APPLIED MICROBIOLOGYVol. 12, No. 1, p. 57-62 January, 1964Copyright ( 1964 by the American Society for Microbiology

Printed in U.S.A.

Degradation of Wood Preservatives by Fungi

CATHERINE G. DUNCAN AND FLORA J. DEVERALL

Forest Products Laboratory,' Forest Service, U.S. Department of Agriculture, Madison, Wisconsin

Received for publication 19 September 1963

ABSTRACT

DUNCAN, CATHERINE G. (Forest Products Laboratory, ForestService, U.S. Department of Agriculture, Madison, Wis.), ANDFLORA J. DEVERALL. Degradation of wood preservatives byfungi. Appl. Microbiol. 12:57-62. 1964.-Wood-inhabitingfungi, not necessarily responsible for major decay, are shown tobe capable of degrading a toxic compound into a less potentform, thus rendering it less effective in protecting wood fromdecay by less-tolerant basidiomycetous wood-destroyers. Sweet-gum or pine sapwood blocks treated with preservatives (am-moniacal copper arsenate, fluor-chrome-arsenate-dinitrophenol, acreosote or pentachlorophenol) were exposed progressively to twodifferent wood-inhabiting fungi with sterilization between thefirst and second exposure. The fungus in the first exposurewas usually an Ascomycete or a Fungi Imperfecti-Chaetomiumglobosum, Phoma, Orbicula, Graphium, Pestalozzia, or Trichodermaspecies, isolated from wood below the ground. In one experi-ment, the fungus in the first exposure was a basidiomycete,Lenzites trabea or Polyporus versicolor. The second fungus, a prom-inent Basidiomycete-Coniophora puteana, Lentinus lepideus, orLenzites trabea-was the bioassay fungus, since its purpose was toshow whether the first fungus had degraded the preservative.Generally, the treated block, except where exposed to anotherfungus, remained virtually untouched by the bioassay fungus.Clearly, therefore, the first fungus had rendered the preservativeineffective but without appreciably decaying the wood itselfChemical analyses of treated blocks indicated that in the firstexposure the fungi had substantially depleted sodium arsenateand pentachlorophenol.

Microbial associations, in addition to being numerousand varied, are frequently so complex and sensitive toslight changes in environment that it is difficult to discoverthe primary phenomena that contribute to the end result.Certain fungi, with changes in environment, may be antag-onistic to each other, stimulate the growth and parasiticcapabilities of one another, or live together seeminglyunaffected by each other's presence except for spatial andnutritional limitations (Osteraas, 1956). Since a changein the biotic environment may modify the relationshipbetween the organisms, efforts to separate a single affinityand disregard the influence of others in a mixed populationmay not always give a true picture of natural associations.However, individual relationships must be established toanalyze a complicated association among microorganisms.An example of an intricate group of microorganisms

living in close association may be found in wood that has

1 Maintained at Madison, Wis., in cooperation with the Univer-sity of Wisconsin.

been exposed to the atmosphere or the soil for a long periodof time. Prominent members of such an association arethose fungi (Basidiomycetes, Ascomycetes, and FungiImperfecti) capable of utilizing the sugars and hemicellu-loses of the wood ray cells or the cellulose and lignin ofthe tracheids, fibers, and vessels.

Little is known of the effect of a preservative in woodupon such a fungus association. In the addition of a pre-servative, it is expected that those fungi mainly respons-ible for the decay of wood, namely, the Basidiomycetes,will be retarded or killed. The preservative tolerance ofmany economically important wood-destroyers has beenstudied; however, little is known of the factors involvedin a specific tolerance. The question of how commonly thetolerance of a preservative by a decay fungus may beattributed to degradation of the preservative, throughthe metabolism of the wood-attacking organism or anassociate, is of particular significance. Are wood-inhabitingorganisms prevalently capable of metabolizing a toxicchemical into a less potent derivative? The experimentsreported here indicate that degradation of preservativesmay be brought about by several fungi. These includespecies that are not primarily responsible for decay but,because of this effect, may render treated items susceptibleto decay by typical wood-destroying fungi.

