In Silico  Genotyping of  Escherichia coli  Isolates for Extraintestinal Virulence Genes by Use of Whole-Genome Sequencing Data

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Abstract

ABSTRACT

Extraintestinal pathogenic Escherichia coli (ExPEC) is the leading cause in humans of urinary tract infection and bacteremia. The previously published web tool VirulenceFinder ( http://cge.cbs.dtu.dk/services/VirulenceFinder/) uses whole-genome sequencing (WGS) data for in silico characterization of E. coli isolates and enables researchers and clinical health personnel to quickly extract and interpret virulence-relevant information from WGS data. In this study, 38 ExPEC-associated virulence genes were added to the existing E. coli VirulenceFinder database. In total, 14,441 alleles were downloaded. A total of 1,890 distinct alleles were added to the database after removal of redundant sequences and analysis of the remaining alleles for open reading frames (ORFs). The database now contains 139 genes—of which 44 are related to ExPEC—and 2,826 corresponding alleles. Construction of the database included validation against 27 primer pairs from previous studies, a search for serotype-specific P fimbriae papA alleles, and a BLASTn confirmation of seven genes ( etsC, iucC, kpsE, neuC, sitA, tcpC, and terC) not covered by the primers. The augmented database was evaluated using (i) a panel of nine control strains and (ii) 288 human-source E. coli strains classified by PCR as ExPEC and non-ExPEC. We observed very high concordance (average, 93.4%) between PCR and WGS findings, but WGS identified more alleles. In conclusion, the addition of 38 ExPEC-associated genes and the associated alleles to the E. coli VirulenceFinder database allows for a more complete characterization of E. coli isolates based on WGS data, which has become increasingly important considering the plasticity of the E. coli genome.

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Rapid and precise alignment of raw reads against redundant databases with KMA

Philip T L C Clausen, Frank Aarestrup, Ole Lund (2018)

Background As the cost of sequencing has declined, clinical diagnostics based on next generation sequencing (NGS) have become reality. Diagnostics based on sequencing will require rapid and precise mapping against redundant databases because some of the most important determinants, such as antimicrobial resistance and core genome multilocus sequence typing (MLST) alleles, are highly similar to one another. In order to facilitate this, a novel mapping method, KMA (k-mer alignment), was designed. KMA is able to map raw reads directly against redundant databases, it also scales well for large redundant databases. KMA uses k-mer seeding to speed up mapping and the Needleman-Wunsch algorithm to accurately align extensions from k-mer seeds. Multi-mapping reads are resolved using a novel sorting scheme (ConClave scheme), ensuring an accurate selection of templates. Results The functionality of KMA was compared with SRST2, MGmapper, BWA-MEM, Bowtie2, Minimap2 and Salmon, using both simulated data and a dataset of Escherichia coli mapped against resistance genes and core genome MLST alleles. KMA outperforms current methods with respect to both accuracy and speed, while using a comparable amount of memory. Conclusion With KMA, it was possible map raw reads directly against redundant databases with high accuracy, speed and memory efficiency. Electronic supplementary material The online version of this article (10.1186/s12859-018-2336-6) contains supplementary material, which is available to authorized users.

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ClermonTyping: an easy-to-use and accurate in silico method for Escherichia genus strain phylotyping

Johann Beghain, Antoine Bridier-Nahmias, Hervé Le Nagard … (2018)

The genus Escherichia is composed of Escherichia albertii, E. fergusonii, five cryptic Escherichia clades and E. coli sensu stricto. Furthermore, the E. coli species can be divided into seven main phylogroups termed A, B1, B2, C, D, E and F. As specific lifestyles and/or hosts can be attributed to these species/phylogroups, their identification is meaningful for epidemiological studies. Classical phenotypic tests fail to identify non-sensu stricto E. coli as well as phylogroups. Clermont and colleagues have developed PCR assays that allow the identification of most of these species/phylogroups, the triplex/quadruplex PCR for E. coli phylogroup determination being the most popular. With the growing availability of whole genome sequences, we have developed the ClermonTyping method and its associated web-interface, the ClermonTyper, that allows a given strain sequence to be assigned to E. albertii, E. fergusonii, Escherichia clades I–V, E. coli sensu stricto as well as to the seven main E. coli phylogroups. The ClermonTyping is based on the concept of in vitro PCR assays and maintains the principles of ease of use and speed that prevailed during the development of the in vitro assays. This in silico approach shows 99.4 % concordance with the in vitro PCR assays and 98.8 % with the Mash genome-clustering tool. The very few discrepancies result from various errors occurring mainly from horizontal gene transfers or SNPs in the primers. We propose the ClermonTyper as a freely available resource to the scientific community at: http://clermontyping.iame-research.center/.

