Genome-Wide Identification of Epigenetic Regulators in Quercus suber L.

Modifications of DNA and histones, including methylation and acetylation, are critical for the epigenetic regulation of gene expression during plant development, particularly during environmental adaptation processes. However, information on the enzymes catalyzing all these modifications in trees, such as Quercus suber L., is still not available. In this study, eight DNA methyltransferases (DNA Mtases) and three DNA demethylases (DDMEs) were identified in Q. suber. Histone modifiers involved in methylation (35), demethylation (26), acetylation (8), and deacetylation (22) were also identified in Q. suber. In silico analysis showed that some Q. suber DNA Mtases, DDMEs and histone modifiers have the typical domains found in the plant model Arabidopsis, which might suggest a conserved functional role. Additional phylogenetic analyses of the DNA and histone modifier proteins were performed using several plant species homologs, enabling the classification of the Q. suber proteins. A link between the expression levels of each gene in different Q. suber tissues (buds, flowers, acorns, embryos, cork, and roots) with the functions already known for their closest homologs in other species was also established. Therefore, the data generated here will be important for future studies exploring the role of epigenetic regulators in this economically important species.

Physiological Response of Pedunculate Oak Trees to Gall-Inducing Cynipidae.

Gall-inducing Cynipidae (Hymenoptera) manipulate the leaves of their host plants and induce local resistance, resulting in a diversity of physiological changes. In this study, three gall morphotypes caused by the asexual generation of Cynips quercusfolii L., Neuroterus numismalis (Fourc.) and Neuroterus quercusbaccarum L. (Hymenoptera: Cynipidae) on pedunculate oaks (Quercus robur L. (Fagales: Fagaceae)), were used as a model to examine physiological alterations in galls and foliar tissues, compared to non-galled tissues. Our goal was to investigate whether plant physiological response to insect feeding on the same host plant varies depending on gall-wasp species. In particular, the cytoplasmic membrane condition, hydrogen peroxide (H2O2) concentration and changes in antioxidative enzyme activities, including guaiacol peroxidase (GPX) and ascorbate peroxidase (APX) were examined in this study. All cynipid species increased H2O2 levels in the leaves with galls, while the level of H2O2 in galls depended on the species. The presence of galls of all species on oak leaves caused an increase of electrolyte leakage and lipid peroxidation level. A significant induction of GPX activity was observed in the leaves with galls of all species, indicating stress induction. Conversely, the decrease in APX activity in both leaves with galls and galled tissues exposed to feeding of all cynipid species.

Identification of UGT84A13 as a candidate enzyme for the first committed step of gallotannin biosynthesis in pedunculate oak (Quercus robur).

A cDNA encoding the ester-forming hydroxybenzoic acid glucosyltransferase UGT84A13 was isolated from a cDNA library of Quercus robur swelling buds and young leaves. The enzyme displayed high sequence identity to resveratrol/hydroxycinnamate and hydroxybenzoate/hydroxycinnamate glucosyltransferases from Vitis species and clustered to the phylogenetic group L of plant glucosyltransferases, mainly involved in the formation of 1-O-ß-D-glucose esters. In silico transcriptome analysis confirmed expression of UGT84A13 in Quercus tissues which were previously shown to exhibit UDP-glucose:gallic acid glucosyltransferase activity. UGT84A13 was functionally expressed in Escherichia coli as N-terminal His-tagged protein. In vitro kinetic measurements with the purified recombinant enzyme revealed a clear preference for hydroxybenzoic acids as glucosyl acceptor in comparison to hydroxycinnamic acids. Of the preferred in vitro substrates, protocatechuic, vanillic and gallic acid, only the latter and its corresponding 1-O-ß-D-glucose ester were found to be accumulated in young oak leaves. This indicates that in planta UGT84A13 catalyzes the formation of , 1-O-galloyl-ß-D-glucose, the first committed step of gallotannin biosynthesis.

The diversification of terpene emissions in Mediterranean oaks: lessons from a study of Quercus suber, Quercus canariensis and its hybrid Quercus afares.

