Fungi in the rhizosphere of common oak and its stumps and their possible effect on infection by Armillaria

Applied Soil Ecology 17 (2001) 215–227

Fungi in the rhizosphere of common oak and its stumps and their possible effect on infection by Armillaria H. Kwa´sna Department of Forest Pathology, Agricultural University, ul. Wojska Polskiego 71 c, 60-625 Pozna´n, Poland Received 26 October 2000; received in revised form 26 March 2001; accepted 27 March 2001

Abstract About 16 fungal communities were isolated from the rhizospheres of thick (0.5–1 cm diameter) and thin (0.5–1 mm diameter) roots of living trees and stumps of common oak (Quercus robur). The density of fungi was 2–5× greater on thick roots from stumps than from living trees. The diversity of fungi was similar in the living trees and stumps. Some of the fungal species whose density was greater in rhizospheres of stumps than of living trees, e.g. Chrysosporium merdarium, C. pannorum, Cylindrocarpon destructans, C. didymum, Mortierella gracilis, M. hygrophila, M. microspora var. macrocystis, M. vinacea, Penicillium adametzii, P. daleae, P. janczewskii, Phialophora cyclaminis, Pseudogymnoascus roseus and Sporothrix schenckii, stimulated the formation of rhizomorphs of Armillaria ostoyae and A. gallica in oak-wood segments. It is presumed that the increase in density of fungi stimulating the rhizomorph production may favour the infection of oak stumps by A. ostoyae and A. gallica. © 2001 Elsevier Science B.V. All rights reserved. Keywords: Armillaria; Oak; Rhizosphere fungi; Stimulating effect; Forestry

1. Introduction The genus Armillaria (Fr.:Fr.) Staude includes economically important fungal pathogens causing root and butt rot in conifers and broadleaved trees throughout the northern temperate zone (Shaw and Kile, 1991). The pathogen invades root bark tissue and colonizes the cambium. Armillaria can be highly aggressive, rapidly advancing through the inner bark to the collar, where it girdles and kills the tree. Armillaria is essentially a pathogen of areas with a hardwood forest history (Peace, 1962). Lethal infections of coniferous stands are often seen on sites where broadleaved trees had been present (Rishbeth, 1982; Łakomy, 1998).

E-mail address: [email protected] (H. Kwa´sna).

In Poland, conifers, especially Pinus sylvestris L., are infected mainly by A. ostoyae (Romagn.) Herink. Oak, especially Quercus robur L., is often attacked by A. gallica Marxmüller and Romagn. Although, A. mellea (Vahl:Fr.) Kummer is the most aggressive to Quercus in Europe, generally it is absent from, or very rare in Poland (Guillaumin et al., 1993; Z˙ ółciak, 1999a,b). In mixed stands of Pinus and Quercus, particularly in young stands, the extent of disease and production of Armillaria rhizomorphs was substantially greater where oak stumps were left after felling (Łakomy, 1998). It was observed that the mycelium of Armillaria was easily harboured by oak stumps which act as a source of nutrients and a prolonged ‘reservoir’ of inoculum. On the surface of stumps, in autumn, the pathogen produces abundant basidiomes. It remains alive for a long time in wood. Armillaria initiates rhizomorphs continuously from oak stumps. Their

0929-1393/01/$ – see front matter © 2001 Elsevier Science B.V. All rights reserved. PII: S 0 9 2 9 - 1 3 9 3 ( 0 1 ) 0 0 1 3 7 - 8

216

H. Kwa´sna / Applied Soil Ecology 17 (2001) 215–227

presence increases the danger of infection for the neighbouring trees or the following plantations. The importance of rhizomorphs in the disease cycle of Armillaria has prompted investigations on the effects of microfungi from rhizospheres and roots of trees on rhizomorph formation. Studies by Kwa´sna and Łakomy (1998) proved that the individual microfungi inhabiting birch stump roots can stimulate the formation and growth of rhizomorphs of A. ostoyae. The purpose of the present study was to determine the composition and changes in microfungal communities in the rhizosphere of stump roots of common oak (Q. robur) 2 years after thinning, and the effects of the most commonly occurring fungi on the formation of rhizomorphs of A. ostoyae and A. gallica. An evaluation of the possible effects of changes in the composition of fungal communities in oak rhizospheres on the infection of stumps by A. ostoyae and A. gallica is attempted. 2. Materials and methods 2.1. Isolation of fungi Samples were collected from two stands, of 30-year-old and 50-year-old common oaks, in the 4700 ha conifer–hardwood complex in the Huta Pusta Forest District (western Poland, 17◦ 10 E, 52◦ 50 N) in divisions 118 a,b and 131 d. Today, this region forms a mosaic of virgin areas mixed with forests in various stages of secondary succession which arose as a result of logging. In both stands, the vegetation was dominant second-growth Scots pine (Pinus sylvestris L.) mixed with the common oak. In the 30-year-old stand, sporadically there was birch (Betula pendula Roth.) in the understory. In the 50-year-old stand, there was ground cover mainly of Festuca, Poa, Calamagrostis whilst, in the 30-year-old stand, there was also Rubus. The afforestation rate was 1.0 in the 30- and 0.6 in the 50-year-old stand. The area is flat. The soil is brown, leached, slightly loamy, deep and fresh, stratified with a light, strongly sandy clay. The pH of the soil was 4.25 in the 30- and 4.15 in the 50-year-old stand. The fungi were isolated from the rhizospheres of common oak trees in mixed stands with Scots pine affected by A. ostoyae. The roots were collected from five randomly selected, apparently healthy, 30-year-old and five 50-year-old co-dominant

