Study On Isolation, Identification And Conservation Of Lactobacilli From Poultry

In order to prepare the best probiotic of poultry, the experiment isolated fiveEnterococcus and two Lactobacilli from the blind intestinal mocous and feces ofhealthy Beijing You Fowl. According to the experiments of OD_600nm and pH, weselected 1 strain as the better lactobacilli, and confirm the best culture condition. Then,the experiment carried out some experiments including optimizing culture medium,cultivating in 50 litre fermenter, freeze-dyring with different cry-protective agent, andso on. At last, we made the production of the freeze-dry powder of M8 inbroilers'feed in order to investigate its effect on the microflora of broilers. The resultsshowed:(1) 41 strains bacterias species from the blind intestinal mocous and feces ofhealthy Beijing You Fowl. Though the observation of colonial morphology and Gramstaining, 7 strains were identified physiologically and biochemistrically according tothe diagnostic characteristics of bacteria and were preliminarily defined as Lactic acidbacteria. The result showed:M1,M2 were defined as E.scecorum; F5,F8,F11 weredefined as E. faecalis; M3 was defined as L. brevis and M8 was defined as L.pentosus.(2) The experiment got the growth curves and pH of Lactobacilli M3 and M8 inbreezy and sealed bottles. OD_600nm of M3 reached to 1.154 and pH reached to 4.16from 5.86 in breezy bottles, and in the sealed bottles, OD_600nm reached to 1.157 andpH reached to 4.14 from 5.85. On the other hands, OD_600nm of M8 reached to 1.011and pH reached to 4.36 from 5.64 in breezy bottles, and in the sealed bottles, OD_600nmreached to 2.083 and pH reached to 3.42 from 5.64. Therefore, M8 was the betterLactobacilli and its optimum culture condition was in sealed bottles.(3) We also got the growth OD_600nm and pH of M8 in sealed bottles with differentlevels of inulin. With the four levels of inulin, M8 turned into the stationary phaseunder 20h, and reached to highest value about 28h The best percent of inulin was3%,, and the OD_600nm reached to 2.937 and pH reached to 3.40 from 5.81.(4) In 50 litre fermenter, M8 turned into the stationary phase about 20h, andreached to the highest value with 23h, which the OD_600nm reached to 2.502. Then, M8grew slowly under 27h and its OD_600nm became more lower, therefore the best time of harvest was 20h.(5) When the culture of M8 stored as liquid at room temperature or 4℃, theviable cell number reached to 5.11×10⁸cfu/ml from 1.1×10⁹cfu/ml when it was storedat room temperature (p＜0.01). on the other hands, the viable cell number reached to6.79×10⁷cfu/ml from 1.37×10⁹cfu/ml (p＜0.01) at 4℃, Therefore, the culture of M8stored as liquid were not the best method of conservation.(6) The result indicated that the cry-protective agent had obvious effect onincreasing the survival rate of M8. In the experiments, we have used 15 kinds ofdifferent cry-protective agent. The different cry-protective agent had differentsynergistic accumulation, and the survival rate of M8 were 96.64% using thecomponents of 18% skim milk+1% saccharose +1% glycerin+2% amylum.(7) Compared with the Groupâ… (control group) (p＜0.01) and the Groupâ…¡(antibiotic group) (p＜0.05), the Groupâ…¢(freeze-dry powder of M8) adding intobasal diets could increase the number of the Lactic acid bacteria of blind intestinalmucous; the E.coil of blind intestinal mocous (p＞0.05),the Lactic acid bacteria andthe E.coil of feces had no different from the group of freeze-dry powder.