Chemistry and Microscopy of Wood Decay by Some Higher Ascomycetes

Vol. 43 (1989) No. 1 Chemistry and Microscopy of Wood Decay by Ascomycetes 11 Holzforschung OFFPRINT 43 (1989) 11-18

Chemistry and Microscopy of Wood Decay by Some Higher Ascomycetes By Thomas Nilsson and Geoffrey Daniel Department of Forest Products, The Swedish University of Agricultural Sciences, Box 7008, S-75007 Uppsala, Sweden and T. Kent Kirk and John R. Obst Forest Products Laboratory1), Forest Service, U.S. Department of Agriculture, Madison, Wisconsin 53705-2398, U.S.A.

Keywords Summary White-rot Chemical and microscopic features of wood decay by several ascomycetes in axenic culture are described. Soft-rot The tested ascomycetes caused significant weight losses in birch wood. Daldinia concentrica was espe- Lignin biodegradation cially active, causing a weight loss of 62.9% after 2 months. While lignin and carbohydrates were both Syringyl:guaiacyl ratio degraded, carbohydrates were preferentially attacked. As measured by alkaline nitrobenzene oxidation, Daldinia syringylpropane units of the lignin were removed selectively. All of the tested ascomycetes eroded fiber Hypoxylon cells walls beginning from the lumen. Some also caused cavities in the secondary walls, typical of soft-rot Xylaria decay. Pine wood was generally resistant to decay; some of those species which were capable of forming Libertella soft-rot cavities in birch caused significant weight loss in pine. Alstonia scholaris, a tropical hardwood Alstonia scholaris with a guaiacyl-rich lignin, was resistant to degradation. Betula verrucosa Populus tremula Pinus sylvestris

Introduction remnants of the middle lamella were seen. The vessels, ray parenchyma, and vertical parenchyma in lime showed no signs of degra- Ascomycetes and Fungi imperfecti generally cause dation even in advanced decay. In elm the vessels and ray paren- soft-rot decay of wood (Savory 1954; Corbett 1965; chyma in the earlywood but not latewood resisted degradation. Duncan and Eslyn 1966; Nilsson 1973). Xylariaceous From staining experiments the author concluded that removal of ascomycetes from genera such as Daldinia, Hypoxy all substance from the fiber walls in both woods was preceded by localized delignification. Later, larger areas of the walls were delig- lon, and Xylaria have often been regarded as white- nified. The decayed wood was lighter in color than the normal rot fungi (Campbell and Wiertelak 1935; Cartwright wood, indicating a bleaching effect. and Findlay 1946; Kirk 1971; Rogers 1979). Past re- Laboratory decay studies with axenic cultures were search on wood decay by the ascomycetes has been later conducted by several investigators. Blaisdell limited; only one report was found on chemical ana- (1939) found significant weight losses in three differ- lyses of wood decayed in axenic culture. The purpose ent hardwoods by Daldinia concentrica, Hypoxylon of the present study was to describe the chemical and sp., and Xylaria sp. Cartwright and Findlay (1946) re- microscopic features of attack of woods by several ported that x. polymorpha caused a weight loss of higher ascomycetes in axenic culture. 14% in beech wood blocks after 4 months. Similarly, The earliest reports were of chemical and microscopic studies of decay of aspen wood to 16% weight loss by X. digitata naturally decayed hardwoods. Thus Campbell and Wiertelak H. rubiginosum (1935) analyzed lime (Tilia vulgaris) wood decayed by Ustulina vul and after 3 months was reported by garis Tul. (= Hypoxylon deustum (Hoffm.: Fr.) Grev.). They esti- Rajagopalan (1966). Weight losses from 10 to 26% in mated from chemical analyses that about one-third of the cellulose aspen and red oak woods in 3 months by D. concen and one-fourth of the lignin had been lost through fungal attack. trica, H. atropunctatum, H. mediterraneum, H. Wilkins (1936, 1939) gave detailed microscopic descriptions of ad- pruinatum, and H. punctulatum were reported by vanced decay by U. vulgaris in standing trees of lime and elm. At- tack in lime, more pronounced in latewood than in earlywood fi- Merrill, French, and Wood (1964). Weight losses by a bers, led to a thinning of secondary cell walls. The middle lamella white-rot fungus and a brown-rot fungus included for in both lime and elm persisted, but in the most advanced decay only comparison were 50 to 70%. Microscopic examination of aspen blocks decayed by H. pruinatum re- 1) The Laboratory is maintained in cooperation with the University vealed an erosion form of attack in fiber, vessel, and of Wisconsin. ray parenchyma walls; the middle lamella was not af-

