Antonie van Leeuwenhoek 78: 129–139, 2000.

© 2000 Kluwer Academic Publishers. Printed in the Netherlands.

129

Screening of basidiomycetes for antimicrobial activities

Inmaculada Suay, Francisco Arenal, Francisco J. Asensio, Angela Basilio,

M. Angeles Cabello, M. Teresa Dı́ez, Juan B. Garcı́a, Antonio González del Val,
Julián Gorrochategui, Pilar Hernández, Fernando Peláez & M. Francisca Vicente∗
Centro de Investigación Básica, Merck Sharp & Dohme de España S.A., Josefa Valcárcel 38, E-28027 Madrid,
Spain (∗ Author for correspondence; E-mail: [email protected])

Received 14 December 1998; accepted 14 June 2000

Key words: antibacterial and antifungal activities, basidiomycetes

Abstract
As a part of a screening programme developed to evaluate the antimicrobial activity of basidiomycetes, 317 isol-
ates representing 204 species collected in Spain were screened against a range of human clinical pathogens and
laboratory controls. Extracts from 45% of the isolates, representing 109 species, showed antimicrobial activity. An-
tibacterial activity was more pronounced than antifungal activity. The proportion of extracts from basidiomycetes
showing antimicrobial activity was similar to or above that obtained for representative orders of Ascomycetes, such
as Pezizales and Xylariales, but lower than that produced by members of the orders Diaporthales, Eurotiales, Hy-
pocreales, Leotiales and Sordariales. Suprageneric taxa (orders and families) did not show pronounced differences
in their antimicrobial activities though such differences were observed at the genus level, suggesting that the ability
to produce these bioactive compounds is not homogenously distributed amongst the basidiomycetes. Isolates from
some species showed large differences in their ability to produce metabolites with antimicrobial activity, possibly
reflecting genetic differences at the infraspecific level.

Introduction have been reinvestigated, mainly due to the increasing

difficulty and cost of isolating novel bioactive com-
Basidiomycetes produce a large number of secondary pounds from members of the order Actinomycetales.
metabolites (Anke 1989) which show antibacterial, an- Advances in fermentation technology, separation tech-
tifungal, antiviral, cytotoxic and hallucinogenic activ- niques, as well as in the methodology used in struc-
ity or which can be a source of plant growth regulators tural determinations, have made other microorganisms
or flavours (Heim 1963; Korzybski et al. 1967; Bene- attractive as possible sources of new bioactive second-
dict & Brady 1972; Anke 1985; Hautzel & Anke 1990; ary metabolites. Difficulties such as the slow growth
Janssens et al. 1992; Breheret et al. 1997; Marumoto rate of basidiomycetes and the low yields of products
et al. 1997). derived from these are now far outweighed by the
Florey et al. (1949) detected diverse antibiotic opportunity of finding new antibiotics with differ-
activity in analyses of either fruiting bodies or my- ent structural types, as well as compounds with new
celial cultures of over 2,000 fungal species. One ex- modes of action (Brizuela et al. 1998). The fact that
ample, pleuromutilin, was used in the treatment of basidiomycetes have been insufficiently investigated,
mycoplasma-related diseases in cattle (Kavanagh et together with the broad range of structural types of
al. 1952). However, interest in the metabolites pro- antibiotics which are produced by these organisms,
duced by basidiomycetes declined as streptomycetes suggests that they may be a source of new and useful
were considered to be more prolific and an easily ma- bioactive compounds (Anke 1989).
nipulated source of antibiotics (Anke 1989). It is only The antibacterial and antifungal activities of a
recently that basidiomycetes and other higher fungi broad range of basidiomycetes are considered in this
130

