Recent developments in activities, utilization and sources of cellulase

One of the latest sources of alternative energy, bioethanol, has been the focus of modem research, The production of bioethanol is commonly restricted by the activity of cellulase. Therefore, cellulase has become one of the critical issues in the conversion of lignocelluloses to bioethanol. This article is an overview of the sources and factors affecting enzyme activity, as well as methods of evaluation and utilizations of cellulase. We conclude that a combination of cellulases from various strains can enhance hydrolysis of substrates. Large enough amounts of cellobiase or sufficient cellobiase activity can reduce the inhibition to exoglucanase activity of cellobiose. Characterization and exploitation of cellulase should focus on a definite substrate. Promotion and mixed incubation of strains can reduce the cost of industrial utilization of cellulase.

Salt-tolerant mechanism of marine Aspergillus niger cellulase cocktail and improvement of its activity

The cellulase cocktail produced by marine Aspergillus niger exhibits a property of salt-tolerance,which is of great potential in cellulose degradation in high salt environment.In order to explain the mechanism on the salttolerance of the cellulase cocktail produced by marine A.niger,six cellulase components(AnCel6,AnCel7A,AnCel7B,AnEGL,AnBGL1 and AnBGL2)were obtained by directed expression.Studies on their enzymatic properties revealed that oneβ-glucosidase(AnBGL2)and one endoglucanase(AnEGL)exhibited an outstanding salttolerant property,and one cellobiohydrolase(AnCel7B)exhibited a certain salt-tolerant property.Subsequent study revealed that the salt-tolerant An EGL and AnCel7B endowed the cellulase cocktail with stronger salttolerant property,while the salt-tolerant An BGL2 had no positive effect.Moreover,after overexpression of AnCel6,AnCel7A,AnCel7B and AnEGL,the activity of cellulase cocktail increased by 80%,70%,63%and 68%,respectively.However,the activity of cellulase cocktail was not improved after overexpression of AnBGL1 and AnBGL2.After mixed-strain fermentation with cellobiohydrolase recombinants(cel6 a,cel7a and cel7b recombinants)and endoglucanase recombinant(egl recombinant),the the activity of cellulase cocktail increased by 114%,102%and91%,respectively.

Influence of lactic acid bacteria, cellulase, cellulase-producing Bacillus pumilus and their combinations on alfalfa silage quality

This study assessed the effects of lactic acid bacteria(LAB), cellulase, cellulase-producing Bacillus pumilus and their combinations on the fermentation characteristics, chemical composition, bacterial community and in vitro digestibility of alfalfa silage. A completely randomized design involving a 8(silage additives)×3 or 2(silage days) factorial arrangement of treatments was adopted in the present study. The 8 silage additive treatments were: untreated alfalfa(control); two commercial additives(GFJ and Chikuso-1); an originally selected LAB(Lactobacillus plantarum a214) isolated from alfalfa silage; a cellulase-producing Bacillus(CB) isolated from fresh alfalfa; cellulase(C); and the combined additives(a214+C and a214+CB). Silage fermentation characteristics, chemical composition and microorganism populations were analysed after 30, 60 and 65 days(60 days followed by exposure to air for five additional days). In vitro digestibility was analysed for 30 and 60 days. Compared with the other treatments, selected LAB a214, a214 combined with either C or CB, and Chikuso-1 had the decreased(P<0.001) pH and increased(P<0.001) lactic acid concentrations during the ensiling process, and there were no differences(P>0.05) among them. Fiber degradation was not significant(P≥0.054) in any C or CB treatments. The a214 treatment showed the highest(P=0.009) in vitro digestibility of dry matter(595.0 g kg–1DM) after ensiling and the highest abundance of Lactobacillus(69.42 and 79.81%, respectively) on days 60 and 65, compared to all of other treatments. Overall, the silage quality of alfalfa was improved with the addition of a214, which indicates its potential as an alfalfa silage inoculant.

