Evaluation of a novel potential probiotics Bacillus spp. isolated from the gut of healthy gibel carp (Carassius auratus gibelio): Effects on the growth performance, nonspecific immunity and disease resistance against CyHV-2 infection

Aims: For sustainable aquaculture, probiotics are viable alternatives to antibiotics. However, most commercial probiotics originate from non-aquatic environments, limiting their efficacy in fish. This study aimed to isolate and evaluate indigenous probiotics from the intestinal microbiota of wild and cultured gibel carp (Carassius auratus gibelio) to provide effective, host-specific probiotics. Methods and Results: A total of 112 bacterial isolates (47 from cultured, 65 from wild carp) were obtained; 45 showed enzyme-producing activity. Isolates SY01 and SY02 outstandingly produced amylase, protease, and cellulase, and were non-pathogenic to gibel carp. They were identified as Bacillus subtilis (SY01) and B. amyloliquefaciens (SY02) via morphological, biochemical, and 16S rRNA sequencing analyses. Dietary supplementation with 1×10⁷ CFU/g for 30 days significantly improved growth, digestive enzyme activity, and serum antioxidant capacity (CAT, T-AOC, SOD), and upregulated immune-related genes (ITLN1, IL11, IFN). After CyHV-2 challenge, survival rates were 71% (SY01), 57% (SY02) vs. 14% (control). Conclusions: The two host-derived probiotics enhanced gibel carp’s growth, digestion, antioxidant status, and disease resistance, promising as antibiotic alternatives. This study provides a scientific basis for indigenous probiotic application in sustainable, eco-friendly aquaculture. Impact Statement As a key freshwater aquaculture species in China, gibel carp (Carassius auratus gibelio) faces major threats from Cyprinid herpesvirus 2 (CyHV-2), with no effective control measures available. Probiotics are a promising alternative to antibiotics, but non-host-derived strains are often less effective. This study addresses this by isolating and identifying host-specific Bacillus strains from gibel carp intestines. These native probiotics lay a new foundation for antibiotic substitution in aquaculture and offer a novel microecological strategy to prevent CyHV-2 outbreaks. Their use is expected to enhance the sustainability and biosafety of gibel carp farming.