report within a taxonomic context. Comparative data Merck Culture Collection. All of the bacteria, ex-
from some groups of ascomycetes are also presented. cept B. subtilis strain MB964, were resistant to
known antimicrobial agents; S. aureus MB5393 was
methicillin-resistant, E. faecium MB5571 was res-
Materials and methods istant to vancomycin and ß-lactam antibiotics, M.
smegmatis MB2233 was resistant to aminoglycosides,
Isolation of fungi macrolides and penicillin and the two Gram negat-
ive bacterial strains were resistant to cephalosporins
The test strains were isolated from fruiting bodies col- (up to the third generation), macrolides and penicillin
lected at different locations at Spain between 1995 and in the case of P. aeruginosa MB979 also to imi-
and 1997. Axenic cultures were obtained from the penem (Zak 1980; Davis & Stone, 1986; Sunderam
inner fragments of the living tissues of the fruit- et al. 1986; Al-Obeid et al. 1990). Aspergillus fu-
bodies using a isolation medium supplemented with migatus MF5668 and Candida albicans MY1055 are
benomyl and dichloran to inhibit the growth of non- major causes of systematic fungal infections, partic-
basidiomycetous fungi as described by Worrall (1991). ularly in immunosuppressed patients, including those
All of the strains were maintained as potato dex- with acquired immunodeficiency syndrome (Polak &
trose agar slants (Difco) and were preserved as frozen Hartman 1991; Vartivarian et al. 1993).
agar plugs in glycerol (10%, v/v) at −80 ◦ C in the The B. subtilis MB964 assay plates were prepared
Centro de Investigación Básica de España (CIBE) by adding 1 ml (1.5 × 108 cfu ml−1 ) of a spore sus-
Culture Collection. The species were grouped into pension (Difco) directly to 1 liter of cooled nutrient
suprageneric taxa following the criteria established by agar yeast extract medium (NAYE, nutrient agar 23 g
Hawksworth et al. (1995). l−1 , yeast extract 2 g l−1 ). Inocula for the remaining
bacterial strains were prepared by adding the contents
Fungal fermentations of thawed cryovials to the relevant medium and incub-
ating overnight. The Enterococcus faecium strain was
Inocula for the fermentation studies were prepared by grown in static cultures of brain heart infusion broth
aseptically transferring the upper part of the growth (BHI, 37 g l−1 ) at 37 ◦ C, Mycobacterium smegmatis
on agar slants of the test strains into unbaffled 250 ml MB2233 in nutrient broth supplemented with Tween
Erlenmeyer flasks fitted with cellulose plugs and con- 80 (NBT, nutrient broth 8 g l−1 and 10 ml l−1 of
taining 50 ml of the seed medium described by Peláez 20% solution Tween 80) at 37 ◦ C, Pseudomonas aer-
et al. (1998). The inoculated flasks were shaken at 220 uginosa MB979 and Serratia marcescens MB252 in
revolutions per minute (rpm) at 25 ◦ C for 3 to 4 days. nutrient broth yeast extract (NBYE, nutrient broth 8 g
Two ml. aliquots of the resultant cultures were used l−1 and yeast extract 2 g l−1 ) at 37 ◦ C (static) or 28
◦ C (shaken) respectively. The bacterial assay plates
to inoculate additional Erlenmeyer flasks which con-
tained the production medium CYS80 (sucrose, 80 g were prepared by inoculating the same cooled me-
l−1 ; yellow corn meal, 50 g l−1 and yeast extract [Di- dia plus agar (15 g l−1 ) for each microorganism with
fco], 1 g l−1 ). The production flasks were shaken at 3.3% of the inoculum adjusted to an optical density
220 rpm for 21 days at 25 ◦ C. (OD) range of 0.22–0.35 at 660 nm. Thawed cryovials
of the stock inoculum suspensions of the yeasts were
Evaluation of antimicrobial activity used to seed Sabouraud dextrose broth (SDB, 30 g
l−1 ) in culture inoculation flasks which were incub-
Test microorganisms and assay plates. In vitro anti- ated overnight at 28 ◦ C; the C. albicans MY1055 and
microbial susceptibility tests were performed using a S. cerevisiae W303 suspensions were adjusted to 40%
panel of strains which included both human clinical transmittance at 660 nm. This suspension was added to
pathogens and laboratory control strains. The panel yeast nitrogen base-dextrose (YNB-D, yeast nitrogen
consisted of Aspergillus fumigatus MB5668, Bacillus base 6.75 g l−1 , dextrose 1 g l−1 ) in the proportion of
subtilis MB964, Candida albicans MY1055, Entero- 10 ml l−1 . The Aspergillus fumigatus MF5668 stock
coccus faecium MB5571, Mycobacterium smegmatis conidial suspension was adjusted to 3.5 × 109 cfu
MB2233, Pseudomonas aeruginosa MB979, Serra- ml−1 using quantitative colony counts. The conidial
tia marcescens MB252, Saccharomyces cerevisiae suspension was diluted into yeast nitrogen base broth
W303 and Staphylococcus aureus MB5393 from the (YNB; yeast nitrogen base, 6.75 g l−1 ) to 65% trans-
 131
Table 1. Distribution of antimicrobial activities among several orders encompassing basidiomycetes.
Orders are sorted according to the number of isolates tested

Order Number of isolates Active Isolates with Isolates with

isolates (%) antibacterial antifungal
activity (%) activity (%)

