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Published on: January 22, 2021
Proteomic analysis of ceftazidime and meropenem-exposed Pseudomonas aeruginosa ATCC 9027
Hong Loan Ngo1,2, Thuc Quyen Huynh1,2,3, Nguyen Bao Vy Tran1,2
1School of Biotechnology, International University, Ho Chi Minh City, Vietnam.
Background:
Pseudomonas aeruginosa is well known for its intrinsic ability to resist a wide range of antibiotics, thus complicates treatment. Thus, understanding the response of the pathogen to antibiotics is important for developing new therapies. In this study, proteomic response of P. aeruginosa to the commonly used anti-pseudomonas antibiotics, ceftazidime (Caz) and meropenem (Mem) was investigated.
Methods:
P. aeruginosa ATCC 9027, an antibiotic-susceptible strain, was exposed to sub-MIC values of antibiotics either Caz or Mem for 14 days to obtain E1 strains and then cultured in antibiotic-free environments for 10 days to obtain E2 strains. Proteomes of the initial and E1, E2 strains were identified and comparatively analyzed using isobaric tags for relative and absolute quantitation (iTRAQ) in cooperation with nano LC-MS/MS. Noted up and down-regulated proteins were confirmed with quantitative reverse transcriptase PCR (qRT-PCR).
Results:
Overall, 1039 and 1041 proteins were identified in Caz and Mem-exposed strains, respectively. Upon antibiotic exposure, there were 7-10% up-regulated (Caz: 71, Mem: 85) and down-regulated (Caz: 106, Mem: 69) proteins (1.5-fold change cut-off). For both Caz and Mem, the DEPs were primarily the ones involved in metabolic process, membrane, virulence, protein synthesis, and antibiotic resistance in which proteins involved in antibiotics resistance tended to be up-regulated while proteins involved in protein synthesis and metabolic process were down-regulated. Noted proteins included beta-lactamase AmpC which was up-regulated and OprD which was down-regulated in both the antibiotic-exposed strains. Besides, biofilm formation related proteins TssC1 and Hcp1 in Caz- exposed strains and the membrane/ periplasmic proteins Azu and PagL in Mem-exposed strains were found significantly down-regulated. qRT-PCR results confirmed the expression change of AmpC, Hcp1 and OprD proteins.
Conclusion:
Exposure of Pseudomonas aeruginosa to sub-MIC values of Caz and Mem resulted in around 10% change in its proteome. Not only proteins with confirmed roles in antibiotic resistance mechanisms changed their expression but also virulence- associated proteins. Both Caz and Mem response involved up-regulation of AmpC and down-regulation of OprD. While TssC1 and Hcp1 were responsible for Caz response, Azu and PagL were more likely involved in Mem response.
Insights
Pseudomonas aeruginosa proteomic responses to ceftazidime (Caz) and meropenem (Mem) revealed significant changes in antibiotic resistance and virulence proteins. Understanding these changes is crucial for developing effective anti-pseudomonas therapies.
Area of Science:
- Microbiology
- Proteomics
- Drug Discovery
Background:
- Pseudomonas aeruginosa exhibits intrinsic antibiotic resistance, complicating treatment strategies.
- Understanding the pathogen's proteomic response to antibiotics is vital for developing novel therapeutic approaches.
Purpose of the Study:
- To investigate the proteomic response of P. aeruginosa to ceftazidime (Caz) and meropenem (Mem).
- To identify differentially expressed proteins (DEPs) involved in antibiotic resistance and virulence.
Main Methods:
- Antibiotic-susceptible P. aeruginosa ATCC 9027 was exposed to sub-inhibitory concentrations of Caz or Mem for 14 days.
- Proteomic analysis using isobaric tags for relative and absolute quantitation (iTRAQ) and nano LC-MS/MS.
- Quantitative reverse transcriptase PCR (qRT-PCR) was used to validate protein expression changes.
Main Results:
- Exposure to Caz and Mem resulted in approximately 10% changes in the P. aeruginosa proteome.
- Key proteins involved in antibiotic resistance (e.g., AmpC) were upregulated, while those in protein synthesis and metabolic processes were downregulated.
- Specific proteins like OprD, TssC1, Hcp1, Azu, and PagL showed altered expression patterns in response to Caz and Mem.
Conclusions:
- Sub-MIC exposure to Caz and Mem induces significant proteomic alterations in P. aeruginosa, affecting both antibiotic resistance and virulence.
- Both antibiotics led to AmpC upregulation and OprD downregulation, suggesting a common resistance mechanism.
- Distinct protein changes were observed for Caz (TssC1, Hcp1) and Mem (Azu, PagL) responses, highlighting drug-specific adaptations.
