Screening And Culture Conditions Of High Efficient Ammonifying Bacteria In The Rhizosphere Of Camellia Oleifera

The long-term use of chemical fertilizers in large quantities has led to soil consolidation,nutrient disorders,reduced product quality and environmental pollution,while microbial fertilizers can improve a series of negative effects brought about by chemical fertilizers,and the screening of suitable strains is the basis for the development of microbial fertilizers.Camellia oleifera is an important woody oil tree in China and is a typical ammonium-preferred plant.Development of microbial fertilizers for Camellia oleifera is an important measure to improve yield,quality and woodland environment.In this study,we firstly isolated and purified inter-rhizosphere soil ammoniating bacteria from oil tea,then screened out the strains with high conversion capacity,and studied the morphology,physiological and biochemical characteristics,optimal culture conditions,and the effect of microbial agents of the highly efficient strains,in order to provide reference materials for the development of special microbial fertilizers for oil tea and to improve the efficiency of nitrogen utilization in oil tea.The results of the study are as follows.1.160 strains of ammoniated bacteria were screened by traditional media isolation and purification methods,followed by primary screening by means of a nano reagent colour development reaction to obtain 25 strains of ammoniated bacteria with darker colour development.The activity of the ammoniated bacteria was determined and the bacteria numbered AHB,AHO,AHT and AHX were more active.16 S rDNA sequencing showed that the sequence lengths of AHB,AHO,AHT and AHX were 1385 bp,1413 bp,1390 bp and 1418 bp respectively,belonging to the Chryseobacterium sp.,Stenotrophomonas sp.,Arthrobacter sp.and Bacillus sp.2.The effects of culture conditions on the growth of the four strains of efficient ammonifying bacteria were investigated in six aspects,including pH,temperature,rotational speed,inoculum,carbon source and nitrogen source,etc.The results showed that:all four strains could grow at pH 5-9,and the optimum pH was 7-8,indicating that the four strains of ammonifying bacteria were suitable for growth under neutral conditions;the optimum rotational speed for all four strains was 200-250 r/min,and the optimum The optimum speed of all four strains was 200-250 r/min,and the optimum temperature was around 25℃,but strain AHX was more tolerant to high temperature than the other strains;the optimum inoculum level of the four strains was 2%-3%,the optimum carbon and nitrogen sources of AHB were glucose and urea,the optimum carbon and nitrogen sources of AHO were mannitol and potassium nitrate,the utilisation of AHT for sucrose and potassium nitrate was significantly higher than other carbon and nitrogen sources;the utilisation of AHX for all five carbon sources was higher,and the optimum nitrogen source was Potassium nitrate was the most suitable nitrogen source.3.Morphological,physiological and biochemical tests were performed on strains AHB,AHO,AHT and AHX.The results showed that all four strains of ammoniated bacteria were Gram-positive,all were motile,their contact enzyme,V-P and gelatin liquefaction tests were positive,they had the ability to produce ammonia,they did not have the ability to starch hydrolysis and nitrate reduction,and the indole test was negative.The milk litmus test showed that all four strains of ammoniacal bacteria were acid-producing.The antagonism test showed that the four strains of bacteria were not antagonistic to each other and could be cultured together.4.The effect of different fungicides on the physicochemical properties of inter-root soil of oil tea seedlings was investigated in a pot experiment.The results showed that:the fungicide inoculation treatment significantly reduced the pH and total nitrogen content of the inter-root soil,and the pH of the AHT treatment decreased the most,at 10.69%(p<0.05).The fertility of the soil was improved by inoculating the soil with fungicides from 0 to 60 d,which was significantly higher than that of CK(p<0.05).In terms of enzyme activities,the enzyme activities of the fungicide inoculation treatment were significantly higher than those of CK(p<0.05),and the activities of urease,catalase,nitrate reductase and protease all reached their maximum values after 60 d of inoculation.