Effect of Cellulase and Lactic Acid Bacteria on Fermentation Quality and Chemical Composition of Wheat Straw Silage

The object of this study was to determine the effect of cellulase and lactic acid bacteria (LAB) on fermentation quality and chemical composition of wheat straw silage. Silages were prepared using a small-scale fermentation system and the moisture level was adjusted to 60% of fresh matter (FM) with deionized water. Treatments were designed as: control silage without additives, LAB inoculant Lactobacillus casei Z3-1 (1.0 × 106 cfu·g-1 of FM), commercial inoculant L. plantarum FG 1 (1.0 × 106 cfu·g-1 of FM), Z3-1 + cellulase and FG 1 + cellulase. The neutral detergent fiber (NDF), acid detergent fiber (ADF) and crude protein (CP) contents of the wheat straw prior to ensiling were 76.93%, 48.52% and 4.63% of dry matter (DM), respectively. After 30 days of fermentation, the silages treated with LAB and LAB + cellulase had a lower (P < 0.05) pH and higher (P < 0.05) lactic acid content than the control, and the coliform bacteria, yeast and mold were inhibited at the early stage of fermentation. Besides, silages treated with cellulase had lower (P < 0.05) values of ADF and NDF than the control. The results confirmed that the addition of cellulase and LAB contributed to improving the fermentation quality of wheat straw silage.

Effects of cellulase and xylanase enzymes mixed with increasing doses of Salix babylonica extract on in vitro rumen gas production kinetics of a mixture of corn silage with concentrate

An in vitro gas production （GP） technique was used to investigate the effects of combining different doses of Salix babylonica extract （SB） with exogenous fibrolytic enzymes （EZ） based on xylanase （X） and cellulase （C）, or their mixture （XC; 1：1 v/v） on in vitro fermentation characteristics of a total mixed ration of corn silage and concentrate mixture （50：50, w/w） as substrate. Four levels of SB （0, 0.6, 1.2 and 1.8 mL g-1 dry matter （DM）） and four supplemental styles of EZ （1 μL g-1 DM; control （no enzymes）, X, C and XC （1：1, v/v） were used in a 4×4 factorial arrangement. In vitro GP （mL g-1 DM） were recorded at 2, 4, 6, 8, 10, 12, 24, 36, 48 and 72 h of incubation. After 72 h, the incubation process was stopped and supernatant pH was determined, and then filtered to determine dry matter degradability （DMD）. Fermentation parameters, such as the 24 h gas yield （GY24）, in vitro organic matter digestibility （OMD）, metabolizable energy （ME）, short chain fatty acid concentrations （SCFA）, and microbial crude protein production （MCP） were also estimated. Results indicated that there was a SBxEZ interaction （P〈0.0001） for the asymptotic gas production （b）, the rate of gas production （c）, GP from 6 to 72 h, GP2 （P=0.0095）, and GP4 （P=0.02）. The SB and different combination of enzymes supplementation influenced （P〈0.001） in vitro GP parameters after 12 h of incubation; the highest doses of SB （i.e., 1.8 mL g-1 DM）, in the absence of any EZ, quadratically increased （P〈0.05） the initial delay before GP begins （L） and GP at different incubation times, with lowering b （quadratic effect, P〈0.0001 ） and c （quadratic effect, P〈0.0001 ; linear effect, P=0.0018）. The GP was the lowest （P〈0.05） when the highest SB level was combined with cellulose. There were SBxEZ interactions （P〈0.001） for OMD, ME, the partitioning factor at 72 h of incubation （PF72）, GY24, SCFA, MCP （P=0.0143）, and pH （P=0.0008）. The OMD, ME, GY24 and SCFA with supplementation of SB extract at 1.8 mL g-1 DM were higher （P〈0.001） than the other treatments, however,PF72 was lower （quadratic effect, P=0.0194） than the other levels. Both C and X had no effect （P〉0.05） on OMD, pH, ME, GY24, SCFA and MP. The combination of SB with EZ increased （P〈0.001） OMD, ME, SCFA, PFz2 and GP24, whereas there was no impact on pH. It could be concluded that addition of SB extract, C, and X effectively improved the in vitro rumen fermentation, and the combination of enzyme with SB extract at the level of 1.2 mL g-1 was more effective than the other treatments.