Holzforschung / Vol. 43 / 1989 / No. 1 1989 Walter de Gruyter · Berlin · New York 12 T. Nilsson, G. Daniel, T.K. Kirk and J.R. Obst Holzforschung fected, and no soft-rot cavities were reported. No Wood species chemical analyses of the laboratory-decayed woods The following wood species were used in the experiments: birch were reported by any of the foregoing authors. In (Betula verrucosa Ehrh.), Scots pine (Pinus sylvestris L.), Euro- other work, however, Kistler and Merrill (1968) pean aspen (Populus tremula L.), and a tropical hardwood Alstonia scholaris (L.) R.Br. The latter has an exceptionally high lignin con- found a lignin loss of 25% at a weight loss of 25% in tent of approximately 30% and, as shown here, a guaiacyl-rich lig- red oak decayed by Strumella coryneoidea. Weight nin. losses up to 40% were also observed. The fungus For the majority of the decay experiments, sapwood blocks of birch caused a gradual thinning of the secondary cell walls. and pine measuring 5 × 15 × 30 mm (5- × 15-mm transverse face) The xylariaceous ascomycetes occur primarily on an- were employed. In one test involving Daldinia concentrica and Tra metes versicolor the block size was 10 × 20 × 20 mm (20- × 20-mm giosperm wood (Rogers 1979), and apparently no one transverse face). has studied the effects of these fungi on gymnosperm woods. The group is important in degrading woody Decay materials on the forest floor. The wood blocks were dried at 105°C and weighed. They were then buried, transverse face downwards, in a layer of moist commercial In the present study we conducted both chemical and potting soil in 100-ml Erlemeyer flasks. The flasks were equipped microscopic analyses of birch wood decayed by sev- with cotton plugs, sterilized by autoclaving, cooled, inoculated with eral species of the Xylariaceae. Pine, aspen, and suspensions of mycelium, and incubated for 2 and 4 months at Alstonia woods were included for comparison with 25°C. The initial moisture content of the wood blocks was certain fungi. 30-40%.In a separate experiment with D. concentrica and T. ver sicolor on aspen birch, and Alstonia scholaris wood blocks, the incubation period was 10 weeks. After incubation, the wood blocks were removed and cleaned of Material and Methods adhering soil and mycelium. They were then dried at 105°C to con- Fungal species stant weight. Weight losses were calculated as percentage of the original weight. The studied species and their origins are given in Table 1.

Table 1. Species and strains of fungi studied. and their origins

Species Strain Origin

Ascomycetes Daldinia concentrica (Bolt.: Fr.) Ces. and De Not C62 Forintek1) D. eschscholzii (Ehrenb.) Rehm Candoussau J.D.R.2) D. occidentalis Child 9597 Petrini3) Hypoxylon confluens (Tode: Fr.) Westend. CBS335.70 CBS4) H. deustum (Hoffm.: Fr.) Grev. CBS 723.69 CBS (Synonym: Ustulina vulgaris Tul.) H. fuscum (Pers.: Fr.) Fr. C144 Forintek H. mammatum (Wahlenb.) J.H. Miller 324A, 238A5) Forintek H. multiforme (Fr.) Fr. CBS 856.72 CBS H. nummularium Bull.: Fr. CBS 969.70 CBS H. rubiginosum Pers.: Fr. fide Miller C194, 328A6) Forintek Libertella betulina Desm. [anamorph of BH SLU7) Diatrype stigma (Hoffm.: Fr.) Fr.] Xylaria acuta Peck Cooke J.D.R. X. arbuscula Sacc. Samuels J.D.R. X. carpophila (Pers.: Fr.) Fr. 9612 Petrini X. feejeensis (Berk.) Fr. 9613 Petrini X. hypoxylon (L.: Fr.) Grev. 74426 SLU X. longipes Nits. CBS 148.73 CBS X. polymorpha (Pers.: Fr.) Grev. Proffer J.D.R. Basidiomycete Trametes versicolor (Fr.) Pilat (= Coriolus versicolor (Fr.) Quel.)

