Antimicrobial resistance (AMR) is a worldwide health crisis resulting in growing economic burden and increasing mortality1,2. The most important strategies to minimize AMR are the development of rapid and accurate diagnostic protocols for antimicrobial susceptibility testing (AST), which facilitating the timing of prescription to use the effective antibiotics3,4. Therefore, numerous efforts are exerted on developing rapid, sensitive and acute detection and AST for bacterial infection.
Traditional bacterial growth-based protocols are considered as the gold standard methods for bacterial detection and AST. These protocols show ideal repeatability, high standardization and good reliability. However, the isolation and identification procedure of bacterial detection usually require tedious process of 24–48 h5. The subsequent broth dilution or disk diffusion for AST performance require another bacterial growth cultured with given concentrations of various antibiotics, which demands almost another 24–48 h6,7. Lack of timely and accurate results of bacterial AST results in frequent empiric antibiotic therapy, causing irrational antibiotic use and the development of AMR8. Therefore, some other bacterial growth-based protocols are reported to aim at shortening the time of AST, such as microscopy detection9–11, electrochemical sensor12–14, phase-shift spectroscopy detection15, fluorescent detection16 and microfluidic devices based on slipchip technique17, surface-enhanced raman scattering18. Nevertheless, since lack of capacity of isolating the given bacterial species, they also require time-consuming pretreatment procedure for bacterial culture, isolation and identification. Polymerase chain reaction-based protocols gradually attract attention to bacterial detection and performance of AST due to the advantages of rapidity, sensitivity and culture-free process20–22. However, they suffer from well-trained personnel, complicated molecular manipulation, prerequisite precise resistant gene information and frequent gene mutation.
With the increasing global AMR, bacteriophages as the natural enemies to the bacteria have regained interest in solving the crisis caused by multi-resistant bacteria. Each strain of bacteria has one or more corresponding bacteriophages. Bacteriophages can highly specifically recognize their target bacteria even in harsh environment. Typical lifecycle of virulent bacteriophage involves the following steps: specific absorption on the bacterial cell wall, DNA injection into the bacterial cell, progenies replication and the lysis of bacteria cell for releasing progenies. Bacteriophages functional proteins (BFP) such as tail fiber protein (TFP), tailspike protein (TSP) and endolysin are the essential recognition elements which are responsible for absorption, injection and lysis, respectively23. Therefore, BFPs assumes the ideal molecular recognition attributes of high specificity, robustness, good anti-interference capability and universality to each bacterium23. Unfortunately, just like the bacteriophage entity, TSP and endolysin both have the inherent lytic activity which could be unfavorable for bacteria capture and sample manipulation.
In the previous work, we had utilized to the Escherichia coli (E. coli) expression system to produce the TFP of Pseudomonas aeruginosa (P. aeruginosa)24. This recombinant TFP can specifically recognize P. aeruginosa without lytic activity. To investigate the application of TFP in AST detection, TFP-functionalized magnetic particles were utilized to specifically capture P. aeruginosa, and fluorescein isothiocyanate (FITC) labeled magainin II was utilized as the fluorescent tracer. A reverse assaying protocol (RAP) combined magnetic separation was developed to specific, rapid and sensitive detection and AST of P. aeruginosa.