oaks (September 1995) and their 2-year-old stumps (September 1997) after the deliberate felling of the trees, within a 30 m × 50 m area. The representativeness of the results was ensured by sampling in two locations (I and II) situated 200 m apart in each stand. Three root complexes of approximately 30 cm length and 5–10 mm in diameter were excavated from the B-horizon (30–50 cm) around each of five trees and, 2 years later, from around the stumps left after these trees had been felled, in each location. The roots were kept at 4◦ C for 5 days. Ten segments of 1 cm long, thick (0.5–1 cm diameter) roots and 10–12 segments of 2 cm long, thin (0.5–1 mm diameter) roots (=0.20 g) were excised from three or four root complexes from each tree and were separately washed in 20 ml of sterile distilled water for 3 min. The suspension was diluted (1:10) and 0.1 ml was placed centrally on agar medium (KH2 PO4 , 1 g; MgSO4 ·7H2 O, 0.5 g; peptone, 5 g; dextrose, 10 g; 0.3% rose-bengal, 10 ml; streptomycin sulphate, 0.1 g; agar, 20 g; distilled water, 1 l) in the center of a Petri dish (90 mm × 15 mm) and spread carefully over the entire surface. Thirty replicates (six for each tree) were used per trial. Fungi were incubated at 22◦ C for 7–10 days. The plates were examined microscopically and sporulating fungi were identified. Penicillium spp. and non-sporulating colonies were transferred to potato dextrose agar (PDA, Difco, 39 g; distilled water, 1 l) slants and incubated at room temperature under diffused daylight until sporulation occurred. Fungi were identified according to their morphology on PDA, SNA (glucose, 0.2 g; sucrose, 0.2 g; KH2 PO4 , 1 g; KNO3 , 1 g; MgSO4 ·7H2 O, 0.5 g; KCl, 0.5 g; agar, 15 g; streptomycin sulphate, 0.1 g; distilled water, 1 l) (Nirenberg, 1976), Czapek yeast extract agar (KH2 PO4 , 1 g; Czapek concentrate, 10 ml; powdered yeast extract, 5 g; sucrose, 30 g; agar, 15 g; distilled water, 1 l) and 2% malt extract agar (powdered malt extract, 20 g; peptone, 1 g; glucose, 20 g; agar, 20 g; distilled water, 1 l). Czapek concentrate included: NaNO3 , 30 g; KCl, 5 g; MgSO4 ·7H2 O, 5 g; FeSO4 ·7H2 O, 0.1 g; ZnSO4 ·7H2 O, 0.1 g; CuSO4 ·5H2 O, 0.05 g; distilled water, 100 ml. Some isolates with sterile black mycelia were induced to sporulate under UV light (310–420 nm for 12 h a day) at 20◦ C, or on 2% malt extract agar kept at 5◦ C in high humidity for 12–15 months.

H. Kwa´sna / Applied Soil Ecology 17 (2001) 215–227

The cultures of A. osloyae and A. gallica were isolated in 1994 from basidiomes fruiting in severely diseased 6-year-old Scots pine in the Huta Pusta Forest District. The density of a fungal community or of a population of a single species was defined as the number of isolates on a determined number of roots. The diversity was defined as the number of species on that number of roots. The statistical significance of differences between numbers of fungi in various root rhizosphere fungal communities was determined by χ 2 -tests. 2.2. Growth in oak sections The most frequently occurring fungi in the rhizosphere (‘test’ fungi) were tested for their interaction with A. ostoyae and A. gallica. Sections (2 cm diameter) from freshly cut live branches of 20-year-old Q. robur were cut into 1 or 5 cm lengths, washed with 70% ethanol and autoclaved twice at 121◦ C for 60 min. The 1 cm sections were immersed into cultures of A. ostoyae or A. gallica growing on 8% malt agar (Difco malt extract, 80 g; agar, 15 g; streptomycin sulphate, 0.1 g; distilled water, 1 l) and incubated in the dark for 30 days at 22◦ C. The 5 cm sections were inoculated with ‘test’ fungi by immersing each in a jar containing a single fungal culture growing on PDA at the bottom. Jars were filled with wet, sterilized sand, closed and left for 2 months in the dark at 22◦ C. When each 1 cm section was covered with a thin mat of white-cream to light brown Armillaria mycelium it was used to inoculate a 5 cm section infested by a ‘test’ fungus. The sections were joined by nailing them together with 2 cm long nails. The control treatment consisted of a sterile 5 cm section attached to an Armillaria-infested section. The attached sections were put into wet, sterilized sand in jars and incubated in the dark at 22◦ C. After 2 months all sections were carefully removed from jars and inserted into plastic bags containing 0.7 kg of a substrate consisting of forest soil, sand, peat and humus (1:1:1:1). After 2 months at 22◦ C, the number of rhizomorphs, their length, number of living initials, and dry weight were assessed. Each ‘test’ fungus/Armillaria treatment and the control treatment were replicated four times. ANOVA was performed to determine significant (P ≤ 0.05 and ≤ 0.001) effects of the ‘test’ fungi

217

on the formation of rhizomorphs of A. ostoyae and A. gallica.

