鲫肠道乳酸菌的分离及益生特性

Isolation and probiotic properties of lactic acid bacteria from the gut of crucian carp(Carassius auratus)

为获得鱼源益生乳酸菌,本研究从健康鲫肠道内分离鉴定得到38株乳酸菌,并选择其中8株乳酸菌进行体外益生特性分析。结果显示,8株乳酸菌均能在p H=4.5和10%鲫胆汁环境中存活,对嗜水气单胞菌、豚鼠气单胞菌、无乳链球菌和副溶血弧菌均有较强的抑菌能力,但菌体表面疏水性和自凝集能力具有菌株特异性。选择5株疏水性较高(63%~89%)、自凝集能力强(37%~45%)的乳酸乳球菌进行体外黏附能力测定,结果显示,黏附能力在菌株间具有差异性(4.5%~7.9%),但均能显著降低嗜水气单胞菌NJ-35的体外黏附率,黏附抑制率达30%~35%;最后对筛选出的3株高黏附力的乳酸乳球菌进行安全性评价,发现3株菌对鲫均无致病力。鱼源乳酸菌的筛选,为鲫养殖生产中益生菌的实际应用提供了理论依据。

As an important freshwater aquaculture species in China,Carassius auratus culture is currently suffering from serious losses due to infectious diseases.The use of antibiotics in preventing and controlling the bacterial diseases has caused the evolution of resistant strains of aquatic pathogens.This is a serious constraint on aquaculture development.Therefore,some alternative measures need to be developed to improve the quality and sustainability of aquaculture production.Probiotics can be a substitute for antibiotics.Lactobacillus sp.from the host or the culture environment have shown to have ability to better adapt to the culture environment and colonize in the host gut,which help them exhibit their probiotic effects better.This study intends to screen probiotic candidates from C.auratus according to their in vitro probiotic properties,which include acid and bile tolerance ability,antimicrobial activity,cell surface hydrophobicity,aggregation ability,in vitro adhesive ability and pathogenicity in crucian carp.Based on biochemical tests and 16S rRNA gene sequence analysis,thirty-eight strains of Lactobacillus sp.were isolated from the gut of C.auratus,which belonged to Lactococcus,Leuconostos,Carnobacterium and Enterococcus faecium,respectively,and eight of them were selected to evaluate the in vitro probiotic properties.The results showed that all eight strains of Lactobacillus sp.bacteria could tolerate acid(pH 4.6)and bile(10%bile of C.auratus),and exhibit good antibacterial activities against four pathogens including Aeromonas hydrophila,A.caviae,Streptococcus agalactiae and Vibrio parahaemolyticus.Five of eight strains showed high cell surface properties,i.e.hydrophobicity(63%-89%)and autoaggregation(37%-45%),and their adhesive rates to C.auratus intestinal mucus were ranged from 4.5%to 7.9%.These five isolates differed in coaggregation ability,but exhibited good adhesion inhibition rates for the binding of A.hydrophila to C.auratus intestinal mucus,with the inhibition rates of 30%to 35%.Three L.lactis isolates 16-7,16-19 and16-24 with a higher adhesive ability showed no pathogenicity when administered by intraperitoneal injection to C.auratus.Based on our data,the three L.lactis isolates could be considered as potential probiotic candidates for C.auratus farming.Further in vivo evaluation needs to be performed in the future study.