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Rapid Detection of vanA Resistance Gene from E. faecalis Clinical Isolates Using Duplex Loop-Mediated Isothermal
Mohsen Azizi1, Hamid Motamedi1,2, Hadi Hossainpour1,2
1Department of Microbiology, School of Medicine, Kermanshah University of Medical Sciences, Kermanshah, Iran.
Abstract:
Today, the spread of vancomycin-resistant strains isolated from Enterococcus faecalis (E. faecalis) has become a major health concern worldwide. Therefore, it is essential to provide a rapid and sensitive assay for identifying vanA gene for timely and appropriate antimicrobial control of resistant enterococcal infections. For this purpose, a cross-sectional study was performed on different clinical specimens of enterococci from Imam Reza hospital, Kermanshah, Iran. The antimicrobial susceptibility testing was determined by disk diffusion and MIC methods. Triplex-PCR and duplex-LAMP assays were also used to identify vanA E. faecalis resistance gene isolates. The results of this study shown that out of 108 Enterococcus isolates, 86, 18, 2, 1, and one isolates of E. faecalis, E. faecium, E. avium, E. psudoavium, and E. raffinosus were identified, respectively. On the other hand, E. faecalis was confirmed in 87 and 88 isolates using duplex-LAMP and triplex PCR, respectively. The LAMP primer set designed in this study can reliably identify seven distinct regions of the vanA gene, and finally the sensitivity, specificity, and the positive and negative predictive values of LAMP assay were shown to be 94.19%, 72.73%, 76.19%, and 93.10%, respectively. In general, sample processing, isothermal reaction and result reporting were completed using the LAMP assay in 75 minutes. Our findings suggest that LAMP assay has been approved as an alternative to the vancomycin resistance Enterococcus genotype (vanA and vanB) compared to other methods and has the advantage of being rapid, time-consuming, and easy for diagnosis.
Insights
A new LAMP assay rapidly and accurately detects the vanA gene in Enterococcus faecalis, offering a faster alternative for identifying vancomycin-resistant enterococcal infections worldwide.
Area of Science:
- Microbiology
- Molecular Biology
- Clinical Diagnostics
Background:
- Vancomycin-resistant Enterococcus faecalis (VRE) is a global health threat.
- Rapid identification of the vanA gene is crucial for managing VRE infections.
Purpose of the Study:
- To develop and evaluate a rapid and sensitive assay for identifying the vanA gene in E. faecalis.
- To compare the performance of the developed assay with existing methods.
Main Methods:
- Antimicrobial susceptibility testing (disk diffusion, MIC).
- Triplex-PCR and duplex-LAMP assays for vanA gene detection.
- Analysis of clinical enterococcal isolates.
Main Results:
- Out of 108 Enterococcus isolates, E. faecalis was the most prevalent (86 isolates).
- Duplex-LAMP and triplex-PCR confirmed E. faecalis in 87 and 88 isolates, respectively.
- The LAMP assay demonstrated high sensitivity (94.19%), specificity (72.73%), and completed results in 75 minutes.
Conclusions:
- The developed LAMP assay is a rapid, sensitive, and efficient alternative for detecting the vanA gene.
- This assay facilitates timely diagnosis and management of vancomycin-resistant enterococcal infections.
- LAMP assay offers advantages in speed, simplicity, and cost-effectiveness for clinical diagnostics.
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