AGARICALES 110 48 42 25
PORIALES 91 49 42 21
STEREALES 34 44 41 20
HYMENOCHAETALES 19 11 0 11
BOLETALES 13 46 46 8
GANODERMATALES 11 73 73 0
LYCOPERDALES 10 30 30 10
THELEPHORALES 5 40 40 20
SCHIZOPHYLLALES 4 25 25 0
SCLERODERMATALES 4 0 0 0
NIDULARIALES 3 33 33 0
DACRYMYCETALES 3 33 33 0
CORTINARIALES 3 67 0 67
HERICIALES 3 67 67 33
CANTHARELLALES 2 50 50 0
RUSSULALES 1 100 100 0
HYMENOGASTRALES 1 0 0 0
TOTAL 317 45,1 40 20

mittance at 660 nm; 10 ml of this inoculum was added the same proportion of active isolates was considered
to 1 liter of yeast nitrogen base-dextrose broth. In all in each of the taxonomic categories.
cases, 100 ml aliquots of the seeded agar media were
poured into Nunc square plates (24 × 24 cm).
Results and Discussion
Antimicrobial activity assays. The methanol extracts
from the fermentation broths were prepared by mix- The main purpose of the study was to evaluate and
ing 2 ml of the production culture with 2 ml of 100% compare the ability of different kinds of basidiomy-
methanol, shaking for 15 min and then centrifuging for cetes to produce bioactive substances. A total of 317
15 min at 3000 rpm (×1500 g). Two ml aliquots of the strains, representing 204 species and 17 orders, were
methanol extracts were evaporated to half their volume screened for the production of antimicrobial activ-
in a nitrogen flow in order to increase the concentra- ities. The frequency of the resultant antimicrobial
tion of the metabolites and reduce any toxic effect due activities was considered to be an indicator of the cap-
to the solvent. Methanol extracts (25 µl) were applied ability of the different taxonomic groups of fungi to
to the surface of the seeded assay plates, which were produce bioactive secondary metabolites of potential
incubated at either 28 ◦ C (yeasts) or 37 ◦ C (bacteria). therapeutic interest.
Inhibition zones around the application points were Just over 45% of the test strains showed anti-
measured after 24 hours. microbial activity (Table 1). The highest proportion
of activities occurred with members of the Ganoder-
Data analyses. Statistical analyses of the distribu- matales (73% of the isolates tested). Members of the
tion of the antimicrobial activities were carried out orders Agaricales, Boletales, Poriales and Stereales
using the chi-square test, taking into consideration contained similar percentages of active isolates (in
only those taxa in which ten or more isolates were the range of 46–49%). A lower percentage of activ-
tested. The actual data were contrasted with the ex- ity was shown by the representatives of the orders
pected distribution under the null hypothesis: that is, Hymenochaetales and Lycoperdales (30 and 11%, re-
132
Table 2. Distribution of antimicrobial activities amongst basidiomycetes. The families are sorted alphabetically and
the orders according to the number of isolates tested

Order Family Number of Number of Isolates with Isolates with

isolates active antibacterial antifungal
strains activity activity

AGARICALES Agaricaceae 9 2 2 0
Amanitaceae 3 1 1 0
Bolbitiaceae 8 3 3 0
Coprinaceae 12 8 8 4
Entolomataceae 5 0 0 0
Pluteaceae 1 0 0 0
Strophariaceae 14 6 6 3
Tricholomataceae 58 33 26 21

PORIALES Coriolaceae 69 35 30 17
Lentinaceae 7 3 3 1
Polyporaceae 15 7 6 1

STEREALES Aleurodiscaceae 1 1 1 0
Atheliaceae 2 0 0 0
Botryobasidiaceae 1 1 0 1
Hyphodermataceae 5 3 3 2
Meruliaceae 6 1 1 0
Peniophoraceae 5 2 2 0
Sistotremataceae 1 0 0 0
Steccherinaceae 2 0 0 0
Stereaceae 11 7 7 4

HYMENOCHAETALES Hymanochaetaceae 19 2 0 2

BOLETALES Boletaceae 2 2 2 0
Coniophoraceae 3 1 1 0
Hygrophoropsidaceae 1 0 0 0
Paxillaceae 4 2 2 1
Rhizopogonaceae 3 1 1 0

GANODERMATALES Ganodermataceae 11 8 8 0

LYCOPERDALES Lycoperdaceae 9 3 3 1
Mycenastraceae 1 0 0 0

THELEPHORALES Bankeraceae 2 1 1 0
Thelephoraceae 3 1 1 1

SCHIZOPHYLLALES Schizophyllaceae 4 1 1 0

SCLERODERMATALES Astraeceae 2 0 0 0
Sclerodermataceae 1 0 0 0
Sphaerobolaceae 1 0 0 0

NIDULARIALES Nidulariaceae 3 1 1 0

Continued on p. 133
 133
Table 2. (Continued)

Order Family Number of Number of Isolates with Isolates with

isolates active antibacterial antifungal
strains activity activity

DACRYMYCETALES Dacrymycetaceae 3 1 1 0

CORTINARIALES Crepidotaceae 3 2 0 2
HERICIALES Auriscalpiaceae 1 0 0 0
Gloeocystidiellaceae 1 1 1 0
Lentinellaceae 1 1 1 1

