title: Highly Specific Bacteriophage-Affinity Strategy for Rapid Separation and Sensitive Detection of Viable Pseudomonas aeruginosa.
abstract: A virulent bacteriophage highly specific to Pseudomonas aeruginosa (P. aeruginosa) was isolated from hospital sewage using a lambda bacteriophage isolation protocol. The bacteriophage, named as PAP1, was used to functionalize tosyl-activated magnetic beads to establish a bacteriophage-affinity strategy for separation and detection of viable P. aeruginosa. Recognition of the target bacteria by tail fibers and baseplate of the bacteriophage led to capture of P. aeruginosa onto the magnetic beads. After a replication cycle of about 100 min, the progenies lysed the target bacteria and released the intracellular adenosine triphosphate. Subsequently, firefly luciferase-adenosine triphosphate bioluminescence system was used to quantitate the amount of P. aeruginosa. This bacteriophage-affinity strategy for viable P. aeruginosa detection showed a linear range of 6.0 × 102to 3.0 × 105CFU mL-1, with a detection limit of 2.0 × 102CFU mL-1. The whole process for separation and detection could be completed after bacteria capture, bacteriophage replication, and bacteria lysis within 2 h. Since the isolated bacteriophage recognized the target bacteria with very high specificity, the proposed strategy did not show any signal response to all of the tested interfering bacteria. Furthermore, it excluded the interference from inactivated P. aeruginosa because the bacteriophage could replicate only in viable cells. The proposed strategy had been applied for detection of P. aeruginosa in glucose injection, human urine, and rat plasma. In the further work, this facile bacteriophage-affinity strategy could be extended for detection of other pathogens by utilizing virulent bacteriophage specific to other targets.
authors.affiliation: Key Laboratory of Luminescence and Real-Time Analytical Chemistry (Ministry of Education), College of Pharmaceutical Sciences, Southwest University , Chongqing 400716, China.|Department of Pharmacy, Affiliated Hospital of Zunyi Medical College , Zunyi 563000, China.
authors.forename: Yong
authors.initials: Y
authors.lastname: He
authors.affiliation: Key Laboratory of Luminescence and Real-Time Analytical Chemistry (Ministry of Education), College of Pharmaceutical Sciences, Southwest University , Chongqing 400716, China.
authors.forename: Mengyao
authors.initials: M
authors.lastname: Wang
authors.affiliation: Key Laboratory of Luminescence and Real-Time Analytical Chemistry (Ministry of Education), College of Pharmaceutical Sciences, Southwest University , Chongqing 400716, China.
authors.forename: Enci
authors.initials: E
authors.lastname: Fan
authors.affiliation: Key Laboratory of Luminescence and Real-Time Analytical Chemistry (Ministry of Education), College of Pharmaceutical Sciences, Southwest University , Chongqing 400716, China.
authors.forename: Hui
authors.initials: H
authors.lastname: Ouyang
authors.affiliation: Key Laboratory of Luminescence and Real-Time Analytical Chemistry (Ministry of Education), College of Pharmaceutical Sciences, Southwest University , Chongqing 400716, China.
authors.forename: Huan
authors.initials: H
authors.lastname: Yue
authors.affiliation: Key Laboratory of Luminescence and Real-Time Analytical Chemistry (Ministry of Education), College of Pharmaceutical Sciences, Southwest University , Chongqing 400716, China.
authors.forename: Xiaoxiao
authors.initials: X
authors.lastname: Su
authors.affiliation: Key Laboratory of Luminescence and Real-Time Analytical Chemistry (Ministry of Education), College of Pharmaceutical Sciences, Southwest University , Chongqing 400716, China.
authors.forename: Guojian
authors.initials: G
authors.lastname: Liao
authors.affiliation: Key Laboratory of Luminescence and Real-Time Analytical Chemistry (Ministry of Education), College of Pharmaceutical Sciences, Southwest University , Chongqing 400716, China.
authors.forename: Lin
authors.initials: L
authors.lastname: Wang
authors.affiliation: Department of Microbiology, College of Basic Medical Science, Third Military Medical University , Chongqing 400038, China.
authors.forename: Shuguang
authors.initials: S
authors.lastname: Lu
authors.affiliation: Key Laboratory of Luminescence and Real-Time Analytical Chemistry (Ministry of Education), College of Pharmaceutical Sciences, Southwest University , Chongqing 400716, China.
authors.forename: Zhifeng
authors.identifier: 0000-0002-8238-7987
authors.initials: Z
authors.lastname: Fu
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D015374:Biosensing Techniques / Q000379:methods*; D006090:Gram-Negative Bacteria / Q000302:isolation & purification; D006094:Gram-Positive Bacteria / Q000302:isolation & purification; D006801:Humans; D018189:Immunomagnetic Separation / Q000379:methods*; D057230:Limit of Detection; D049449:Luminescence; D018613:Microscopy, Confocal; D008855:Microscopy, Electron, Scanning; D046529:Microscopy, Electron, Transmission; D008856:Microscopy, Fluorescence; D017105:Pseudomonas Phages / Q000472:pathogenicity / Q000502:physiology* / Q000648:ultrastructure; D011550:Pseudomonas aeruginosa / Q000302:isolation & purification* / Q000378:metabolism; D017207:Rats, Sprague-Dawley; D014774:Virulence
chemical_list: D000255:Adenosine Triphosphate