| Methanotrophs are belonging to microorganisms which are widespread in nature and ruminant animals’ gastrointestinal tract and grow with methane as the sole carbon source and energy. They can oxidize methane into carbon dioxide and water under the action of adding oxygenase alone into methane, and can provide the energy needed for their own growth. As an important greenhouse gas, methane has not only caused the pollution of atmospheric environment, but also feed energy loss in animal production and production performance descends. Because of its unique way of metabolism of methane, the green and pollution-free metabolic product, methanotrophs is now widely used in industrial production, agriculture, breeding and control of greenhouse gases.As a new type promoter and an alternative to antibiotics, microecologic preparation not only plays an important role in the aspects of the prevention and treatment of animal diseases, promoting animal growth, adjusting intestinal flora, and improving production performance, but also has been greatly welcomed by masses of farmers because of its green and safe, no pollution, no drug residues, etc. In addition, microecologic preparation can help improve livestock and poultry production and product quality, it is of great significance to human health, environmental protection and maintaining the sustainable development of ecological farming.In this study, a strain of methanotrophs that efficiently utilizes methane has been identified and isolated from cow manure, and its viable microecologic preparation has been applied to clinical practical production. Through detecting methane contents in rumen gas and serological indexes at different points in time after feeding viable preparations, the author evaluated clinical laboratory effect of the methanotrophs viable microecologic preparation used in actual production.By multiple concentration and purification of healthy cow manure samples on methanotrophs selective medium, this experiment achieved success in separation and screening of pure strains of methanotrophs, then the author chose one strain of higher metabolic rate of methane as the research object, and named it as M17. After morphological observation, physical and chemical properties analysis and molecular biology identification, the results showed that the strain, belonging to Methylobacterium genus, is gram negative bacteria, and under the electron microscope scanning mirror it is cylindrical and even short rod which appears 0.3 ~ 0.5μm in diameter and about 2.0 ~ 2.5μm in length; the results of H2 S production test and indole test are negative; the results of methyl red experiment, contact enzyme, citrate utilization test, starch hydrolysis, gelatin liquefaction test are positive; the strain can make use of various carbon sources such as methanol, glucose, sucrose, maltose and so on as the substrate for its growth, but its growth is best when taking methanol as carbon source; its optimum growth temperature is 35℃, and the optimum p H is 7.0.In addition, in order to confirm the methanotrophs application effect in the practical production, in this study the author has made methanotrophs viable preparations by using methanotrophs M17. The author chose 12 healthy cows on dairy farms, 6 cows for one control group, feeding at 6:00 AM and 18:00 PM every day, free drinking water, and no gavage of viable preparations for the testing cows in this control group; the other 6 cows for one treatment group, the breeding conditions are the same as the control group, but the testing cows in this group need gavaging viable preparations 500 ml at 6:00 AM before every morning feeding. There are two experimental groups carrying out the rumen gas collection. Experimental group one collected the gas samples at 6:00 AM before every morning feeding on day 0, day2, day 4, day 6, day 8, day10, day 12, day 14, and day 16, and observed the overall change of methane contents in rumen gas; Experimental group two collected gas samples between 6:00 AM and 18:00 PM’s feeding intervals on day 16, that is setting 0 h, 3 h, 6 h, 9 h, 12 h, a total of five sampling time points, of which 0 h is no feeding and no gavaging time, 3 h, 6 h, 9 h, 12 h are time respectively 3 h, 6 h, 9 h, 12 h after feeding and gavaging viable preparations; the control group set the same time points, respectively 3 h, 6 h, 9 h, 12 h before and after feeding. The results are used to observe the cyclical changes of methane contents in rumen gas. Gas chromatograph was used to measure methane contents in the gas samples. Safety evaluation of the effect of viable microecologic preparation would be detected by serological indexes of serum diagnosis which was obtained from 10 ml vein blood from the tail of per testing cow before every morning gavage.The results of experimental group one showed that the methane contents in rumen gas decreased on day 8 when compared with that on day 0 after gavaging the testing cows for 16 days, but the difference was not big; the methane contents decreased significantly on day 14, and the difference is also significant. With the feed fermentation in rumen after feeding, experimental group two drew a conclusion that in both the treatment group and the control group the methane contents in rumen gas is increased significantly in 6h to 9h after feeding; the methane content is declined in 12 h after feeding; compared with the control group, the overall methane contents in rumen of the treatment group dropped significantly. Compared with the data before gavage, there are no obvious changes for serological indexes of ALT, AST, ALB, GLB, TP, direct bilirubin, indirect bilirubin and total bilirubin. Research proves that feeding methanotrophs viable microecologic preparation can reduce the rumen methane emissions, and that is safe and reliable for dairy production. |
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