Literature. Although numerous accounts have appearedin literature pertaining to the inactivation of toxicants bymicroorganisms in carrying on their chemical activities,there is little reference to those directly concerned in wooddecay (Foster, 1949; Horsfall, 1956; Gottlieb, 1957). It isof interest to cite incidents that indicate inactivation ofchemical compounds used in the preservation of woodproducts such as arsenic, copper, mercury, and phenolcompounds.

Arsenic, for example, is an ingredient in several of thewater-soluble salt preservatives. A classic example ofbiological detoxification is that in which the fungi asso-ciated with wallpaper containing arsenical pigments gaverise to a volatile toxic gas (Gozio, 1892). The gas wasidentified as trimethylarsine (Challenger, 1945). It is nowknown that a considerable number of fungi can breakdown arsenic compounds, while many others can toleratethem but cannot destroy them (Thom and Raper, 1932).

Furthermore, Madhosingh (1961a) noted the invariableassociation of Coprinus micaceus and Fusarium oxysporiumin white birch and poplar fence posts treated with a water-soluble preservative mixture containing arsenic. Coprinuswas found to be much less tolerant of the preservative

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than Fusarium. It was suggested, therefore, that Fusariummay have reduced the toxicity of the wood preservative,thereby facilitating the entry and spread of Coprinus.Copper tolerance is often associated with tolerance to

hydrogen or other cations. For instance, the copper toler-ance of the acid-forming brown-rot fungi is generallygreater than that of white-rot fungi, and can be increasedby a lowering of the pH of the substrate (Young, 1961).Rabanus (1939) suggested that one of the acids (oxalic)precipitated the copper into the insoluble, oxalate form,thus rendering the copper fungicidally inert. The associa-tion of copper tolerance to hydrogen ions, however, sug-gested to Horsfall (1956) that the mechanism of coppertolerance is a general exclusion of metallic cations fromthe cell.

Stored pulp impregnated with phenyl mercury acetatecan be deteriorated by Stereum sanguinolentum and Penio-phora pitya. This is made possible by preceding infection ofPencillium roqueforti, which reduces the concentrationby absorbing relatively large amounts of the biologicallyactive mercury. The phenyl mercury acetate therebyremoved from the solution is rendered ineffective as aninhibitor (Russell, 1955; Kiessling, 1961).

Phenols may be rapidly reduced by microorganisms.Dinitrophenol was shown by Madhosingh (1961b) to bereduced by Fusarium oxysporium to less toxic components.Stemphylium sarcinae forme, resistant to many quinones,possesses a laccase enzyme system which can oxidasephenols to quinones and polymerize the quinones to a non-toxic black pigment (Rich and Horsfall, 1954).

Recently, Lyr (1963) showed that wood-destroyingBasidiomycetes that secrete laccase into the culture me-dium are able to overcome the toxic effects of penta-chlorophenol.

Creosote, one of the oldest wood preservatives, is amost potent fungicide of complex composition, containingphenols, aromatic hydrocarbons, nitrogen bases, and othertoxicants. In spite of this, Hormodendrum resinae has beenfound to be invariably associated with creosote-impreg-nated wood (Christensen et al., 1942). 1larsden (1951,1954) suggested that, over long periods of time, thisfungus may actually degrade the preservative value sincecreosote may serve as its sole source of nitrogen and carbon.Pseudomonas cresotensis, a marine bacterium frequentingcreosoted pilings, likewise has a high resistance to creosote(O'Neill, Drisko, and Hochman, 1961).

It has also been shown that some of the natural decay-retardant substances in heartwood, such as pinosylvinmonomethyl ether and thujaplicin, as well as chloro-phenols, may be detoxified through oxidation by suchenzymes as laccase, tyrosinase, and peroxidase, producedby many fungi (Lyr, 1:962).

MIATERIALS AND MIETHODSFive experiments were conducted during a 3-year period,

each varying to some extent in the methods used. Details

of the methods used are shown in Table 1, including a de-scription of the test specimens, the preservative treat-ments, and the culture techniques with the fungi involved.

General plan of all the experiments. Each experiment (Athrough E) included two separate exposures to fungi insequence, designated exposures 1 and 2. In exposure 1, theblocks were subjected to pure cultures of Fungi Imperfectior Ascomycete fungi, known or reported to cause soft rotof wood. In addition, exposure 1 of experiment A includeda white- and a brown-rot Basidiomycete fungus. In ex-posure 2, the blocks were steamed to kill the originalinfection from exposure 1, and then subjected to decay bya Basidiomycete brown-rot fungus.

