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Updated: Jul 21, 2025
Author Spotlight: Advancing Antibiotic Resistance Research Using an Efflux-Deficient Bacterial Strain and a Single-Copy Gene Expression System
Published on: January 5, 2024
Phenotypic and genetic insights into efflux pump mechanism in Mycoplasma anserisalpingitidis
Eszter Zsófia Nagy1,2, Áron Botond Kovács1,2, Enikő Wehmann1,2
1Veterinary Medical Research Institute, Budapest, Hungary.
Introduction:
Mycoplasma anserisalpingitidis is one of the most important waterfowl-pathogenic mycoplasmas. Due to inadequate antibiotic treatment, many strains with high minimal inhibitory concentration (MIC) values for multiple drugs have been isolated lately. Decreased antibiotic susceptibility in several Mycoplasma species are known to be associated with mutations in topoisomerase and ribosomal genes, but other strategies such as active efflux pump mechanisms were also described. The scope of this study was the phenotypic and genetic characterization of the active efflux mechanism in M. anserisalpingitidis.
Methods:
We measured the MIC values in the presence and absence of different efflux pump inhibitors (EPIs), such as carbonyl cyanide m-chlorophenylhydrazine (CCCP), orthovanadate (OV), and reserpine (RSP). Moreover, bioinformatic tools were utilized to detect putative regulatory sequences of membrane transport proteins coding genes, while comparative genome analysis was performed to reveal potential markers of antibiotic resistance.
Results:
Out of the three examined EPIs, CCCP decreased the MICs at least two-fold below the original MICs (in 23 cases out of 36 strains). In the presence of OV or RSP, MIC value differences could be seen only if modified dilution series (10% decrease steps were used instead of two-fold dilutions) were applied (in 24/36 cases with OV and 9/36 with RSP). During comparative genome analysis, non-synonymous single nucleotide polymorphisms (nsSNPs) were identified in genes encoding ABC membrane transport proteins, which were displayed in higher percentages in M. anserisalpingitidis strains with increased MICs. In terms of other genes, a nsSNP was identified in DNA gyrase subunit A (gyrA) gene which can be related to decreased susceptibility to enrofloxacin. The present study is the first to highlight the importance of efflux pump mechanisms in M. anserisalpingitidis.
Discussion:
Considering the observed effects of the EPI CCCP against this bacterium, it can be assumed, that the use of EPIs would increase the efficiency of targeted antibiotic therapy in the future control of this pathogen. However, further research is required to obtain a more comprehensive understanding of efflux pump mechanism in this bacterium.
Insights
Efflux pump inhibitors, particularly CCCP, show promise in increasing antibiotic effectiveness against Mycoplasma anserisalpingitidis. This study highlights the role of efflux pumps in antibiotic resistance for this waterfowl pathogen.
Area of Science:
- Veterinary Microbiology
- Molecular Biology
- Antimicrobial Resistance
Background:
- Mycoplasma anserisalpingitidis is a significant pathogen in waterfowl, with increasing antibiotic resistance observed.
- Reduced antibiotic susceptibility is linked to genetic mutations and efflux pump mechanisms.
Purpose of the Study:
- To phenotypically and genetically characterize active efflux mechanisms in Mycoplasma anserisalpingitidis.
- To investigate the role of efflux pumps in the reduced antibiotic susceptibility of M. anserisalpingitidis strains.
Main Methods:
- Minimal Inhibitory Concentrations (MICs) were determined with and without efflux pump inhibitors (EPIs) like CCCP, OV, and RSP.
- Bioinformatic analysis identified regulatory sequences of membrane transport protein genes.
- Comparative genome analysis revealed potential antibiotic resistance markers.
Main Results:
- CCCP significantly reduced MICs in 23 out of 36 M. anserisalpingitidis strains.
- Non-synonymous single nucleotide polymorphisms (nsSNPs) were found in ABC membrane transport protein genes, correlating with higher MICs.
- A nsSNP in the gyrA gene was associated with reduced enrofloxacin susceptibility.
Conclusions:
- Efflux pump mechanisms play a crucial role in Mycoplasma anserisalpingitidis antibiotic resistance.
- Using EPIs like CCCP could enhance antibiotic therapy efficacy against this pathogen.
- Further research is needed for a comprehensive understanding of efflux pump mechanisms in M. anserisalpingitidis.