Purification and characterization of the kinetic parameters of cellulase produced from wheat straw by Trichoderma viride under SSF and its detergent compatibility

This paper reports the purification and characterization of kinetic parameters of cellulase produced from Trichoderma viride under still culture solid state fermentation technique using cheap and an easily available agricultural waste material, wheat straw as growth supported substrate. Trichoderma viride was cultured in fermentation medium of wheat straw under some previously optimized growth conditions and maximum activity of 398±2.43U/mL obtained after stipulated fermentation time period. Cellulase was purified 2.33 fold with specific activity of 105U/mg in comparison to crude enzyme extract using ammonium sulfate precipitation, dialysis and Sephadex-G-100 column chromatography. The enzyme was shown to have a relative low molecular weight of 58kDa by sodium dodecyl sulphate poly-acrylamide gel electrophoresis. The purified enzyme displayed 6.5 and 55oC as an optimum pH and temperature respectively. Using carboxymethyl cellulose as substrate, the enzyme showed maximum activity (Vmax) of 148U/mL with its corresponding KM value of 68μM. Among activators/inhibitors SDS, EDTA, and Hg2+ showed inhibitory effect on purified cellulase whereas, the enzyme activated by Co2+ and Mn2+ at a concentration of 1mM. The purified cellulase was compatible with four local detergent brands with up to 20 days of shelf life at room temperature suggesting its potential as a detergent additive for improved washing therefore, it is concluded that it may be potentially useful for industrial purposes especially for detergent and laundry industry.

Oscillating Cellulase Adsorption and Enhanced Lignocellulose Hydrolysis upon Ultrasound Treatment

We investigated the effects of ultrasound treatment on cellulase adsorption and lignocellulose hydrolysis.The activity of cellulase remained constant upon lowpower ultrasound treatment(<120 W) and decreased using high-power ultrasound(>280 W).Oscillating cellulase adsorption occurred upon ultrasound treatment with any intensity.The maxima for desorption and adsorption were41.9 and 83.1%,respectively,during 1 h of 90 W ultrasound treatment at 50 °C.A comparison between the shorttime with long-time ultrasound experiments indicated that ultrasound treatment tended to desorb cellulase from substrate.However,ultrasound treatment also led to further surface erosion of biomass,which increased cellulase accessibility.These joint actions of ultrasound treatment induced the oscillating adsorption of cellulase.The increase in cellulase accessibility caused by ultrasound treatment led to a significant enhancement in lignocellulose hydrolysis.

Cellulase production by Aspergillus unguis in solid state fermentation

Lignocellulosic substrates are a good carbon source and provide rich growth media for a variety of microorganisms which prodLuce industrially important enzymes. Cellulases are a group of hydrolytic enzymes such as filter paperase (FPase), carboxymethyl cellulase(CMCase) andβ-glucosidase-responsible for release of sugars in the bioconversion of the lignocellulosic biomass into a variety of value-added products. This study examined cellulase production by a newly isolated Aspergillus unguis on individual lignocellulosic substrates in solid state fermentation (SSF). The maximum peak production of enzymes varied from one substrate to another, however,based on the next best solid support and local availability of groundnut fodder supported maximum enzyme yields compared with other solid supports used in this study.Groundnut fodder supported significant production of FPase (5.9 FPU/g of substrate), CMCase (1.1 U/g of substrate) andβ-glucosidase activity (6.5 U/g of substrate) in SSF. Considerable secretion of protein (27.0 mg/g of substrate) on groundnut fodder was recorded. Constant increment of protein content in groundnut fodder due to cultivation of A. unguis is an interesting observation and it has implications for the improvement of nutritive value of groundnut fodder for cattle.

Research Progress of Cellulase

Cellulase is a complex enzyme that can decompose cellulose into glucose,and it could effectively treat cellulose waste. In this paper,it aims to explore development status and research progress of cellulase,and introduce concept and action mechanism of cellulase,research situation of cellulase in molecular aspect,application of cellulase,and development of cellulase is also prospected.

Screening and characterization of a novel ruminal cellulase gene(Umcel-1) from a metagenomic library of gayal(Bos frontalis)

Gayal is a rare semi-wild bovine species found in the Indo-China. They can graze grasses, including bamboo leaves, as well as reeds and other plant species, and grow to higher mature live weights than Yunnan Yellow cattle maintained in similar harsh environments. The aim of this study was to identify specific cellulase in the gayal rumen. A metagenomic fosmid library was constructed using genomic DNA isolated from the ruminal contents of four adult gayals. This library contained 38400 clones with an average insert size of 35.5 kb. The Umcel-1 gene was isolated from this library. Investigation of the cellulase activity of 24 random clones led to the identification of the Umcel-1 gene, which exhibited the most potent cellulase activity. Sequencing the Umcel-1 gene revealed that it contained an open reading frame of 942 base pairs that encoded a product of 313 amino acids. The putative gene Umcel-1 product belonged to the glycosyl hydrolase family 5 and showed the highest homology to the cellulase （GenBank accession no. YP_004310852.1 ） from Clostridium lentocellum DSM 5427, with 44% identity and 62% similarity. The Umcel-1 gene was heterologously expressed in Escherichia coil BL21, and recombinant Umcel-1 was purified. The activity of purified recombinant Umcel-1 was assessed, and the results revealed that it hydrolyzed carboxymethyl cellulose with optimal activity at pH 5.5 and 45~C. To our knowledge, this study provides the first evidence for a cellulase produced by bacteria in gayal rumen.

