Dataset Information

Comparison of single-nucleotide variants identified by Illumina and Oxford Nanopore technologies in the context of a potential outbreak of Shiga toxin-producing Escherichia coli.

ABSTRACT:

Background

We aimed to compare Illumina and Oxford Nanopore Technology sequencing data from the 2 isolates of Shiga toxin-producing Escherichia coli (STEC) O157:H7 to determine whether concordant single-nucleotide variants were identified and whether inference of relatedness was consistent with the 2 technologies.

Results

For the Illumina workflow, the time from DNA extraction to availability of results was ∼40 hours, whereas with the ONT workflow serotyping and Shiga toxin subtyping variant identification were available within 7 hours. After optimization of the ONT variant filtering, on average 95% of the discrepant positions between the technologies were accounted for by methylated positions found in the described 5-methylcytosine motif sequences, CC(A/T)GG. Of the few discrepant variants (6 and 7 difference for the 2 isolates) identified by the 2 technologies, it is likely that both methodologies contain false calls.

Conclusions

Despite these discrepancies, Illumina and Oxford Nanopore Technology sequences from the same case were placed on the same phylogenetic location against a dense reference database of STEC O157:H7 genomes sequenced using the Illumina workflow. Robust single-nucleotide polymorphism typing using MinION-based variant calling is possible, and we provide evidence that the 2 technologies can be used interchangeably to type STEC O157:H7 in a public health setting.

SUBMITTER: Greig DR

PROVIDER: S-EPMC6703438 | biostudies-literature | 2019 Aug

REPOSITORIES: biostudies-literature

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Publications

Comparison of single-nucleotide variants identified by Illumina and Oxford Nanopore technologies in the context of a potential outbreak of Shiga toxin-producing Escherichia coli.

Greig David R DR Jenkins Claire C Gharbia Saheer S Dallman Timothy J TJ

GigaScience 20190801 8

<h4>Background</h4>We aimed to compare Illumina and Oxford Nanopore Technology sequencing data from the 2 isolates of Shiga toxin-producing Escherichia coli (STEC) O157:H7 to determine whether concordant single-nucleotide variants were identified and whether inference of relatedness was consistent with the 2 technologies.<h4>Results</h4>For the Illumina workflow, the time from DNA extraction to availability of results was ∼40 hours, whereas with the ONT workflow serotyping and Shiga toxin subtyp ...[more]

PMID: 31433830

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Project description:Illumina MiSeq is the current standard for characterizing microbial communities in soil. The newer alternative, Oxford Nanopore Technologies MinION sequencer, is quickly gaining popularity because of the low initial cost and longer sequence reads. However, the accuracy of MinION, per base, is much lower than MiSeq (95% versus 99.9%). The effects of this difference in base-calling accuracy on taxonomic and diversity estimates remains unclear. We compared the effects of platform, primers, and bioinformatics on mock community and agricultural soil samples using short MiSeq, and short and full-length MinION 16S rRNA amplicon sequencing. For all three methods, we found that taxonomic assignments of the mock community at both the genus and species level matched expectations with minimal deviation (genus: 80.9-90.5%; species: 70.9-85.2% Bray-Curtis similarity); however, the short MiSeq with error correction (DADA2) resulted in the correct estimate of mock community species richness and much lower alpha diversity for soils. Several filtering strategies were tested to improve these estimates with varying results. The sequencing platform also had a significant influence on the relative abundances of taxa with MiSeq resulting in significantly higher abundances Actinobacteria, Chloroflexi, and Gemmatimonadetes and lower abundances of Acidobacteria, Bacteroides, Firmicutes, Proteobacteria, and Verrucomicrobia compared to the MinION platform. When comparing agricultural soils from two different sites (Fort Collins, CO and Pendleton, OR), methods varied in the taxa identified as significantly different between sites. At all taxonomic levels, the full-length MinION method had the highest similarity to the short MiSeq method with DADA2 correction with 73.2%, 69.3%, 74.1%, 79.3%, 79.4%, and 82.28% of the taxa at the phyla, class, order, family, genus, and species levels, respectively, showing similar patterns in differences between the sites. In summary, although both platforms appear suitable for 16S rRNA microbial community composition, biases for different taxa may make the comparison between studies problematic; and even with a single study (i.e., comparing sites or treatments), the sequencing platform can influence the differentially abundant taxa identified.

Dataset Information

Comparison of single-nucleotide variants identified by Illumina and Oxford Nanopore technologies in the context of a potential outbreak of Shiga toxin-producing Escherichia coli.

Background

Results

Conclusions

Publications

Comparison of single-nucleotide variants identified by Illumina and Oxford Nanopore technologies in the context of a potential outbreak of Shiga toxin-producing Escherichia coli.

Similar Datasets

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