Abstract：Anaerobic fungi (Neocallimastigomycota) are important fiber-degrading functional microorganisms in the herbivores' digestive tract. The purpose of the present study was to isolate anaerobic rumen fungi with high activities of plant cell wall degrading enzymes for practical applications to improve the utilization of roughage. Twelve anaerobic fungi strains were isolated from the ruminal contents of Xinong Saanen dairy goat (Capra hircus) using the Hungate roll tube technique. The fungi strains were identified using the morphological characteristics, nucleotide sequence analysis of internal transcribed spacer region and 28S rDNA D1/D2 region. Meanwhile, activities of xylanase, carboxymethyl cellulase (CMCase), avicelase, acetyl esterase (AE) and β-glucanase of 12 fungi cultures were assayed, and the enzymatic characteristics of xylanase and AE produced by fungus with the highest enzyme activities were also analysed. The results showed that 12 anaerobic fungi strains belonged to Piromyces, named Piromyces sp. CN1~ Piromyces sp. CN12, respectively. The ITS gene Genbank number is KY368100~KY368111. The xylanase, CMCase and AE activities of Piromyces sp. CN6 were 1655.3, 93.4 and 152.8 mU which were significantly higher than those of other fungi strains (P＜0.05). The avicelase activity of Piromyces sp. CN3 was the highest among these fungi, but its activity showed no difference with that of Piromyces sp. CN6 (P＞0.05). There was no significant difference on β-glucanase activity produced by fungi strains (P＞0.05). The xylanase activities had extremely significant positive correlation with CMCase and AE activities (P＜0.01), and the xylanase activities had significant positive with avicelase activity (P＜0.05). The optimum temperature of CMCase activity was 50 ℃, and the optimum pH was 5.0. The CMCase was stable at 40 ℃ and pH 5.0~8.0. K＋, Ca2+and Co2+ had active effects on its enzyme activity, whereas Zn2+, Cu2+, Mg2+, Fe2+ and Mn2+ had inhibitory be promoted on it. The AE activity preformed optimally at 50 ℃ and pH 9.0, and it was stable at 40 ℃ and pH 5.0~10.0. The AE activity could be promoted by Mg2+, K+ and Ca2+ and inhibited by Zn2+, Fe2+, Co2+ and Mn2+. Piromyces sp. CN6 isolated from rumen of Xinong Saanen dairy goats showed the highest plant cell wall degrading enzymes activity. Through enzyme activity assays and enzymological characterization, the enzyme system information of rumen fungi can be enriched and improved, which can provide basis for further application.
