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Determination of Diphtheria Toxin in Bacterial Cultures by Enzyme Immunoassay
Since diphtheria toxin (DT) is the main virulence factor of Corynebacterium diphtheriae and C. ulcerans, the detection of DT in corynebacterial cultures is of utmost importance in the laboratory diagnosis of diphtheria. The need to measure the level of DT production (LTP) arises when studying the vi...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2022
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9498204/ https://www.ncbi.nlm.nih.gov/pubmed/36140605 http://dx.doi.org/10.3390/diagnostics12092204 |
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author | Simonova, Maria A. Melnikov, Vyacheslav G. Lakhtina, Olga E. Komaleva, Ravilya L. Berger, Anja Sing, Andreas Zavriev, Sergey K. |
author_facet | Simonova, Maria A. Melnikov, Vyacheslav G. Lakhtina, Olga E. Komaleva, Ravilya L. Berger, Anja Sing, Andreas Zavriev, Sergey K. |
author_sort | Simonova, Maria A. |
collection | PubMed |
description | Since diphtheria toxin (DT) is the main virulence factor of Corynebacterium diphtheriae and C. ulcerans, the detection of DT in corynebacterial cultures is of utmost importance in the laboratory diagnosis of diphtheria. The need to measure the level of DT production (LTP) arises when studying the virulence of a strain for the purpose of diphtheria agent monitoring. To determine the LTP of diphtheria agents, an immunoassay based on monoclonal antibodies (mAbs) has been developed. A pair of mAbs specific to the fragment B of DT was selected, which makes it possible to detect DT in a sandwich ELISA with a detection limit of DT less than 1 ng/mL. Sandwich ELISA was used to analyze 218 liquid culture supernatants of high-, low- and non-toxigenic strains of various corynebacteria. It was shown that the results of ELISA are in good agreement with the results of PCR and the Elek test for the tox gene and DT detection, respectively. The diagnostic sensitivity of the assay was approximately 99%, and specificity was 100%. It has been found that strains of C. ulcerans, on average, produce 10 times less DT than C. diphtheriae. The mAbs used in the ELISA proved to be quite discriminatory and could be further used for the design of the LFIA, a method that can reduce the labor and cost of laboratory diagnosis of diphtheria. |
format | Online Article Text |
id | pubmed-9498204 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-94982042022-09-23 Determination of Diphtheria Toxin in Bacterial Cultures by Enzyme Immunoassay Simonova, Maria A. Melnikov, Vyacheslav G. Lakhtina, Olga E. Komaleva, Ravilya L. Berger, Anja Sing, Andreas Zavriev, Sergey K. Diagnostics (Basel) Article Since diphtheria toxin (DT) is the main virulence factor of Corynebacterium diphtheriae and C. ulcerans, the detection of DT in corynebacterial cultures is of utmost importance in the laboratory diagnosis of diphtheria. The need to measure the level of DT production (LTP) arises when studying the virulence of a strain for the purpose of diphtheria agent monitoring. To determine the LTP of diphtheria agents, an immunoassay based on monoclonal antibodies (mAbs) has been developed. A pair of mAbs specific to the fragment B of DT was selected, which makes it possible to detect DT in a sandwich ELISA with a detection limit of DT less than 1 ng/mL. Sandwich ELISA was used to analyze 218 liquid culture supernatants of high-, low- and non-toxigenic strains of various corynebacteria. It was shown that the results of ELISA are in good agreement with the results of PCR and the Elek test for the tox gene and DT detection, respectively. The diagnostic sensitivity of the assay was approximately 99%, and specificity was 100%. It has been found that strains of C. ulcerans, on average, produce 10 times less DT than C. diphtheriae. The mAbs used in the ELISA proved to be quite discriminatory and could be further used for the design of the LFIA, a method that can reduce the labor and cost of laboratory diagnosis of diphtheria. MDPI 2022-09-11 /pmc/articles/PMC9498204/ /pubmed/36140605 http://dx.doi.org/10.3390/diagnostics12092204 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Simonova, Maria A. Melnikov, Vyacheslav G. Lakhtina, Olga E. Komaleva, Ravilya L. Berger, Anja Sing, Andreas Zavriev, Sergey K. Determination of Diphtheria Toxin in Bacterial Cultures by Enzyme Immunoassay |
title | Determination of Diphtheria Toxin in Bacterial Cultures by Enzyme Immunoassay |
title_full | Determination of Diphtheria Toxin in Bacterial Cultures by Enzyme Immunoassay |
title_fullStr | Determination of Diphtheria Toxin in Bacterial Cultures by Enzyme Immunoassay |
title_full_unstemmed | Determination of Diphtheria Toxin in Bacterial Cultures by Enzyme Immunoassay |
title_short | Determination of Diphtheria Toxin in Bacterial Cultures by Enzyme Immunoassay |
title_sort | determination of diphtheria toxin in bacterial cultures by enzyme immunoassay |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9498204/ https://www.ncbi.nlm.nih.gov/pubmed/36140605 http://dx.doi.org/10.3390/diagnostics12092204 |
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