3. Results Sixteen fungal assemblages were isolated from rhizospheres of thick (0.5–1 cm diameter) and thin (0.5–1 mm diameter) roots of living trees and stumps of common oak in 30- and 50-year-old stands, from two locations (I and II) in each stand (Table 1). A total of 172 different species of fungi was isolated. The most common species are presented in Table 1. All of them were identified to species or genus except for 22 species which did not sporulate and occurred rarely. Each fungal community was represented by 40–56 species and consisted of 384–3286 isolates. The most frequently isolated fungi were: Absidia cylindrospora, Aspergillus spp., Chrysosporium spp., Cylindrocarpon spp., Gymnoascus reessii, Mortierella spp., particularly M. gracilis, M. hygrophila, M. microspora var. macrocystis and M. vinacea, Mucor hiemalis, Penicillium spp., particularly P. adametzii, P. citrinum, P. daleae, P. janczewskii and P. steckii, Pseudogymnoascus roseus, Sesquicillium candelabrum, Sporothrix schenckii and Tolypocladium niveum. The genus Trichoderma was represented by 10 species but T. koningii, T. polysporum and T. viride occurred most often. The diversity of fungi (number of species) was similar in the rhizospheres of thick and thin roots of the living trees and stumps (Tables 2 and 3). The density (number of isolates) of fungi on thick stump roots was significantly greater (two to five times) than on living tree roots (Table 2). The difference in density on thin roots of living trees and stumps in the 30-year-old stand was more variable. Density was significantly greater on thin roots of stumps than of living trees in location II in both stands, but the reverse was the case for location I in the 30-year-old stand (Table 3). Chrysosporium spp., G. reessii and P. roseus are grouped in Tables 2–5 because of their morphological similarities and taxonomical affiliation. Gymnoascus and Pseudogymnoascus may be the teleomorphic genus of a heterogeneous assemblage of many species within Chrysosporium (Domsch et al., 1980). The density of Chrysosporium spp. + G. reessii + P. roseus, as well as of Mucorales and Penicillium spp., was

218

H. Kwa´sna / Applied Soil Ecology 17 (2001) 215–227

Table 1 Number of isolates of the most common fungi in the rhizosphere of thick and thin roots of oak (Q. robur L.) and its stumps Fungus

Absidia cylindrospora Hagem Acremonium bacillisporum (Onions & Barron) W. Gams A. fusidioides (Nicot) W. Gams A. strictum W. Gams Aspergillus kanagawaensis Nehira A. niveus Blochwitz A. versicolor Tiraboschi Casdida albicans Robin (Berkhout) Chloridium virescens var. chlamydosporum (V. Beyma) W. Gams et Hoi.-Jech. Chrysosporium merdarium (L. ex G.) Carm C. pannorum (Link) Hughes Cladosporium herbarum Link ex Fr. Cylindrocarpon destructans (Zinssm.) Scholten C. didymum (Hartig) Wollenw. Exophiaia sp. Gymnoascus reessii Baranetzky Hormiactis candida Höhn. Monodictis putredinis (Wallr.) Hughes Mortierella gracilis Linn. M. hygrophila Linn. M. isabellina Oudemans et Koning M. microspora Wolf var. macrocystis (Gams) Linn., W. Gams M. nana Linn. M. turficola Ling-Young M. vinacea Dixon-Stewart Mortierella sp. Mucor hiemalis Wehmer M. plumbeus Bonorden M. racemosus Fres. Mycelium radicis atrovirens alpha Melin Parcilomyces farinosus (Holmskiod) A.H. S Brown & G. Sm. Penicillium adametziii Zaleski Penicillium brevicompactum Dierckx P. chrysogenum Thom P. citrinum Thom Penicillium daleae Zaleski P. herquei Bainier & Sartory P. janczewskii Zaleski P. raistrickii G. Sm. P. spinulosum Thom P. steckii Zaleski P. roseus Raillo Sesquicillium candelabrum (Bonorden) W. Gams Sporothrix schenckiiHectoen et Perkins Tolypocladium niveum (Rostrup) Bissett