Isolation and characteristics of one marine psychrotrophic cellulase-generating bacterium Ar/w/b/75°/10/5 from Chuckchi Sea,Arctic

Microorganisms living in polar zones play an important part as the potential source of organic activity materials with low temperature characteristics in the bio-technological applications. A psychrotrophic bacterium (strain Ar/w/b/75°/10/5) , producing cellulose at low temperatures during late-exponential and early-stationary phases of cell growth, was isolated from sea ice-covered surface water in Chuckchi Sea, Arctic. This bacterium, with rod cells, was Gram-negative, slightly halophilic. Colony growing on agar plate was in black. Optimum growth temperature was 15℃. No cell growth was observed at 351 or above. Optimum salt concentration for cell growth was between 2 and 3 % of sodium chloride in media. Maximal cellulase activity was detected at a temperature of 35℃ and pH8. Cellulase was irreversibly inactivated when incubated at 55℃ within 30 min. Enzyme can be kept stable at the temperature no higher than 25℃. Of special interest was that this bacterium produced various extracellular enzymes including cellulase, amylase, agar hydrolase and protease, at low or moderate temperature conditions, which is certainly of it potential value for applications.

Effects of Exogenous NSP Enzymes(Xylanase, β-glucanase and Cellulase) on Morphology and Functions of Digestive Tract in Growing Pigs Fed with Paddy-Based Diets

Ninety Landrace×Jia 35±0. 40 kg weight growing pigs were randomly allotted to three treatments , each of which was replicated three times with ten pigs per replicate. The pigs were reared on either a conventional corn-based diet (control I ) or a paddy-based diet (control I ) or a paddy diet supplemented with 0.2% NSP enzymes (test group). All pigs were given ad libitum access to both feed and water. The results of feeding trial showed that supplementation of NSP enzymes significantly increased ADG by 8.78% (P< 0.05) and decreased F/G by 9. 42% (P<0. 05) over the control group Ⅱ. No significant differences were found in ADG and F/G between control group I and the test group. The digestive trial showed that adding NSP enzymes significantly improved apparent digestibility of CP, EE and CF by 18. 76 (P<0. 01), 16.04 (P <0.05) and 108. 57%(P<0. 05), respectively, compared to control Ⅱ. The activities of proteolytic enzyme and α-amylase in duodenal contents were increased by 99. 07 (P<0. 01) and 18. 41% (P<0. 05) with the addition of NSP enzymes. No significant differences between test and control Ⅱ group were found in activities of the pepsin in the gastric content, the trypsin and lipase in duodenal contents. the disaccharidase and y-glutany transferase (γ-GT) in intestinal mucosa, but there was a tendency towards higher activities associated with the NSP enzymes diet(P>0. 05). The lengths of the villi within the duodenal, jejunal and ileal sections of the small intestine of pigs receiving the NSP enzymes diet were increased by 23. 68 (P<0. 05), 56. 00 (P<0. 01) and 76. 90%(P<0. 01) respectively, relative to the pigs in controlⅡ.

Optimization of Cultural Condition and Synergistic Effect of Lactose with Carboxymethyl Cellulose on Cellulase Production by <i>Bacillus</i>sp. Isolated from Fecal Matter of Elephant (<i>Elephas maximus</i>)

A cellulase producing bacterium (E3 strain) was isolated from fecal matter of elephant and identified as Bacillus sp. using 16S rDNA sequenced based molecular phylogenetic approach. While studying the effect of substrates like Carboxymethyl cellulose (CMC), avicel, starch, maltose, sucrose, glucose, fructose, galactose and lactose on cellulase production, it was found that CMC was best carbon source induced cellulase production followed by lactose in this bacterial strain. A positive synergistic effect of lactose with CMC was also observed with enhancement of 5 - 6 times in cellulase production. The optimum cellulase production was recorded with 1% CMC and 1% lactose when added individually in the Omeliansky’s medium. The results showed that addition lactose with CMC greatly enhances the production and activity of various cellulase enzymes. The optimal fermentation conditions for the biosynthesis of cellulase by this strain were found to be temperature: 37℃, pH 7.0. The nitrogen source NH4Cl at 0.15% was optimum for cellulase production by this bacterium.

