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Characterisation of pellicle-forming ability in clinical carbapenem-resistant Acinetobacter baumannii
Heng Kang Ng1, Suat Moi Puah1, Cindy Shuan Ju Teh2
1Department of Biomedical Science, Faculty of Medicine, Universiti Malaya, Kuala Lumpur, Malaysia.
Background:
Acinetobacter baumannii was reported to have resistance towards carbapenems and the ability to form an air-liquid biofilm (pellicle) which contributes to their virulence. The GacSA two-component system has been previously shown to play a role in pellicle formation. Therefore, this study aims to detect the presence of gacA and gacS genes in carbapenem-resistant Acinetobacter baumannii (CRAB) isolates recovered from patients in intensive care units and to investigate their pellicle forming ability.
Methods:
The gacS and gacA genes were screened in 96 clinical CRAB isolates using PCR assay. Pellicle formation assay was performed in Mueller Hinton medium and Luria Bertani medium using borosilicate glass tubes and polypropylene plastic tubes. The biomass of the pellicle was quantitated using the crystal violet staining assay. The selected isolates were further assessed for their motility using semi-solid agar and monitored in real-time using real-time cell analyser (RTCA).
Results:
All 96 clinical CRAB isolates carried the gacS and gacA genes, however, only four isolates (AB21, AB34, AB69 and AB97) displayed the ability of pellicle-formation phenotypically. These four pellicle-forming isolates produced robust pellicles in Mueller Hinton medium with better performance in borosilicate glass tubes in which biomass with OD570 ranging from 1.984 ± 0.383 to 2.272 ± 0.376 was recorded. The decrease in cell index starting from 13 hours obtained from the impedance-based RTCA showed that pellicle-forming isolates had entered the growth stage of pellicle development.
Conclusion:
These four pellicle-forming clinical CRAB isolates could be potentially more virulent, therefore further investigation is warranted to provide insights into their pathogenic mechanisms.
Insights
All carbapenem-resistant Acinetobacter baumannii isolates possessed the gacS and gacA genes. Four isolates demonstrated pellicle formation, suggesting increased virulence and potential for further pathogenic mechanism investigation.
Area of Science:
- Microbiology
- Molecular Biology
- Infectious Diseases
Background:
- Acinetobacter baumannii exhibits carbapenem resistance and forms virulence-contributing air-liquid biofilms (pellicles).
- The GacSA two-component system is implicated in pellicle formation in A. baumannii.
Purpose of the Study:
- To detect gacS and gacA genes in carbapenem-resistant A. baumannii (CRAB) isolates from intensive care units.
- To investigate the pellicle-forming ability of these CRAB isolates.
Main Methods:
- PCR assay to screen for gacS and gacA genes in 96 CRAB isolates.
- Pellicle formation assay using Mueller Hinton and Luria Bertani media in glass and plastic tubes.
- Quantification of pellicle biomass via crystal violet staining and real-time cell analysis (RTCA).
Main Results:
- All 96 CRAB isolates harbored both gacS and gacA genes.
- Four isolates (AB21, AB34, AB69, AB97) exhibited pellicle formation.
- Pellicle formation was robust in Mueller Hinton medium, particularly in borosilicate glass tubes, with biomass OD570 ranging from 1.984 to 2.272.
Conclusions:
- The presence of gacS and gacA genes is common in CRAB isolates.
- A subset of CRAB isolates demonstrates pellicle-forming ability, indicating potential for enhanced virulence.
- Further research is needed to elucidate the pathogenic mechanisms of these pellicle-forming CRAB isolates.
