Hybrid catfish has gained importance in the catfish industry due to superior growth performance and enhanced resistance to most pathogens. However, hybrid catfish are more susceptible to Edwardsiella piscicida than channel catfish. Beta-glucan, a component of yeast cell walls, is widely used in aquaculture as an immunostimulant, and it is a promising product for the channel catfish industry to reduce the impact of diseases. This study aimed to evaluate the single-cell RNA expression profiles of hybrid catfish leukocytes following exposure to beta-glucan and subsequent infection with E. piscicida. Hybrid catfish (~60 g) were stocked in four 30 L tanks (8 fish per tank). Fish were exposed to 60 µg beta-glucan/g body weight via gavage, while control fish received saline solution 0.85%. Two weeks later, fish were injected intraperitoneally with 100 µL of a sublethal dose of E. piscicida (estimated concentration of 7.64 × 105 CFU/g of body weight) or saline solution 0.85%. Leukocytes were isolated from the anterior kidney at 12 and 24 hours post-infection (hpi) and processed using the GEM-X Single Cell 3’ Chip Kit v4 (10x Chromium) for library construction. Four treatment groups were analyzed at 12 hpi based on first + secondary exposures: beta-glucan + E. piscicida, beta-glucan + no infection, saline + E. piscicida, and saline + no infection. The 24 hpi samples were not processed due to insufficient cell numbers.
Erythroid and myeloid cell counts were reduced in infected groups. Decreased granulocytes likely reflected neutrophil mobilization during early innate responses. There was a lower count of lymphoid cells in the non-infected groups. In the beta-glucan + no infection group, upregulated genes included those involved in energy metabolism and immune regulation: TKT, fbp1a, TXN, PRDX1, PRDX2, aldocb, ATP5PB, 143b1, ctss2.1, npsn, mpx, timp2b, NCF2, rac2, LCP1, CD63, MMP1, and ARPC3. Beta-glucan + E. piscicida fish upregulated genes linked to cell recruitment and inflammatory signaling: ccl34b.1, irf1b, fosab, CXCR4, ATP6V0C, RPL38, RPS29, lygl2, RPS21, rpl39, and rps24. The saline-infected group showed increased expression of genes associated with antigen presentation, lymphocyte activation, cell adhesion, and ribosomal structure: CD83, alcamb, rpl6, RPS27A, rps6, RPL27, and RPL19.
In conclusion, beta-glucan modulated leukocyte transcriptional profiles, enhancing expression of genes involved in metabolic reprogramming and innate immune activation. These findings provide insight into beta-glucan induced cellular mechanisms underlying immunostimulant efficacy in hybrid catfish.