166 Notes and brief articlesalbo-atrum and Fusarium oxysporum f.sp. ly copersici.Physiological Plant Pathology 5, 135-156.

DADE, H . A. & WRIGHT, J. (1931). Note in Bulletin N o.23 Yearbook 1930 Department of Agriculture, GoldCoast, pp . 249-250.

HOFFMAN, R. M. & TURNER, J. G. (1982). Partialpurification of proteins from pea leaflets that inhibitAscochyta pisi endopolygalacturonase. PhysiologicalPlant Pathology 20,173-187.

HORTON, J. C. & KEEN, N . T. (1966). Sugar repression ofendopolygalacturonase and cellulase synthesis duringpathogenesis by Pyrenochaeta terrestris as a resistancemechanism in onion pink root. Phytopathology 56,908--916.

JONES, T. M., ANDERSON, A. J. & ALBERSHEIM, P . (1972).Host pathogen interactions. IV. Studies on thepolysaccharide-degrading enzymes secreted by Pus-

arium oxysporum f.sp . ly copersici. Phy siological PlantPathology 2, 153-166.

MIEGE, J. (1957). Influence de quelques caracteres destubercules-semences sur les levees et l'amendement designames cultivees. J. Agrtc , trop. Bot. appl. 4, 315.

OGUNDANA, S. K., NAQVI, S. H. Z. & EKUNDAYO, J. A.(1970) . Fungi associated with soft rot of yams(Dioscorea spp.) in storage in Nigeria. Transactions ofthe British Mycological Society 54, 445-451.

OGUNDANA, S. K., NAQVI, S. H. Z. & EKUNDAYO, J. A.(1971 ). Studies on soft rot of yams in storage.Transactionsofthe British Mycological Society 56, 73-80.

OKAFOR, N . (1966) . Microbial rotting of stored yams(Dioscorea spp .) in Nigeria. Experimental Agriculture 2,

179-182.PIENING, L. J . (1962). A check list of fungi recorded in

Ghana. Bulletin of the Ministry of Agriculture, Ghana.No .2, pp. 130.

DEGRADATION OF LIGNIN BY AQUATIC AND AERO-AQUATICHYPHOMYCETES

BY P. J. FISHER, R. A. DAVEY AND J. WEBSTER

Department of Biological Sciences, University of Exeter

Lignin is a generic name for the complex aromaticpolymers that are major components of vascularplant tissue, and in terms of weight it is probablysecond only to cellulose among renewable organicmaterials (Kirk, Higuchi & Chang, 1979). In termsof energy content, it might well be the single mostabundant plant polymer. Higher fungi are primaryutilizers of complex sources of carbon in theterrestrial environment. The white-rot fungi, inparticular, are known to decompose lignin, celluloseand hemicellulose (Setliff & Endy, 1979).

Since aero-aquatic Hyphomycetes are dominantmembers of the mycoflora colonizing leaf litter andwood in static and stagnant water (Fisher, 1977),and aquatic Hyphomycetes colonize similar sub-strates in fresh-water streams (Barlocher & Ken-drick, 1974; Sanders & Anderson, 1979; GarethJones, 1981), aero-aquatic and aquatic Hyphomy-cetes were investigated for ligninolytic activity.Assessment of lignin degradation by fungi is diffi-cult, since it can range from simple demethylationof side chains to complete depolymerization of themolecule and disruption of benzene rings. In ad-dition, pure preparations of lignins are necessarilyhighly modified during the extraction procedure,and interpretations of microbial attack in terms ofintact lignins may not be valid. It is, however,possible to grow fungi on lignin agar preparationsand to test for dephenolization of the lignins in the

vicinity of the colony (Sundman & Nase, 1971).This approach gives direct evidence of ability todegrade lignin at least partially.

