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| A Rapid Sensitive Detection Method of Pathogen Edwardsiella tarda |
| DIAO Jing,YANG Xiu-sheng,YE Hai-bin,YU Xiao-qing,XU La,LI Tian-bao,WANG Yong-qiang |
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Abstract Edw ardsiella tarda as an etiological agent of edwardsiellosis ,a serious systematic disease ,is responsible for large economic losses in aquaculture .Recently ,a number of E .tarda strains isolated have exhibited resistance to multiple antibiotics ,and lead to difficulty to effective clinic therapy .In order to exert effective and in-time therapeutic measures against edwardsiellosis ,rapid antimicrobial susceptibility testing methods are urgently needed to be developed for scientific and reasonable selection of antibiotics in therapy .Based on MH broth ,enrichment broth for E.tarda strain JF-1 was optimized by addition of fish peptone and inorganic salt ions ,and Alamar Blue was employed as the color indicator ,and 11 kinds of antibiotics were inoculated into the 96-well microplates at 8 different concentrations by doubling dilution for construction of rapid antibiotic sensitivity testing (AST) plate which contained AST region ,growth control region and blank region .Compared with the color-developing method and spectrophotometric assay ,the strain JF-1 suspension at 1 × 105 cells/mL as optimal inoculation concentration was inoculated into the prepared rapid AST plate ,and detected by a spectrophotometric assay and tube dilution method for AST in parallel as comparison .12 h post inoculation , the MIC data obtained by color-developing method on the rapid AST plate was in accordance with the results acquired by spectrophotometric assay , and showed a little deviation with the results obtained by tube dilution method at 24 h post inoculation . The results showed that the rapid AST method developed based on color-developing was well supported by spectrophotometric assay and tube dilution method ,although there is a little deviation between the results obtained from tube dilution method and the rapid AST method . The wells in the microplate have intrinsically a large surface-to-volume (S/V) ratio resulting in a higher oxygen dissolving efficiency ,which would greatly benefit the bacterial propagation and show a less susceptibility to a certain antibiotic concentration compared to the tube dilution method .The findings could be further applied to the establishment of rapid AST kit for E .tarda ,and provides a basic reference for development of simple , accurate ,effective and rapid AST approaches for aquaculture .
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Received: 25 January 2014
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