Test specimens. The test blocks, although of variablesizes, were of sweetgum sapwood in four bioassays (experi-ments A, B, D, E) and pine sapwood in one (experimentC). Sweetgum was emphasized because, as an angio-spermous wood, it is more readily attacked by soft-rotfungi than is a gymnospermous wood.

Preservative treatments. The test blocks, in accordancewith the standard method for testing preservatives (Ameri-can Society for Testing and Materials, 1961), were treatedto refusal with varying concentrations of a preservativesolution to give a range of retentions (pounds per cubicfoot) in the wood. The retentions were intended to fallabove and below the approximate threshold previouslydetermined for the respective preservative.Four standard and frequently used preservative solu-

tions were tested. Two were salt mixtures, one containingcopper and arsenic in an ammoniacal solution, and theother fluoride, chromate, dinitrophenol, and arsenic inwater. The other two preservatives were creosote and a5 % pentachlorophenol in a low-boiling petroleum. Bothwere diluted in toluene for the treatments.

After treatment, all blocks were conditioned at 30%relative humidity and 27 C for 1 month. A similar condi-tioning period also followed exposures 1 and 2.

Substrate for pure culture testing. The substrate for thefungus was either a mineral-vitamin-agar or soil. The agarsubstrate was used only in exposure 1, experiment A.Where soil was the substrate, the technique in its use wasdifferent with the Basidiomycete fungi (exposure 2) thanwith the Fungi Imperfecti and Ascomycetes (exposure 1).With the Basidiomycete wood-decay fungi, the techniquewas essentially the same as described by the AmericanSociety for Testing and Alaterials (1961). Certain modi-fications in this standard, however, were necessary withthe soft-rot fungi, primarily because of their need for ahigher wood moisture and supplementary minerals forrapid decay.

Modifications of the procedure, to provide the greatermoisture and mineral requirements, were: (i) use of soilwith a greater water-holding capacity, and a soil moisturecontent up to 110 C( of this capacity; (ii) addition of nitro-gen, phosphorus, and magnesium to the soil water (asg per liter: N-H4NO3, 6; K2HP'04, 4; KH2PO4, 5; M\IgSO4.7

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DEGRADATION OF WOOD PRESERVATIVES BY FUNGI

TABLE 1. Materials and procedures used in the various experiments to determine the capacity of fungi to decompose preservatives

No. of Concn Reten-Expt Expos- repli- of tions of Pure culture Incu-no. ure Species Size* cates Toxic components treating preserv testing Fungi used bation

*[Sno.pecies Size3 per solution ative in substrate u periondvariable ~~~~~~~woodt

sn. % lb/fl3 weeks

A 1 Sweetgum } X 1 4 Copper as CU2(OH)2, 0.8 0.36 Mineral-vitamin- Lenzites trabea (617) 6sapwood X 1 arsenic as As203, NH3, 1.6 0.73 filter paper Polyporus versicolor (697)

and CH3COOH (1.8: 2.4 1.1 agar$ Chaetomiumglobosum (S70B)1.3:2.8:0.05%) in an 3.2 1.5 Graphium sp. (R47F)ammoniacal solution 4.3 2.0 Orbicula sp. (P36)

Phoma sp. (R2)

A 2 Sweetgum ½ X ½ 4 Soil-block tech- Coniophora puteana (5127) 12sapwood X 1 nique with

moisture equalto 150% ofwater holdingcapacity

B 1 Sweetgum 38 X 3i 4 Fluoride as NaF, ar- 0.5 0.22 Soil-burial Phoma sp. (R2) 6sapwood X 2 senic as Na2HAsO4, 1.0 .42 Graphium sp. (R47F)

chromate as Na2CrO4, 1.5 .68and dinitrophenol 2.0 .92(1:1:1.5:0.5 ratio) in 2.5 1.12water

B 2 Sweetgum Ys X 3/ 8 Soil-block Lenzites trabea (617) 12sapwood X 1 (ASTM D1413)

C 1 Pine sap- 38 X Y 4 Fluoride as NaF, ar- 0.5 0.17 Soil-burial Phoma sp. (R2) 6wood X 21/ senic as Na2HAsO4, 1.0 .35 Graphium sp. (R47F)

chromate as Na2CrO4, 1.5 .56and dinitrophenol 2.0 .77(1:1:1.5:0.5 ratio) 2.5 .93in water