1) Obtained from Keith A. Seifert, Forintek Canada Corp., Ottawa, Canada. 2) Obtained from Jack D. Rogers, Department of Plant Pathology, Washington State University, Pullman, Washington, U.S.A. 3) Obtained from Liliane Petrini, Benglen, Switzerland. 4) Obtained from Centraalbureau voor Schimmelcultures, Baarn, The Netherlands. 5) Received as Hypoxylon pruinatum. 6) Received as Hypoxylon perforatum. 7) Obtained from the culture collection at Department of Forest Products, Swedish University of Agricultural Sciences, Uppsala, Sweden Vol. 43 (1989) No. 1 Chemistry and Microscopy of Wood Decay by Ascomycetes 13

For microscopy, another type of decay procedure was used (Nilsson Table 2. Average *) percentage weight loss of birch and pine wood 1973). Sterilized birch wood blocks (5 × 5 × 10 mm) were placed decayed for 2 and 4 months by 10 species of ascomycetes on mycelia growing on malt extract or cellulose/mineral salts medium on agar slopes. Test fungus Average weight loss, % In a separate experiment, thin transverse sections (approx. 10 µm) of pine sapwood were exposed directly to the mycelium of D. con 2 months 4 months centrica growing on malt agar plates. Birch Pine Birch Pine Transmission electron microscopy (T.E.M.) Daldinia concentrica 62.9 2.5 76.6 2.4 Small pieces of birch wood from blocks degraded by either D. con Hypoxylon fuscum 20.1 2.4 40.3 2.3 centrica X. polymorpha or were fixed at room temperature in 3% H. mammatum (324A) 20.4 2.6 39.9 4.7 v/v glutaraldehyde and 2% paraformaldehyde in 0.1M sodium H. mammatum (238A) 23.7 2.2 45.7 1.4 cacodylate buffer (pH 7.2). After washing in buffer (3 × 30 min) H. multiforme 22.7 1.4 49.0 2.5 the samples were post-fixed in 1% w/v osmium tetroxide (3 hr, H. rubiginosum (194) 21.0 1.5 34.7 1.1 room temperature) also in buffer. Thereafter, the samples were H. rubiginosum (328A) 8.9 2.2 21.2 2.0 washed in deionized water and dehydrated in a graded ethanol series to propylene oxide and then flat embedded in Spurr’s (1969) Libertella betulina 21.0 2.5 45.6 2.1 resin. Selected material was sectioned on a LKB Ultramicrotome I Xylaria acuta 18.3 12.4 23.4 18.4 fitted with a diamond knife, with sections collected on carbon- X. arbuscula 9.1 3.3 14.9 6.7 stabilized parlodion-coated copper grids. After post-staining with X. hypoxylon 16.1 2.2 40.6 3.3 lead citrate (Reynolds 1963) and uranyl acetate, sections were vie- X. polymorpha 21.4 12.0 26.6 18.8 wed using a Philips 201 T.E.M. operated at 60 kV. *) Average of 10 wood blocks Light microscopy Thin transverse or longitudinal sections were cut from decayed wood blocks. The sections were stained with either aqueous saf- ranine or an aniline blue-lactic acid solution. Polarized light was also employed. The second decay experiment (Table 3) showed that D. concentrica was as effective as T. versicolor in de- Lignin and wood sugar analyses caying aspen wood, but less active in degrading birch The wood samples were ground to pass a 40-mesh screen and the wood. The weight losses of Alstonia wood blocks wood meal dried in a vacuum. The samples were then analyzed, were much lower for both of these fungi but lowest without extraction, for sulfuric acid (Klason) lignin (Effland 1977). for D. concentrica (4.1%). The hydrolysate was then analyzed for individual sugars using high performance liquid chromatography (Wentz, Marey, and Gray Table 3. Average *) percentage weight loss of aspen, birch, and 1982). Alstonia scholaris wood blocks exposed for 10 weeks to Daldinia concentrica and Trametes versicolor Determination of syringyl:guaiacyl (S:G) ratio in lignin The S:G ratio of the lignin in the wood samples was determined Test fungus Aspen Birch Alstonia from the yields of syringaldehyde and vanillin produced on nitrobenzene oxidation. About 40 mg of sample were oxidized with D. concentrica 46.8 39.2 4.1 0.5 mi of nitrobenzene and 5 ml of 2N sodium hydroxide in stainless T. versicolor 46.0 58.2 14.7 steel bombs for 2.5 hours at 165°C. The alkaline solution was washed with chloroform to remove neutrals, acidified, and then ex- *) Average of 10 wood blocks tracted with chloroform to give the aldehydes. Quantitation was by gas chromatography on a 60-m DB-5 capillary column (J & W Sci- entific, Inc., Folsom, CA) using isovanillin as an internal standard.