CANTHARELLALES Sparassidaceae 2 1 1 0

RUSSULALES Russulaceae 1 1 1 0

HYMENOGASTRALES Gastrosporiaceae 1 0 0 0

spectively), though the differences recorded between ates showed antibacterial activity and 52 antifungal
the orders were not statistically significant according properties.
to the Chi-square test. No activity was observed with We detected at least one antimicrobial activity in
the representatives of the orders Hymenogastrales and 109 species belonging to 78 genera (Table 3). Eighty
Sclerodermatales, though few strains were studied. two out of 143 test strains which showed activity only
The test strains showed greater antibacterial than inhibited bacteria. In contrast, only 19 strains spe-
antifungal activities with the exception of members cifically inhibited the fungi. The remaining 42 strains
of the order Hymenochaetales. Antifungal activities showed activity against both bacteria and fungi. Activ-
were most frequently found in members of the or- ities against Gram negative bacteria were much less
der Agaricales; the highest percentage of antibacterial common than against Gram positive (25 vs. 119 active
activities was found in the representatives of the order strains, respectively). However, these results may be
Ganodermatales. The differences between the propor- influenced by the fact that the Gram-negative strains
tions of antibacterial activities observed for each of the were highly resistant to many known antibiotics. Only
fungal orders were not statistically significant. Valid eighteen strains showed activity against both groups of
comparisons of the antifungal activities could only be bacteria. These results are consistent with the known
made for the representatives of the orders Agaricales, differences in susceptibility among these organisms
Poriales, Stereales and Hymenochaetales, as members (Peláez et al. 1998).
of the remaining taxa produced too few antifungal It is apparent from Table 3 that representatives
metabolites. The differences between representatives from the different genera showed marked differences
of the orders cited above were not statistically signi- in their ability to show antimicrobial activities. Mem-
ficant. bers of the genera Collybia, Ganoderma, Polyporus
Antimicrobial activity was observed in 32 out and Psathyrella were especially productive (75–100%
of the 44 families that were included in the study of their species showing activity) while Mycena and
(Table 2). However, more representatives of these taxa Trametes strains gave intermediate results (33–50%
need to be studied before any firm conclusions can of species showing activity). In contrast, none of the
be drawn. Nevertheless, it was interesting that the representatives of the genera Agaricus, Panaeolus,
highest activity rates were obtained with the represent- Phellinus and Tricholoma showed activity (members
atives of the families Coprinaceae, Ganodermataceae, of at least three species of the genera cited above were
Stereaceae and Tricholomataceae. Most of these activ- studied). The differences between the representatives
ities were antibacterial. Thus, 91 out of the 106 active of all of the genera might reflect the diverse metabolic
isolates representing families with ten or more isol-
134

Table 3. Antimicrobial activities of representatives of fungal species belonging to the basidiomycetes. The taxa are cited in
alphabetic order

Order Family Species Strain Code PSE SER ENT MYC STAP BAC CAN SAC ASP

AGARICALES Agaricaceae Macrolepiota procera F-074,605 – – + – ++ – – – –

Macrolepiota rhacodes F-051,131 – – – – + – – – –

Amanitaceae Amanita muscaria F-051,823 – – – – – + – – –

Bolbitiaceae Agrocybe semiorbicularis F-057,370 – – – – +++ – – – –

Agrocybe sp. F-074,597 – +++ – – – – – – –
Bolbitius vitellinus F-072,745 – – – – – ++ – – –

Coprinaceae Coprinus episcopalis F-050,152 – – – – +++ +++ – – +

Coprinus episcopalis F-065,192 – – – – +++ +++ – – +++
Coprinus episcopalis F-061,559 – – – – +++ +++ – – –
Psathyrella calcarea F-065,700 – – – – – +++ +++ – –
Psathyrella gracilis F-064,461 – – – – +++ +++ – + –
Psathyrella lacrimabunda F-064,469 – – – – ++ +++ – – –
Psathyrella sylvestris F-073,271 – – + – +++ – – – –
Psathyrella sp. F-074,635 – – – – +++ + – – –

Strophariaceae Hypholoma fasciculare F-058,158 – – + – + – – – –

Panaeolus semiovatus F-064,729 – – – – – ++ – – –
Pholiota carbonaria F-064,714 – – – – – +++ – – –
Pholiota destruens F-051,464 – – – – – ++ – – +++
Stropharia aeruginosa F-051,826 – + – – – +++ ++ – ++
Stropharia coronilla F-065,698 – – ++ – +++ +++ ++ – –