Thick roots

Thin roots

Living trees

Stumps

30-year

50-years

30-year

I

I

I

14 4

II

II

II

Living trees

Stumps

50-years

30-year

50-years

30-year

50-years

I

I

I

I

I

II

II

II

II

II

2 0

16 1

23 0

23 0

14 0

10 0

47 0

11 8

6 4

44 20

1 7

8 1

7 0

21 0

10 0

0 0 2 0 1 0 22 11 0 0 0 0 4 1

0 0 0 5 1 0 0

0 0 0 0 0 0 0

0 0 159 0 0 0 0

0 0 0 0 8 1 39

0 0 0 0 4 1 4

0 0 1 17 0 13 1

0 4 0 4 24 1 0

0 5 0 21 0 1 1

0 0 0 3 0 1 1

0 1 0 3 0 20 0

24 0 12 0 70 0 1

40 0 1 0 14 0 3

1 0 0 8 9 2 0

1 0 1 0 13 11 0

11 10 0 3 0 25 0 0 5 3 3 2 16

15 5 2 0 0 21 0 0 0 0 0 1 12

8 3 4 0 0 38 0 0 1 10 1 3 8

11 187 1 6 2 0 18 0 0 2 6 15 53

5 4 3 18 9 9 48 6 51 4 9 10 0 0 0 24 6 3 53 1 0 0 2 0 10 310 1100 0 0 0 0 0 0 109 174 194 1 18 25 0 0 0 0 52 0 1 0 0 86 0 0 0 0 0 6 13 2 1 20 4 20 3 1 17 2 1 0 2 1 0 0 12 59 31 12 24 45 15

45 127 50 32 271 9 13 9 8 37 1 2 0 0 2 0 0 0 1 0 2 0 324 0 1 0 0 0 0 0 0 20 2 9 19 0 0 0 0 0 0 0 1 0 0 7 8 25 5 74 9 2 2 19 1 0 4 2 3 2 0 13 17 116 19

0 0 1 0 46 42 0 0 1 1 0 1 0 1 3 3 0 1

0 1 8 0 1 0 0 1 7

3 1 67 0 27 0 0 1 1

4 0 47 15 0 0 0 3 6

1 0 58 1 0 0 0 3 0

0 0 17 5 14 1 0 0 1

8 11 6 1 0 13 1 0 0 1 0 0 7

43 2 0 73 32 5 26 4 1 2 0 0 4 1

21 0 0 43 30 1 52 2 23 2 3 0 5 6

17 7 2 12 0 0 32 98 76 56 5 3 54 107 3 2 6 6 2 9 0 0 0 3 16 2 17 23

11 0 42 7 0 0 0 2 0

5 0 55 1 0 0 0 0 102

0 3 37 1 31 0 1 0 0

73 68 13 17 24 0 0 0 0 22 2 2 0 2 0 4 3 48 7 1 86 58 177 71 1 0 3 5 0 2 161 74 112 114 126 2 0 0 0 11 0 0 7 3 0 1 6 0 0 730 23 8 0 0 1 7 70 4 1562 0 27 245 287 8 16 0 3 28 5 0

0 0 18 0 5 1 4 0 1

1 7 28 1 9 0 0 0 3

7 0 38 12 0 0 0 2 41

9 22 4 130 0 3 1 0 0 0 0 2 75 144 107 16 7 304 19 24 4 0 8 3 33 48 193 190 3 2 0 1 1 13 2 1 3 10 296 0 11 0 0 10 2 0 0 10 16 2 11 11 20 165 16

10 0 26 0 0 0 0 3 4

3 0 33 0 5 0 0 0 0

0 0 67 0 0 0 0 0 0

52 37 41 3 1 31 10 5 20 6 177 47 34 55 109 2 11 1 30 104 218 0 2 0 3 7 7 0 1 1 33 5 10 3 126 1 4 29 5 22 25 38

H. Kwa´sna / Applied Soil Ecology 17 (2001) 215–227

219

Table 1 (Continued) Fungus

Thick roots

Thin roots

Living trees

Stumps

30-year

50-years

30-year

I

I

I

II

Trichoderma koningii Oudemans 1 0 T. polysporum (Link et Pers.) Rifai 2 0 T. viride Pers. ex Fr. 0 0 Verticillium bulbillosum W. Gams et Malla 0 18 V. lamellicola (F.E.V. Sm.) W. Gams 21 0 Zygorhynchus moelleri Vuill. 10 5 Sterile mycelia 0 0

II 0 2 2 0 3 0 0

0 0 0 0 7 9 0

II 5 0 0 2 0 0 0

Living trees

Stumps

50-years

30-year

50-years

30-year

50-years

I

I

I

I

I

0 2 2 2 2 1 8

II 6 42 3 1 0 1 0

0 0 2 4 0 1 1

II 0 17 0 7 1 4 0

1 3 4 2 0 3 1

II 0 0 1 2 0 1 4

6 8 6 3 1 0 18

II 3 4 4 3 4 0 13

1 0 0 13 0 1 9

II 15 2 3 2 0 0 18

1 0 4 34 0 0 6

Table 2 Effects of felling of oaks on fungal diversity (total number of species), density (total number of isolates) and on number of selected fungal taxa in the rhizospheres of thick roots Parameters