Performance and Nutrient Utilization of Layers Fed Diet Supplemented with Microbial Phytase and Cellulase

A 31 week feeding trial was conducted to investigated the effects of dietary supplementation of microbial phytase and cellulase on performance,nutrients utilization and tibia quality of laying hens fed maize and soybean meal diets.192 18 week old Hisex layers were used in the trial A 2×2×2 factorial design was used in the experiment with three factors of two levels each:0 38% and 0 16% of dietary non phytate P(nP).0 and 300 U·kg -1 of phytase (Ph),and 0 and 0 10% of cellulase (Ce).The results showed that supplementation of 300 U·kg -1 phytase significantly improved utilization of dietary crude ash,CP,Ca,total P and copper (P<0 05),and improved tibia breaking strength (P<0 05).No effect of phytase on performance was observed.Addition of 0 10% cellulase decreased feed intake (P<0 05),increased utilization of CF (P<0 05) and Ca(P<0 01),and decreased total tibia ash weight (P<0 05).300 U·kg -1 phytase and 0 10% cellulase exhibited obvious positive interactions to enhance utilization of dietary phytic P and copper (P<0 05).0 16% nP did not reduce performance of the layers,but improved egg shell quality at 20 wks,increased utilization of dietary total P,phytic P and Copper (P<0 01),decreased utilization of dietary CP,increased tibia breaking strength and Ca,Mn contents of tibia(P<0 01)

植物细胞脱壁酶Cellulase ZA-P的制备和应用效果

Cellulase ZA-P的产生菌为李氏木霉(Trichoderma reesei ZA-1)。研究出适于此菌产酶的液体发酵培养基配方和最佳发酵条件。其发酵液的滤纸酶活力达1.0～2.5IU/ml。发酵液经调节pH值,硫酸铵低浓度盐析和聚苯胺酚树脂脱色等处理后酶液得到纯化,最后经一种简便经济的脱盐干燥方法获得精制酶粉。Cellulase ZA-P可用于蚕豆、大小麦、番茄、烟草和青菜等多种科属植物的原生质体制备。在水稻和番茄上进行的测定试验表明,Cellulase ZA-P能分离产生大量的原生质体,其中番茄原生质体的成活率大于90％,经培养后能正常分裂形成细胞团,并进一步发育成植株。对细胞脱壁效果,原生质体成活率及原生质体分裂频率等方面与Onozuka R-10及RS的效果相当,而对原生质体无毒害。

Application of Statistically Based Experimental Designs to Optimize Cellulase Production and Identification of Gene

A natural bacterial strain identified as Bacillus amyloliquefaciens MBAA3 using 16S rDNA partial genome sequencing has been studied for optimization of cellulase production.Statistical screening of media components for production of cellulase by B.amyloliquefaciens MBAA3 was carried out by Plackett–Burman design.Plackett–Burman design showed CMC,MgSO4 and pH as significant components influencing the cellulase production from the media components screened by Plackett-Burman fractional factorial design.The optimum concentrations of these significant parameters were determined employing the response surface central composite design,involving three factors and five levels was adopted to acquire the best medium for the production of cellulase enzyme revealed concentration of CMC(1.84 g),MgSO4(0.275 g),and pH(8.5)in media for highest enzyme production.Response surface counter plots revealed that middle level of MgSO4 and middle level of CMC,higher level of CMC and lower level of pH and higher level of MgSO4 with lower level of pH increase the production of cellulase.After optimization cellulase activity increased by 6.81 fold.Presence of cellulase gene in MBAA3 was conformed by the amplification of genomic DNA of MBAA3.A PCR product of cellulase gene of 1500 bp was successfully amplified.The amplified gene was conformed by sequencing the amplified product and sequence was deposited in the gene bank under the accession number KF929416.