C 2 Pine sap- 8 X /3 8 Soil-block Lenzites trabea (617) 12wood x 1jI (ASTM D1413)

D 1 Sweetgum Y X 34 6 Coal tar creosote di- 2 0.6sapwood X 2 luted with toluene 4 1.4 Soil-burial Pestalozzia (P40) 12

6 1.88 2.312 3.616 4.8

D 2 Sweetgum Y8 X 34 12 20 6.0 Soil-block Lentinus lepideus (534) 12sapwood X 1 26 7.8 (ASTM D1413)

E 1 Sweetgum 3 X 34 6 Pentachlorophenol in 0.1 0.03 Soil-burial Trichoderma sp. (P42) 12sapwood X 2 light solvent diluted 0.2 .07

with toluene 0.3 .090.4 .120.6 .180.8 .24

E 2 Sweetgum Y8 X 3i 12 1.0 .30 Soil-block Lenzites trabea (617) 12sapwood X 1 1.3 .39 (ASTM D1413)

* Length is measured along the grain.t Figures shown are initial retentions. At 4 weeks after blocks were treated, preparatory to exposing them to the fungi, they were

leached. Treated blocks, separated by retention groups, were covered with distilled water (approximately 25 ml of water per block)and impregnated to refusal. Water was changed hourly during an 8-hr period on first day and every 2 hr in a similar period on secondday. Blocks were removed from leach after 48 hr, surface dried, placed in petri dishes, and steamed for 20 min (100 C). They werethen ready for the culture chamber.

I Mineral-vitamin-agar ingredients (in g per liter): NH4NO3, 6.0; K2HPO4, 4.0; KH2PO4, 5.0; MgSO4-7H20, 4.0. Microquantitiesof the vitamins-thiamine, biotin, and pyridoxine-and compounds containing the elements Ca, Mn, Fe, Zn, Ba, Co, and Cu.

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H20, 4); (iii) elimination of the strip of wood (feederblock) between the test block and the soil; (iv) saturationof the test block with water; and (v) aseptic burial of thetest block vertically in the soil that had been thoroughlypenetrated by a 2- to 3-week growth of fungus.To reveal any change in weight of the blocks not due

to decay, replicate blocks were placed in uninoculated soilas controls.

Fungi. The fungi used in exposure 1 were: the Basidio-mycetes Lenzites trabea Pers. ex Fr. (617), and Polyporusversicolor L. ex Fr. (697); the Ascomycetes Chaetomiumglobosum Kunze ex Fr. (S70B); and the Fungi ImperfectiGraphium sp. (R47F), Orbicula sp. (P36), Phomna sp. (R2),Pestalozzia sp. (P40), and Trichoderma sp. (P42). In ex-posure 2, to bioassay the blocks for evidence of preserva-tive degradation in exposure 1, the BasidiomycetesLentinus lepideus Fr. (534), Coniophora puteana (Fr.)Karst. (5127), and L. tr-abea were used.

Except for C. puteana all of the Basidiomycetes werestandard test fungi of exceptional tolerance to one or morepreservatives, namely, L. lepideus to creosote, Lenzitestrabea and Polyporus versicolor to pentachlorophenol, andLentinus trabea to arsenic. C. puteana is used in Canadaand Europe as a test fungus because of its tolerance topreservatives. It was used here primarily because it hasfrequently been isolated from exceptionally wet woodwhich also contained soft rot.

All the soft-rot fungi were common isolates from thebelow-ground portion of treated and untreated wood.They were also isolated from the very wet parts of coolingtowers. In agar plate toxicity tests, they have shown moretolerance to arsenic, chromate, fluoride, and zinc thanhave Basidiomycetes (Duncan, 1960a, b).

RESULTS AND DISCUSSION

Experiments A, B, and C. The results of these experi-ments are shown graphically in Fig. 1 and 2. The preserva-tive used was one of two salt mixtures, differing in com-position but with arsenic as a common toxic component.

