Environmental determinants of antibiotic resistance and virulence of Acinetobacter baumannii and development of novel molecular tools for genetic manipulations of Acinetobacter baumannii
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Date
2019-08
Authors
De Silva, Pawuththuwadura Malaka Pulinda
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Abstract
Acinetobacter baumannii is a notorious pathogen in health- care settings around the world, primarily due to its reduced susceptibility to antibiotics. A. baumanniiIt also shows impressive capabilities to adapt to harsh conditions in clinical settings, which contributes to its persistence in such conditions. Following their traditional role, the Two Component Systems (TCSs) present in A. baumannii play a crucial role in sensing and adapting to the changing environmental conditions. Out of the TCSs present in A. baumannii, AdeRS TCS has been shown to modulate a key RND efflux pump, AdeABC which can result in antibiotic resistance. In-frame deletion of adeRS resulted in reduced expression of AdeB as demonstrated previously confirming the regulatory role of AdeRS in expression of AdeABC. DFurther, deletion of adeRS affected biofilm formation and motility of A. baumannii demonstrating the multi-faceted nature of regulatory networks controlled by AdeRS. Interestingly, the adeRS deletion mutant was sensitive to saline stress and surface associated motility was adversely affected at higher NaCl concentrations suggesting that AdeRS could be a part of salinity stress adaptation in A. baumannii.
Investigations into the adaptation of A. baumannii to incubation temperatures of 28˚C and 37˚C revealed potentially clinically relevant phenotypic changes. Specifically, motility was reduced at 28˚C while biofilm formation significantly increased at 28˚C. Proteomic analysis showed an upregulation of 366 proteins and downregulation of 263 proteins at 28˚C.
An uncharacterized orphan response regulator, A1S_2006 displayed significantly higher expression levels in a MDR A. baumannii clinical isolate AB030. A deletion mutant of A1S_2006 displayed decreased biofilm formation, motility, and virulence in Galleria mellonella model. However, deletion of A1S_2006 resulted in significantly higher attachment of A. baumannii in A549 cells. RNA-Seq analysis of the deletion mutant revealed upregulation of 116 genes and downregulation of 42 genes.
Finally, to address the lack of molecular tools available for genetic manipulations of A. baumannii, a set of novel mini-Tn7 based vectors were created by using apramycin and zeocin resistance markers to rejuvenate the current mini-Tn7 based vectors. These can be used for genetic manipulations in clinical isolates with reduced susceptibility to commonly used markers. These results show the role of environmental determinants in modulating clinically relevant characteristics of A. baumannii as well as adding new components to the molecular toolkit for A. baumannii.
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Keywords
Acinetobacter baumannii, Antimicrobial resistance, Two component systems
Citation
De Silva, P. M., & Kumar, A. (2019). Signal Transduction Proteins in Acinetobacter baumannii: Role in Antibiotic Resistance, Virulence, and Potential as Drug Targets. Front Microbiol, 10, 49. doi:10.3389/fmicb.2019.00049
De Silva, P. M., & Kumar, A. (2017). Effect of Sodium Chloride on Surface-Associated Motility of Acinetobacter baumannii and the Role of AdeRS Two-Component System. J Membr Biol. doi:10.1007/s00232-017-9985-7
De Silva, P. M., Chong, P., Fernando, D. M., Westmacott, G., & Kumar, A. (2018). Effect of Incubation Temperature on Antibiotic Resistance and Virulence Factors of Acinetobacter baumannii ATCC 17978. Antimicrob Agents Chemother, 62(1). doi:10.1128/AAC.01514-17
Ducas-Mowchun, K., De Silva, P. M., Crisostomo, L., Fernando, D. M., Chao, T. C., Pelka, P., . . . Kumar, A. (2019). Next Generation of Tn7-Based Single-Copy Insertion Elements for Use in Multi- and Pan-Drug-Resistant Strains of Acinetobacter baumannii. Appl Environ Microbiol, 85(11). doi:10.1128/AEM.00066-19
De Silva, P. M., & Kumar, A. (2017). Effect of Sodium Chloride on Surface-Associated Motility of Acinetobacter baumannii and the Role of AdeRS Two-Component System. J Membr Biol. doi:10.1007/s00232-017-9985-7
De Silva, P. M., Chong, P., Fernando, D. M., Westmacott, G., & Kumar, A. (2018). Effect of Incubation Temperature on Antibiotic Resistance and Virulence Factors of Acinetobacter baumannii ATCC 17978. Antimicrob Agents Chemother, 62(1). doi:10.1128/AAC.01514-17
Ducas-Mowchun, K., De Silva, P. M., Crisostomo, L., Fernando, D. M., Chao, T. C., Pelka, P., . . . Kumar, A. (2019). Next Generation of Tn7-Based Single-Copy Insertion Elements for Use in Multi- and Pan-Drug-Resistant Strains of Acinetobacter baumannii. Appl Environ Microbiol, 85(11). doi:10.1128/AEM.00066-19