Interspecific gene flow is common in oaks. In the Mediterranean, this process produced geographical differentiations and new species, which may have contributed to the diversification of the production of volatile terpenes in the oak species of this region. The endemic North African deciduous oak Quercus afares (Pomel) is considered to be a stabilized hybrid between the evergreen Quercus suber (L.) and the deciduous Quercus canariensis (Willd.), presumably being monoterpene and isoprene emitters, respectively. In a common garden experiment, we examined the terpene emission capacities, terpene synthase (TPS) activities and nuclear genetic markers in 52 trees of these three oak species. All but one of the Q. suber and Q. canariensis trees were found to be genetically pure, whereas most Q. afares trees possessed a mixed genotype with a predominance of Q. suber alleles. Analysis of the foliar terpene emissions and TPS activities revealed that all the Q. canariensis trees strongly produced isoprene while all the Q. suber trees were strong monoterpene producers. Quercus afares trees produced monoterpenes as well but at more variable and significantly lower rates, and with a monoterpene pattern different than that observed in Q. suber. Among 17 individuals tested, one Q. afares tree emitted only an insignificant amount of terpenes. No mixed isoprene/monoterpene emitter was detected. Our results suggest that the capacity and pattern of volatile terpene production in Algerian Q. afares populations have strongly diverged from those of its parental species and became quantitatively and qualitatively reduced, including the complete suppression of isoprene production.

Increased drying rate lowers the critical water content for survival in embryonic axes of English oak (Quercus robur L.) seeds.

The potential to cryopreserve embryonic axes of desiccation-sensitive (recalcitrant) seeds is limited by damage during the desiccation necessary for low temperature survival, but the basis of this injury and how to reduce it is not well understood. The effects of drying rate on the viability, respiratory metabolism and free radical-mediated processes were therefore investigated during dehydration of Quercus robur L. embryonic axes. Viability, assessed by evidence of germination and tetrazolium staining, showed a sharp decline at 0.27 and 0.8 g/g during rapid (<12 h) or slow (3 d) dehydration, respectively. Rapid dehydration therefore lowered the critical water content for survival. At any given water content rapid dehydration was associated with higher activities of the free radical processing enzymes, superoxide dismutase, catalase and glutathione reductase and lower levels of hydroperoxide and membrane damage. Rapid dehydration was also associated with lower malate dehydrogenase activity, and a reduced decline in phosphofructokinase activity and in levels of the oxidized form of nicotinamide dinucleotide. Ageing may have contributed to increased damage during slow dehydration, since viability declined even in hydrated storage after 3 d. The results presented are consistent with rapid dehydration reducing the accumulation of damage resulting from desiccation induced aqueous-based deleterious reactions.

Seasonal changes in the excess energy dissipation from Photosystem II antennae in overwintering evergreen broad-leaved trees Quercus myrsinaefolia and Machilus thunbergii.

We monitored chlorophyll (Chl) fluorescence, pigment concentration and the de-epoxidation state of the xanthophyll cycle (DPS(1)) in two warm temperate broad-leaved evergreen species (Quercus myrsinaefolia and Machilus thunbergii). Reduction of the maximal quantum yield of Photosystem II (PSII) (calculated from Fv/Fm, variable to maximal Chl a fluorescence) and retention of a high DPS were observed in both species in the winter, and can be interpreted as acclimation to winter. In particular, the acclimation of PSII in these species can be chiefly attributed to thermal dissipation, which is correlated with the retention of high zeaxanthin. Furthermore, we attempted to divide the fate of the absorbed light energy by the PSII antennae into three components: (i) PSII photochemistry (represented by its quantum yield, ΦPSII), (ii) dissipation by down-regulation via non-photochemical quenching (ΦNPQ) and (iii) other non-photochemical processes (ΦONP). The estimated energy allocation of the absorbed light indicated that the proportion of ΦPSII decreased, whereas that of ΦNPQ+ΦONP increased during winter. This result suggests that the excess energy absorbed in the PSII complexes is safely dissipated from the PSII antennae. Based on these results, we conclude that thermal dissipation from the PSII antennae plays an important role in two overwintering broad-leaved evergreen trees growing in Japan.

A method to quantify transesterification activities of lipases in litters.