In experiment A (Fig. 1), C. puteana caused no decayin blocks with as little as 0.36 lb/ft3 of preservative saltmixture unless they had previously been subjected to oneof four soft-rot fungi, isolates of Chaetomium, Graphium,Orbicula, or Phoma; a brown-rot fungus, Lenzites trabea;or a white-rot fungus, Polyporus versicolor. When blockswere previously exposed to one of these fungi, C. puteananot only attacked the blocks treated with 0.36 lb/ft3 butthose having a preservative concentration 5.5 timesgreater. All six fungi caused a change in the preservative,as evidenced by the degree of attack in exposure 2.

In experiment B (Fig. 2), it was further indicated thattwo of the soft-rot fungi, a Phoma and a Graphium species,used in experiment A, could also degrade another preserva-tive salt mixture containing arsenic. The change in pre-servative enabled L. trabea in exposure 2 to attack woodtreated with a considerably higher concentration than

when the soft rotters were absent from the soil inexposure 1.

Experiment C was similar to experiment B in all respectsexcept that the preservative salt mixture was tested inpine sapwood instead of sweetgum sapwood. This variation

k 70

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N 60H

.0 50

0" 301t

'R 40

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S/OI.,

FUNGUS INEXPOSURE I:LENZIrESrRABEA

EXPERIMENT A

LEGEND:0 BLOCKS PREVIOUSLY EXPOSED rO A FUNGUS IN EXPOSURE -.

E BLOCKS NOr PREVIOUSLY EXPOSED rO A FUNGUS IN EXPOSURE IAND SERVING AS CONrROLS.

FUNGUS INEXPOSURE r:POLYPORUS

_ VEfRSICOLOR

FUNGUS INEXPOSURE I:CHAAErOM/UMGLONOSUM

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FUNGUS IN FUNGUS IN FUNGUS IN -

EXPOSURE t: EXPOSURE 1: EXPOSURE r:GRAPHIUM SP ORB1CULA SP PHONA SP.

a/ 04 0.9O./ 04 ON a/ 0.4 oN al 04 o0 al a4 09 a/ 04 ON

02 06 0.2 06 a2 06 02 06 02 O6 02 06RErENTrION OF PRESERVArIVE (POUNDS PER CUaIC FOOr)

FIG. 1. Weight losses in copper arsenate-treated blocks of sweetgumsapwood resulting from decay caused by Coniophora puteana duringexperiment A, exposure 2. Some of the blocks were previously exposedto other fungi (exposure 1). Where the weight losses for such blocksare greater than those for corresponding blocks not subjected to afungus in exposure 1, degradation of the preservative is indicated.

_ AND SER VING AS CONTRsOL s

1-k60 _U1 EXPERIMENr

(SWEETGUM SAPWOOD)

O5 50 FUNGUS IN EXPOSURE r: FUNGUS IN EXPOSURE I:

PHOMA SP GRAPH/UM SP

4.12 a22 a42 069 092 1.12 0.DI 035 0.56 077 a93 017 035 056 077 0.93RErENrION OF PRESERVArIVE (POUNDS PER CURIC FOOr)

FIG. 2. Weight losses in fluor-chrome-arsenate-dinitrophenol-treated blocks of sweetgum and pine sapwood resulting from decaycaused by Lenzites trabea during experiments B and C, exposure 2.Some of the blocks were previously exposed to other fungi (exposure 1).Where the weight losses for such blocks are greater than those for corre-sponding blocks not subjected to a fungus in exposure 1, degradationof the preservative is indicated.

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DEGRADATION OF WOOD PRESERVATIVES BY FUNGI

was injected into the tests, since preservative effectivenessis sometimes influenced by the test wood (Duncan, 1958).Soft-rot fungi under laboratory conditions do not attack a

gymnospermous wood as readily as an angiospermouswood, even though the nonresistant sapwood of both isused. This difference in attack is indicated by the weightlosses in Fig. 2. However, this inability to attack the pinewood did not prevent preservation degradation.