The fungi were grown in Petri dishes on amedium of the following composition; basalmedium-sucrose 30 g, NaN03 2'0 g, KCl 0'5 g;KH2S04 1'0 g; MgS04.7H20 0'5 g; FeS040'1 g;yeast extract 0'5 g; agar 15'0 g; distilled water1000 em". An addition of o·5 g 1- 1 of either of thefollowing industrial lignins was made to the basalmedium: Indulin AT from Westvaco Polychemic-als, U.S.A.; Peritan Na from Norcem Ltd,Norway. Brauns (1952) describes commercialIndulin as a mixture of thio and alkali pine ligninscontaining 13% sulphur and 14% methoxyl.Peritan Na is the more modified of the two ligninsand is also known as sodium lignosulphonate. Thelignins were mixed to a paste with a little distilledwater before adding to melted basal medium priorto autoclaving. The Petri dishes containing 15 em"lignin agars were inoculated with 8 mm diam agardisks taken from z-week-old cultures of theappropriate fungi, and incubated from 1 to 2 weeksuntil approximately 15 mm diam colonies had beenformed . Freshly mixed ferricyanide reagent wasprepared by mixing 1 % solutions of FeCI3 andK3Fe(CN)6 in a 1 : 1 ratio. The aerial mycelium wasscraped off each plate, and the plates were floodedwith the ferricyanide reagent and stored in the dark

Trans. Br. mycol. Soc. 80 (1) (1983) Printed in Great Britain

Notes and brief articles

Table 1. Dephenolization of the lignins, Indulin AT and Peritan Na

Aero-aquatic hyphomycetesAegerita candida Pers ,Aegeritoidea oiridis (Bayliss-Elliot) Abdullah ined .Clathrosphaerina zalewskii BeverwijkHelicodendron conglomeratum Glen-BottH. fractum FisherH. fuscum (Berk. & Curt.) LinderH . giganteum Glen-BoltH. hyalinum LinderH . intestinale AbdullahH. luteq-album Glen-BonH . paradoxum PeyronelH . triglitziense (Jaap) LinderH. uiesterdtjkiae BeverwijkHelicoma dennisii EllisHelicoma sp.Helicoon ellipticum (Peck) MorganH. fuscosporum LinderH. pluriseptatum BeverwijkH. richonis (Boudier) LinderH. sessile MorganHelicosporium phragmitis von HohnelSpirosphaera fioriformis BeverwijkS . minuta Henneben

Aquatic hyphomycetesAnguillospora furtioa Descals ined.A. longissima (Sacc. & Syd .) IngoldArticulospora tetracladia IngoldClauariopsis aquatica de WildemanDaetylella aquatica (Ingold) RanzoniDendrospora tenella Descals & WebsterDimorphospora foliicola TubakiGyoerffyella rotula (von Hohnel) MarvanovaHeliscus lugdunensis Sacco& TherryLemonniera aquatica de WildemanL. comUla RanzoniLunulospora curvula IngoldMargaritispora aquatica IngoldScorpiosporium gracile (Ingold) IqbalTetracladium setigerum (Grove) IngoldTricladium giganteum Iqbal

+ = positive result; - = negative result.

Indulin

+

++++

++

++++

+++

+

++

+

Peritan

+

+

+

+

+++

+++

+

+

+

+

for 10 min. Sterile control plates received similartreatment. The excess reagent was decanted and theplates placed over a white sheet of paper. Thecontrols gave a dark blue-green colour whiledephenolization of lignin was shown by a yellow-green colour under and around the colonies.

Many of the aero-aquatic and some of the aquaticHyphomycetes tested caused partial dephenoliza-tion of the lignins in the test medium. It has beenshown, at least during the early stages of leafdecomposition in streams, that fungi are more

dominant members of the microflora than bacteria(Kaushik & Hynes, 1968; Willoughby, 1974).Aquatic Hyphomycetes, in particular, have beenrepeatedly observed on decaying leaf litter andwood in streams (Nilsson, 1964; Ingold, 1966;Sanders & Anderson, 1979), and ecological studieshave suggested a major role of these fungi inprocessing of leaf material in water (Nilsson, 1964;Barlocher & Kendrick, 1974; Suberkropp & Klug,1976). Willoughby & Archer (1973) reported 130fungi from timber in freshwater habitats, including

Trans. Br. mycol. Soc . 80 (1) (1983) Printed in Great Britain

168 Notes and brief articlesthe aquatic Hyphomycetes Heliscus lugdunensis,Anguil/ospora longissima and Clavariopsis aquaticaas some of the most common colonizers. Leightley& Eaton (1977) showed that Heliscus lugdunensisdegraded wood and produced cellulase, xylanaseand mannase under laboratory conditions. Similar-ly, aero-aquatic Hyphomycetes have repeatedlybeen isolated from decaying leaf litter and woodtaken from static water, where they are oftenpresent in great abundance (Linder, 1929; Glen-Bon, 1951; Fisher, 1977). The present study showsthat ligninolytic activity of members of both groupsmay contribute to the decomposition of lignin-conta ining detritus in water .