Tricholomataceae Armillaria ostoyae F-051,829 – – + – – – – – –

Armillaria tabescens F-064,487 – – + – – – – – –
Clitocybe nebularis F-051,471 – – – – ++ +++ – – +
Collybia butyracea F-065,711 – – – – – – – – ++
Collybia dryophila F-058,983 – – – – + +++ – – +
Collybia fusipes F-074,604 ++ + – – – + – – –
Crinipellis stipitaria F-055,126 – – – – +++ +++ – +++ ++
Flammulina velutipes F-056,759 – – – – – ++ – – –
Hohenbuehelia mastrucata F-052,950 – – – – + ++ – – –
Hohenbuehelia mastrucata F-051,466 – – – – + + – – –
Hohenbuehelia mastrucata F-065,578 – – – – ++ ++ – – +
Lepista luscina F-064,459 – – +++ – +++ +++ + – +
Lepista nuda F-052,039 – – ++ – +++ – +++ – –
Lepista nuda F-065,697 – + +++ – +++ – +++ – –
Lepista personata F-065,696 – +++ + – +++ – – – –
Leucopaxillus lepistoides F-074,617 – + – – – ++ + – –
Marasmius androsaceus F-065,573 – – +++ – – – + – –
Marasmius corbariensis F-066,985 – – – – – – + – +
Marasmius oreades F-061,534 – – – – – +++ – – –
Marasmius quercophilus F-059,865 – – – – + ++ – – –
Mycena aurantiomarginata F-052,038 – – – – – – + – +
Mycena cf. alcalina F-051,133 – – – – – – +++ – +++
Mycena leucogala F-072,698 – – – – – +++ – – –
 135
Table 3. (Continued)

Order Family Species Strain Code PSE SER ENT MYC STAP BAC CAN SAC ASP

Mycena maculata F-065,581 - – – – – – + –

Oudemansiella longipes F-052,947 – + – – – – ++ – +++
Pseudoclitocybe expallens F-065,703 – – – – – – + – –
Psilocybe semilanceata F-079,215 – – + – +++ – – – –
Rhodotus palmatus F-051,461 – – – – – ++ – + +
Rickenella fibula F-073,712 – – – – ++ ++ – - –
Strobilurus tenacellus F-072,719 – – – – – – – – ++
Strobilurus tenacellus F-072,740 – – – – – – + – +
Strobilurus tenacellus F-072,758 – – – – + – + – +
Xeromphalina junipericola F-054,096 +++ ++ – – – ++ +++ – +

BOLETALES Boletaceae Suillus luteus F-051,470 – – – – – +++ – – –

Suillus variegatus F-051,473 – – – – + +++ – – –

Coniophoraceae Coniophora arida F-066,171 +++ ++ – – – – – – –

Paxillaceae Paxillus involutus F-062,746 – – +++ – +++ +++ – – ++

Paxillus panuoides F-034,827 – – – – – ++ – – –

Rhizopogonaceae Rhizopogon luteolus F-074,590 + + – + + – – – –

CANTHARELLALES Sparassidaceae Sparassis crispa F-051,838 – – – – – +++ – – –

CORTINARIALES Crepidotaceae Crepidotus variabilis F-067,348 – – – – – – + – –

Cyphellopsis anomala F-051,832 – – – – – – +++ – +++

GANODERMATALES Ganodermataceae Ganoderma lucidum F-053,664 – – – – – +++ – – –

Ganoderma lucidum F-050,187 – – – – – ++ – – –
Ganoderma lucidum F-055,100 – – – – – +++ – – –
Ganoderma pfeifferi F-043,710 – – – – – ++ – – –
Ganoderma pfeifferi F-051,842 – – – – – +++ – – –
Ganoderma pfeifferi F-047,996 – – – – – +++ – – –
Ganoderma resinaceum F-057,297 – – – – – +++ – – –
Ganoderma resinaceum F-051,138 – – – – – ++ – – –

HERICIALES Gloeocystidiellaceae Gloeocystidiellum porosum F-062,760 +++ +++ – - ++ ++ – – –

Lentinellaceae Lentinellus omphalodes F-054,100 – – – – ++ +++ – ++ +

HYMENOCHAETALES Hymenochaetaceae Inonotus hispidus F-065,713 – – – – – – – – +++

Onnia tomentosa F-064,431 – – – – – – – – +

PORIALES Coriolaceae Abortiporus biennis F-050,189 – – – – – – – ++ –

Cerrena unicolor F-051,137 – – – – – +++ – + –
Cerrena unicolor F-053,575 – – – – + +++ ++ – +++
Coriolopsis bursina F-061,554 – – – - +++ +++ – – +
Coriolopsis rigida F-061,555 – – – – ++ + – – +
Daedalea quercina F-065,714 – – – – – +++ – – –
Daedaleopsis confragosa F-043,711 – – – – – +++ – – –
Daedaleopsis confragosa F-052,602 – – – – – – – +++ +++
Fomes fomentarius F-051,847 +++ ++ – – – – – – –
136

Table 3. (Continued)

