Carbapenemase-Encoding Gene Dynamics in Carbapenem-Resistant Enterobacter cloacae: Insights from Guangdong Hospitals

Study Background and Research Question

The global rise of carbapenem-resistant Enterobacteriaceae remains a critical threat to healthcare systems, particularly as multidrug-resistant organisms proliferate in hospital settings. Among these, Enterobacter cloacae has become increasingly prevalent as a cause of nosocomial infections. The COVID-19 pandemic exacerbated this challenge by increasing antibiotic consumption, disrupting routine infection control, and presenting complex patient profiles. However, there is a scarcity of detailed data on the molecular characteristics and transmission dynamics of carbapenemase-encoding genes (CEGs) in Enterobacter cloacae during this period. The recent study by Chen et al. addresses this gap by systematically characterizing CEGs in carbapenem-resistant E. cloacae (CREC) isolates from eight teaching hospitals in Guangdong province, China, collected between December 2022 and June 2024 (Chen et al., 2025).

Key Innovation from the Reference Study

The central innovation of this research lies in its comprehensive molecular profiling of CEGs, combined with an in-depth analysis of their horizontal and vertical transmission potential in a real-world, post-pandemic hospital environment. By simultaneously examining plasmid- and chromosome-associated resistance determinants, the study provides new insights into how multidrug resistance propagates within and between healthcare institutions. The work also correlates resistance phenotypes with epidemiological factors, such as patient age, gender, clinical department, and sample type, offering a granular understanding of CREC dissemination in clinical settings.

Methods and Experimental Design Insights

Chen et al. analyzed 54 non-duplicate CREC isolates obtained over an 18-month period from eight tertiary hospitals. The isolates underwent molecular characterization using the variable temperature sodium dodecyl sulfate (SDS) plasmid elimination method and PCR to detect the presence and location of key carbapenemase genes (notably bla_NDM-1, bla_IMP, and bla_KPC-2). Broth microdilution assays determined antimicrobial susceptibility profiles, and plasmid conjugation experiments assessed the transferability of CEGs. Genetic diversity was mapped using ERIC-PCR and analyzed with NTSYS software, while the prevalence of mobile genetic elements was also quantified.

Protocol Parameters

• Sample collection: Non-duplicate CREC isolates from multiple hospital departments, December 2022–June 2024.
• CEG detection: PCR targeting bla_NDM-1, bla_IMP, bla_KPC-2; localization via plasmid elimination method.
• Antimicrobial susceptibility: Broth microdilution following CLSI guidelines; focus on imipenem, cefepime, gentamicin, ceftazidime/avibactam, ciprofloxacin, and levofloxacin.
• Plasmid transferability: Conjugation experiments to measure horizontal gene transfer rate.
• Genotyping: ERIC-PCR for strain diversity; NTSYS software for cluster analysis.
• Mobile genetic elements: PCR screening for integrons, insertion sequences (e.g., ISEcp1), and other mobility markers.

Core Findings and Why They Matter

The study found that 85.2% of CREC isolates harbored at least one CEG. The bla_NDM-1 gene was the most prevalent, detected either on both chromosomes and plasmids (33.3%) or exclusively on plasmids (46.3%). A smaller proportion carried bla_IMP or a combination of bla_NDM-1 and bla_KPC-2. The high rate of CEG-positive isolates correlated with extensive multidrug resistance, especially against cefepime, imipenem, and other key antibiotics (Chen et al., 2025).

Conjugation experiments revealed a 95.7% success rate for CEG transfer, underscoring the risk of rapid horizontal gene spread. Mobile genetic element analysis showed that the insertion sequence ISEcp1 was highly prevalent (87.0%), and 40.7% of isolates carried four types of mobile elements simultaneously. Genotyping revealed 17 distinct clusters, with dominant types found across several hospitals and departments, suggesting both local expansion and inter-institutional transmission.

From an epidemiological perspective, detection rates were highest in male and elderly patients, in respiratory medicine, and in sputum samples. These findings highlight specific clinical contexts where enhanced surveillance and control may be most urgent.

Comparison with Existing Internal Articles

This study’s molecular approach complements and extends findings from recent research on cephalosporin antibiotics in resistance modeling. For example, investigations into Cefepime (BMY-28142) have showcased its role in resistance profiling, particularly within central nervous system (CNS) infection models, and emphasized the importance of tracking carbapenemase gene transmission dynamics. Similarly, Cefepime’s blood-brain barrier penetration and broad-spectrum activity have been leveraged to design robust multidrug resistance assays. These resources offer practical protocols and troubleshooting advice for researchers aiming to simulate the complex resistance patterns observed in clinical CREC isolates.

In contrast, studies on novel β-lactam/β-lactamase inhibitor combinations (e.g., ceftolozane-tazobactam) and siderophore cephalosporins (cefiderocol) have focused on overcoming resistance in Pseudomonas and Enterobacterales, providing alternative model systems for antimicrobial activity against Gram-positive and Gram-negative bacteria. Together, these studies build a toolkit for researchers investigating CNS infection models, neurotoxicity, and the efficacy of blood-brain barrier-crossing antibiotics.

Limitations and Transferability

While Chen et al.'s study delivers a robust molecular epidemiology of CREC in a high-burden region, several limitations must be considered. The sample size, though multicentric, is modest and may not capture the full diversity of CEGs across China or other global regions. The analysis is also limited to a discrete timeframe during the COVID-19 pandemic, which may have introduced atypical selection pressures. Furthermore, phenotypic resistance was not exhaustively correlated with clinical outcomes or in vivo infection models.

Transferability of these findings is strong for hospital-based surveillance and in vitro antimicrobial resistance modeling but should be cautiously extended to community-acquired strains or non-Chinese settings without further validation.

Research Support Resources

For researchers aiming to replicate or extend this work, access to reliable reagents and validated protocols is crucial. Cefepime (BMY-28142) (SKU BA1013, APExBIO) offers a well-characterized, broad-spectrum cephalosporin antibiotic suitable for resistance profiling and modeling CNS infection scenarios, particularly when studying antimicrobial activity against Gram-positive and Gram-negative bacteria or neurotoxicity in bacterial infection models. Its established ability to cross the blood-brain barrier makes it a practical choice for multidrug resistance studies in CNS contexts. Solutions should be prepared fresh, following manufacturer guidance, and handled with care due to neurotoxicity potential.