We would like to thank Dr J. Hedger forsupplying the commerciallignins used in this studyand for his advice.

REFERENCES

BARLOCHER, F . & KENDRICK, B. (1974) . Dynamics of thefungal population on leaves in a stream . Journal ofEcology 62., 761-791.

BRAUNS, F . E. (1952). The chemistry of lignin . In WoodChemistry (ed. L. E. Wise & E. J. Jahn), pp. 409-539.New York: Reinhold.

FISHER, P. J. (1977). New methods of detecting andstudying saprophytic behaviour of aero-aquatic hypho-mycetes from stagnant water . Transac tions of the BritishMycological Society 68, 407-411.

GLEN-BoTT, J. I. (1951). Helicodendron giganteum n.sp.and other aerial sporing Hyphomycetes of submergedleaves. Transactions of the British Mycological Society34, 275-279.

INGOLD, C. T. (1966). The tetraradiate aquatic fungalspore. Mycologia 58, 43-56.

JONES, E. B. G. (1981) . Observations on the ecology of

lignicolous aquatic hyphomycetes. In The FungalCommunity (ed. D . T . Wicklow & G. C. Carroll), pp .731-742. New York : Marcel Dekker.

KAUSHIK, N . K. & HYNES, H. B. N . (1968). Experimentalstudy on the role of autumn shed leaves in aquaticenvironments. Journal of Ecology 56, 229-243.

KIRK, T. K., HIGUCHI, T. & CHANG, H. (1979). LigninBiodegradation .. Microbiology, Chemistry and PotentialApplication , vol. 1. Florida: CRC Press.

LEIGHTLEY, L. E. & EATON, R. A. (1977). Mechanisms ofdecay of timber by aquatic micro-organisms. BritishWood Preserving Association, Annual Convention, pp .1-26.

LINDER, D. H. (1929). A monograph of the helicosporousfungi imperfecti. Annals of the Missouri BotanicalGardens 16, 227-388.

NILSSON, S. (1964).Freshwaterhyphomycetes, taxonomy,morphology and ecology. Symbolae Botanicae Upsali-enses 18, 1-130.

SANDERS, P. F. & ANDERSON, J. M. (1979). Coloniza-tion of wood blocks by aquatic hyphomycetes.Transactions of the British Mycological Society 73,103-107.

SETLIFF, E. & ENDY, W. (1979). Screening white-rot fungifor their capacity to delignify wood. In LigninBiodegradation, vol. 1 (ed. T. K. Kirk, T. Higuchi &C. Chang), pp . 135-151. Florida: CRC Press.

SUBERKROPP, K. & KLUG, M. J. (1976). Fungi andbacteria associated with leaves during process ing in awoodland stream. Ecology 57, 707-719.

SUNDMAN, V. & NASE, L. (1971). A simple plate test forthe direct visualisation of biological lignin degradation.Paper and Timber 53, 67-71.

WILLOUGHBY, L. G. (1974). Decomposition of litter infresh water. In B iology of Plant Lifter Decomposition,vol. 2 (ed. C. H. Dickinson & G . J. F. Pugh), pp .659-681. London: Academic Press .

WILLOUGHBY, L. G. & ARCHER, J. F . (1973). The fungalspora of a freshwater stream and its colonization patternon wood. Freshwater Biology 3, 219-239.

PEPTIDASE ACTIVITIES OF FUNGI AS ASSAYED BY NOVELFLUOROGENIC SUBSTRATES

BY N. BLAKE, A. BARKER*, M. TROTTERt AND G. W. GOODAY*

*Department of Chemistry and tMedical School, University of Birmingham,and *Department of Microbiology, University of Aberdeen

The use of model peptide substrates is enabling thecharacterization of hitherto unrecognized peptidaseactivit ies in fungi (Wolf, 1982). Some of thesesubstrates may be specific enough to be useful fordeveloping techniques for diagnosis of fungalinfections such as those caused by dermatophytes.Here we present results of a survey of activities of

culture filtrates of a range of fungi on ninefluorogenic amides.

The fluorophore in each case was the dimethylester of the amine, 5-amidoisophthalic acid (AlE).The excitation and emission wave-lengths of thiscompound are 350 and 440 om respectively.Syntheses of the amides will be described elsewhere.

Trans . Br. mycol. Soc. 80 (1) (1983) Printed in Great Britain