Chemical analyses (Table 4) showed that at 20% Results weight loss about 20% of the lignin was lost. One ex- rubiginosum In the first experiment, the tested ascomycetes caused ception was the wood decayed by H. significant weight losses in birch wood (Table 2). D. (strain 328A), in which the loss of lignin was only concentrica was the most effective, causing a weight 11.7% at a weight loss of 21.2%. At higher weight los- loss of 62.9% after 2 months. The other ascomycetes ses the lignin losses lagged considerably behind those were less active, causing weight losses ranging from for carbohydrates for all of the fungi. The highest lig- D. concentrica 8.9 to 23.7% after 2 months, and from 14.9 to 49.0% nin loss, 44%, was obtained with at a in 4 months. X. acuta and X. polymorpha also caused weight loss of 76.6%. significant weight losses in pine wood, 12 and 18%, The degradation of cellulose (glucan, measured as respectively, after 2 and 4 months. Small weight losses glucose) and xylan (xylose) was simultaneous in sev- were also obtained in pine after 4 months with X. ar eral cases, but in other samples the xylan was de- buscula (6.7%) and H. mammatum (4.7%). The graded in preference to cellulose. No difference in other ascomycetes produced very low weight losses in chemical composition of the degraded birch wood was pine wood. found between the cavity-forming species and those 14 T. Nilsson, G. Daniel, T.K. Kirk and J.R. Obst Holzforschung

Table 4. Chemical analyses of birch wood samples decayed by ratios. The sample with the highest lignin loss, 44%, seven species of ascomycetes had the lowest S:G ratio.

