| China is a large agricultural country with the production of crop straw 800 million tons per year, accounting for 20% ~30% of total output in the world. Most of them are burned in the farmland after grain harvest as waste, which not only waste the resources, but also cause the pollution of the environment. Straw returning is a beneficial measure in farmland, which can increase soil fertility and crops production generally.The decomposition of corn stalk returning to soil is very difficult and slow. It has been the hot topic how to accelerate the decomposition of corn stalk in soil around the world. Isolation of cellulose-decomposing bacteria or fungi was conducted using method of cellulose-Congo red stain in soil with straw returning which was collected from wheat-corn rotation farmland in Hebei province, its application of beneficial soil rapidiy available phospprus has a decomposition release and can increase the absorption of plant phosphorus in the soil. On the basis of optimize the fermentation process, the conditions for enzyme production and enzymatic properties. The main results showed as follows:1. Eight strains of fungi or 20 strains of bacteria were isolated using method of cellulose-Congo red stain. One fungus and three strains of bacteria were screened with hydrolysis circle determination and enzyme activity analysis.The 18SrDNA gene sequence showed that the fungus strain isolated was identified as Penicillium oxalicum. The isolated strain was named Penicillium oxalicum HB1 (P.oxalicum HB1), preserved in the China General Microbiological Culture Collection Center (CGMCC), preserved No. of CGMCC NO.4842. Moreover, B1 and B2 are strains with Bacillus and B3 is a strain without Bacillus. The growth curve and CMC and FPA enzyme activities were studied further for HB1 and three strains of bacteria. The results showed that the enzymic activities of CMC and FPA of HB1 were 672.8U and 774.6U on the sodium carboxymethyl cellulose (CMC-Na) medium when inoculum size kept at 1% and grew on the condition of 30℃and shaking at 180rpm for 72h. CMC enzyme activities of B1 and B3 are close to the peak at 12h after cultured, up to 31.2U and 24.4U, respectively. CMC enzyme activities of B2 are the highest and reached 36.3U at 18h after cultured. For another enzyme-FPA, the enzyme activities of B1 strain had reached the peak at 12h after cultured and maximum value is 34.1U. However, the FPA enzyme activities of B2 and B3 strains is the highest at 8h after cultured and reached 39.2U and 63.6U, respectively. Comparison Penicillium oxalicum HB1 to 2 strains of bacteria in enzyme activities and growth conditions, CMC and FPA enzymes activities of Penicillium oxalicum HB1 were highest. Therefore, further study was conducted for Penicillium oxalicum HB1.2. The study of fermentation optimization and CMC enzyme activities of Penicillium oxalicum HB1The results showed that CMC enzyme activities were high when HB1 grew on the medium with 3% N as beef extract peptone, 5% inoculum size, incubation temperature at 28℃～35℃, pH at 4～7 and incubation time for 48h～96h. The further experiment was conducted on the base of this result to explore fermentation optimization.The best combination of pH, temperature, solid-liquid ratio and incubation time was solid-liquid ratio of pH=6.5, 30℃, 1:10 and incubation time of 48h.CMC enzyme activity was higheat when HB1 grew in the medium with pH5 and kept at 50℃. In addition, CMC enzyme activity was stable on the conditions of pH4~5 and 40℃～50℃.3. The ability evaluate of Penicillium oxalicum HB1 decomposing corn strawsThe incubation experiment illustrated that the enzymic activities of CMC and FPA and the total activity of celluase were 282.9U, 618.3U and 376.1U respectively when HB1 grew on the medium withcorn straw, 1% of inoculum size, shaking at 30℃and 180rpm for 72h. When the HB1 strain was cultured in agar medium with powder of corn stalk together for 10 days, the 87.3% of core stalk was decomposed and 1.90 times higher than that in the control; When the HB1 strain were incubated in soil with corn stalk for 30 days, the decomposition rates were 83.5% and 1.62 times higher than that in the control; Finally, the pot experiment with wheat plants was carried out to investigate the effect of HB1 strain on corn stalk decomposition in the soil for 50 days. The result showed that the straw decomposition rate was 70.8% and 15.1% higher than that in the control.4. The functions of HB1 on fixed P release in soil and beneficial effects on plant growthWhen the HB1 strain were incubated in soil with corn stalk for 30 days, the soil available phosphorus (Olsen-P) concentrations were 98.6mg/kg and 77.1mg/kg in HB1 application treatment and control, respectively. When incubation time expanded to 50d, P concentrations in wheat shoots were 12.6g/kg and 7.62g/kg for treatment and control, and P concentration in wheat roots were 25.9g/kg and 21.1g/kg in treatment and control.moreover,There were significant differences in P concentrations between control and HB1 treatment(P<0.05). Meanwhile, the biomass of wheat increased by 16.8% in treatment comparing to the control when plant growth for 30d.Penicillium oxalicum HB1 not only accelerates the decomposition of corn stalk in soil, but also enhances the available P released from soil. So it is necessary to do further research on its application in farmland. |
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