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Title: Flavobacterium branchiophilum and F. succinicans associated with bacterial gill disease in rainbow trout Oncorhynchus mykiss (Walbaum) in water recirculation aquaculture systems

Author
item GOOD, CHRISTOPHER - Freshwater Institute
item DAVIDSON, JOHN - Freshwater Institute
item Wiens, Gregory - Greg
item Welch, Timothy - Tim
item SUMMERFELT, STEVEN - Freshwater Institute

Submitted to: Journal of Fish Diseases
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 2/21/2014
Publication Date: 4/10/2014
Citation: Good, C., Davidson, J., Wiens, G.D., Welch, T.J., Summerfelt, S. 2014. Flavobacterium branchiophilum and F. succinicans associated with bacterial gill disease in rainbow trout Oncorhynchus mykiss (Walbaum) in water recirculation aquaculture systems. Journal of Fish Diseases. DOI: 10.1111/jfd.12249.

Interpretive Summary: Sought to identify bacterial species associated with bacterial gill disease (BGD) in rainbow trout Oncorhynchus mykiss in replicated water recirculation aquaculture systems (WRAS). By manipulating environmental conditions, over time we were able to instigate BGD in three WRAS, and during each BGD episode gill samples were collected from affected fish, unaffected fish in the same system, and comparison fish in ozonated WRAS and flow-through tanks. Statistical analyses revealed that Flavobacterium branchiophilum was the species most associated with BGD; Flavobacterium succinicans, while not at the same abundance levels as F. branchiophilum, was nonetheless significantly associated with BGD. Our results support the assertion that F. branchiophilum is the major bacterial species associated with BGD, and provides evidence that F. succinicans might also play a role in the development and progression of this disease.

Technical Abstract: Raised rainbow trout Oncorhynchus mykiss in six replicated water recirculation aquaculture systems (WRAS), and manipulated environmental conditions to promote bacterial gill disease (BGD). For each episode of BGD, gill tissue was sampling from affected fish, unaffected fish within the same WRAS, and comparison fish from nearby ozonated and flow-through systems. Bacterial DNA was then extracted from frozen gill samples and used for PCR amplification of the V1-V3 regions of 16S rRNA genes; purified amplicons were pooled and subjected to pyrosequencing using the Roche 454 GS-FLX Titanium chemistry. Operational Taxonomic Unit (OTU) pipeline yielded individual spreadsheets for each OTU-cutoff from 97% (Species), 95% (Genus), 90% (family/class) and 80% (phylum) with OTU abundance counts for each sample. Despite bacterial DNA contamination during sample processing, all Flavobacterium sequences were considered unrelated to the processing contamination, and were analyzed independently of the other identified bacterial genera. Overall, there were 75 species-level OTUs of the Flavobacterium genus, with a total of 6,305 reads among all samples. Of the 75 Flavobacterium OTUs, 5,253 reads (83.3%) were associated with two OTUs; best hit-to-species, using BLASTn against the NCBI non-redundant nucleotide database, revealed that these two most prevalent species were F. branchiophilum and F. succinicans. Both species were significantly associated (p<0.05 by ANOVA) with BGD-affected fish; additionally, F. branchiophilum was significantly more prevalent in random non-affected fish sampled from BGD tanks than those sampled from ozonated WRAS and flow-through tanks. These results confirm that F. branchiophilum is the dominant bacterial species present on the gills of rainbow trout affected by natural, environmentally-induced BGD. The detection of F. succinicans at significantly higher levels in BGD-affected fish, in conjunction with significantly higher levels of F. branchiophilum, is also novel, and supports the assertion that F. succinicans may be a commensal species with the potential to act as an opportunistic pathogen under certain conditions (e.g. BGD).