Lipases are glycerol ester hydrolases (EC 3.1.1.3) produced by a wide range of microorganisms. They catalyse the hydrolysis of different esters but this reaction is reversible, depending on the water content of the reaction medium, via esterification and transesterification. The synthetic activity of lipases can be of major importance in natural ecosystems since it can be involved in carbon stockage in soils or litters. Here, the detection of transesterification activities of lipases in litter is reported for the first time. We used two different litters: litter of Quercus pubescens (QP) and litter of both Q. pubescens and Q. ilex. Different p-nitrophenyl esters and pentanol were used to test transesterification in a reaction medium with an organic solvent (heptane). We showed that these activities were proportional to the amount of litter, the incubation time and the substrate concentration and that they increased with temperature. Furthermore, the lipases from the litters studied were very thermostable since they were still active after 2 h at 70 degrees C. These activities showed common properties of lipases: the highest activities were obtained with a medium acyl-chain substrate p-nitrophenyl caprylate and transesterification activities were correlated to water activity, a(w). The following parameters are recommended to quantify transesterification activities in litter: 10 mM of p-nitrophenyl caprylate, 1 g of litter, 500 microL of pentanol, q.s.p. 4 mL of heptane incubated at 30 degrees C for 2 h.

Photosynthesis, non-photochemical pathways and activities of antioxidant enzymes in a resilient evergreen oak under different climatic conditions from a valley-savanna in Southwest China.

Plants in the savanna-valleys in Southwest China are annually exposed to different combinations of multiple stresses from the hot-rainy, to chill-dry, and to warm-dry seasons. This study monitored seasonal changes in photosynthesis and photoprotection in an evergreen oak (Cyclobalanopsis helferiana) from one of these valleys for four years during which usual and abnormal drought occurred. In general, during the study period with decreasing xylem water potential (Psix), photosynthetic gas exchange, quantum yield of photosystem II (PSII) photochemistry and activities of most of the measured antioxidant enzymes decreased, while activities of the xanthophyll cycle and associated non-photochemical energy dissipation and glutathione peroxidase (GP) (EC 1.11.1.9) increased. In a fairly severe chill period, high concentration of reactive oxygen species induced high activities of most of the antioxidant enzymes and relatively stronger decrease in gas exchange. In the most severe dry period, even when predawn Psix decreased down to -4 MPa, considerable Pn (maximum photosynthetic rate) (4 micromol m(-2) s(-1)) was still maintained in midmorning. At this time, most of the antioxidant enzyme activities decreased to the lowest values, whereas the xanthophyll cycle and associated non-photochemical energy dissipation and GP activities increased to their highest levels. High predawn antheraxanthin and zeaxanthin contents were observed in the severe and very severe drought periods. Superoxide dismutase maintained high and fairly constant activity (1500-1800 U mg(-1) protein) and predawn maximum photochemistry efficiency of PSII was always above 0.8 throughout the whole study period. These results indicated that the photosynthetic apparatus of the oak leaves was highly capable of maintaining its function under the multiple stresses in different seasons in the present valley-savanna.

[Fine spatial structure of allozyme genotypes in isolated population of pedunculate oak Quercus robur L. (Fagaceae)].

The extent and spatial pattern of genetic variation at polymorphic allozyme loci in a population of pedunculate oak Quercus robur from the Bashkir Transural region was investigated using autocorrelation analysis. In the plantation examined, statistically significant local concentration of most of the alleles in two-dimensional space was identified. The measures for protection of this small population located outside of the western border of the species range, in the mountain--steppe habitat, and characterized by specific gene pool, are suggested.

Hydrolysis of glycosidically bound flavour compounds from oak wood by Oenococcus oeni.

Malolactic fermentation (MLF), which is conducted by lactic acid bacteria (LAB), has a significant influence on the stability and organoleptic quality of wine. Recent studies have shown that when MLF is carried out in oak wood barrels, LAB were also able to interact with wood and increase volatile compound contents such as vanillin during MLF. The release of these compounds indicates that LAB may convert vanillin precursors present in oak wood. In this work, the effect of commercial glycosidases on the released vanillin was firstly studied. This aldehyde is present in wood extracts in monoglycosidic forms where the major glycones are arabinose and xylose. Other aglycons released during MLF in barrels, syringaldehyde and whisky-lactones, can be considered as other sources of aroma. Secondly, strains selected with high activities toward glycoside substrates could hydrolyse vanillin glycoside precursors from oak wood with the same efficiency as commercial enzymes.

Effect of aluminum on nitrate reductase and photosynthetic activities in Quercus serrata seedlings.

