High-resolution melting analysis of gyrA codon 84 and grlA codon 80 mutations conferring resistance to...

Area of Science:

• Veterinary Microbiology
• Molecular Diagnostics
• Antimicrobial Resistance

Background:

• Staphylococcus pseudintermedius is a significant opportunistic pathogen in companion animals.
• Fluoroquinolone (FQ) resistance is a growing concern in treating S. pseudintermedius infections.
• Accurate and rapid susceptibility testing is crucial for effective antimicrobial therapy.

Purpose of the Study:

• To develop and validate a high-resolution melting analysis (HRMA) protocol for detecting FQ resistance in S. pseudintermedius.
• To correlate HRMA results with known FQ susceptibility and specific gyrA/grlA gene mutations.
• To assess the potential of HRMA as a tool for guiding FQ therapy in veterinary medicine.

Main Methods:

• A novel high-resolution melting analysis (HRMA) protocol was designed.
• The HRMA assay was tested on 42 clinical isolates of S. pseudintermedius.
• Isolates had pre-determined fluoroquinolone (FQ) susceptibility and mutation status at gyrA codon 84 and grlA codon 80.

Main Results:

• The HRMA protocol successfully differentiated between FQ-sensitive and FQ-resistant S. pseudintermedius strains.
• The assay confirmed that the primary mutation site for FQ resistance is at position 251 (Ser84Leu) of the gyrA gene.
• HRMA demonstrated potential for predicting FQ resistance based on genetic markers.

Conclusions:

• HRMA is a promising method for rapid FQ susceptibility profiling of S. pseudintermedius.
• This molecular approach can aid in guiding FQ therapy for canine and feline infections.
• Further validation with a larger number of isolates is recommended to confirm the assay's resistance-predictive power.