Living trees 30-year-old

2-year-old stumps 50-year-old

30-year-old

50-year-old

I

II

I

II

I

II

I

II

Number of species χ2 P

47 0.563 n.s.a

45 0 n.s.a

47 0.177 n.s.a

42 0 n.s.a

40 0.563 n.s.a

45 0 n.s.a

43 0.177 n.s.a

42 0 n.s.a

Number of isolates χ2 P

406 255.6 <0.00l

384 272.1 <0.001

402 469.9 <0.001

591 1873.3 <0.001

1007 255.6 <0.001

997 272.1 <0.001

1295 469.9 <0.001

3286 1873.3 <0.001

Number of isolates Chrysosporium spp. + G. reessii + P. roseus χ2 P

21 182.7 <0.001

4 16.7 <0.001

20 1.3 n.s.a

13 47.3 <0.001

239 182.7 <0.001

62 16.7 <0.001

28 1.3 n.s.a

79 47.3 <0.001

Mucorales χ2 P

82 38.5 <0.001

81 27 <0.001

43 40.7 <0.001

157 0.2 n.s.a

183 38.5 <0.001

162 27 <0.001

126 40.7 <0.001

166 0.2 n.s.a

Penicillium spp. χ2 P

201 33.9 <0.001

179 4 <0.05

249 23.3 <0.001

326 21.8 <0.001

336 33.9 <0.001

219 4 <0.05

369 23.3 <0.001

217 21.8 <0.001

Trichoderma spp χ2 P

3 0.5 n.s.a

4 0 n.s.a

4 42.1 <0.001

0 2 n.s.a

5 0.5 n.s.a

4 0 n.s.a

53 42.1 <0.001

2 2 n.s.a

a

n.s.: The ratio for living trees:stumps is not significantly different from 1:1 at P ≤ 0.05.

usually significantly greater in the rhizospheres of thick roots of stumps than of living trees (Table 2). On thin roots, the densities of Chrysosporium spp. + G. reessii + P. roseus and of Mucorales in the 50-year-old stand were also greater on stumps. Mucorales in location I of the 30-year-old stand and Penicillium

spp. in the 50-year-old stand and in location I of the 30-year-old stand were less dense on stumps (Table 3). Effects on Trichoderma spp. were more variable. The location of the sample (I or II) did not affect the number of species isolated from either thick or thin roots (Tables 4 and 5). There were significant

220

H. Kwa´sna / Applied Soil Ecology 17 (2001) 215–227

Table 3 Effects of felling of oaks on diversity (total number of species), density (total number of isolates) and on number of selected fungal taxa in the rhizospheres of thin roots Parameters

Living trees 30-year-old

2-year-old stumps 50-year-old

30-year-old

50-year-old

I

II

I

II

I

II

I

II

Number of species χ2 P

43 0.18 n.s.a

51 0 n.s.a

45 1.2 n.s.a

48 0.01 n.s.a

47 0.18 n.s.a

51 0 n.s.a

56 1.2 n.s.a

47 0.01 n.s.a

Number of isolates χ2 P

1303 83.6 <0.00l

548 52.3 <0.001

985 1.1 n.s.a

1044 4.9 <0.05

876 83.6 <0.001

815 52.3 <0.001

938 1.1 n.s.a

1148 4.9 <0.05

Number of isolates Chrysosporium spp.+ G. reessii + P. roseus χ2 P

24 110.8 <0.001

29 8 <0.05

22 14.1 <0.001

54 204.8 <0.001

171 110.8 <0.001

94 8 <0.05

55 14.1 <0.001

337 204.8 <0.001

Mucorales χ2 P

136 8.1 <0.05

114 1.9 n.s.a

134 21.3 <0.001

74 42.3 <0.001

93 8.1 <0.05

94 1.9 n.s.a

221 21.3 <0.001

177 42.3 0.001

Penicillium spp. χ2 P

918 234.2 <0.001

137 0.17 n.s.a

547 21.2 <0.001

663 23.1 <0.001

369 234.2 <0.001

144 0.17 n.s.a

405 21.2 <0.001

499 23.1 <0.001

Trichoderma spp χ2 P

17 1.28 n.s.a

13 10.3 <0.001

1 21.2 <0.001

24 12.5 <0.001

11 1.28 n.s.a

1 10.3 <0.001

24 21.2 <0.001

5 12.5 <0.001

a

n.s.: The ratio for living trees:stumps is not significantly different from 1:1 at P ≤ 0.05.

differences between locations in densities (number of isolates) from the rhizospheres of thick roots of living trees and of stumps in the 50-year-old stand (Table 4) and also in the rhizospheres of thin roots of living trees in the 30-year-old stand and of stumps in the 50-year-old stand (Table 5). The densities of the selected fungal taxa in the two locations within stands were often, but not always, significantly different (Tables 4 and 5). Acremonium bacillisporum, A. strictum, Exophiala sp., Mortierella turficola, Mucor hiemalis, M. plumbeus, M. racemosus and P. steckii were detected only or mostly in rhizospheres of living trees. Acremonium fusidioides, A. kanagawaensis, C. didymum, G. reessii, P. chrysogenum and S. candelabrum were found only or mostly in rhizospheres of the stump roots. The densities of the most commonly occurring fungi were usually greater in rhizospheres of stump roots

