Mycobacterium tuberculosis β-lactamase variant reduces sensitivity to ampicillin/avibactam in a zebrafish-Mycobacterium marinum model of tuberculosis

Ilona van Alen, Mayra A. Aguirre García, Janneke J. Maaskant, Coenraad P. Kuijl, Wilbert Bitter, Annemarie H. Meijer, Marcellus Ubbink*

*Corresponding author for this work

Research output: Contribution to JournalArticleAcademicpeer-review

Abstract

The β-lactamase of Mycobacterium tuberculosis, BlaC, hydrolyzes β-lactam antibiotics, hindering the use of these antibiotics for the treatment of tuberculosis. Inhibitors, such as avibactam, can reversibly inhibit the enzyme, allowing for the development of combination therapies using both antibiotic and inhibitor. However, laboratory evolution studies using Escherichia coli resulted in the discovery of single amino acid variants of BlaC that reduce the sensitivity for inhibitors or show higher catalytic efficiency against antibiotics. Here, we tested these BlaC variants under more physiological conditions using the M. marinum infection model of zebrafish, which recapitulates hallmark features of tuberculosis, including the intracellular persistence of mycobacteria in macrophages and the induction of granuloma formation. To this end, the M. tuberculosis blaC gene was integrated into the chromosome of a blaC frameshift mutant of M. marinum. Subsequently, the resulting strains were used to infect zebrafish embryos in order to test the combinatorial effect of ampicillin and avibactam. The results show that embryos infected with an M. marinum strain producing BlaC show lower infection levels after treatment than untreated embryos. Additionally, BlaC K234R showed higher infection levels after treatment than those infected with bacteria producing the wild-type enzyme, demonstrating that the zebrafish host is less sensitive to the combinatorial therapy of β-lactam antibiotic and inhibitor. These findings are of interest for future development of combination therapies to treat tuberculosis.

Original languageEnglish
Article number15406
Number of pages9
JournalScientific Reports
Volume13
DOIs
Publication statusPublished - 16 Sept 2023

Bibliographical note

Funding Information:
This study was supported by internal funding, by the Netherlands Organization for Scientific Research (NWO), Grant Number 711.017.013, and by the Marie Skłodowska-Curie Innovative Training Network INFLANET (Grant agreement 955576). The authors declare no competing interests.

Publisher Copyright:
© 2023, Springer Nature Limited.

Funding

This study was supported by internal funding, by the Netherlands Organization for Scientific Research (NWO), Grant Number 711.017.013, and by the Marie Skłodowska-Curie Innovative Training Network INFLANET (Grant agreement 955576). The authors declare no competing interests.

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