We investigated the physiological effect of rhizospheric aluminum (Al) on the activities of nitrate reductase and photosynthesis in Quercus serrata seedlings. The seedlings were cultured hydroponically in nutrient solution with or without 1 mM AlCl(3) (pH 4.0) for 14 days. After Al treatment for 3 days, the number of primordia of tertiary lateral roots on secondary lateral roots appeared to increase. As a result, the biomass of the roots significantly increased (by 5%) after Al treatment for 14 days. The uptake of NO(3)(-) by the seedlings from the culture medium was stimulated to 125% by Al treatment for 3 days. Al treatments for 7 and 14 days increased the nitrate reductase activities in the roots to 300% and 170%, respectively. Al treatment had no effect on photosynthetic activity or shoot biomass even after 14 days, although the chlorophyll content was slightly increased by Al treatment. These results suggest that the stimulation of root growth by Al might be closely related to metabolic changes including the increase in nitrate reductase activity in the leaves and roots.

Response of transglutaminase activity and bound putrescine to changes in light intensity under natural or controlled conditions in Quercus ilex leaves.

In order to further study a previously observed relationship between polyamine (PA) content and changes in irradiation, we examined the level of free and bound PAs, the activity of transglutaminase (TGase, EC 2.3.2.13) and chlorophyll fluorescence in holm oak (Quercus ilex L.) leaves in response to different levels of light intensity and amount. A diurnal trend of free and bound putrescine (F-Put and B-Put, respectively) and TGase activity was observed in plants under natural conditions in the forest, with the highest value corresponding to the maximum light intensity and amount of light received by the leaves. In another set of experiments, potted Q. ilex plants in experimental fields were subjected to a range of periods of natural photosynthetic photon flux density (PPFD) by covering or not covering the whole trees. Under a natural photoperiod (uncovered leaves), B-Put content and TGase activity paralleled the diurnal PPFD pattern, reaching a maximum at the highest PPFD; prior to this maximum, free PAs showed a significant rise. Plants that were in darkness until midday and suddenly exposed to high light intensity showed enhanced TGase activity, resulting in the maximum accumulation of B-Put. The involvement of the accumulation of B-Put reflected in the changes of the B-Put/bound spermidine ratio during the photoprotective responses to high light stress in forest plants is discussed in relation to the chlorophyll fluorescence parameters observed.

Differential expression of two class III chitinases in two types of roots of Quercus robur during pre-mycorrhizal interactions with Piloderma croceum.

Expression of two plant chitinase genes, representing members of class III chitinases, was studied in Quercus robur roots during interactions in a pre-mycorrhizal stage with the ectomycorrhizal fungus Piloderma croceum. Chitinase gene expression was compared in lateral roots destined to form ectomycorrhiza, and in principal roots that are not directly involved in mycorrhizal interactions. The transcript level of the first chitinase (QrchitIII-1) was upregulated in lateral roots, whereas no significant differential expression was observed in principal roots. The second chitinase (QrchitIII-2) was regulated neither in lateral nor in principal roots in presence of the fungus. Because P. croceum did not induce significant chitinase responses in principal roots, the enhanced expression of QrchitIII-1 in lateral roots after inoculation may be related to some steps in symbiosis ontogenesis.

Inferring colonization history from analyses of spatial genetic structure within populations of Pinus strobus and Quercus rubra.

Many factors interact to determine genetic structure within populations including adult density, the mating system, colonization history, natural selection, and the mechanism and spatial patterns of gene dispersal. We examined spatial genetic structure within colonizing populations of Quercus rubra seedlings and Pinus strobus juveniles and adults in an aspen-white pine forest in northern Michigan, USA. A 20-year spatially explicit demographic study of the forest enables us to interpret the results in light of recent colonization of the site for both species. We assayed 217 Q. rubra seedlings and 171 P. strobus individuals at 11 polymorphic loci using nine allozyme systems. Plant genotypes and locations were used in an analysis of spatial genetic structure. Q. rubra and P. strobus showed similar observed levels of heterozygosity, but Q. rubra seedlings have less heterozygosity than expected. Q. rubra seedlings show spatial genetic clumping of individuals on a scale to 25 m and levels of genetic relatedness expected from the clumped dispersion of half-siblings. In contrast, P. strobus has low levels of genetic relatedness at the smallest distance class and positive spatial genetic structure at scales < 10 m within the plot. The low density of adult Q. rubra outside the study plot and limited, spatially clumped rodent dispersal of acorns is likely responsible for the observed pattern of spatial genetic structure and the observed heterozygote deficit (i.e. a Wahlund effect). We attribute weaker patterns observed in P. strobus to the longer dispersal distance of seeds and the historical overlap of seed shadows from adults outside of the plot coupled with the overlap of seed shadows from younger, more recently established reproductive adults. The study demonstrates the utility of long-term demographic data in interpreting mechanisms responsible for generating contemporary patterns of genetic structure within populations.

