ArticlePloS one2015
Back to Basics--The Influence of DNA Extraction and Primer Choice on Phylogenetic Analysis of Activated Sludge Communities.
Article in PloS one, 2015. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 201 papers, 4 of them syntheses that pooled it.
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Who cites it
201 citing papers in PubMed, 4 syntheses or guidelines pooled it, 558 citations in OpenAlex.
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- Characteristics of the gut microbiome in women with gestational diabetes mellitus: A systematic review.PloS one · 2022Pooled it
- Gut microbiota variations in patients diagnosed with major depressive disorder-A systematic review.Brain and behavior · 2021Pooled it
- Gut microbiota profiles of autism spectrum disorder and attention deficit/hyperactivity disorder: A systematic literature review.Gut microbes · 2020Pooled it
- Probiotics Partly Suppress the Impact of Sugar Stress on the Oral Microbiota-A Randomized, Double-Blinded, Placebo-Controlled Trial.Nutrients · 2023Trial
- Cell recovery and DNA extraction methods skew relative abundance estimations of foodborne pathogens and spoilage organisms on stainless steel surfaces.Microbiology spectrum · 2026Article
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- A Scalable and Cost-Effective In-Line Barcoding Strategy for Standardized 16S rRNA Gene Amplicon Sequencing: Performance Evaluation and Bias Assessment.Molecular ecology resources · 2026Article
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- Enhanced resistance and resilience of anaerobic digestion microbiome after single and dual short-term disturbances.Scientific reports · 2026Article
- Impact of Isosorbide Diesters from Coconut and Sunflower Fatty Acids on Pediatric Atopic Dermatitis and the Skin Microbiome: A Randomized, Double-Blind, Vehicle-Controlled Trial.Journal of clinical medicine · 2026Article
- Oral health and oral microbiota composition in patients with Parkinson's disease: a case‒control study.Journal of oral microbiology · 2026Article
- Microbiota-Metabolome Alterations Suggest Pro-Inflammatory Phenotype in Congenital Central Hypoventilation Syndrome.International journal of microbiology · 2026Article
- Diet supplementation with the macroalgaeFrontiers in nutrition · 2026Article
- Microbial community patterns in two geochemically contrasting zones within the alkaline lake Bagno dell'Acqua (Pantelleria Island, Italy).Frontiers in microbiology · 2026Article
- Environmental distribution, physiology and genomic adaptations of arctic ammonia-oxidizing archaea.Frontiers in microbiology · 2026Article
- State-Specific Extraction of Environmental DNA: Spike-and-Recovery Controls to Validate and Optimise Extraction Protocols.Environmental microbiology · 2025Article
- Mucosal washes are useful for sampling intestinal mucus-associated microbiota despite low biomass.Gut microbes · 2025Article
141 more citing papers are in PubMed but not listed here.
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Authors and funding
5 authors at 1 institution in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
DNA extraction and primer choice have a large effect on the observed community structure in all microbial amplicon sequencing analyses. Although the biases are well known, no comprehensive analysis has been conducted in activated sludge communities. In this study we systematically explored the impact of a number of parameters on the observed microbial community: bead beating intensity, primer choice, extracellular DNA removal, and various PCR settings. In total, 176 samples were subjected to 16S rRNA amplicon sequencing, and selected samples were investigated through metagenomics and metatranscriptomics. Quantitative fluorescence in situ hybridization was used as a DNA extraction-independent method for qualitative comparison. In general, an effect on the observed community was found on all parameters tested, although bead beating and primer choice had the largest effect. The effect of bead beating intensity correlated with cell-wall strength as seen by a large increase in DNA from Gram-positive bacteria (up to 400%). However, significant differences were present at lower phylogenetic levels within the same phylum, suggesting that additional factors are at play. The best primer set based on in silico analysis was found to underestimate a number of important bacterial groups. For 16S rRNA gene analysis in activated sludge we recommend using the FastDNA SPIN Kit for Soil with four times the normal bead beating and V1-3 primers.
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