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Volume 12
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10.3390/diagnostics12092204
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Article

Determination of Diphtheria Toxin in Bacterial Cultures by Enzyme Immunoassay

by
Maria A. Simonova
1,†,
Vyacheslav G. Melnikov
2,*,†,
Olga E. Lakhtina
1,
Ravilya L. Komaleva
1,
Anja Berger
2,3,
Andreas Sing
2,3,‡ and
Sergey K. Zavriev
1,‡
1
Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry of Russian Academy of Sciences, Miklukho-Maklaya 16/10, 117997 Moscow, Russia
2
National Conciliary Laboratory on Diphtheria, Veterinaerstrasse 2, 85764 Oberschleissheim, Germany
3
Department of Public Health Microbiology, Bavarian Health and Food Safety Authority, Veterinaerstrasse 2, 85764 Oberschleissheim, Germany
*
Author to whom correspondence should be addressed.
†
These authors contributed equally to this work.
‡
These authors contributed equally to this work.
Diagnostics 2022, 12(9), 2204; https://doi.org/10.3390/diagnostics12092204
Submission received: 13 August 2022 / Revised: 7 September 2022 / Accepted: 8 September 2022 / Published: 11 September 2022
(This article belongs to the Special Issue Laboratory Diagnosis in Microbial Diseases)
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Abstract

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.
Keywords: Corynebacterium diphtheriae; Corynebacterium ulcerans; diphtheria toxin; ELISA; monoclonal antibodies; level of toxin production Corynebacterium diphtheriae; Corynebacterium ulcerans; diphtheria toxin; ELISA; monoclonal antibodies; level of toxin production

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MDPI and ACS Style

Simonova, M.A.; Melnikov, V.G.; Lakhtina, O.E.; Komaleva, R.L.; Berger, A.; Sing, A.; Zavriev, S.K. Determination of Diphtheria Toxin in Bacterial Cultures by Enzyme Immunoassay. Diagnostics 2022, 12, 2204. https://doi.org/10.3390/diagnostics12092204

AMA Style

Simonova MA, Melnikov VG, Lakhtina OE, Komaleva RL, Berger A, Sing A, Zavriev SK. Determination of Diphtheria Toxin in Bacterial Cultures by Enzyme Immunoassay. Diagnostics. 2022; 12(9):2204. https://doi.org/10.3390/diagnostics12092204

Chicago/Turabian Style

Simonova, Maria A., Vyacheslav G. Melnikov, Olga E. Lakhtina, Ravilya L. Komaleva, Anja Berger, Andreas Sing, and Sergey K. Zavriev. 2022. "Determination of Diphtheria Toxin in Bacterial Cultures by Enzyme Immunoassay" Diagnostics 12, no. 9: 2204. https://doi.org/10.3390/diagnostics12092204

APA Style

Simonova, M. A., Melnikov, V. G., Lakhtina, O. E., Komaleva, R. L., Berger, A., Sing, A., & Zavriev, S. K. (2022). Determination of Diphtheria Toxin in Bacterial Cultures by Enzyme Immunoassay. Diagnostics, 12(9), 2204. https://doi.org/10.3390/diagnostics12092204

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