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Medical and economic impact of extraintestinal infections due to Escherichia coli: focus on an increasingly important endemic problem.

Thomas Russo, James Johnson (2003)

Escherichia coli is probably the best-known bacterial species and one of the most frequently isolated organisms from clinical specimens. Despite this, underappreciation and misunderstandings exist among medical professionals and the lay public alike regarding E. coli as an extraintestinal pathogen. Underappreciated features include (i) the wide variety of extraintestinal infections E. coli can cause, (ii) the high incidence and associated morbidity, mortality, and costs of these diverse clinical syndromes, (iii) the pathogenic potential of different groups of E. coli strains for causing intestinal versus extraintestinal disease, and (iv) increasing antimicrobial resistance. In this era in which health news often sensationalizes uncommon infection syndromes or pathogens, the strains of E. coli that cause extraintestinal infection are an increasingly important endemic problem and underappreciated "killers". Billions of health care dollars, millions of work days, and hundreds of thousands of lives are lost each year to extraintestinal infections due to E. coli. New treatments and prevention measures will be needed for improved outcomes and a diminished disease burden.

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Author and article information

Contributors

John P. Dekker: Role: Editor

Journal

Journal ID (nlm-ta): J Clin Microbiol

Journal ID (iso-abbrev): J. Clin. Microbiol

Journal ID (hwp): jcm

Journal ID (pmc): jcm

Journal ID (publisher-id): JCM

Title: Journal of Clinical Microbiology

Publisher: American Society for Microbiology (1752 N St., N.W., Washington, DC )

ISSN (Print): 0095-1137

ISSN (Electronic): 1098-660X

Publication date (Electronic preprint): 15 July 2020

Publication date (Electronic): 22 September 2020

Publication date Collection: October 2020

Publication date PMC-release: 22 September 2020

Volume: 58

Issue: 10

Electronic Location Identifier: e01269-20

Affiliations

[a ]Research Group for Genomic Epidemiology, Technical University of Denmark, Kongens Lyngby, Denmark

[b ]Minneapolis Veterans Affairs Health Care System, Minneapolis, Minnesota, USA

[c ]University of Minnesota, Minneapolis, Minnesota, USA

[d ]The International Centre for Reference and Research on Escherichia and Klebsiella, Department of Bacteria, Parasites and Fungi, Statens Serum Institut, Copenhagen, Denmark

National Institute of Allergy and Infectious Diseases

Author notes

Address correspondence to Flemming Scheutz, fsc@ 123456ssi.dk .

Citation Malberg Tetzschner AM, Johnson JR, Johnston BD, Lund O, Scheutz F. 2020. In silico genotyping of Escherichia coli isolates for extraintestinal virulence genes by use of whole-genome sequencing data. J Clin Microbiol 58:e01269-20. https://doi.org/10.1128/JCM.01269-20.

Author information

Brian D. Johnston https://orcid.org/0000-0001-7347-9276

Ole Lund https://orcid.org/0000-0003-1108-0491

Flemming Scheutz https://orcid.org/0000-0002-3931-4846

Article

Publisher ID: 01269-20

DOI: 10.1128/JCM.01269-20

PMC ID: 7512150

PubMed ID: 32669379

SO-VID: 5eb1387b-7dc6-4426-ae6b-dc5a7cf56104

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This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license.

History

Date received : 25 May 2020

Date revision requested : 23 June 2020

Date accepted : 8 July 2020

Page count

supplementary-material: 2, Figures: 0, Tables: 3, Equations: 0, References: 36, Pages: 13, Words: 9551

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cover-date October 2020

ScienceOpen disciplines: Microbiology & Virology

Keywords: expec,in silico,virulence typing,whole-genome sequencing

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ScienceOpen disciplines: Microbiology & Virology

Keywords: expec, in silico, virulence typing, whole-genome sequencing

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In Silico Genotyping of Escherichia coli Isolates for Extraintestinal Virulence Genes by Use of Whole-Genome Sequencing Data

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Microbial Genomics

Most cited references 27

Rapid and precise alignment of raw reads against redundant databases with KMA

ClermonTyping: an easy-to-use and accurate in silico method for Escherichia genus strain phylotyping

Medical and economic impact of extraintestinal infections due to Escherichia coli: focus on an increasingly important endemic problem.

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Cited by 105

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