Abstract
Abstract Background Edwardsiella tarda, the etiologic agent of edwardsiellosis, is a devastating fish pathogen prevailing in worldwide aquaculture industries and accounting for severe economic losses. There is a raising concern about E. tarda being a significant zoonotic pathogen, and it is urgent to develop a rapid detection of this pathogen. This is the first study to develop a test strip for rapid detection of E. tarda in turbot. Results Mouse monoclonal antibodies (MAbs) and rabbit polyclonal antibody (PAb) against E. tarda were generated from immunization of mice and rabbits with a virulent isolate of E. tarda EIB202. Two MAbs specific to isolates of E. tarda were obtained, and one of them (25C1) was selected to conjugate with colloidal gold as the detector antibody. Rabbit PAb was used as the capture antibody. It was found the strip had no cross-reactivity with non-E. tarda bacterial microbes and the limit of detection (LOD) was 1 × 105 colony-forming units (CFU)/ml. The detection could be visually observed by the naked eye within 5 min. This test strip was verified with a similar detection limit and much less analysis time compared with a dot blot immunoassay (1 × 105 CFU/ml for LOD and 120 min for reaction time). When the samples were mixed with turbot tissue homogenates, strong immunoreactivity was observed over 105 CFU/ml, which suggested that the turbot tissue homogenates did not affect the detection of the strip. Pre-enrichment with homogenized turbot tissue for 12 h could increase the detection limit of the E. tarda present in the sample up to 1 to 10 CFU/ml. In practice, in detecting 20 turbot ascite samples infected by E. tarda, the immunochromatographic test strip showed a high accuracy (100% positive). Conclusions The immunochromatographic test strip offers great promise for a rapid, simple, and economical method of E. tarda on-site detection, and with different antibodies, it might be used to detect other aquatic pathogens.
Highlights
Edwardsiella tarda, the etiologic agent of edwardsiellosis, is a devastating fish pathogen prevailing in worldwide aquaculture industries and accounting for severe economic losses
E. tarda strains were grown in tryptic soy broth (TSB) (Difco, Detroit, MI, USA) or tryptic soy agar (TSA) (Difco, Detroit, MI, USA) at 28°C, while Escherichia coli strains were cultured in Luria-Bertani (LB) broth at 37°C
Preparation and characteristics of antibodies against E. tarda In order to obtain monoclonal antibodies (MAbs), BALB/c mice were immunized with the formalin-killed bacteria and purified outer membrane proteins (OMPs) of E. tarda, respectively
Summary
Edwardsiella tarda, the etiologic agent of edwardsiellosis, is a devastating fish pathogen prevailing in worldwide aquaculture industries and accounting for severe economic losses. There is a raising concern about E. tarda being a significant zoonotic pathogen, and it is urgent to develop a rapid detection of this pathogen. This is the first study to develop a test strip for rapid detection of E. tarda in turbot. There is a raising concern about E. tarda being a significant fish pathogen, and it is urgent to develop a rapid detection of this pathogen. The traditional diagnostic test available for identification of E. tarda in diseased fish is bacterial culture and identification using. The infecting pathogen of diseased fish could be determined on-site in several minutes without any instrument
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