Order Family Species Strain Code PSE SER ENT MYC STAP BAC CAN SAC ASP

Fomes fomentarius F-065,712 – – – – – ++ – – –

Fomitopsis pinicola F-052,945 + +++ – – – +++ – – –
Fomitopsis pinicola F-048,825 – – – – – +++ – – –
Fomitopsis pinicola F-076,863 ++ ++ – – + ++ – – –
Gloeophyllum odoratum F-052,032 – – – – + ++ – + +
Gloeophyllum sepiarium F-051,468 – – – – – – – +++ +++
Heteroporus biennis F-053,574 – – – – + +++ + – +
Ischnoderma benzoinum F-053,569 – – – – – ++ ++ +++ +
Ischnoderma benzoinum F-064,485 – – – – + – + – +
Laetiporus sulphureus F-052,031 – ++ – – +++ ++ – – –
Leptoporus mollis F-054,768 – – – – +++ – – – –
Lenzites elegans F-057,298 – – – – + ++ – – –
Meripilus giganteus F-050,635 – – – – – + – – –
Meripilus giganteus F-076,892 – – – – – +++ ++ – ++
Meruliopsis corium F-060,150 – – – – + – – – –
Meruliopsis corium F-063,931 +++ ++ – – +++ ++ – – –
Perenniporia fraxinea F-065,717 – + – – – – – – –
Phaeolus schweinitzii F-051,846 – – – – + – – – +
Piptoporus betulinus F-048,838 +++ +++ – + ++ +++ – – –
Piptoporus betulinus F-051,849 +++ +++ – – – ++ – – –
Piptoporus betulinus F-054,152 +++ +++ + + +++ – + +
Piptoporus betulinus F-062,745 – – – – – +++ – – +
Pycnoporus cinnabarinus F-076,859 + – – – + + – – –
Pycnoporus sanguineus F-057,299 – – – – – – – + –
Trametes gibbosa F-052,946 – + – – – – – – –
Trametes versicolor F-053,576 – – – – – – – – +

Lentinaceae Pleurotus eryngii F-073,291 – – – – – + – – –

Pleurotus eryngii F-073,223 – – + – +++ ++ – – +
Pleurotus eryngii F-064,460 – – – – – ++ – – –

Polyporaceae Dichomitus squalens F-061,527 – – – – – – ++ – –

Polyporus arcularius F-073,224 – – – + – – – – –
Polyporus arcularius F-057,302 – – – – ++ – – – –
Polyporus arcularius F-057,363 – – – – ++ – – – –
Polyporus arcularius F-074,576 – + + – + – – – –
Polyporus meridionalis F-054,150 – – – – – +++ – – –
Polyporus rhizophilus F-074,637 – – – – + – – – –

SCHIZOPHYLLALES Schizophyllaceae Schizophyllum commune F-056,755 – – – – + – – – –

STEREALES Aleurodiscaceae Aleurodiscus botryosus F-061,553 – – – – – ++ – – –

Botryobasidiaceae Botryobasidium sp. F-061,523 – – – – – – ++ – –

Hyphodermataceae Cerocorticium confluens F-058,082 – – – – ++ – + – –

Hyphoderma definitum F-061,525 – – – – + – – – –
Hyphodontia subalutacea F-061,526 – – – – + – – – –
 137

Table 3. (Continued)

Order Family Species Strain Code PSE SER ENT MYC STAP BAC CAN SAC ASP

Meruliaceae Chondrostereum purpureum F-076,897 ++ +++ – – – – – – –

Peniophoraceae Peniophora incarnata F-061,552 – – – – – ++ – – –

Peniophora quercina F-053,433 +++ +++ – + + – – – –

Stereaceae Stereum hirsutum F-056,757 – – – – + +++ – – –

Stereum hirsutum F-055,125 - – – – + ++ – + –
Stereum hirsutum F-054,102 - – – – – +++ – – +
Stereum hirsutum F-052,037 - – – – + +++ – – +
Stereum hirsutum F-052,035 - – – – ++ – – – –
Stereum hirsutum F-048,834 - – – – + – – – –
Stereum insignitum F-065,721 – – – – – + – – +

DACRYMYCETALES Dacrymycetaceae Calocera viscosa F-051,462 – – – – - ++ – – –

LYCOPERDALES Lycoperdaceae Bovista plumbea F-048,839 – – – – – + – ++ –

Bovista plumbea F-073,702 - – – – – + – – –
Lycoperdon echinatum F-074,659 – – + – + – – – –

NIDULARIALES Nidulariaceae Crucibulum laeve F-065,576 – – – – + – – – –

RUSSULALES Russulaceae Lactarius deliciosus F-052,030 – – – + +++ +++ – – –

THELEPHORALES Bankeraceae Bankera violascens F-052,944 – – – – – +++ – – –

Thelephoraceae Hydnellum ferrugineum F-054,098 – – – – ++ – + – +
TOTAL ACTIVITIES 15 24 16 6 66 84 28 14 41