植物细胞脱壁酶Cellulase ZA-P的性质分析

植物细胞脱壁酶 Cellulase ZA-P 是一种灰白色粉末,酶溶液的颜色极淡,透明,无沉淀,在45℃以下、pH4～7范围内其纤维素酶活力有较好的稳定性,粉剂贮藏1年以上其滤纸酶活力不变。Cellulase ZA-P 的滤纸酶活力为176.7IU/g,CMC 酶活力为1192.9IU/g,棉花酶活力为22.2IU/g,半纤维素酶活力为322IU/g,几丁质酶活力为4.0IU/g。果胶酶、蛋白酶、RNA 酶和过氧化氢酶的含量均很低,未测出α一淀粉酶酶活。制备原生质体的效果、物理性状以及酶系分析结果相似于日本 Yakult Honsha Co 的 Cellulase Onozuka R-10。CellulaseZA-P 对植物原生质体无毒害,适于制备各种植物的原生质体。

Application of response surface methodology to the modeling of cellulase purification by solvent extraction

Central composite design (CCD)sp. JS14 in a solvent extraction was established with Response surface methodology (RSM). Solvent concentration, pH, temperature and retention time were selected as process variables to evaluate the purification impact factor in solvent precipitation, including the purification fold and % recovery. An experimental space with 13 purification fold and 23 recovery percentage recovery is achieved through the optimized condition based on the model. The molecular weight of the purified enzyme was estimated to be 32.5 KDa. Optimum activity of purified enzyme was at pH and temperature 6.5℃ and 40℃ respectively. Enzyme showed maximum activity with carboxymethyl cellulose as substrate with compare to rice husk, wheat straw and sucrose. The purified cellulase activity was inhibited by Na+, Cl- Mg2+ Tween 80 and EDTA.

Cellulase Producing Bacteria from the Wood-Yards on Kallai River Bank

This study evaluates the influence of growth parameters such as pH, temperature, Carboxy Methyl Cellulose (CMC) concentration and agitation on cellulase production from three bacterial strains, viz., Achromobacter xylosoxidans BSS4, Bacillus sp. BSS3 and Pseudomonas sp. BSS2 isolated from the wood-yards on Kallai river bank in Kerala. Production of cellulase by these isolates was detected using basal salt medium (BSM) with 0.5% CMC as supplement, and CMCase activity was confirmed by iodine test. Dinitrosalicylic acid method was employed for assaying the cellulase production by measuring the amount of glucose liberated in μmol/mL/min. Maximum enzyme production from Pseudomonas sp. BSS2 was at pH 8, 37℃ with 1% CMC and 150 rpm, and cellulase production increased from initial 49.84 U/mL to 91.28 U/mL after optimization. The highest enzyme activity from Bacillus sp. BSS3 was at pH 9, 37℃ with 1% CMC, 150 rpm, and cellulase production increased from initial 26.05 U/mL to 104.68 U/mL after optimization. The maximum enzyme production from A. xylosoxidans BSS4 was at pH 7, 40℃ with 0.5% CMC and 150 rpm, and cellulase production increased from initial 55.28 U/mL to 68.37 U/mL after optimization. Thus among the three isolates, Bacillus sp. BSS3 showed maximum enzyme yield which can be explored for further scale up studies with an industrial perspective.

Cellulase in Anoplophora glabripennis adults emerging from different host tree species

In order to investigate different kinds of cellulase in insect pests, we selected male and female adults of Anoplophora glabripennis （Motschulsky） emerging from four different host species （Populus alba var. pyramidalis, P. nigra var. thevestina （Dode） Bean., P. simonii × P. pyramidliscr cv. Opera 8277 Hsu. and Salix matsudana f. lobato-glandulosa） as our research material. The enzyme activitives of three kinds of cellulase in the intestines of the adult insects were measured. The Cx-cellulase isozymes were detected with a CMC-incorporated polyacrylamide gel. The results show that： there are no statistically significant differences between the enzyme activities of males and females emerging from the different host species. The order of magnitude in activity is： Cx-cellulase 〉 β-glycosidase 〉 C1-cellulase. When the adults emerge from the same host species, there are no statistically significant differences between male and female enzyme activities of β-glycosidase and C1-cellulase, but the enzyme activity of Cx-cellulase of males is clearly higher than that of females. The patterns and migration of Cx-cellulase isozymes of males and females emerging from differ-ent hosts trees are clearly not different, and neither are they different when emerging from the same hosts.