Long-term crowding stress causes compromised nonspecific immunity and increases apoptosis of spleen in grass carp (Ctenopharyngodon idella).

Crowding stress is one of the most common environmental stressors in intensive aquaculture. To investigate the influences of long-term crowding stress on nonspecific immune responses and apoptosis in fish, grass carp (Ctenopharyngodon idella) were cultured at low (0.9 kg m-2), medium (2.97 kg m-2) and high (5.9 kg m-2) stocking densities for 10 weeks in the present study. The results showed that elevation of stocking densities caused splenic tissue damages and inflammatory responses, which are characterized with the formation of melano-macrophage centers and the increase of granulocytes as well as significant upregulation of inflammatory cytokine genes (il1β and tnfα). The remarkable decline in the activities of serum lysozyme, acid phosphatase and alkaline phosphatase under high stocking density further confirmed that increased stocking density affected fish nonspecific immune response negatively. Moreover, the transcriptional levels of splenic apoptotic-related genes caspase-8, fasl and caspase-3 increased significantly while the mRNA levels of bax, bcl2, apaf1 and caspase-9 remained unchanged. This result showed that increased stocking density caused splenic cell apoptosis, which were closely associated with the FasL signaling pathway. Our findings revealed that crowding stress could influence fish nonspecific immune response negatively and increase inappropriate apoptosis of the spleen, which would make fish more susceptible to pathogens and ultimately impair fish survival. The breeding density utilized in this study also provides some reference values in intensive aquaculture systems from the perspective of fish health and welfare.