Activities were classified according to the diameter of the inhibition zones around the point of application of the sample. +++: more or
equal than 15 mm; ++: less than 15 mm; +: hazy/very hazy inhibition zone; –: without activity.
PSE=Pseudomonas aeruginosa MB979, SER=Serratia marcescens MB252, ENT=Enterococcus faecium MB5571, MYC=Mycobacterium
smegmatis MB2233, STAP=Staphylococcus aureus MB5393,
BAC=Bacillus subtilis MB964, CAN= Candida albicans MY1055, SAC= Saccharomyces cerevisiae W303, ASP= Aspergillus fumigatus
MF5668.
No activities were found in representatives of the following species:
Agaricus augustus, Agaricus bitorquis, Agaricus semiotus, Agaricus sylvicola, Agaricus xanthoderma, Agrocybe praecox, Amanita
malleata, Amanita rubescens, Astraeus hygrometricus, Auriculariopsis ampla, Auriscalpium vulgare, Calocera cornea, Calvatia cyath-
iformis, Calvatia utriformis, Clitocybe odora, Clitopilus prunulus, Collybia distorta, Coniophora puteana, Conocybe pubescens, Coprinus
domesticus, Dacrymyces stillatus, Dichomitus campestris, Entoloma conferendum, Entoloma papillatum, Faerberia carbonaria, Fibu-
lomyces mutabilis, Gloeophyllum striatum, Hemimycena cephalotricha, Hexagonia purpurea, Hydnellum aurantiacum, Hygrophoropsis
aurantiaca, Inonotus tamaricis, Irpex lacteus, Laccaria amethystina, Laccaria farinacea, Lentinus tigrinus, Lenzites betulinum, Leuco-
agaricus naucinus, Leucopaxillus cutrefactus, Lopharia spadicea, Lycoperdon perlatum, Macrolepiota permixta, Melanoleuca brevipes,
Mycena amicta, Mycena corticola, Mycena flavoalba, Mycenastrum corium, Onnia triqueter, Omphalotus olearius, Panaeolus acuminatus,
Panaeolus ater, Panaeolus sphinctrinus, Peniophora fraxinea, Phanerochaete sanguinea, Phellinus igniarius, Phellinus merillii, Phellinus
pini, Phellinus pomaceus, Phellinus torulosus, Phellinus tuberculosus, Phellodon melaleucus, Phlebiopsis roumeguerii, Pisolithus tinc-
torius, Pluteus cervinus, Polyporus squamosus, Psathyrella candoleana, Pterula multifida, Resupinatus aplicatus, Rhizopogon roseolus,
Rigidoporus ulmarius, Schizopora paradoxa, Sistotrema muscicola, Sphaeorobolus strellatus, Spongipellis pachyodon, Spongipellis
spumens, Steccherinum ochraceum, Thelephora caryophyllea, Trametes hirsuta, Trametes junipericola, Trametes multicolor, Trametes
trogii, Trichaptum biformis, Tricholoma imbricatum, Tricholoma portentosum, Tricholoma terreum.
In addition, no activities were detected in representatives of four undetermined species of the genera Hemimycena, Hohenbuehelia, Mycena
and Rickenella.
138
Table 4. Antimicrobial activities shown by representatives of
different fungal groups
cularius strains. It is possible that the differences may
reflect genetic divergence at the infraspecific level but
Fungal group Number of Number of Percentage of they may also be attributable, in part, to experimental
isolates active isolates active isolates error and flask-to-flask variation. Significant differ-
ences in the production of metabolites among conspe-
Eurotiales1 175 131 74.9
cific isolates of non-basidiomycetes species have been
Sordariales2 94 69 73.4
Hypocreales3 207 141 68.1
reported (e.g. Möller et al. 1997; Peláez et al. 1998).
Diaporthales4 190 113 59.5
It is apparent from Table 4 that basidiomycetes
Leotiales5 112 62 55.4 compare favorably with ascomycetes as a source of
Dothideales6 1041 561 53.9 biologically active natural products. Their potential
Xylariales7 873 328 37.6 was similar or even slightly above that found for
Pezizales8 47 14 29.8 members of the Pezizales and Xylariales. In contrast,
Basidiomycetes 317 143 45.1 representatives of other ascomycetous groups, such as
the Diaporthales, Eurotiales, Hypocreales, Leotiales
The ascomycetes were tested under the same conditions
described in the materials and methods section. and Sordariales showed a higher ability to synthesize
1 Aspergillus, Emericella, Emericellopsis, Eupenicillium, a range of biologically active metabolites. Dreyfuss
Neosartorya, Paecilomyces, Penicillium and Talaromyces & Chapela (1994) calculated the “creativity index” of
species. several fungal groups as the ratio between the num-
2 Arthrinium, Chaetomium, Coniochaeta, Corynas-
cus,Emilmuelleria, Gelasinospora, Lasiosphaeria, Podospora, ber of known metabolites and species estimated for
Sordaria, Triangularia and Zopfiella species. each group. They obtained a difference of two orders
