茶花鸡源罗伊特氏黏液乳杆菌CHF7-2的益生特性及全基因组测序分析

Probiotic characterization and whole genome sequencing of Limosilactobacillus reuteri CHF7-2 from Chahua chicken

摘要:

目的 解析茶花鸡源罗伊特氏黏液乳杆菌CHF7-2的益生特性和安全性，为开发利用该菌株作为饲用微生态制剂提供理论依据。方法 通过体外试验检测菌株CHF7-2的黏附性能、产酶性能、抑菌功效和抗氧化活性。利用PacBio Sequel II和Illumina NovaSeq 6000平台对菌株CHF7-2进行全基因组测序，并采用多种生物信息学工具和数据库对其基因组进行注释，从分子层面探究其益生机制和安全性。结果 体外试验研究表明，菌株CHF7-2展现出良好的益生特性和安全性：具有较强的表面疏水性、自凝聚性和一定的抗氧化能力；能有效抑制大肠杆菌K88、金黄色葡萄球菌ATCC 49521、鸡伤寒沙门氏菌CICC 21510和猪霍乱沙门氏菌CVCC 3383的生长；能够产生蛋白酶和脂肪酶；不产生溶血环，表明其饲用安全性。全基因组测序分析结果显示，菌株CHF7-2的基因组大小为2 116 761 bp，平均G+C含量为38.8%，编码基因数量为2 067个。同时，在CHF7-2基因组中发现了Ⅲ类细菌素EnlA合成基因簇，以及多个与耐酸、耐胆盐、耐热胁迫、耐冷胁迫、黏附、抗氧化和有机酸合成相关的抗应激和益生基因，且基因组中未发现毒力和耐药基因。结论 罗伊特氏黏液乳杆菌CHF7-2是一株具有潜力的益生菌，有望成为饲用微生态制剂的重要候选菌株。

关键词:罗伊特氏黏液乳杆菌;全基因组测序;益生特性;微生态制剂

Abstract:

Objective To study the probiotic properties and safety of Limosilactobacillus reuteri CHF7-2 isolated from Chahua chicken, providing a theoretical basis for the development of this strain as a microecological feed additive.Methods In vitro assays were conducted to evaluate the adhension, enzyme production, antibacterial activity, and antioxidant activity of strain CHF7-2. PacBio Sequel II and Illumina NovaSeq 6000 were used for whole genome sequencing of strain CHF7-2. Bioinformatics tools and databases were then used for genome annotation to explore the probiotic mechanisms and safety of this strain at the molecular level.Results Strain CHF7-2 possessed significant probiotic properties and safety. It exhibited high surface hydrophobicity, self-coagulation, and antioxidant activity. The strain inhibited the growth of Escherichia coli K88, Staphylococcus aureus ATCC 49521, Salmonella gallinarum CICC 21510, and Salmonella choleraesuis CVCC 3383. Strain CHF7-2 produced protease and lipase, and it was safe for use in feed since it did not exhibit hemolytic activity. Whole genome sequencing revealed that the genome size of CHF7-2 was 2 116 761 bp, with the G+C content of 38.8%, encoding 2 067 genes. Additionally, the genome carried the biosynthetic gene cluster of the class III bacteriocin EnlA and multiple genes involved in the acid tolerance, bile salt tolerance, heat stress tolerance, cold stress tolerance, adhesion, antioxidation, and organic acid synthesis, with no virulence or antibiotic resistance genes detected.Conclusion L. reuteri CHF7-2 is a potential probiotic strain and a promising candidate for use as a microecological feed additive.

Key words:Limosilactobacillus reuteri;whole genome sequencing;probiotic properties;microecological feed additive