Seasonal changes in photosynthesis and photoprotection in a Quercus ilex subsp. ballota woodland located in its upper altitudinal extreme in the Iberian Peninsula.

Quercus ilex L. subsp. ballota (Desf.) Samp., a Mediterranean evergreen species growing in a continental Mediterranean climate, did not experience water stress and showed greater sensitivity to winter stress than to summer stress over a 12-month period. Net CO2 assimilation rates and photosystem II (PSII) efficiency decreased markedly during the cold months and recovered completely in spring. Lutein, neoxanthin and beta-carotene to chlorophyll (Chl) molar ratios all showed the same trend throughout the year, increasing from September to March. This increase was a result of increases in carotenoid concentrations, because Chl concentration per unit leaf area remained stable, and was higher at the end than at the beginning of the first growing season. Lutein-epoxide was a minor component of the total lutein pool. Thermal energy dissipation and non-photochemical quenching (NPQ) were associated with the de-epoxidated forms of the xanthophyll cycle pigments in the warm months. Photosynthetic rates decreased slightly at midday in summer. These changes were accompanied by decreases in maximum potential PSII efficiency (which recovered during the night), actual and intrinsic PSII efficiencies, photochemical quenching and increases in NPQ. Overall, our data indicate down-regulation of photosynthesis during the summer. The diurnal de-epoxidation of violaxanthin to antheraxanthin and zeaxanthin occurred throughout the year, except in January. Antioxidant enzymatic activity increased in the winter months, especially during the coldest months, highlighting its key role in photoprotection against photo-oxidation. Structural and functional modifications protected PSII from permanent damage and allowed 1-year-old leaves to photosynthesize at high rates when temperatures increased in spring.

Differential activity of peroxidase isozymes in response to wounding, gypsy moth, and plant hormones in northern red oak (Quercus rubra L.).

We measured total peroxidase activity and the activities of peroxidase isoforms in leaves of red oak (Quercus rubra L.) seedlings exposed to wounding and plant hormones in the greenhouse. Activity of specific peroxidase isoforms was induced differentially by gypsy moth wounding, mechanical wounding, and the wound-associated plant hormone jasmonic acid. Activity of one isoform was enhanced modestly by treatment with salicylate. A study of peroxidase activity in naturally occurring galls elicited on red oak leaves by 12 hymenopteran and dipteran insect species found 16 POD isoforms, 11 of which were differentially induced or suppressed in galls compared with leaves. In both studies, total peroxidase activity as measured spectrophotometrically was not clearly related to activity of these isoforms. These results indicate that red oak seedlings and trees may respond specifically to wounding, particular insects, and plant signals through changes in the activities of individual isozymes.

Changes in pectins and MAPKs related to cell development during early microspore embryogenesis in Quercus suber L.

The occurrence and significance of changes in cell wall components and signalling molecules has been investigated during early microspore embryogenesis in cork oak (Quercus suber L.) in relation to cell proliferation and cell differentiation. Microspore embryogenesis has been induced in in vitro anther cultures of Q. suber by the application of a stress treatment of 33 degrees C. After the treatment, microspores at the responsive developmental stage of vacuolate microspore switched towards proliferation and the embryogenesis pathway to further produce haploid plantlets. Ultrastructural and immunocytochemical analysis revealed changes in cell organisation after induction at different developmental stages, the cellular features displayed being in relation to the activation of proliferative activity and the beginning of differentiation in young and late proembryos. Immunogold labelling with JIM5 and JIM7 antibodies showed a different presence of pectin and level of its esterification in cell walls at different developmental stages. Non-esterified pectins were found in higher proportions in cells of late proembryos, suggesting that pectin de-esterification could be related to the beginning of differentiation. The presence and subcellular distribution of Erk 1/2 MAPK homologues have been investigated by immunoblotting, immunofluorescence and immunogold labelling. The results showed an increase in the expression of these proteins with a high presence in the nucleus, during early microspore proembryos development. The reported changes during early microspore embryogenesis are modulated in relation to proliferation and differentiation events. These findings provided new evidences for a role of MAPK signalling pathways in early microspore embryogenesis, specifically in proliferation, and would confer information for the cell fate and the direction of the cell development.

