Isolation And Identification Of Lactic Acid Bacteria From The Intestine Of Piglets And Preliminar Study On Their Antibacterial Activity

Eleven strains of lactic acid bacteria(LAB) were isolated from the piglet intestinal and their probiotic roles were investigated by in vitro experiments.The results were described in the following four sections.In the first section,a litter of piglets which had edema disease from a pig farm in Taizhou were used to isolate E.coil and LAB.A strain of pathogenic E.coil(O138) was isolated from mesenteric lymphaden of affected piglets.A group of eleven LAB strains were isolated from duodenal and jejunal mucosas of recovered piglets from edema.L1,L2,L3 and L4 grew more rapidly than the other strains,and the pH value in the culture of these four LAB strains showed faster decline.According to their morphology,and physiological and biochemical characteristics,L1,L2,L3 and L4 were preliminarily identified as Lactobacillus murinus,Lactobacillus acetotolerans,Lactobacillus intestinalis and Lactobacillus rhamnosus.In the second section,effects of gastrointestinal factors(low pH,bile salt and pepsin) on the survival of strain of L1,L2,L3 and L4,and heat tolerance of strains were investigated.The results showed that L1,L2,L3 and L4 could survive well at pH3.5 and pH4 for three hours,but could not when pH was at or below 2.5,while pH6.4 was served as the control.While the group that had no bile in the culture was served as the control,the survival counts of L1 and L2 declined from 10⁹ to 10⁸,L3 from 10¹⁰ to 10⁹,L4 fom10⁸ to 10⁷ after incubated in the medium with 0.08%bile for 10 minutes.The survival counts of L1 and L2 declined from 10⁹ to 10⁸,L3 from 10¹⁰ to 10⁸,and L4 from 10⁸ to 10⁷ when incubated in the medium with 0.4%bile for 10 minutes.There was no effect of pepsin on the growth of four strains.When the group treated with 37℃for 20 minutes was served as the control,treatment with 50℃showed no effect on survival of L1,L2,L3 and L4,while treatment with 65℃decreased the survival of L1,L2,L3 and L4 significantly.The survival counts of L4 was not detected when treated with 80℃,but the survival counts of L1 and L3 were 10⁴.In the third section,lactic acid production and antagonistic property of the four strains of LAB(L1,L2,L3and L4) isolated from piglet intestine were investigated.The results showed that among all strains,L3 exhibited the rapidest speed in production of lactic acid and the highest yield of lactic acid at 48 hours with the concentration of 60.48mmol9L^-1. Accordingly,the pH value in the culture of L3 showed fast decline,with the final value significantly lower than those of other culture.The inhibitory effects of L3 on pathogens were more significant than those of the other strains.The inhibitory effect was more significant when pH in the medium was low.When subtracting the effect of the acid,the culture supernatants of four LAB only showed 10%inhibitory effect,which suggested that their inhibitory effect was mainly caused by lactic acid.The inhibitory effect of the culture supernatants was above 95%after heat treatment and above 90%when treated with proteases.In the fourth section,fermentation property and kinetic analysis of the Inhibition of in vitro growth of Pathogenic Escherichia coli K88,O138 by Lactobacillus(Lactobacillus intestinalis) isolated from porcine intestine mucus were investigated.The results of fermentation showed that L3 result in a pH of 3.90 within 12h and a large amount of lactic acid production,with the concentration of 104.08mmol·L.Kinetic analysis of the antibacterial activity of L3 metabolic product toward K88,O138 showed that CFUS display strong antibacterial activity toward K88,O138;the antibacterial activity of L1 CFUS was higher than the lactic acid control samples and was mainly due to the production of lactic acid;K88,O138 had the tolerance property toward pH4.5.