than of living tree roots. Changes in frequency (in %) of the most common fungal species on stump roots are presented in Table 6. Fungi whose densities were greater in stump rhizospheres had statistically significant effects on the rhizomorph characteristics of A. ostoyae and A. gallica (Tables 7 and 8). ANOVA showed that C. pannorum, C. destructans 1 and 2, M. microspora var. macrocystis, P. adametzii 4 and 6, P. daleae 1, P. janczewskii 1 and P. roseus caused significant increases in number, length and weight of A. ostoyae rhizomorphs (P ≤ 0.05 and ≤ 0.001). Cylindrocarpon destructans 1 and 3, C. didymum 1, M. hygrophila, P. adametzii and P. daleae 2 and 3 increased significantly the number of A. gallica rhizomorph apices, their length and weight (P ≤ 0.05and ≤ 0.001). Chrysosporium merdarium and C. didymum 3 caused significant increases in length and number of rhizomorph initials, and C. destructans 2, M. gracilis, M microspora var. macro-

H. Kwa´sna / Applied Soil Ecology 17 (2001) 215–227

221

Table 4 Effects of the location (I and II) on the diversity (total number of species), density (total number of isolates) and on number of selected fungal taxa in the rhizospheres of thick roots Parameters

Living trees

2-year-old stumps

30-year-old

50-year-old

I

II

I

II

I

II

I

II

Number of species χ2 P

47 0.04 n.s.a

45 0.04 n.s.a

47 0.28 n.s.a

42 0.28 n.s.a

40 0.29 n.s.a

45 0.29 n.s.a

43 0.01 n.s.a

42 0.01 n.s.a

Number of isolates χ2 P

406 0.61 n.s.a

384 0.61 n.s.a

402 35.9 <0.001

591 35.9 <0.001

1007 0.05 n.s.a

997 0.05 n.s.a

1295 865.3 <0.001

3286 865.3 <0.001

Number of isolates Chrysosporium spp.+ G. reessii + P. roseus χ2 P

21 0.2 n.s.a

24 0.2 n.s.a

20 1.48 n.s.a

13 1.48 n.s.a

239 104.1 <0.001

62 104.1 <0.001

28 24.3 <0.001

79 24.3 <0.001

Mucorales χ2 P

82 0.006 n.s.a

81 0.006 n.s.a

43 64.98 <0.001

157 64.98 <0.001

183 1.28 n.s.a

162 1.28 n.s.a

126 5.48 <0.05

166 5.48 <0.05

Penicillium spp. χ2 P

201 0.58 n.s.a

179 0.58 n.s.a

249 10.3 <0.05

326 10.3 <0.05

336 24.7 <0.001

219 24.7 <0.001

369 39.4 <0.001

217 39.4 <0.001

Trichoderma spp χ2 P

3 0.14 n.s.a

4 0.14 n.s.a

4 4 <0.05

0 4 <0.05

5 0.11 n.s.a

4 0.11 n.s.a

53 47.3 <0.001

2 47.3 <0.001

a

30-year-old

50-year-old

The ratio for location I:location II is not significantly different from 1:1 at P ≤ 0.05.

cystis and P. roseus increased rhizomorph length. Some fungi caused partial inhibition of rhizomorph formation. Compared to the control, the rhizosphere fungi only rarely caused increases in the number of rhizomorphs of A. ostoyae or A. gallica (Tables 7 and 8). Rhizomorphs were produced from one or several points, mostly at the ends of segments. In the controls, A. ostoyae formed, on average, only one very short rhizomorph.

4. Discussion In the temperate zone, the intensive development of butt and root rot caused by Armillaria is often observed in conifers (particularly young plantations) planted after oaks. Damage is generally unimportant or much reduced in crops replacing conifers (Childs and Zeller, 1929; Redfern and Filip, 1991; Łakomy, 1998). Oak stumps are an important source of nutri-

ent for Armillaria. They are generally a satisfactory substrate for rhizomorph production until decay is far advanced. Rishbeth (1972) reported that Armillaria produced rhizomorphs on wood from English oak stumps for at least 40 years after the trees were cut. Usually, the yield of rhizomorphs increased with the period since felling and was greatest after 14 years, although the amount varied considerably between stumps (Rishbeth, 1972). A. ostoyae and A. gallica are the species that occur most often on oak stumps in Poland. A. ostoyae is considered as highly pathogenic to following conifers and A. gallica mainly to following hardwoods. The latter, although it has been generally regarded as a weak pathogen on both coniferous and hardwood hosts, infects mainly weakened broadleaved trees in Poland. Although, according to Mohammed and Guillaumin (1989), A. gallica may act in synergy with A. ostoyae, intensive studies on Armillaria in Poland did not confirm this (Łakomy, 1998).

