Extreme Environmental Tolerance Of LAB From Different Materials From The Qinghai-Tibet Plateau And Preliminary Study On The Probability Of Excellent Strains

In this study,representative strains of lactic acid bacteria(LAB)isolated from gut of Qinghai lake-naked carp(Gymnocyprisprzewalskii),plant surface and soil of the Qinghai-Tibet Plateau were used to study the tolerability under extreme environmental conditions,then the probabilistic properties of some excellent LAB strains were studied in orded to screen LAB with potential probiotic features.According to the morphology,physiological and biochemical characteristics of LAB,combined with 16 SrRNA gene sequence analysis,LAB from gut of different age Qinghai lake-naked carp were generally classified as 4 genera,Lactobacillus(L.)?Leuconostoc(Leu.)?Lactococcus(Lc.)?Pediococcu(P.),respectively and which belong to 8 species,Lc.lactis ? P.acidilactici ? P.pentosaceus ?Leu.mesenteroides ? L.paraplantarum ? L.sakei ? L.fuchuensis ? L.casei,among them,bacilli were much more than cocci obviously.A total of 48 strains of LAB were randomly selected to detect the tolerance under extreme environmental.The growth of LAB at low pH,high salt concentration and extreme temperature was measured by plate counting method and turbidity.The results showed that all LAB can not survive under extreme acid environment pH1.While in pH2,the growth of most LAB were significantly inhibited,but,some of the survival rate were 70%,F2-2,F9-10,F12-66 isolated from gut of lake-naked carp showed strong acid tolerance.The survival rate was generally high in pH3 and LAB even can grow.At 16% of the high salt(NaCl)concentration,the survival rate of multiple LAB were more than 50% and even can grow.High temperature as50 ?caused a serious threat on the survival of LAB.Most LAB can not survive and only a very small number showed slowly growth in the first 12 hours.Most LAB can grow well at 10 ?,but can not grow at 0?.However,F2-1,F11-67 and M1-8 have OD value of 0.35 after 168 h of culture,showing slowly growth Trends,of which 2were P.pentosaceus.The antibiotics susceptibility of LAB and gastrointestinal environment were detected by plate diffusion method and plated counting method.The results showed that LAB were mainly resistant to vancomycin,gentamicin,polymyxin B,ciprofloxacin and penicillin G and sensitive to chloramphenicol,ampicillina and tetracycline.Most of the LAB on the gastrointestinal environment tolerance is relatively poor,the LAB survival rate is relatively low or even all death after 7 h of artificial gastrointestinal environment,which N20,N7,N6,N1 and qz1242 tolerance were relatively strong,survival can up to 61.11%,the lowest was 31.95%.Most of the strains with better tolerability were Lactobacillus,the highest survival rate was L.plantarum isolated from the soil.The LAB with good gastrointestinal tolerance were tested by plate count method to detect tolerance under different bile salt concentrations.Bile salts showed inhibitory effects on the growth of LAB and gradually increased with the increase of their concentration and the prolongation of culture time.In the 0.1% bile salt concentration,the survival rate of LAB was relatively high,even up to 100%,a small part of LAB could survive in 0.2% bile salt medium,while the bile salt concentration increased to 0.3%,the survival rate of most LAB have poor tolerance as 0.Among them,N22 and N19 isolated from soil had a certain degree of bile salt tolerance,viable count were 2.70 logcfu/ml at 0.3% bile salt concentration after 4 h cultured.