Antibiotic-affinity strategy for bioluminescent detection of viable Gram-positive bacteria using daptomycin as recognition agent

•A bioluminescent protocol was developed for the detection of viable Gram-positive bacteria.•Daptomycin was used to functionalize magnetic beads for Gram-positive bacteria capture.•It could exclude the interference from both Gram-negative bacteria and dead Gram-positive bacteria.

A bioluminescent method was proposed for rapid detection of viable Gram-positive bacteria based on a novel antibiotic-affinity strategy on a magnetic beads (MBs) platform. Daptomycin, a highly efficient lipopeptide antibiotic for Gram-positive bacteria, was used as a recognition agent to functionalize MBs. The daptomycin-functionalized MBs showed high capture and concentration efficiency for Gram-positive bacteria due to the strong binding between daptomycin and bacterial cell membrane in the presence of Ca2+ ion. The captured bacteria were lysed by hexadecyl trimethyl ammonium bromide solution, followed by a bioluminescent detection of the released intracellular adenosine triphosphate. Four Gram-positive bacteria, including Staphylococcus aureus, Streptococcus mutans, Bacillus subtilis and Staphylococcus epidermidis, were detected as model bacteria by this method. Under the optimal conditions, the bacteria could be detected within a linear range of 1.0 × 102-3.0 × 106 CFU mL−1, with a detection limit of 33 CFU mL−1. The whole detection procedure could be completed within 20 min. Gram-negative bacteria and dead Gram-positive bacteria showed negligible interference to the detection of viable Gram-positive bacteria. The proposed method was successfully applied to quantify the amount of viable Gram-positive bacteria in cheese, milk, lake water, human urine and physiological saline injection with acceptable recovery values ranging from 75.0% to 120.0%. The strategy possessed some advantages such as high sensitivity, short assay time and simple operation, thus showed great promise for food hygiene, environment monitoring, clinical diagnosis and drug safety.

Graphical abstractA bioluminescent method was developed for the detection of viable Gram-positive bacteria captured by daptomycin-functionalized magnetic beads.Download high-res image (256KB)Download full-size image