Replication, lies and lesser‐known truths regarding experimental design in environmental microbiology

Lennon, Jay T.

doi:10.1111/j.1462-2920.2011.02445.x

Cited by 42 publications

(25 citation statements)

References 20 publications

(22 reference statements)

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“…Also, replicated samples are not available for the vast majority of studies on microbial communities using molecular methods that have high cost and time-consuming nature (Prosser 2010). This limitation does not necessarily restrict microbial diversity analyses nor the power of comparative statistical tests, given that appropriate methods as well as hypotheses are applied (Lennon 2011;Smetia et al 2013). Moreover, pooled samples could reduce the effects of fine scale spatial heterogeneity on microbial communities (Berthrong et al 2013).…”

Section: Soil Samplingmentioning

confidence: 98%

Long-term oil contamination causes similar changes in microbial communities of two distinct soils

Liao

Wang

Jiang

et al. 2015

Appl Microbiol Biotechnol

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Since total petroleum hydrocarbons (TPH) are toxic and persistent in environments, studying the impact of oil contamination on microbial communities in different soils is vital to oil production engineering, effective soil management and pollution control. This study analyzed the impact of oil contamination on the structure, activity and function in carbon metabolism of microbial communities of Chernozem soil from Daqing oil field and Cinnamon soil from Huabei oil field through both culture-dependent techniques and a culture-independent technique-pyrosequencing. Results revealed that pristine microbial communities in these two soils presented disparate patterns, where Cinnamon soil showed higher abundance of alkane, (polycyclic aromatic hydrocarbons) PAHs and TPH degraders, number of cultivable microbes, bacterial richness, bacterial biodiversity, and stronger microbial activity and function in carbon metabolism than Chernozem soil. It suggested that complicated properties of microbes and soils resulted in the difference in soil microbial patterns. However, the changes of microbial communities caused by oil contamination were similar in respect of two dominant phenomena. Firstly, the microbial community structures were greatly changed, with higher abundance, higher bacterial biodiversity, occurrence of Candidate_division_BRC1 and TAO6, disappearance of BD1-5 and Candidate_division_OD1, dominance of Streptomyces, higher percentage of hydrocarbon-degrading groups, and lower percentage of nitrogen-transforming groups. Secondly, microbial activity and function in carbon metabolism were significantly enhanced. Based on the characteristics of microbial communities in the two soils, appropriate strategy for in situ bioremediation was provided for each oil field. This research underscored the usefulness of combination of culture-dependent techniques and next-generation sequencing techniques both to unravel the microbial patterns and understand the ecological impact of contamination.

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Section: Soil Samplingmentioning

confidence: 98%

Long-term oil contamination causes similar changes in microbial communities of two distinct soils

Liao

Wang

Jiang

et al. 2015

Appl Microbiol Biotechnol

View full text Add to dashboard Cite

show abstract

“…The collecting of field samples to cover the targeted variation and enable statistically robust conclusions at the desired scale of inference represents a major challenge, and optimal strategies concerning the number and spatial distribution of samples have been discussed extensively (Petersen et al ., 2005; Prosser, 2010; Lennon, 2011). Collection of fungal samples calls for some additional consideration, because of the indeterminate growth of mycelia and the multitude of contrasting morphologies and trophic strategies that coexist and interact in fungal communities.…”

Section: Samplingmentioning

confidence: 99%

Fungal community analysis by high‐throughput sequencing of amplified markers – a user's guide

et al. 2013

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Novel high-throughput sequencing methods outperform earlier approaches in terms of resolution and magnitude. They enable identification and relative quantification of community members and offer new insights into fungal community ecology. These methods are currently taking over as the primary tool to assess fungal communities of plant-associated endophytes, pathogens, and mycorrhizal symbionts, as well as free-living saprotrophs.Taking advantage of the collective experience of six research groups, we here review the different stages involved in fungal community analysis, from field sampling via laboratory procedures to bioinformatics and data interpretation. We discuss potential pitfalls, alternatives, and solutions.Highlighted topics are challenges involved in: obtaining representative DNA/RNA samples and replicates that encompass the targeted variation in community composition, selection of marker regions and primers, options for amplification and multiplexing, handling of sequencing errors, and taxonomic identification.Without awareness of methodological biases, limitations of markers, and bioinformatics challenges, large-scale sequencing projects risk yielding artificial results and misleading conclusions.

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“…Debris samples were kept at þ 4 1C during transport to the laboratory, which occurred within 12 h. Samples were then divided into two aliquots: one was stored at À 20 1C for DNA extraction and the other at þ 4 1C for the analyses of OC and pH. No replicated samples were taken at each site because multivariate regression-like analyses (see Statistical methods) were used to investigate the change in the structure of microbial communities along an ecological gradient (Lennon, 2011).…”

Section: Study Sites and Environmental Datamentioning

confidence: 99%

Bacterial community structure on two alpine debris-covered glaciers and biogeography of Polaromonas phylotypes

et al. 2013

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High-elevation cold environments are considered ideal places to test hypotheses about mechanisms of bacterial colonization and succession, and about bacterial biogeography. Debris-covered glaciers (glaciers whose ablation area is mainly covered by a continuous layer of rock debris fallen from the surrounding mountains) have never been investigated in this respect so far. We used the Illumina technology to analyse the V5 and V6 hypervariable regions of the bacterial 16S rRNA gene amplified from 38 samples collected in July and September 2009 at different distances from the terminus on two debris-covered glaciers (Miage and Belvedere-Italian Alps). Heterotrophic taxa-dominated communities and bacterial community structure changed according to ice ablation rate, organic carbon content of the debris and distance from the glacier terminus. Bacterial communities therefore change during downwards debris transport, and organic carbon of these recently exposed substrates is probably provided more by allochthonous deposition of organic matter than by primary production by autotrophic organisms. We also investigated whether phylotypes of the genus Polaromonas, which is ubiquitous in cold environments, do present a biogeographical distribution by analysing the sequences retrieved in this study together with others available in the literature. We found that the genetic distance among phylotypes increased with geographic distance; however, more focused analyses using discrete distance classes revealed that both sequences collected at sites o100 km and at sites 9400-13 500 km to each other were more similar than those collected at other distance classes. Evidences of biogeographic distribution of Polaromonas phylotypes were therefore contrasting.

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Replication, lies and lesser‐known truths regarding experimental design in environmental microbiology

Cited by 42 publications

References 20 publications

Long-term oil contamination causes similar changes in microbial communities of two distinct soils

Long-term oil contamination causes similar changes in microbial communities of two distinct soils

Fungal community analysis by high‐throughput sequencing of amplified markers – a user's guide

Bacterial community structure on two alpine debris-covered glaciers and biogeography of Polaromonas phylotypes

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