New Monoclonal Antibodies against a Novel Subtype of Shiga Toxin 1 Produced by <named-content content-type="genus-species">Enterobacter cloacae</named-content> and Their Use in Analysis of Human Serum

ABSTRACT Shiga toxin (Stx) is a major virulence factor of several bacterial pathogens that cause potentially fatal illness, including Escherichia coli and Shigella spp. The continual emergence of new subtypes of Stxs presents challenges for the clinical diagnosis of infections caused by Stx-producin...

Descripción completa

Guardado en:

Detalles Bibliográficos
Autores principales:	Craig Skinner, Stephanie Patfield, Rowaida Khalil, Qiulian Kong, Xiaohua He
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2016
Materias:	Enterobacter Shiga toxins Stx1e immunoassays monoclonal antibodies Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/0b2e261d5f4442cd85123d59dad25783
Etiquetas:	Agregar Etiqueta Sin Etiquetas, Sea el primero en etiquetar este registro!

id	oai:doaj.org-article:0b2e261d5f4442cd85123d59dad25783
record_format	dspace
spelling	oai:doaj.org-article:0b2e261d5f4442cd85123d59dad257832021-11-15T15:21:38ZNew Monoclonal Antibodies against a Novel Subtype of Shiga Toxin 1 Produced by <named-content content-type="genus-species">Enterobacter cloacae</named-content> and Their Use in Analysis of Human Serum10.1128/mSphere.00099-152379-5042https://doaj.org/article/0b2e261d5f4442cd85123d59dad257832016-02-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mSphere.00099-15https://doaj.org/toc/2379-5042ABSTRACT Shiga toxin (Stx) is a major virulence factor of several bacterial pathogens that cause potentially fatal illness, including Escherichia coli and Shigella spp. The continual emergence of new subtypes of Stxs presents challenges for the clinical diagnosis of infections caused by Stx-producing organisms. Here, we report the development of four new monoclonal antibodies (MAbs) against Stx1e, a novel subtype of Stx1 that was produced by an Enterobacter cloacae strain and had limited reactivity with existing anti-Stx1 antibodies. Western blot analysis indicates that these MAbs were Stx1 specific, bound to the A subunit, and had distinct preferences for subtypes of Stx1. Of the four MAbs, Stx1e-2 was capable of partially neutralizing cytotoxicities derived from Stx1e in Vero cells. Enzyme-linked immunosorbent assays assembled with these high-affinity MAbs detected Stx1e at concentrations as low as 4.8 pg/ml in phosphate-buffered saline and 53.6 pg/ml in spiked human serum samples and were also capable of distinguishing Stx1e-producing strains in enriched cultures. These assays may therefore have clinical value in diagnosing Stx1e-producing bacterial infection. Additionally, characteristics of Stx1e, such as the origin of stx1e genes, conditions for toxin expression, receptor binding, and cytotoxicity, were investigated with the new antibodies developed in this study. This information should be useful for further understanding the clinical significance and prevalence of Stx1e-harboring E. cloacae and other organisms. IMPORTANCE Stxs are among the most clinically important virulence factors of Shigella and enterohemorrhagic Escherichia coli. There are many varieties of Stx, and although Stx1a and Stx2a are the most common and widely distributed types of Stx, new variants of Stx are continually emerging. These new variants of Stx can be challenging to detect, since most Stx detection kits are optimized for the detection of Stx1a and Stx2a. Stx1e, recently discovered in an atypical host (Enterobacter cloacae), is undetectable by many Stx assays. To formulate new assays for the detection of Stx1e, we generated four new MAbs that recognize this Stx subtype. Using these antibodies, we generated an assay capable of detecting Stx1e at low picogram-per-milliliter concentrations. This assay is also compatible with a human serum matrix, suggesting that it may have utility for the clinical detection and diagnosis of Stx1e-associated infections.Craig SkinnerStephanie PatfieldRowaida KhalilQiulian KongXiaohua HeAmerican Society for MicrobiologyarticleEnterobacterShiga toxinsStx1eimmunoassaysmonoclonal antibodiesMicrobiologyQR1-502ENmSphere, Vol 1, Iss 1 (2016)
institution	DOAJ
collection	DOAJ
language	EN
topic	Enterobacter Shiga toxins Stx1e immunoassays monoclonal antibodies Microbiology QR1-502
spellingShingle	Enterobacter Shiga toxins Stx1e immunoassays monoclonal antibodies Microbiology QR1-502 Craig Skinner Stephanie Patfield Rowaida Khalil Qiulian Kong Xiaohua He New Monoclonal Antibodies against a Novel Subtype of Shiga Toxin 1 Produced by <named-content content-type="genus-species">Enterobacter cloacae</named-content> and Their Use in Analysis of Human Serum