3 Cylindrocarpon, Cylindrocladium, Fusarium, Nectria and
of magnitude between the creativity indexes of basi-
Trichoderma species.
4 Coryneum, Cytospora and Phomopsis species. diomycetes and representatives of genera classified in
5 Bisporella, Ciboria, Cryptosporiopsis, Dasyscyphus, the Eurotiales or Hypocreales, such as Aspergillus,
Discomycete, Hyaloscypha, Hymenoscyphus, Lachnella, Fusarium, Penicillium and Trichoderma. It is inter-
Lachnum, Mitrula, Mollisia, Neodasyscypha, Phaeohelotium, esting that the results of the present study are in line
Pyrenopeziza, Rustroemia, Spathularia, Tapesia and Vibrissea
species.
with this conclusion, though the differences involved
6 Alternaria, Ascochyta, Bipolaris, Botryosphaeria, Cla- are not so great. It is possible that the low index calcu-
dosporium, Curvularia, Didymosphaeria, Diplodia, Dreschlera, lated for the basidiomycetes by Dreyfuss & Chapela
Epicoccum, Exserohilum, Hormonema, Phoma, Pleospora, (1994) underestimates their real creative capacity as
Preussia, Septoria, Sporormiella and Stemphyllium species.
7 Geniculosporium, Hypoxylon, Nodulisporium and Xylaria most members of this group have still to be included
species. in natural products screening programs; there is also
8 Aleuria, Ascodesmis, Cheilymenia, Chromelosporium, a dearth of information on the structures of bioactive
Cyathipodia, Eleutherascus, Gyromitra, Morchella, Paxina, compounds produced by these organisms.
Peziza, Sarcoscypha, Scutellinia
Some of the results reported in this study are
consistent with those from earlier studies. It is, for
creativity of these fungi, but may be at least partially a instance, known that Sparassis crispa produces bio-
function of the cultivation conditions. active metabolites as the antifungal compounds spara-
Small or large differences were commonly ob- ssol, ScI and ScII (Woodward et al. 1993). Similarly,
served with respect to the ability of strains from the Crinipellis stipitaria produces crinipellins (Kupka et
same species to produce antimicrobial compounds al. 1979); Clitocybe nebularis, the antibacterial antibi-
(Table 3). However, three strains of Hohenbuehelia otic nebularine (Lofgren et al. 1954) and Strobilurus
mastrucata showed very similar patterns of activity, tenacellus, the fungicide strobilurin A (Anke 1995).
as did the eight strains from the three Ganoderma However, it is also apparent from this study that biolo-
species tested. Relatively minor differences in activ- gical activity is shown by representatives of taxa that
ity were shown by the representatives of Fomitop- have not previously been studied.
sis pinicola, Lepista nuda, Stereum hirsutum and The results of the present study lend weight to
Strobilurus tenacellus, whereas remarkable variation those from earlier reports (Anke 1989) which showed
was seen amongst the Bovista plumbea, Cerrena uni- that basidiomycetes produce a great variety of anti-
color, Daedaleopsis confragosa, Fomes fomentarius, microbial activities. It is also clear that the creativity
Ischnoderma benzoinum, Meripilus giganteus, Pipto- of the basidiomycetes in this respect was evenly dis-
porus betulinus, Pleurotus eryngii and Polyporus ar- tributed across suprageneric groups, though some taxa
 139

are a richer source of bioactive activity than others. Hautzel R & Anke T (1990) Screening of basidiomycetes and asco-
It can, therefore, be concluded that while basidiomy- mycetes for plant growth regulating substances. Introduction of
the gibberellic acid induced de-novo synthesis of hydrolytic en-
cetes may not be as easy to grow as bacteria, including zymes in embryoless seeds of Triticum aestivum as test system.
actinomycetes, they may well be worth screening as Z. Naturforsch. 45c: 1093–1098
their diverse biosynthesis abilities may be harnessed Hawksworth DL, Kirk PM, Sutton BC & Pegler DN (1995)
in the search and discovery of new compounds active Ainsworth & Bisby’s Dictionary of the Fungi (8th Ed.) CAB
International: Kew, UK
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Janssens L, De Pooter HL, Schamp NM & Vandamme EJ (1992)
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involved in this study for their technical assistance; Quel. Proc. Natl. Acad. Sci. U.S. 38: 555–560
notably for their help with the assays, the preparation Korzybski T, Kowszyk-Gindifer Z & Kurylowicz W (1967) Antibi-
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