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Development and Standardization of a High-Throughput Bordetella pertussis Growth-Inhibition Assay

Bordetella pertussis, the main causative agent of whooping cough, is a reemerging pathogen, and recent vaccine-resistant strain outbreaks and emergence of macrolides-resistant strains in China raised new concerns for control of the disease. New vaccines and potentially new antibiotics are thus neede...

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Autores principales: Thiriard, Anaïs, Raze, Dominique, Locht, Camille
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7212404/
https://www.ncbi.nlm.nih.gov/pubmed/32425912
http://dx.doi.org/10.3389/fmicb.2020.00777
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author Thiriard, Anaïs
Raze, Dominique
Locht, Camille
author_facet Thiriard, Anaïs
Raze, Dominique
Locht, Camille
author_sort Thiriard, Anaïs
collection PubMed
description Bordetella pertussis, the main causative agent of whooping cough, is a reemerging pathogen, and recent vaccine-resistant strain outbreaks and emergence of macrolides-resistant strains in China raised new concerns for control of the disease. New vaccines and potentially new antibiotics are thus needed. B. pertussis is tedious to culture and requires several days of growth to count isolated colonies on agar-based media, making large-scale screening of new anti-B. pertussis compounds or functional evaluation of large sample sizes of immune sera difficult. Here, we developed a scalable, rapid, high-throughput luminescence-based Bordetella growth inhibition assay (BGIA) to quantify surviving bacteria after treatment with anti-B. pertussis compounds. A strong correlation between luminescence and colony-forming units (r(2) = 0.9345, p < 0.0001) was found and the BGIA showed high sensitivity and reproducibility. We demonstrate here that the BGIA can be used to quantify resistance of B. pertussis to antibiotics, sensitivity to complement and to human serum in an easy-to-operate and fast manner. We have optimized the assay and tested the effects of different B. pertussis strains and growth conditions on serum and complement sensitivity. We also uncovered complement-independent antibody-mediated inhibition of B. pertussis growth. The BGIA can thus effectively be implemented for large-scale serum studies to further investigate anti-B. pertussis immune responses at a functional level, as well as for screening of B. pertussis strains for their resistance to antibiotics or complement, and for high-throughput screening of novel anti-B. pertussis compounds.
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spelling pubmed-72124042020-05-18 Development and Standardization of a High-Throughput Bordetella pertussis Growth-Inhibition Assay Thiriard, Anaïs Raze, Dominique Locht, Camille Front Microbiol Microbiology Bordetella pertussis, the main causative agent of whooping cough, is a reemerging pathogen, and recent vaccine-resistant strain outbreaks and emergence of macrolides-resistant strains in China raised new concerns for control of the disease. New vaccines and potentially new antibiotics are thus needed. B. pertussis is tedious to culture and requires several days of growth to count isolated colonies on agar-based media, making large-scale screening of new anti-B. pertussis compounds or functional evaluation of large sample sizes of immune sera difficult. Here, we developed a scalable, rapid, high-throughput luminescence-based Bordetella growth inhibition assay (BGIA) to quantify surviving bacteria after treatment with anti-B. pertussis compounds. A strong correlation between luminescence and colony-forming units (r(2) = 0.9345, p < 0.0001) was found and the BGIA showed high sensitivity and reproducibility. We demonstrate here that the BGIA can be used to quantify resistance of B. pertussis to antibiotics, sensitivity to complement and to human serum in an easy-to-operate and fast manner. We have optimized the assay and tested the effects of different B. pertussis strains and growth conditions on serum and complement sensitivity. We also uncovered complement-independent antibody-mediated inhibition of B. pertussis growth. The BGIA can thus effectively be implemented for large-scale serum studies to further investigate anti-B. pertussis immune responses at a functional level, as well as for screening of B. pertussis strains for their resistance to antibiotics or complement, and for high-throughput screening of novel anti-B. pertussis compounds. Frontiers Media S.A. 2020-04-23 /pmc/articles/PMC7212404/ /pubmed/32425912 http://dx.doi.org/10.3389/fmicb.2020.00777 Text en Copyright © 2020 Thiriard, Raze and Locht. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Microbiology
Thiriard, Anaïs
Raze, Dominique
Locht, Camille
Development and Standardization of a High-Throughput Bordetella pertussis Growth-Inhibition Assay
title Development and Standardization of a High-Throughput Bordetella pertussis Growth-Inhibition Assay
title_full Development and Standardization of a High-Throughput Bordetella pertussis Growth-Inhibition Assay
title_fullStr Development and Standardization of a High-Throughput Bordetella pertussis Growth-Inhibition Assay
title_full_unstemmed Development and Standardization of a High-Throughput Bordetella pertussis Growth-Inhibition Assay
title_short Development and Standardization of a High-Throughput Bordetella pertussis Growth-Inhibition Assay
title_sort development and standardization of a high-throughput bordetella pertussis growth-inhibition assay
topic Microbiology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7212404/
https://www.ncbi.nlm.nih.gov/pubmed/32425912
http://dx.doi.org/10.3389/fmicb.2020.00777
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