Further work needs to be done before the mechanisminvolved in the degradation of the salt mixtures can beadequately explained. However, a few analyses of treatedblocks offered some clues. For example, the analysis of a

group of blocks from experiments B and C showed thatapproximately one-third more arsenic was present in theblocks that had been kept over sterile, uninoculated soilthan in blocks that had been subjected to fungi, whetherthe preservative was in sweetgum or pine (Table 2). Thisindicated, therefore, that the Phoma and Graphium specieswere both capable of degrading at least the arsenate inthe fluor-chrome-arsenate-dinitrophenol mixture. Withoutan analysis of the copper-arsenate blocks (experiment A),it can only be assumed that the Phoma and Graphiumspecies likewise degraded the arsenic in this mixture.The decrease in effectiveness of copper arsenate by

L. trabea, known to be arsenic tolerant, was noteworthy(Kaufert and Schmitz, 1937). This was demonstrated bythe rather large amount of attack in the treated blocks byC. puteana (Fig. 1). However, this degradation may be theresult of a change in the copper rather than arsenate, sincein an analysis of the fluor-chrome-arsenate blocks (Table 2)no depletion of arsenic was evident after exposure toL. trabea. L. trabea was shown by Baechler and Roth(1956) to be more tolerant of copper sulfate than sodiumarsenate in soil blocks tests.

Experiment D. The exposure of creosote-treated blocksto a Pestalozzia species in experiment D apparently causedlittle change in the preservative effectiveness againstLentinus lepideus, since weight losses in similarly treated

blocks were approximately the same whether Pestalozziahad been present or absent from the soil (Fig. 3).

It should be mentioned that Hormodendrum Xresinae,commonly present in creosoted wood and known to havesome ability to utilize creosote, was not used in this experi-ment. Preliminary tests indicated it was antagonistic toL. lepideus under the soil block test conditions. Blockscontaining 2 lb/ft3 of creosote were readily decayed byL. lepideus alone, or if L. lepideus was permitted to becomeestablished, by a 2-month incubation before inoculationwith Hormodendrum. However, Hormodendrum preventeddecay by L. lepideus if the two different inoculations were

made at the same time, or if a 2-month incubation periodwith Hormodendrum preceded inoculation with L. lepideus.L. lepideus did not even decay Hormodendrum-infectedblocks which had been sterilized. It is unlikely thatHormodendrum causes much competition in nature, sinceit has not been found to be a common saprophyte in thesoil and is unable to survive well there in competition withother organisms (Christensen et al., 1942). Whether antag-onism may occur commonly between Hormodendrum andLentinus in practical situations, it was not believed desir-able to inject the variable of antagonism into the experi-ments.Experiment E. In this experiment, pentachlorophenol

was changed to such an extent (exposure 1) that weightlosses by L. trabea (exposure 2) were greater in treatedblocks exposed to Trichoderma than in those not exposed.Moreover, approximately twice the amount of penta-chlorophenol was required to prevent decay (Fig. 3).Evidence that the decrease in effectiveness was due todegradation of the pentachlorophenol is shown in Table 3.In the blocks analyzed, approximately 43 % of the penta-chlorophenol in the wood at the time of treatment was no

longer present after the exposure to Trichoderma.Summary. In general, the experiments indicated that

probably many wood-inhabiting fungi may change a

preservative, degrading it so as to render it less effective

TABLE 2. Analysis of blocks treated with afluor-chrome-arsenate-dinitrophenol preservative (experiments B and C)

Na2AsO4 by Microscopical observations for detection of decaychemicalanalysis of

Conditions to which treated treated wood Sweetgum Pineblocks were subjected species

Sweet- Pine Hyphae Cell-wall deterioration Hyphae Cell-wall deteriorationgum

Treatment and leaching 0.22 0.15 None None None NoneExposure 1 with no fungus 0.22 0.15 None None None NoneExposure 1 with no fungus and ex- 0.23 0.16 Very few None V"ery few None

posure 2 with Lenzites trabeaExposure 1 with Graphium sp. 0.07 0.05 Many hyphae Only to maximal depth of Many hyphae None

five cells on surfaceExposure 1 with Graphium sp. and 0.08 0.05 Many hyphae Soft rot five cells deep; Many hyphae Thinning of cell walls

exposure 2 with Lenzites trabea of two thinning of cell walls plus of two plus bore holestypes bore holes types

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FUNGUS IN EXPOSURE I:rRICHODERMA SP.

