title: Point-of-care and visual detection of P. aeruginosa and its toxin genes by multiple LAMP and lateral flow nucleic acid biosensor.
abstract: This study describes a simple and sensitive approach for visual and point-of-care detection of P. aeruginosa and its toxin genes based on multiple loop-mediated isothermal amplification (mLAMP) and lateral flow nucleic acid biosensor (LFNAB). Differentiation of the internal standard gene ecfX and toxin genes (ExoS and ExoU) in P. aeruginosa was determined using FITC-, hex-and digoxin-modified primers in the mLAMP process. In the presence of biotin-and FITC- (hex-, digoxin-) modified primers and Bst DNA polymerase large fragments, the mLAMP produced numerous biotin- and FITC- (hex-, digoxin-) attached duplex DNA products. The products were detected by LFNAB through dual immunoreactions (anti-biotin antibodies on the gold nanoparticle (Au-NP) and biotin on the duplex, anti-FITC (hex, digoxin) antibodies on the LFNAB test line and FITC (hex, digoxin) on the duplex). The accumulation of Au-NPs produced a characteristic red band, enabling visual detection of P. aeruginosa and its toxin genes without instrumentation. After systematic optimization of LFNAB preparation and detecting conditions, the current approach was capable of detecting concentrations as low as 20 CFU/mL P. aeruginosa or its toxin genes within 50min without complicated instrument, which is more sensitive than PCR. Therefore, this approach provides a simple, pollution free, sensitive, and low-cost point-of-care test for the detection of P. aeruginosa and its toxin genes.
authors.affiliation: Beijing Laboratory for Food Quality and Safety, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China.
authors.forename: Yuting
authors.initials: Y
authors.lastname: Chen
authors.affiliation: Beijing Advanced Innovation Center for Food Nutrition and Human Health, College of Food Science & Nutritional Engineering, China Agricultural University, Beijing 100083, China.
authors.forename: Nan
authors.initials: N
authors.lastname: Cheng
authors.affiliation: Beijing Advanced Innovation Center for Food Nutrition and Human Health, College of Food Science & Nutritional Engineering, China Agricultural University, Beijing 100083, China; Beijing Laboratory for Food Quality and Safety, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China.
authors.forename: Yuancong
authors.initials: Y
authors.lastname: Xu
authors.affiliation: Beijing Advanced Innovation Center for Food Nutrition and Human Health, College of Food Science & Nutritional Engineering, China Agricultural University, Beijing 100083, China; Beijing Laboratory for Food Quality and Safety, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China.
authors.forename: Kunlun
authors.initials: K
authors.lastname: Huang
authors.affiliation: Beijing Advanced Innovation Center for Food Nutrition and Human Health, College of Food Science & Nutritional Engineering, China Agricultural University, Beijing 100083, China; Beijing Laboratory for Food Quality and Safety, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China.
authors.forename: Yunbo
authors.initials: Y
authors.lastname: Luo
authors.affiliation: Beijing Advanced Innovation Center for Food Nutrition and Human Health, College of Food Science & Nutritional Engineering, China Agricultural University, Beijing 100083, China; Beijing Laboratory for Food Quality and Safety, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China. Electronic address: xuwentao@cau.edu.cn.
authors.forename: Wentao
authors.initials: W
authors.lastname: Xu
mesh_terms: D036002:ADP Ribose Transferases / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics*; D001427:Bacterial Toxins / Q000235:genetics*; D015374:Biosensing Techniques / Q000191:economics / Q000295:instrumentation*; D004269:DNA, Bacterial / Q000235:genetics / Q000302:isolation & purification; D060766:Drinking Water / Q000382:microbiology; D006801:Humans; D057230:Limit of Detection; D021141:Nucleic Acid Amplification Techniques / Q000191:economics / Q000295:instrumentation; D019095:Point-of-Care Systems / Q000191:economics; D011552:Pseudomonas Infections / Q000382:microbiology*; D011550:Pseudomonas aeruginosa / Q000235:genetics* / Q000302:isolation & purification; D015203:Reproducibility of Results; D045483:Rivers / Q000382:microbiology; D014118:Toxins, Biological / Q000235:genetics*; D014871:Water Microbiology
chemical_list: D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004269:DNA, Bacterial; D060766:Drinking Water; D014118:Toxins, Biological; C111749:pseudomonas exoprotein A protein, Pseudomonas aeruginosa; D036002:ADP Ribose Transferases; C444820:exoenzyme S
keywords: Lateral flow nucleic acid biosensor (LFNAB); Multiple loop-mediated isothermal amplification (mLAMP); P. aeruginosa; Toxin genes; Visual detection