A BCWD-resistant line of rainbow trout exhibits higher abundance of IgT+ B cells and heavy chain tau transcripts compared to a susceptible line following challenge with Flavobacterium psychrophilum

Abstract

Bacterial Cold Water Disease (BCWD) is a common, chronic disease in rainbow trout, and is caused by the gram-negative bacterium Flavobacterium psychrophilum (Fp). Through selective breeding, the National Center for Cool and Cold Water Aquaculture has generated a genetic line that is highly resistant to Fp challenge, designated ARS-Fp-R (or R-line), as well as a susceptible “control” line, ARS-Fp-S (S-line). In previous studies, resistance to Fp had been shown to correlate with naive animal spleen size, and further, naïve R-line trout had been shown to have a lower abundance of IgM+ and IgM++ cells compared to S-line fish. Here we wished to first determine whether the abundance of IgT+ and/or IgT++ cells differed between the two lines in naïve fish, and if so, how these patterns differed after in vivo challenge with Fp. Fp challenge was by intramuscular injection of live Fp and tissue collections were on days 5, 6, and/or 28 post-challenge, in two independent challenge experiments. Flow cytometric and gene expression analyses revealed that naïve R-line fish had a higher abundance of IgT+ B cells in their anterior kidney, spleen, and blood, compared to S line fish. Further, that after Fp challenge, this difference was maintained between the two lines. Lastly, abundance of IgT+ B cells and expression of secHCtau correlated with lower Fp pathogen loads in challenged fish. In the anterior kidney, IgM+ B cell abundance correlated with increased Fp loads. Together, these results suggest that IgT+ B lineage cells may have a protective function in the immune response to Fp.

Innate immune cell signatures in a BCWD-Resistant line of rainbow trout before and after in vivo challenge with Flavobacterium psychrophilum

“…A fifth population (Pop 5), mostly seen in PBL and SPL, stained moderately for Q4E, and strongly for Pax5.PD (++), and could represent activated or phagocytic B cells. These cells were identical to a cell type described previously, which stained positive for both IgM and IgT (Zwollo et al, 2017), and which is currently being investigated as part of a different study.…”

“…The Fp challenge on YC15 fish has been described previously (Zwollo et al, 2017). Immune tissues were collected on day 5 post-challenge.…”

“…Relative fold change (RFC) was determined by calculating the 2^-ddCT for each sample using α-tubulin as the control target as reported previously (Schouten et al, 2013). The Taqman assay for Fp loads has been described elsewhere (Marancik and Wiens, 2013) (Zwollo et al, 2017). …”

“…Statistical analysis used the environment R to calculate p values and has been described previously (Zwollo et al, 2017). …”

“…Additionally, R-line fish have a higher abundance of Early B-cell Factor-expressing (EBF) cells compared to S-line trout (Zwollo et al, 2015), although the identity of EBF + cells is more complicated than previously assumed (this study). Further, R-line fish have a higher abundance of IgT + B lineage cells (Zwollo et al, 2017), a lower abundance of IgM + B lineage cells (Zwollo et al, 2015), and higher expression of secreted heavy chain tau transcripts, compared to S-line fish (Zwollo et al, 2017), suggesting that R-line fish have stronger mucosal responses than S-line fish.…”