Whole Genome and Comparative Genomic Analysis of Bacillus sp. KNSH11 Isolated from the Gut of Litopenaeus vannamei Reveals Its Probiotic Potential

In sustainable aquaculture, probiotics offer a promising alternative to antibiotics for improving shrimp health. Bacillus sp. KNSH11, isolated from the intestine of whiteleg shrimp ( Litopenaeus vannamei ), was characterized to evaluate its probiotic potential. The strain, a Gram-positive, rod-shaped bacillus, exhibited exceptional spore formation efficiency (> 99%), ensuring resilience in challenging environments. Functional assays demonstrated that KNSH11 maintained high viability at pH 2–4, in the presence of bile salts, at temperature up to 95°C, and under lysozyme exposure, indicating tolerance to gastrointestinal and processing stresses. Metabolic profiling indicated significant lactic acid production with minimal acetate and propionate, distinguishing it from conventional lactic acid bacteria. KNSH11 also displayed strong antioxidative activities and moderate antibiofilm effects against pathogens. Antibiotic susceptibility testing revealed sensitivity to amoxicillin (30 µg/disc), chloramphenicol (30 µg/disc), kanamycin (30 µg/disc) and tetracyclines (30 µg/disc), but resistance to ampicillin (10 µg/disc) and penicillin (10 µg/disc). Whole genome sequencing (WGS) confirmed the absence of virulence factors and identified mobile genetic elements, a CRISPR/Cas system, and gene clusters potentially encoding bacteriocins. Collectively, these findings suggest that Bacillus sp. KNSH11 is safe, eco-friendly probiotic with significant potential to enhance shrimp health and advance sustainable aquaculture.