Daptomycin-modified magnetic beads integrated with lysostaphin for selective analysis of Staphylococcus

An antibiotic-affinity method was developed for analyzing Staphylococcus on the basis of the strong binding capability of daptomycin towards Gram-positive bacteria cellular membrane, as well as the selective lytic action of lysostaphin towards Staphylococcus. Daptomycin-modified magnetic beads were adopted to enrich Staphylococcus from sample matrix. Afterwards lysostaphin was adopted to lyse Staphylococcus, which can hydrolyze pentaglycine cross-linkers of peptidoglycan composing the cellular wall of Staphylococcus. The concentration of Staphylococcus was quantified by collecting the bioluminescent signal of the released intracellular adenosine triphosphate of the enriched Staphylococcus. Staphylococcus aureus (S. aureus) was analyzed as a model bacterium to study the feasibility of the proof-of-principle work. For bioluminescent analysis of S. aureus with the developed method, the linear range was 5.0 × 10²-5.0 × 10⁶ colony forming units mL^-1, and the limit of detection was 3.8 × 10² colony forming units mL^-1. The analytical procedure consisting of bacterial enrichment, cell lysis and signal collection can be accomplished within 20 min. Some common Gram-positive bacteria and Gram-negative bacteria all indicated very low interference to the analysis of the target bacterium. It has been successfully used to analyze S. aureus in milk as well as physiological saline injection, indicating its application potential for real samples.