Combining immunomagnetic beads and the phage lysin LysGH15 for fluorescence detection of Staphylococcus aureus

Abstract

Background

Staphylococcus aureus (S. aureus) is one of the most important pathogens that causes foodborne illnesses. The development of new methods for the early detection and surveillance of S. aureus is key to preventing and controlling the occurrence of foodborne diseases.

Results

In this study, the C54A mutation of LysGH15 (LysGH15-C54A) of the S. aureus bacteriophage lysin was fused with enhanced green fluorescent protein (EGFP) to construct the recombinant protein LysGH15-C54A-EGFP. A fluorescence (FL) detection protocol using LysGH15-C54A-EGFP as the FL signal carrier and LysGH15-C54A coupled with immunomagnetic beads (LysGH15-C54A-IMBs) as the isolating element and capturing agent for the detection of S. aureus was established on the basis of the high specificity and efficiency of LysGH15-C54A in recognizing S. aureus. LysGH15-C54A-IMBs were able to specifically separate and capture S. aureus, and LysGH15-C54A-EGFP was used to combine the captured surface vacancies of S. aureus to achieve FL detection of S. aureus using two-site recognition. When S. aureus was detected in the concentration range of 10³–10⁶ CFU/mL, the FL method showed good linearity with a detection limit of 85 CFU/mL, and the whole process lasted less than 1 h. The procedure was successfully applied to quantify S. aureus in fresh milk samples, and the recovery values ranged from 93.09 % to 101.04 %.

Significance

Based on these results, we believe that LysGH15-C54A-based FL detection method hold great potential for rapid, accurate, and specific diagnosis of S. aureus, and it has good prospects in practical applications.