Genetic variation of oaks ( Quercus spp.) in Switzerland. 3. Lack of impact of postglacial recolonization history on nuclear gene loci.

Quercus petraea, Quercus pubescens and Quercus robur are closely related and interfertile white oaks native to Switzerland. The three species are known to share identical cpDNA haplotypes, which are indicative of the postglacial recolonization history of populations. Only two haplotypes are common in Switzerland. We compared variation of cpDNA and of isozymes in 28 oak populations from Switzerland in order to assess the impact of the postglacial population history on current genetic structures of nuclear controlled isozyme gene loci. Species delineation was based on Principal Component Analysis of leaf morphological traits. The species status of populations was reflected at isozyme gene loci, but differentiation between populations with different cpDNA haplotypes and hence different recolonization history was very low at enzyme gene loci for all species. Thus, glacial and postglacial population history was not reflected at nuclear gene loci on the temporal and spatial scale covered by the present study. Extensive gene flow through pollen among populations is likely to have blurred a previously existing genetic differentiation at biparentally inherited gene loci that possibly evolved in the different glacial refugia of the above mentioned cpDNA haplotypes.

Relationship of isopentenyl diphosphate (IDP) isomerase activity to isoprene emission of oak leaves.

Oaks emit large amounts of isoprene, a compound that plays an important role in tropospheric chemistry. Isopentenyl diphosphate isomerase (IDI, E.C. 5.3.3.2) catalyzes the isomerization of isopentenyl diphosphate (IDP) to dimethylallyl diphosphate (DMADP), and in isoprene-emitting plants, isoprene synthase (IS) converts the DMADP to isoprene. To study the role of IDI in isoprene biosynthesis of oak leaves, we compared IDI and IS activities in pedunculate oak (Quercus robur L.) and pubescent oak (Quercus pubescens Willd.) with the isoprene emission rates of these species. We developed a non-radioactive enzyme assay to detect IDI activity in crude leaf extracts of Q. robur. The substrate dependency of IDI activity showed biphasic kinetics with Michaelis constants (K(m)(IDP)) of 0.7 +/- 0.2 micro M for a high-affinity phase and 39.5 +/- 6.9 micro M for a low-affinity phase, potentially attributable to different IDI isoforms. Under standard assay conditions, the temperature optimum for IDI activity was about 42 degrees C, but IDI activity was detectable up to 60 degrees C. A sharp pH optimum appeared around pH 7, with 20 mM Mg(2+) also required for IDI activity. Neither IDI activity nor IS activity showed diurnal variation in Q. robur leaves. The sum of IDI activities showed a significant linear correlation with IS activity in both Q. robur and Q. pubescens leaves, and both enzyme activities showed a linear relationship to isoprene emission factors in leaves of these oak species, indicating the possible involvement of IDI in isoprene biosynthesis by oak leaves.

Measurement and characterization of cellulase activity in sclerophyllous forest litter.

Cellulases are enzymatic proteins which hydrolyze cellulose polymers to smaller oligosaccharides, cellobiose and glucose. They consist in three major types of enzymes: endoglucanases (EC 3.2.1.4), cellobiohydrolases (EC 3.2.1.91) and beta-glucosidases (EC 3.2.1.21) which play an essential role in carbon turnover of forest ecosystem. The aim of this study was firstly to determine the parameters (i.e. buffer type, pH, temperature, quantity of litter, incubation time and reagent type) which affect the measurement of cellulase activity in a sclerophyllous forest litter, and secondly to compare two methods for measuring cellulase activity: a direct method and an extraction method. In the direct method, the litter was directly incubated with a buffered solution containing the enzyme substrate, whereas in the extraction method, the cellulases were firstly extracted before measuring their activity. The results were compared with other studies about soil cellulase activity, and it appeared that several parameters (buffer type, pH, temperature and sample quantity) which influence the measurement of cellulase activity differ according to whether a soil or a litter is considered. Concerning the procedure used for the measurement of cellulase activity, results showed that the activity values were higher when using an extraction procedure than when using a direct procedure. The extraction procedure, combined with a concentration stage of the extract, also allowed electrophoretic analysis (PAGE) of the cellulases extracted from the litter. The electrophoretic pattern revealed two cellulase isoenzymes which may be related to the occurrence of two pH-activity peaks of these enzymes when citrate buffer was used for the measurement of cellulase activity in the litter.