcoding on wine

OENO One ◽  
2019 ◽  
Vol 53 (3) ◽  
Author(s):  
Francesco Cerutti ◽  
Diego Cravero ◽  
Antonella Costantini ◽  
Laura Pulcini ◽  
Paola Modesto ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Alternative Methods ◽  
Dna Purification ◽  
Rrna Gene ◽  
Purification Method ◽  
Main Species ◽  
Purification Methods ◽  
Primer Sets ◽  
The 16S Rrna Gene

Aim: The high-throughput sequencing methods have revolutionized the study of the microbiota in different matrices including those of the grapevine production chain. DNA extraction is a crucial step in the sample processing. In this study, we compared different DNA purification methods and two primer sets for 16S rRNA gene metabarcoding to evaluate the best protocol to explore the wine microbiota by metabarcoding.Methods and results: We collected a wine from Barbera grapes after malolactic fermentation previously inoculated by Oenococcus oeni starter. The same sample was used to evaluate the best performing protocol to study the wine microbiota. DNA was purified using nine different methods and then amplified for the 16S rRNA gene with two primer sets (according to Illumina or Earth Microbiome Project protocols). The obtained amplicons were then sequenced in a single sequencing session on an Illumina MiSeq. We evaluated the best protocol considering DNA concentration and purity, alpha (Observed species) and beta diversity from metabarcoding analysis.The sequencing generated 36,031,756 reads in total. Although no statistically significant difference was observed between purification methods or primer sets, better results were obtained with phenol-chloroform DNA purification combined to Earth Microbiome Project primers.Metabarcoding was able to highlight the domination of the inoculum, O. oeni, representing the main species of the analyzed wine microbiota.Conclusion: Our data show that, for the tested wine, metabarcoding output is more influenced by the primer set than by the DNA purification method. Moreover, the metabarcoding detected that O. oeni represents the main species, evidencing the domination of the inoculum done with lyophilized commercial preparation of this species. Other lactic acid bacteria are present at a much lower abundance.Significance and impact of the study: This is the first report applying the 16S rRNA gene metabarcoding to study the microbiota of wine. For this reason, the evaluation of alternative methods for DNA processing is essential for future research using this innovative methodology.

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