Integrative analysis of environmental sequences using MEGAN4

Huson, Daniel H.; Mitra, Suparna; Ruscheweyh, Hans‐Joachim; Weber, Nico; Schuster, Stephan C.

doi:10.1101/gr.120618.111

Cited by 1,265 publications

(1,072 citation statements)

References 37 publications

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“…Taxonomy was assigned to each OTU using MEGAN version 5.10.5 (Huson, Mitra, Ruscheweyh, Weber, & Schuster, 2011) based on 50 hits per OTU generated by BLASTN searches against the NCBI “nt” database excluding environmental sequences (downloaded June 2016). The lowest common ancestor (LCA) algorithm used to assign taxonomy in MEGAN only assigns an OTU to species (or other taxonomic level) if no other species (or taxon) has a blast hit within a specified percentage of the score of the best hit (top percent parameter).…”

Section: Methodsmentioning

confidence: 99%

Effect of marker choice and thermal cycling protocol on zooplankton DNA metabarcoding studies

Clarke

Beard

Swadling

et al. 2017

Ecology and Evolution

156

193

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DNA metabarcoding is a promising approach for rapidly surveying biodiversity and is likely to become an important tool for measuring ecosystem responses to environmental change. Metabarcoding markers need sufficient taxonomic coverage to detect groups of interest, sufficient sequence divergence to resolve species, and will ideally indicate relative abundance of taxa present. We characterized zooplankton assemblages with three different metabarcoding markers (nuclear 18S rDNA, mitochondrial COI, and mitochondrial 16S rDNA) to compare their performance in terms of taxonomic coverage, taxonomic resolution, and correspondence between morphology‐ and DNA‐based identification. COI amplicons sequenced on separate runs showed that operational taxonomic units representing >0.1% of reads per sample were highly reproducible, although slightly more taxa were detected using a lower annealing temperature. Mitochondrial COI and nuclear 18S showed similar taxonomic coverage across zooplankton phyla. However, mitochondrial COI resolved up to threefold more taxa to species compared to 18S. All markers revealed similar patterns of beta‐diversity, although different taxa were identified as the greatest contributors to these patterns for 18S. For calanoid copepod families, all markers displayed a positive relationship between biomass and sequence reads, although the relationship was typically strongest for 18S. The use of COI for metabarcoding has been questioned due to lack of conserved primer‐binding sites. However, our results show the taxonomic coverage and resolution provided by degenerate COI primers, combined with a comparatively well‐developed reference sequence database, make them valuable metabarcoding markers for biodiversity assessment.

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Section: Methodsmentioning

confidence: 99%

Effect of marker choice and thermal cycling protocol on zooplankton DNA metabarcoding studies

Clarke

Beard

Swadling

et al. 2017

Ecology and Evolution

156

193

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“…MEGAN version 5.6.3. (Huson, Mitra, Ruscheweyh, Weber, & Schuster, 2011) was used to screen out unknowns, unidentified sequences and those not resolved to order with the LCA parameters recommended by Salinas‐Ramos, Herrera Montalvo, León‐Regagnon, Arrizabalaga‐Escudero, and Clare (2015). The remaining identified MOTU were used for statistical analysis of diet.…”

Section: Methodsmentioning

confidence: 99%

Spatiotemporal and demographic variation in the diet of New Zealand lesser short‐tailed bats (Mystacina tuberculata)

Czenze

Tucker

Clare

et al. 2018

Ecology and Evolution

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Variation in the diet of generalist insectivores can be affected by site‐specific traits including weather, habitat, and season, as well as demographic traits such as reproductive status and age. We used molecular methods to compare diets of three distinct New Zealand populations of lesser short‐tailed bats, Mystacina tuberculata. Summer diets were compared between a southern cold‐temperate (Eglinton) and a northern population (Puroera). Winter diets were compared between Pureora and a subtropical offshore island population (Hauturu). This also permitted seasonal diet comparisons within the Pureora population. Lepidoptera and Diptera accounted for >80% of MOTUs identified from fecal matter at each site/season. The proportion of orders represented within prey and the Simpson diversity index, differed between sites and seasons within the Pureora population. For the Pureora population, the value of the Simpson diversity index was higher in summer than winter and was higher in Pureora compared to Eglinton. Summer Eglinton samples revealed that juvenile diets appeared to be more diverse than other demographic groups. Lactating females had the lowest dietary diversity during summer in Pureora. In Hauturu, we found a significant negative relationship between mean ambient temperature and prey richness. Our data suggest that M. tuberculata incorporate a narrower diversity of terrestrial insects than previously reported. This provides novel insights into foraging behavior and ecological interactions within different habitats. Our study is the first from the Southern Hemisphere to use molecular techniques to examine spatiotemporal variation in the diet of a generalist insectivore that inhabits a contiguous range with several habitat types and climates.

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“…Each read was taxonomically assigned down to the species level using a 90% minimum confidence threshold. A phylogenetic tree was generated from a unique sequences fasta file using the Megan software, resulting in a Newick tree format [28]. …”

Section: Methodsmentioning

confidence: 99%

In situ substrate-formed biofilms using IDODS mimic supragingival tooth-formed biofilms

Tomás

Prada-López

Quintas

et al. 2018

Journal of Oral Microbiology

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This study aimed to compare the bacterial viability and diversity of a substrate-formed biofilm (SF-biofilm) in situ to a supragingival tooth-formed biofilm (TF-biofilm) in the same group of individuals. The impact of the device/disc position and toothbrushing during the formation of SF-biofilm was also assessed. Two tests were run. In test 1, 15 volunteers wore two hemi-splints carrying six discs of human enamel, glass, and hydroxyapatite for 2 days, and were instructed to not perform any oral hygiene measure. Biofilm samples were collected from the substrates and the contralateral tooth and were analysed using CLSM. In five volunteers, half of the biofilm present on the discs and their contralateral teeth were scraped and analysed using 16S pyrosequencing. In test 2, the microscopic analysis was repeated only on the SF-biofilm samples, and the volunteers were allowed to brush their teeth. Multivariate analyses revealed that the donors had a significant effect on the composition of the biofilm, confirming its subject-dependent character. The bacterial composition of the SF-biofilm was similar to the TF-biofilm, with significant differential abundance detected in very few taxa of low abundance. The toothbrushing during the formation of SF-biofilm was the only factor that conditioned the thickness or bacterial viability.

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Integrative analysis of environmental sequences using MEGAN4

Cited by 1,265 publications

References 37 publications

Effect of marker choice and thermal cycling protocol on zooplankton DNA metabarcoding studies

Effect of marker choice and thermal cycling protocol on zooplankton DNA metabarcoding studies

Spatiotemporal and demographic variation in the diet of New Zealand lesser short‐tailed bats (Mystacina tuberculata)

In situ substrate-formed biofilms using IDODS mimic supragingival tooth-formed biofilms

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