Evaluation of probiotic properties and complete genome analysis of lactic acid bacteria isolated from crested ibis Nipponia nippon feces

Lei Yang ¹ Jie Luo ² Yan Zeng ³ Baoyue Zhang ¹ Yang Wang ¹ Gang Shu ^1* Xiaoling Zhao ⁴ Haohuan Li ¹ Xu Funeng ¹ Wei Zhang ¹ Hualing Fu ¹ Felix Kwame Amevor ⁴ Rui Liu ²

• ¹ Department of Veterinary Medicine, Sichuan Agricultural University, Cheng Du, China
• ² Department of Animal Husbandry and Veterinary Medicine, Tongren Vocational and Technical College, Tongren, China
• ³ institute of Animal Microecology, College of Veterinary Medicine, Sichuan Agriculture University, Chengdu, Sichuan Province, China
• ⁴ Farm Animal Genetic Resources Exploration and Innovation Key Laboratory of Sichuan Province, Sichuan Agricultural University, Chengdu, Sichuan Province, China

This study aims to identify an optimal lactic acid bacterial strain from the feces of healthy crested ibis (Nipponia nippon). From the fecal samples, twenty isolates were obtained. The isolates were subjected to biochemical identification, acid and bile tolerance tests, in vitro inhibition of pathogenic bacteria assays, cell surface hydrophobicity assessment, antibiotic susceptibility test, and hemolytic activity evaluation to determine their probiotic potential. The results indicated that five isolates (D1, D2, D6, E7, D8) could survive in low acid and high bile salt conditions. Except for D8, all four isolates exhibited inhibitory activity against tested pathogens. Isolates E7 showed the least resistance to antibiotics, and only E7 demonstrated moderate hydrophobicity. The E7 strain was further studied in depth and identified as L. plantarum through 16SrDNA sequencing. To assess its safety, mice were fed with the E7 strain. The results showed that no mice died, and there were no adverse effects on the body weight and blood cell composition of the mice.Whole genome sequencing of Lactiplantibacillus plantarum E7 using Nanopore PromrthION48 and the Illumina Novaseq revealed a ring chromosome and two ring plasmids. The chromosome encodes 3024 genes, some associated with cell adhesion, acid and bile salt tolerance, antioxidant enzymes, and secondary metabolites. Plasmids contained fewer coding genes. The VFDB database detected only a few virulence genes related to adherence, stress survival, exoenzyme production, immune modulation, and regulation factors. KEGG database analysis indicated that the genes of this bacterium are primarily involved in carbohydrate metabolism, amino acid metabolism, vitamin and cofactor metabolism, environmental information processing and genetic processing. This study lays a theoretical foundation for the clinical application and development of probiotics.