Novel multiplex real-time PCR assays reveal a high prevalence of diarrhoeagenic Escherichia coli pathotypes in healthy and diarrhoeal children in the south of Vietnam.
Adolescent
Child
Child, Preschool
Diarrhea
/ epidemiology
Enteropathogenic Escherichia coli
/ classification
Escherichia coli Infections
/ diagnosis
Female
Humans
Infant
Infant, Newborn
Male
Multiplex Polymerase Chain Reaction
Prevalence
Real-Time Polymerase Chain Reaction
/ methods
Sensitivity and Specificity
Vietnam
/ epidemiology
Virulence Factors, Bordetella
Co-infection
Diarrhoea children
EAEC
EHEC
EIEC
EPEC
ETEC
Healthy children
Multiplex real-time PCR
Journal
BMC microbiology
ISSN: 1471-2180
Titre abrégé: BMC Microbiol
Pays: England
ID NLM: 100966981
Informations de publication
Date de publication:
03 07 2020
03 07 2020
Historique:
received:
18
03
2020
accepted:
25
06
2020
entrez:
5
7
2020
pubmed:
6
7
2020
medline:
20
7
2021
Statut:
epublish
Résumé
Diarrhoeagenic Escherichia coli (DEC) infections are common in children in low-middle income countries (LMICs). However, detecting the various DEC pathotypes is complex as they cannot be differentiated by classical microbiology. We developed four multiplex real-time PCR assays were to detect virulence markers of six DEC pathotypes; specificity was tested using DEC controls and other enteric pathogens. PCR amplicons from the six E. coli pathotypes were purified and amplified to be used to optimize PCR reactions and to calculate reproducibility. After validation, these assays were applied to clinical samples from healthy and diarrhoeal Vietnamese children and associated with clinical data. The multiplex real-time PCRs were found to be reproducible, and specific. At least one DEC variant was detected in 34.7% (978/2815) of the faecal samples from diarrhoeal children; EAEC, EIEC and atypical EPEC were most frequent Notably, 41.2% (205/498) of samples from non-diarrhoeal children was positive with a DEC pathotype. In this population, only EIEC, which was detected in 34.3% (99/289) of diarrhoeal samples vs. 0.8% (4/498) non-diarrhoeal samples (p < 0.001), was significantly associated with diarrhoea. Multiplex real-time PCR when applied to clinical samples is an efficient and high-throughput approach to DEC pathotypes. This approach revealed high carriage rates of DEC pathotypes among Vietnamese children. We describe a novel diagnostic approach for DEC, which provides baseline data for future surveillance studies assessing DEC burden in LMICs.
Sections du résumé
BACKGROUND
Diarrhoeagenic Escherichia coli (DEC) infections are common in children in low-middle income countries (LMICs). However, detecting the various DEC pathotypes is complex as they cannot be differentiated by classical microbiology. We developed four multiplex real-time PCR assays were to detect virulence markers of six DEC pathotypes; specificity was tested using DEC controls and other enteric pathogens. PCR amplicons from the six E. coli pathotypes were purified and amplified to be used to optimize PCR reactions and to calculate reproducibility. After validation, these assays were applied to clinical samples from healthy and diarrhoeal Vietnamese children and associated with clinical data.
RESULTS
The multiplex real-time PCRs were found to be reproducible, and specific. At least one DEC variant was detected in 34.7% (978/2815) of the faecal samples from diarrhoeal children; EAEC, EIEC and atypical EPEC were most frequent Notably, 41.2% (205/498) of samples from non-diarrhoeal children was positive with a DEC pathotype. In this population, only EIEC, which was detected in 34.3% (99/289) of diarrhoeal samples vs. 0.8% (4/498) non-diarrhoeal samples (p < 0.001), was significantly associated with diarrhoea. Multiplex real-time PCR when applied to clinical samples is an efficient and high-throughput approach to DEC pathotypes.
CONCLUSIONS
This approach revealed high carriage rates of DEC pathotypes among Vietnamese children. We describe a novel diagnostic approach for DEC, which provides baseline data for future surveillance studies assessing DEC burden in LMICs.
Identifiants
pubmed: 32620076
doi: 10.1186/s12866-020-01878-5
pii: 10.1186/s12866-020-01878-5
pmc: PMC7333254
doi:
Substances chimiques
Virulence Factors, Bordetella
0
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
192Subventions
Organisme : Wellcome Trust
Pays : United Kingdom
Organisme : Wellcome Trust
ID : 215515/Z/19/Z
Pays : United Kingdom
Commentaires et corrections
Type : ErratumIn
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