222

H. Kwa´sna / Applied Soil Ecology 17 (2001) 215–227

Table 5 Effects of the location (I and II) on the diversity (total number of species), density (total number of isolates) and on number of selected fungal taxa in the rhizospheres of thin roots Parameters

Living trees 30-year-old

2-year-old stumps 50-year-old

30-year-old

50-year-old

I

II

I

II

I

II

I

II

Number of species χ2 P

43 0.68 n.s.a

51 0.68 n.s.a

45 0.09 n.s.a

48 0.09 n.s.a

47 0.16 n.s.a

51 0.16 n.s.a

56 0.79 n.s.a

47 0.79 n.s.a

Number of isolates χ2 P

1303 307.9 <0.001

548 307.9 <0.001

985 1.7 n.s.a

1044 1.7 n.s.a

876 2.2 n.s.a

815 2.2 n.s.a

938 21.1 <0.001

1148 21.1 <0.001

Number of isolates Chrysosporium spp.+ G. reessii + P. roseus χ2 P

24 0.47 n.s.a

29 0.47 n.s.a

22 13.8 <0.001

54 13.8 <0.001

171 22.4 <0.001

94 22.4 <0.001

55 202.9 <0.001

337 202.9 <0.001

Mucorales χ2 P

136 1.94 n.s.a

114 1.94 n.s.a

134 17.3 <0.001

74 17.3 <0.001

93 0.005 n.s.a

94 0.005 n.s.a

221 32 <0.001

177 32 <0.001

Penicillium spp. χ2 P

918 578.2 <0.001

137 578.2 <0.001

547 11.1 <0.001

663 11.1 <0.001

369 98.7 <0.001

144 98.7 <0.001

405 9.8 <0.05

499 9.8 <0.05

Trichoderma spp χ2 P

17 0.53 n.s.a

13 9.53 n.s.a

1 21.16 <0.001

24 21.16 <0.001

11 8.3 <0.05

1 8.3 <0.05

24 12.4 <0.001

5 12.4 <0.001

a

n.s.: The ratio for location I:location II is not significantly different from 1:1 at P ≤ 0.05.

The growing roots of trees exude a mixture of 18 amino acids, 8–10 organic acids, 3–4 carbohydrates, mucilage and other substances. Together with sloughed-off root cap and other cells, these exudates cause the rhizosphere effect (Rovira, 1965; Smith, 1976) which, besides the physical and chemical changes, includes the creation of a zone of enhanced microbiological activity due to the germination of the dormant spores and increase in fungal vigour. Despite the degradation and mineralization process of dead stump roots, increases in the density and sometimes in the diversity of rhizosphere fungi occurred. The density of fungi was two to five times greater on the surface of thick roots of 2-year-old oak stumps than on the roots of living trees. A similar effect was observed previously on birch (Kwa´sna, 1996a,b) and on Scots pine (Kwa´sna, 1997). An increase was observed in both the 30- and the 50-year-old stands, although both stands varied in afforestation rate resulting in

differences in temperature and humidity of the soil and, consistently, in the rate of the decomposition process. There was no great difference in the diversity of fungi (measured as the number of species) detected on roots of the living oaks and their 2-year-old stumps, although the decay of stump wood and the presence of a substrate at different stages of decomposition, utilized by different saprotrophes, usually increases the diversity of fungi. A similar observation was made on birch (Kwa´sna, 1996a,b). On Scots pine the diversity of rhizosphere fungi was often greater on stumps than on the living tree roots (Kwa´sna, 1997). It seems that the similar number of fungi on living oaks and its stump roots might indicate a low rate of decomposition of oak stump wood 2 years after felling. This can be explained by the exceptional hardness of oak wood impregnated with tannins. Redfern (1968) and Rishbeth (1972) noticed that the degradation of 2–6 in.

H. Kwa´sna / Applied Soil Ecology 17 (2001) 215–227

223

Table 6 Differences (in %) between living trees and stumps in frequency of the most commonly occurring fungi in the rhizospheres of thick and thin roots of common oak Fungus

A. cylindrospora A. kanagawaensis A. versicolor C. merdarium C. pannorum C. didymum G. reessii M. gracilis M. hygrophila M. microspora var. macrocystis M. nana M. vinacea P. adametzii P. chrysogenum P. daleae P. janczewskii P. roseus S. candelabrum S. schenckii

Thick roots

Thin roots

30-year-old

50-year-old

30-year-old

50-year-old

+332.1 +7900 p −8.5b +990 p +850 +100 +284.4 +462.9 p +25.2 +146.8 p +131.1 +280.8 +83.4 p +2687.5

a +33.4

−5.3 p +95.8 +530.3 +112.5 p p +370 −44.1 −54 p +23.6 +459.7 p +1342.9 −4.1 +550 +25 −45.7

+423.9 p p +378.5 +145.5 −50 p +491.1 +855.6 +337 +100 +155.9 +496.6 p +195.8 +64.75 p p +109.2

pc +150 −67.9 +810 p p −35 p +104.2 +33.4 +203.6 +59.7 p +79.8 +56.9 0 +25983 +996.9

a

‘+’:increase in the fungus frequency in %. ‘−’:decrease in the fungus frequency in %. c ‘p’:fungus is present in rhizosphere of stumps while absent in rhizosphere of living trees roots. b

diameter Q. robur roots is very slow and takes >5 years after felling. Environmental conditions cannot be excluded as causes of the increase in density of fungi on decomposed roots. Diurnal and seasonal fluctuations in temperature and changes in soil humidity might have a direct and immediate impact on all aspects of fungal life. Temperature imposes physiological limits on and controls the rates of spore germination, fungal growth and reproduction. In temperate zone habitats, seasonal changes in temperature can potentially alter the temporal partitioning of resources among fungal species with similar substrate-utilization potentials. Temperature also affects the outcome of competitive interactions among fungi. On a large scale, however, microfungal community size and composition change in response to the type and availability of nutrients. Generally, the density and diversity of fungi increase in the presence of organic substrates (Wicklow and Whittingham, 1974).

