Submitted to: World Aquaculture Society Meeting
Publication Type: Abstract Only
Publication Acceptance Date: 8/5/2003
Publication Date: 3/7/2004
Citation: Welker, T.L., Arias, C.R., Shoemaker, C.A., Klesius, P.H. 2004. Detection of flavobacterium columnare by pcr. World Aquaculture Society Meeting.
Technical Abstract: Flavobacterium columnare is the causal agent of columnaris disease, probably the most important bacterial disease of freshwater fish species. This bacterium is ubiquitous in fresh water environments around the world, affecting both wild and cultured fish. Columnaris disease is especially important in the Southeastern U.S., where negatively impacts the catfish industry. Currently, F. columnare is considered the most important bacterial pathogen in commercial culture of channel catfish. However, little is known about the ecology and epidemiology of F. columnare in catfish aquaculture. Flavobacterium columnare presents a fastidious growing character on standard culture medium, and it is easily overgrown by accompanying microbiota. Therefore, isolation and identification of F. columnare from environmental samples is cumbersome and usually underestimates F. columnare numbers. A sensitive, specific, and rapid identification method of F. columnare from environmental samples is needed in order to assess F. columnare's negative impact on catfish aquaculture. Polymerase chain reaction (PCR) constitutes on the best tool to carry out specific, sensitive, and reliable identification of bacteria from numerous samples in a short period of time. Our group has developed a PCR-based assay to detect F. columnare in fish and water samples less than 8 h. Flavobacterium columnare specific PCR primers were designed using the Intergenic Spacer Region (ISR) contained in the ribosomic operon. Primers specificity was tested against 12 strains belonging to other Flavabacterium species, as well as, other related genera. Sensitivity of the PCR assay was tested by adding known numbers of F. columnare cells to water samples prior to DNA extraction. Our PCR assay was able to detect as low as 10 CFU/ml. Once the sensitivity and specificity of the detection method was establish we applied it to artificially infected fish and water samples. Correlation between classical culture methods and PCR detection was good in all the experiments. However, PCR proved to be a more sensitive and much quicker method than standard procedures for detection of F. columnare from aquatic samples.