Quantification of live Gram-positive bacteria via employing artificial antibacterial peptide-coated magnetic spheres as isolation carriers

A novel artificial antibacterial peptide was prepared by minor modification of the amino acids sequence of a natural peptide protonectin. Distinct from protonectin with natural broad-spectrum antibacterial activity, the designed peptide named as Phe(11)-protonectin displayed obviously improved selectivity towards Gram-positive (G(+)) bacteria. Adopting Phe(11)-protonectin as the recognition reagent, a bioluminescent method was established to quantify live G(+) bacteria. Phe(11)-protonectin-coated magnetic spheres were employed as carriers for isolating G(+) bacteria from sample matrixes. The isolated bacteria were lysed to release intracellular adenosine triphosphate, followed by bioluminescent signal collection for quantifying the amount of live G(+) bacteria. Staphylococcus aureus, Streptococcus mutans and Bacillus cereus were adopted as model bacteria to explore the feasibility of this method. The linear range for G(+ )bacteria was 2.3 x 10(3) - 1.2 x 10(7) CFU mL(-1), and the limit of detection was 2.2 x 10(2) CFU-1 (3 sigma). The quantification process composed of bacteria isolation, bacteria lysis and signal detection was completed within 33 min. Both Gram-negative and dead G(+) bacteria displayed very minor interference to the method for quantifying live G(+) bacteria. It has been successfully employed for four types of sample matrixes including peach juice, glucose injection, human urine and lake water, revealing its application prospect in food and drug safety control, clinical diagnosis and hygiene monitoring.