Ascomycete Percent weight loss*) Microscopic examination showed two distinct types of degradation in the fibers of decayed birch blocks. One Wood Lignin Glucose Xylose was formation of typical soft-rot cavities in the sec- (glucan) (xylan) ondary cell walls. Cavity formation also occurred in pine (Fig. 1). The other was a form of erosion starting Daldinia concentrica 62.9 37.9 76.3 77.4 76.6 44.0 91.3 91.6 from the lumen (Fig. 2). Using the terminology of Hypoxylon fuscum 21.1 20.8 28.8 35.0 Corbett (1965), the two forms of decay are Type 1and 40.3 24.0 56.7 59.4 Type 2, respectively. Table 6 shows the types of decay H. mammatum (324A) 20.4 22.5 20.6 29.1 associated with the test fungi. All of them caused Type 39.9 29.2 46.4 53.9 2 form of attack. H. mammarum and all the Xylaria H. mammatum (238A) 23.7 20.5 29.3 33.0 45.7 26.7 56.2 62.6 species also formed typical soft-rot cavities (Type 1). H. multiforme 26.3 18.2 33.3 34.3 T-branching of hyphae penetrating into the wood cell 49.0 31.6 57.7 59.6 walls (c.f. Corbett and Levy 1963) was observed in all H. rubiginosum (194) 21.0 20.0 22.4 29.2 cavity-forming species. 34.7 23.3 44.9 49.2 H. rubiginosum (328A) 8.9 11.6 6.0 5.0 The erosion type of attack was caused by hyphae 21.2 11.7 26.7 33.5 growing in the fiber cell lumens. The erosion varied Libertella berulina 21.0 22.3 20.8 22.0 from fairly even removal of cell wall material (Fig. 2) 45.6 28.8 56.8 56.3 to more localized attack resulting in irregular troughs Xylaria hypoxylon 16.0 19.1 18.2 17.1 (Fig. 3). The ray cell walls were degraded in the same 20.3 21.1 22.1 28.3 manner, but vessel walls appeared to be little affected *) On basis of original weights by erosion. Soft-rot cavities, however, were formed in the vessel walls by the cavity-forming species. The

erosion of the fiber and ray walls progressed to the S1 layer. This layer and the middle lamella persisted that eroded the wood cell walls (microscopic features even in late stages of attack (Fig. 4). The remaining are described below). layers were birefringent in polarized light on trans- On the basis of alkaline nitrobenzene oxidations, the verse sections, indicating that cellulose was still pre- syringyl:guaiacyl (S:G) ratios of the residual lignin in sent. birch wood decayed by several of the fungi revealed Light microscopic observations of transverse sections that the syringylpropane units had been preferentially from birch wood with fiber degradation with hyphae degraded (Table 5). The samples with lignin losses in the lumen showed that the tertiary wall (S3) in most from 24 to 36% had approximately the same S:G of the fibers persisted, whereas the S2 was extensively

Table 5. Nitrobenzene oxidation of birch wood decayed by four ascomycetes

Weight Lignin Sample Syring- Vanillin SA/V “S:G” loss% loss % weight aldehyde mg (V) molar (= SA/3V) mg (SA)

Sound birch - - 40.3 3.17 0.72 3.68 1.23 Decayed by: H. fuscum 40.3 24.0 40.8 2.74 0.88 2.60 0.87 L. betulina 45.6 28.8 40.5 2.96 1.00 2.47 0.82 H. multiforme 49.0 31.6 40.6 3.24 1.05 2.58 0.86 D. concentrica 76.6 44.0 40.3 3.24 1.74 1.55 0.52

Fig. 1. Light microscopic view of pine latewood tracheids attacked by Xylaria longipes, showing typical soft-rot cavities within the S2 layer. (Bar = 10 µm) Fig. 2-4. Transmission electron microscopic views of birch fibers in transverse section, showing various stages of erosion by Daldinia concentrica.

2. Early stage showing partial erosion of the S2 layer by lumenal hyphae. (Bar = 2 µm)

3. Localized erosion troughs in the S2 layer. Note the hyphae and the presence of abundant extracellular slime. (Bar = 3 µm)

4. Advanced erosion with the S2 being totally eroded and only the S1 and middle lamella regions remaining. (Bar = 2 µm)

Abbreviations used in Figs. 1-4: ML, middle lamella; MLcc, middle lamella cell corner; ES, extracelular slime; S1, S2, secondary cell wall layers; ET, erosion trough; C, cavity; CL, cell lumen; H, hypha. Vol. 43 (1989) No. 1 Chemistry and Microscopy of Wood Decay by Ascomycetes 15

Fig. 1-4. Legend see page 14 16 T. Nilsson, G. Daniel, T.K. Kirk and J.R. Obst Holzforschung

