APPLICATIONS OF Flectobacillus major AND ITS SPHINGOLIPIDS IN THE MODULATION OF FISH HEALTH

Ali Sepahi*, Mariah Sanchez, and Irene Salinas
 
University of New Mexico
Biology Department
Center for Evolutionary and Theoretical Immunology
Albuquerque, NM87131
sepahi@unm.edu

Commensal bacteria influence most physiological systems of animals. Farmed fish have a diverse microbial community living in association with each mucosal surface. In hatchery reared adult rainbow trout (Oncorhynchus mykiss), the skin and gill bacterial microbiome is dominated by the Gram-negative bacterium Flectobacillus major. F. major has the unique ability to synthesize sphingolipids of a rare molecular structure. We hypothesized that F. major sphingolipids influence the immune system of rainbow trout and they can be applied in aquaculture settings to control inflammation as well as infectious diseases.

F. major sphingolipids were extracted and purified according to previously established protocols. Sphingolipids were combined with bovine serum albumin to avoid cell toxicity issues. Trout gills and head-kidney (HK) leucocytes were incubated in vitro with live F. major cells at two different doses (102 cfu/ml and 104 cfu/ml). Changes in immune gene expression were measured 6, 24 and 48 h later. Specifically, IgT, IgM, pro-inflammatory and anti-inflammatory cytokine expression levels were quantified by RT-qPCR. In vivo administration of F. major sphingolipids was performed via two different routes, intraperitoneally (i.p) and intravenously (i.v). Expression of the same immune genes was evaluated 60 hours post-treatment in the gills and head-kidney.

In vitro results show that, compared to unstimulated controls, live F. major stimulates IgT expression in gills but not HK in a sustained and dose-dependent manner, whereas IgM expression stimulation is transient (Figure 1). Measurement of pro-inflammatory and anti-inflammatory cytokine gene expression in vitro and in vivo is currently being analyzed. Future studies will evaluate how the administration of F. major sphingolipids affects disease resistance in rainbow trout.