title: Development of cross-priming amplification assays for rapid and sensitive detection of Aeromonas hydrophila.
abstract: Aeromonas hydrophila has been increasingly implicated as the aetiologic agent of various human diseases. Therefore, reliable laboratory detection and identification of this bacterium has become clinically and epidemiologically desirable. We developed a nearly instrument-free, simple molecular method for rapid detection of Aer. hydrophila using a cross-priming amplification (CPA) assay with the desA gene as the target. The desA gene is crucial for the survival and growth of Aer. hydrophila under iron starvation. The results can be visualized as colour changes without opening the reaction tubes. No false-positive results were observed for the 33 non-Aer. hydrophila strains tested to evaluate assay specificity. The limit of detection for Aer. hydrophila was approximately 200 copies of desA per reaction (on reference plasmids) and 5 × 10(3)  CFU g(-1) Aer. hydrophila in simulated human stool, which is the same sensitivity as a qPCR assay. The performance of the CPA assay was also evaluated with 100 stool specimens from diarrhoea patients and 40 environmental water samples. In conclusion, the simplicity, cost-effectiveness and nearly instrument-free platform of the CPA assay make it practical for use in primary care facilities and smaller clinical laboratories.
  
  
  CONCLUSIONS
  Aeromonas hydrophila is a human pathogen that infects via exposed wounds or ingestion of contaminated water and food. In this study, a CPA-based PCR method was developed for specific, rapid, cost-effective detection of Aer. hydrophila, and the test results could be visualized without opening the reaction tubes. This is the first report on the application of the CPA method for the detection of Aer. hydrophila. This novel method could be practical for use in primary care facilities and smaller clinical laboratories.
authors.affiliation: State Key Laboratory for Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, China CDC, Beijing, China.|Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Hangzhou, China.
authors.forename: S
authors.initials: S
authors.lastname: Meng
authors.affiliation: State Key Laboratory for Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, China CDC, Beijing, China.|Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Hangzhou, China.
authors.forename: Y
authors.initials: Y
authors.lastname: Wang
authors.affiliation: State Key Laboratory for Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, China CDC, Beijing, China.|Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Hangzhou, China.
authors.forename: Y
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Pathogenic Biology Institute, University of South China, Hengyang, Hunan, China.
authors.forename: D
authors.initials: D
authors.lastname: Liu
authors.affiliation: State Key Laboratory for Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, China CDC, Beijing, China.|Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Hangzhou, China.
authors.forename: C
authors.initials: C
authors.lastname: Ye
mesh_terms: D016980:Aeromonas hydrophila / Q000235:genetics / Q000302:isolation & purification*; D001426:Bacterial Proteins / Q000235:genetics*; D005243:Feces / Q000382:microbiology*; D016905:Gram-Negative Bacterial Infections / Q000175:diagnosis* / Q000382:microbiology; D006801:Humans; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
chemical_list: D001426:Bacterial Proteins
keywords: Aeromonas hydrophila; cross-priming amplification; desA gene amplification; detection; quantitative PCR