Table 6. Types of decay caused by the studied ascomycetes. Type in thin transverse sections of pine exposed to D. con 1 decay is cavity formation typical of soft-rot decay. Type 2 decay centrica for 4 months, nor was any degradation de- is cell wall erosion starting from the lumen tected by light microscopy in wood blocks of A. scholaris decayed by the same fungus. T. versicolor, Ascomycete Type 1 Type 2 however, caused typical cell wall thinning through Daldinia concentrica erosion in A. scholaris. D. eschscholzii D. occidentalis Discussion Hypoxylon deustum Soft-rot fungi are generally characterized by their for- H. fuscum mation of cavities within wood cell walls. Attack in H. mammatum (324A) H. mammatum (238A) the form of erosion of fiber cell walls is, however, also H. multiforme very common in birch, but not in pine or spruce H. nummularium (Nilsson 1973). A typical feature of white-rotted an- H. rubiginosum (194) giosperm and gymnosperm woods is the erosion of H. rubiginosum (328A) cell walls (Wilcox 1970). Soft-rot and white-rot ero- Libertella betulina sion cannot always be distinguished from each other Xylaria acuta X. arbuscula microscopically. The ascomycetes tested here differ X. carpophila from typical white-rot fungi by their failure to erode X. feejeensis pine wood tracheids. They were also unable, in con- X. hypoxylon trast to most white-rot fungi, to degrade the middle X. longipes lamella in birch. Even so, H. mammatum and all X. polymorpha Xylaria species formed typical soft-rot cavities in birch and pine wood. Thus their micromorphological decay pattern is similar to that of typical soft-rot fungi. Cavity formation has also been reported for X. degraded. These observations could not reveal digitata (L. ex Fr.) Grev. (Duncan and Eslyn 1966). whether tertiary walls had been affected by degrada- This form of attack appears to be characteristic for the tion. In any event, the degrading enzymes produced genus Xylaria. by lumenal hyphae evidently diffused through the ter- In the present work, another trait common to all tiary walls and then into the S2 layer. Similar decay of patterns have been observed for both soft-rot (Nilsson the fungi also is shared by soft-rot fungi: the preferen- 1973) and white-rot fungi (Nilsson, unpublished). tial utilization of the carbohydrates in birch. This is Several of the ascomycetes produced a mucilage-like also a characteristic of some white-rot fungi (Eslyn substance in the fiber lumens that became attached to and Nakasone 1984). The ability to degrade lignin in the loosened tertiary wall, forming a membrane-like wood is a typical feature of white-rot fungi. However, structure. It was very difficult in later stages of attack significant losses of lignin have also been reported for to distinguish between fungal-derived material and wood attacked by typical soft-rot fungi (Levi and Pre- the remains of the fiber cell wall. ston 1965; Eslyn, Kirk and Effland 1975). Thus chemical effects of decay for some soft-rot fungi are similar Localized delignification in the form of ,,spots" as de- to those of white-rot fungi. scribed by Wilkins (1936, 1939) was not observed in any of the examined wood samples, nor was a general Pine wood was significantly attacked only by the cav- delignification as described for certain white rots ity-forming species, although weight losses were con- (Otjen and Blanchette 1986). siderably lower than those in birch. The ascomycetes that attacked birch wood exclusively through erosion D. concentrica rapidly colonized the birch wood ves- were unable to attack pine significantly. Pine wood is sels and developed abundant dendritic, heavily also much more resistant than birch wood to degrada- melanized mycelium. Hyphae growing in fiber cell lu- tion by white-rot fungi (Peterson and Cowling 1964; mens were also occasionally melanized. Melanin was Highley 1982). Evidence was presented that the dif- also formed around individual hyphae and was even ference in lignin type is responsible for the more facile present on some fiber cell walls. No attack of the ves- white-rot degradation of angiosperm woods than sel walls was observed in spite of the prolific fungal gymnosperm woods (Highley 1982; Faix, Mozuch, growth. and Kirk 1985). Typical gymnosperm lignins are com- Pine wood was affected to a significant degree only by posed almost entirely of guaiacylpropane units (G) the cavity-forming species, mostly in the latewood (Obst and Landucci 1986). In contrast, most angios- tracheids. Even so, degradation was not extensive. perm lignins are formed from both guaiacylpropane Very rarely was a form of erosion observed that was and syringylpropane units (S). The S:G ratio of reminiscent of that in birch. No attack was observed hardwoods varies greatly, but many temperate Vol. 43 (1989) No. 1 Chemistry and Microscopy of Wood Decay by Ascomycetes 17 hardwood lignins have S:G ratios in the range of 0.7 Farrell 1987; Buswell and Odier 1987). These en- to 1.3. zymes oxidize the aromatic nuclei of lignin by one Our results with the nitrobenzene oxidation electron, producing cation radicals which undergo a technique show that the S:G ratio of birch was de- variety of spontaneous degradative reactions. The en- creased markedly due to attack by the ascomycetes, zymes oxidize both guaiacylpropane und syringyl- indicating a preferential attack on the syringylpropane units, although the latter are oxidized more propane units. Calculations, however, showed that readily (Higuchi 1986), reflecting their lower oxida- the losses were not limited to syringylpropane units. tion potentials. It is conceivable that the ascomycetes possess similar but less electropositive peroxidases; Alkaline nitrobenzene oxidation of lignin gives vanillin (V) from G i.e. peroxidases that are more able to oxidize syringyl- units and syringaldehyde (SA) from S units. The yields of these aldehydes have long been used to compare the relative S:G ratios of propane than guaiacylpropane units. angiosperm lignins. Generally, the relative yield of V is considerably lower than that of SA from the same amount of corresponding Acknowledgments monomer units in the lignin, reflecting attachment of many of the G units through the aromatic nuclei to neighboring units. Thus it The authors want to thank Liliane Petrini, Jack D. Rogers and has been suggested that the molar SA/V value, divided by three, Keith A. Seifert for generously providing cultures of several of the more closely reflects the actual S:G ratio (Sarkanen and Hergert ascomycete species used in this study. We are also grateful to Ann- 1971). This adjustment has been re-examined recently (Manders Sofie Hansén and Eva Nilsson for carrying out all the decay exper- 1987)and applied to unique samples (Obst, Landucci, and Manders iments. 1987). including white-rotted hardwoods (Blanchette, Obst. Hedges, and Weliky 1988); the adjustment seems to be valid and References was applied here.

