Investigating the Validity of mCIM and sCIM Phenotypic Methods in Screening Pseudomonas aeruginosa Isolates Producing IMP, VIM, and NDM Metallo-β-Lactamases from Burn Wounds

Background

Pseudomonas aeruginosa is a Gram-negative bacterium widely found in the environment, particularly in hospital settings. It is the second most common pathogen causing burn wound infections, often leading to severe infections and exhibiting high levels of antibiotic resistance, especially to carbapenems. This resistance is primarily due to the production of metallo-β-lactamases (MBLs), such as IMP, VIM, and NDM enzymes, which can hydrolyze carbapenem antibiotics, making treatment extremely challenging. Therefore, rapid and accurate detection of these enzymes is crucial for guiding clinical treatment.

Currently, PCR (polymerase chain reaction) is the gold standard for detecting carbapenemase genes, but its high cost and reliance on specialized equipment limit its widespread use in clinical diagnostics. Thus, the development of an efficient and cost-effective phenotypic detection method has become a research priority. This study aims to evaluate the effectiveness of the Modified Carbapenem Inactivation Method (MCIM) and the Simplified Carbapenem Inactivation Method (SCIM) in screening P. aeruginosa isolates producing IMP, VIM, and NDM metallo-β-lactamases.

Source of the Study

This research was conducted by Hossein Hatami, Shiva Motamedi, Ghazaleh Talebi, and Mojdeh Hakemi-Vala from the School of Public Health and the Department of Microbiology at Shahid Beheshti University of Medical Sciences, Iran, and the Department of Biology at Islamic Azad University, Iran. The paper was published in 2025 in The Journal of Antibiotics, with the DOI 10.1038/s41429-025-00806-x.

Research Process

1. Bacterial Identification

The study collected 40 clinical isolates of P. aeruginosa from burn patients at Shahid Motahari Burn Hospital in Tehran, Iran. All strains were identified using conventional biochemical methods, including Gram staining, oxidase testing, and IMViC tests. The strains were preserved in glycerol-TSB medium at -70°C for subsequent molecular analysis.

2. Antimicrobial Susceptibility Testing (AST)

Antibiotic susceptibility testing was performed using the disk diffusion method following CLSI 2021 guidelines. The antibiotics tested included imipenem, ceftazidime, piperacillin/tazobactam, ciprofloxacin, and amikacin. The results showed that 90% of the isolates were resistant to imipenem, and 95% were classified as multidrug-resistant (MDR).

3. Modified Carbapenem Inactivation Method (MCIM)

The MCIM test was conducted according to CLSI 2023 guidelines. Bacterial suspensions were incubated with meropenem disks for 4 hours, after which the disks were transferred to MHA plates inoculated with E. coli ATCC 25922 and incubated for 18-24 hours. The inhibition zones were then observed. The results showed that 50% of the isolates were carbapenemase-positive, and 2.5% had indeterminate results.

4. Simplified Carbapenem Inactivation Method (SCIM)

The SCIM test is a simplified version of MCIM, where bacteria are directly smeared onto imipenem disks, which are then placed on MHA plates inoculated with E. coli ATCC 25922. However, the SCIM test failed to detect any carbapenemase-positive isolates in this study, indicating a lack of standardization.

5. PCR Detection

Bacterial DNA was extracted using the boiling method, and PCR was performed to detect blaIMP, blaVIM, and blaNDM genes. The results showed that 25% of the isolates carried the blaNDM gene, 12.5% carried blaVIM, and 2.5% carried blaIMP.

6. Statistical Analysis

Statistical analysis was performed using SPSS 27.0 and MedCalc to calculate the sensitivity, specificity, positive predictive value, and negative predictive value of the MCIM test. The sensitivity of MCIM for blaVIM and blaIMP genes was 100%, while for blaNDM, it was 80%, with an overall sensitivity of 87.50%.

Key Findings

1. Antibiotic Resistance: 90% of P. aeruginosa isolates were resistant to imipenem, and 95% were classified as multidrug-resistant.
2. MCIM Test: The overall sensitivity of MCIM for detecting metallo-β-lactamases was 87.50%, with a specificity of 70.83% and a negative predictive value of 89.47%.
3. SCIM Test: The SCIM test failed to detect any carbapenemase-positive isolates in this study, indicating a lack of standardization.
4. PCR Detection: 25% of the isolates carried the blaNDM gene, 12.5% carried blaVIM, and 2.5% carried blaIMP.

Conclusions and Significance

This study highlights the high antibiotic resistance of P. aeruginosa in burn patients, particularly to carbapenems. The MCIM test demonstrated high sensitivity and negative predictive value for detecting metallo-β-lactamases, while the SCIM test failed to provide reliable results due to standardization issues. Although PCR remains the gold standard for detecting carbapenemase genes, MCIM offers a cost-effective and easy-to-use phenotypic screening tool with significant clinical value. Future research should focus on optimizing the SCIM test to improve its reliability in clinical diagnostics.

Research Highlights

1. High Resistance Rates: 90% of P. aeruginosa isolates were resistant to imipenem, and 95% were multidrug-resistant, underscoring the severity of antibiotic resistance in burn patients.
2. Effectiveness of MCIM: The MCIM test demonstrated high sensitivity and negative predictive value for detecting metallo-β-lactamases, providing a rapid screening tool for clinical use.
3. Limitations of SCIM: The SCIM test failed to detect any carbapenemase-positive isolates in this study, indicating a need for further optimization and standardization.
4. Genetic Detection: PCR revealed that 25% of the isolates carried blaNDM, 12.5% carried blaVIM, and 2.5% carried blaIMP, providing critical molecular insights for clinical treatment.

Additional Valuable Information

This study also emphasizes the importance of implementing targeted antimicrobial strategies in burn centers to reduce the spread and infection of multidrug-resistant strains. Future research could further explore the application of the EDTA-modified carbapenem inactivation method (ECIM) in P. aeruginosa to enhance the specificity of metallo-β-lactamase detection.