description	ABSTRACT Shiga toxin (Stx) is a major virulence factor of several bacterial pathogens that cause potentially fatal illness, including Escherichia coli and Shigella spp. The continual emergence of new subtypes of Stxs presents challenges for the clinical diagnosis of infections caused by Stx-producing organisms. Here, we report the development of four new monoclonal antibodies (MAbs) against Stx1e, a novel subtype of Stx1 that was produced by an Enterobacter cloacae strain and had limited reactivity with existing anti-Stx1 antibodies. Western blot analysis indicates that these MAbs were Stx1 specific, bound to the A subunit, and had distinct preferences for subtypes of Stx1. Of the four MAbs, Stx1e-2 was capable of partially neutralizing cytotoxicities derived from Stx1e in Vero cells. Enzyme-linked immunosorbent assays assembled with these high-affinity MAbs detected Stx1e at concentrations as low as 4.8 pg/ml in phosphate-buffered saline and 53.6 pg/ml in spiked human serum samples and were also capable of distinguishing Stx1e-producing strains in enriched cultures. These assays may therefore have clinical value in diagnosing Stx1e-producing bacterial infection. Additionally, characteristics of Stx1e, such as the origin of stx1e genes, conditions for toxin expression, receptor binding, and cytotoxicity, were investigated with the new antibodies developed in this study. This information should be useful for further understanding the clinical significance and prevalence of Stx1e-harboring E. cloacae and other organisms. IMPORTANCE Stxs are among the most clinically important virulence factors of Shigella and enterohemorrhagic Escherichia coli. There are many varieties of Stx, and although Stx1a and Stx2a are the most common and widely distributed types of Stx, new variants of Stx are continually emerging. These new variants of Stx can be challenging to detect, since most Stx detection kits are optimized for the detection of Stx1a and Stx2a. Stx1e, recently discovered in an atypical host (Enterobacter cloacae), is undetectable by many Stx assays. To formulate new assays for the detection of Stx1e, we generated four new MAbs that recognize this Stx subtype. Using these antibodies, we generated an assay capable of detecting Stx1e at low picogram-per-milliliter concentrations. This assay is also compatible with a human serum matrix, suggesting that it may have utility for the clinical detection and diagnosis of Stx1e-associated infections.
format	article
author	Craig Skinner Stephanie Patfield Rowaida Khalil Qiulian Kong Xiaohua He
author_facet	Craig Skinner Stephanie Patfield Rowaida Khalil Qiulian Kong Xiaohua He
author_sort	Craig Skinner
title	New Monoclonal Antibodies against a Novel Subtype of Shiga Toxin 1 Produced by <named-content content-type="genus-species">Enterobacter cloacae</named-content> and Their Use in Analysis of Human Serum
title_short	New Monoclonal Antibodies against a Novel Subtype of Shiga Toxin 1 Produced by <named-content content-type="genus-species">Enterobacter cloacae</named-content> and Their Use in Analysis of Human Serum
title_full	New Monoclonal Antibodies against a Novel Subtype of Shiga Toxin 1 Produced by <named-content content-type="genus-species">Enterobacter cloacae</named-content> and Their Use in Analysis of Human Serum
title_fullStr	New Monoclonal Antibodies against a Novel Subtype of Shiga Toxin 1 Produced by <named-content content-type="genus-species">Enterobacter cloacae</named-content> and Their Use in Analysis of Human Serum
title_full_unstemmed	New Monoclonal Antibodies against a Novel Subtype of Shiga Toxin 1 Produced by <named-content content-type="genus-species">Enterobacter cloacae</named-content> and Their Use in Analysis of Human Serum
title_sort	new monoclonal antibodies against a novel subtype of shiga toxin 1 produced by <named-content content-type="genus-species">enterobacter cloacae</named-content> and their use in analysis of human serum
publisher	American Society for Microbiology
publishDate	2016
url	https://doaj.org/article/0b2e261d5f4442cd85123d59dad25783
work_keys_str_mv	AT craigskinner newmonoclonalantibodiesagainstanovelsubtypeofshigatoxin1producedbynamedcontentcontenttypegenusspeciesenterobactercloacaenamedcontentandtheiruseinanalysisofhumanserum AT stephaniepatfield newmonoclonalantibodiesagainstanovelsubtypeofshigatoxin1producedbynamedcontentcontenttypegenusspeciesenterobactercloacaenamedcontentandtheiruseinanalysisofhumanserum AT rowaidakhalil newmonoclonalantibodiesagainstanovelsubtypeofshigatoxin1producedbynamedcontentcontenttypegenusspeciesenterobactercloacaenamedcontentandtheiruseinanalysisofhumanserum AT qiuliankong newmonoclonalantibodiesagainstanovelsubtypeofshigatoxin1producedbynamedcontentcontenttypegenusspeciesenterobactercloacaenamedcontentandtheiruseinanalysisofhumanserum AT xiaohuahe newmonoclonalantibodiesagainstanovelsubtypeofshigatoxin1producedbynamedcontentcontenttypegenusspeciesenterobactercloacaenamedcontentandtheiruseinanalysisofhumanserum
_version_	1718428111461154816

New Monoclonal Antibodies against a Novel Subtype of Shiga Toxin 1 Produced by <named-content content-type="genus-species">Enterobacter cloacae</named-content> and Their Use in Analysis of Human Serum

Ejemplares similares