Based on lignin analyses amd S:G determinations, the studied ascomycetes degrade mainly the syringylpropane units of lignin. This may explain the resistance of pine and Alstonia woods (the latter was found to have a S:G ratio of only 0.26). The low S:G ratio may contribute to the resistance of the birch middle lamellae and vessel walls in this study. The attack from the lumen into the secondary walls of birch fibers proceeds readily because the lignin in these tis- sues is relatively rich in syringylpropane units (Saka and Goring 1985). As a result, lignin loss parallels weight losses up to about 20% here. After that point, the remaining lignin, richer in guaiacylpropane units than that already degraded, is more resistant to degradation, and further attack is primarily on the exposed polysaccharides. The failure of D. concentrica to attack the thin pine wood sections shows that the resistance is due to the chemical composition of the wood rather than to any morphological features. The cavity-forming species apparently are different. In contrast to those species causing only erosion of wood cells, the cavity-forming species were able to cause significant weight losses in pine wood. Thus cavity formation by certain soft-rot fungi is related to their ability to attack wood with a high lignin content and a low S:G ratio. The rare occurrence of ascomycetes on gymnosperm woods may be due to the inabil- ity of most to degrade guaiacyl lignin; only the cavity- forming species would be able to utilize such sub- strates to any extent. It will be interesting to characterize the ligninolytic enzymes of the xylariaceous ascomycetes. The key enzymes in white-rot fungi seem to be a family of par- ticularly electropositive peroxidases termed “lig- ninases” or “lignin peroxidases” (reviews: Kirk and 18 T. Nilsson, G. Daniel, T.K. Kirk and J.R. Obst Holzforschung