FUNGUS IN EXPOSURE 2: -

LENZITES rRABEA

23 35.6 4. 0o3 006 0,09 0.2 a01 024 0.30 a39RErENTION OF PRESERVArIVE (POUNDS PER CUBIC FOOT)

FIG. 3. Weight losses in creosote or pentachlorophenol-treatedblocks of sweetgum sapwood resuilting from decay cautsed by Lentinuslepideus or Lenzites trabea during experiments D and E, exposure 2.Some of the blocks were previously exposured to other fungi (exposure1). Where the weight losses for such blocks are greater than those forcorresponding blocks not subjected to afunguts in exposure 1, degrada-tion of the preservative is indicated.

TABLE 3. Analysis of blocks treated with a pentachlorophenolsolution (experiment E)

chloro- Microscopical analysis for detectionchlro of decay in sweetgumConditions to which treated phenol

blocks were subjected bychemchemi-cal Cell-wall

analysis Hyphae deterioration

Treatment and leaching 0.40 None NoneExposure 1 with no fungus 0.40 None NoneExposure 1 with no fungus 0.41 V'ery few Noneand exposure 2 withLenzites trabea

Exposure 1 with Tricho- 0.23 Many hyphae Nonederma sp.

Exposure 1 with Tricho- 0.19 Many hyphae Thinning ofderma sp. and exposure 2 of two types cell wallswith Lenzites trabea plus bore

holes

in protecting the wood from decay by other less-tolerantfungi. By chemical analysis, loss of arsenic and pentachlor-ophenol was shown to occur as a result of fungus action.

LITERATURE CITED

AMERICAN SOCIETY FOR TESTING AND MATERIALS. 1961. Standard

method of testing wood preservatives by laboratory soil block

cultures. D1413-61, American Society for Testing and Mate-

rials.

BAECHLER, R. H., AND H. G. ROTH. 1956. Laboratory leaching and

decay tests on pine and oak blocks treated with several pre-servative salts. Proc. Am. Wood Preservers' Assoc. 52:24-34.

CHALLENGER, F. 1945. Biological methylation. Chem. Rev. 36:315-361.

CHRISTENSEN, C. M., F. H. KAUFERT, H. SCHMITZ, AND J. L.ALLISON. 1942. Hormodendrum resinae (Lindau), an inhabitantof wood inpregnated with creosote and coal tar. Am. J. Botany29:552-558.

DUNCAN, C. G. 1958. Evaluating wood preservatives by soil blocktests. 10. Effect of species of wood on preservative thresholdvalues. Proc. Am. Wood Preservers' Assoc. 54:172-177.

DUNCAN, C. G. 1960a. Wood-attacking capacities and physiologyof soft-rot fungi. U.S. Forest Prod. Lab. Rept. 2173.

DUNCAN, C. G. 1960b. Soft rot in wood and toxicity studies oncausal fungi. Proc. Am. Wood Preservers' Assoc. 56:27-35.

FOSTER, J. W. 1949. Chemical activities of fungi. Academic Press,Inc., New York.

GOTTLIEB, D. 1957. The effect of metabolites on antimicrobialagent. Phytopathology 47:59-67.

Gozio, B. 1892. Azione di alcune muffe sui composti fissid'arsenico. Rev. d'Igiene e Sanita Publ. III 8/9:201-230/261-273.

HORSFALL, J. G. 1956. Principles of fungicidal action. ChronicaBotanica Co., Waltham, Mass.

KAUFERT, F., AND H. SCHMITZ. 1937. Studies in wood decay. VI.The effect of arsenic, zinc, and copper on the rate of decay ofwood by certain wood-destroying fungi. Phytopathology 27:780-788.

KIESSLING, H. 1961. Some factors concerning the detoxificationof organic mercurial fungicides. Svensk Papperstid. 64:689-693.

LYR, H. 1962. Detoxification of heartwood toxins and chloro-phenols by higher fungi. Nature 195:289-290.

LYR, H. 1963. Enzymatische Detoxifikation chlorierter Phenole.Phytopathol. Z. 47:73-83.

MADHOSINGH, C. 1961a. Tolerance of some fungi to a water-solublepreservative and its components. Forest Prod. J. 11:20-22.

MADHOSINGH, C. 1961b. The metabolic detoxification of 2,4-dini-trophenol by Fusarium oxysporium. Canad. J. Microbiol.7 :553-567.

MARSDEN, D. H. 1951. Studies of Hormodendrum resinae (Lindau),a common inhabitant of creosoted and coal-tar treated wood.Phytopathology 41:658-659.

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62

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