Different species of fungi can grow and reproduce on various organic substrates after various times of decomposition. On oak stump roots, 2 years after felling, there were mostly A. cylindrospora, Chrysosporium spp., Cylindrocarpon spp., G. reessii, M. gracilis, M. hygrophila, M. microspora var. macrocystis, M. nana, M. vinacea, P. adametzii, P. daleae, P. janczewskii, P. roseus, S. candelabrum and S. schenckii. On birch stump roots there was mainly Zygorhynchus moelleri (Kwa´sna, 1996a,b), and on Scots pine stump roots T. viride predominated (Kwa´sna, 1997). Mortierella gracilis, M. vinacea and P. daleae, were common on all three kinds of stumps. Penicillium spinulosum and P. steckii, common on birch and Scots pine stump roots, occurred mostly on living oak roots. The latter occurred with a high frequency, particularly on thin roots (Table 1). The root washing method used in this study is a modification of the soil dilution plate technique. It allows mainly the identification of fungi that are

224

H. Kwa´sna / Applied Soil Ecology 17 (2001) 215–227

Table 7 The effects of fungal isolates from common oak roots and rhizospheres on rhizomorph production by A. ostoyaea Fungus

Number of rhizomorphs

Number of rhizomorphs apices

Length of rhizomorphs (mm)

Weight of rhizomorph (mg)

A. ostoyae — control C. merdarium C. pannorum C. destructans 1 C. destructans 2 C. destructans 3 C. didymum 1 C. didymum 2 C. didymum 3 C. didymum 4 A. gracilis M. hygrophila M. microspora var. macrocystis M. vinacea P. adametzii 1 P. adametzii 2 P. adametzii 3 P. adametzii 4 P. adametzii 5 P. adametzii 6 P. chrysogenum P. daleae 1 P. daleae 2 P. daleae 3 P. janczewskii 1 P. janczewskii 2 P. cyclaminis P. roseus S. candelabrum S. schenckii

1 3.25 0.25 4.25 2.25 0.5 1.25 0.25 0.5 0 4.0 0 1.75 1.0 0 0 0 1.25 0.25 2.5 0.25 2.7 0.75 0 1.7 1.0 0.25 0.75 0 0.5

1 3.25 0.25 8.75∗ 4.75 0.5 1.25 0.25 0.5 0 5.0 0 18.0∗∗ 1.0 0 0 0 2.0 0.25 5.5 0.5 3.0 1.0 0 1.7 1.0 0.75 3.75 0 0.5

1 5.5 0.5 15.75∗∗ 11.5∗ 0.25 0.875 0.075 1.0 0 5.625 0 37.0∗∗ 11.2∗ 0 0 0 3.5 0.25 14.25∗∗ 1.25 31.0∗∗ 3.0 0 24.0∗ 0.375 2.0 6.25 0 5.0

1 3.25 0.25 37.5∗∗ 17.5∗∗ 0.5 1.5 0.25 0.75 0 5.0 0 46.75∗∗ 0.7 0 0 0 7.5∗ 0.5 17.5∗ 1.25 4.2 6.25 0 1.2 0.5 0.5 7.0∗ 0 0.25

Mean, n = 4. Significantly different at P ≤ 0.05. ∗∗ Significantly different at P ≤ 0.001. a

∗

external on the roots and which can be easily separated. There were mainly spores and fragments of mycelium of externally occurring pathogens as well as saprotrophes. The root fungi harboured by the cork and external bark were detected only if they were also present outside the roots. The method used does not differentiate the externally and internally growing species. It indicates the organisms living in the closest vicitiny to the roots, in the rhizosphere. Macrofungi (Basidiomycotina) which are potential mycorrhizal fungi, are detected with this method only rarely. The studies confirmed some observations made earlier (Kwa´sna, 1996a,b; 1997) that a few typical saprotrophes, e.g. Acremonium, Exophiala, Mucor

and P. steckii, more often colonize the living tissues, while others, e.g. A. kanagawaensis, M spinosa, P. farinosus and P. chrysogenum, occur mainly on dead roots. The density of C. didymum, S. candelabrum and S. schenckii on stump roots was locally very high. Their particularly frequent appearance on the special kind of substrate may result from their nutritional preferences or temporal and spatial heterogeneity in the distribution of nutrients and in microclimate. Many species commonly occurring in oak rhizospheres were studied from the point of view of their effect on rhizomorph production by A. ostoyae and A. gallica. Fungi that live in the oak rhizosphere and whose density was greater on stump roots often

H. Kwa´sna / Applied Soil Ecology 17 (2001) 215–227

225

Table 8 The effects of fungal isolates from common oak rhizosphere on rhizomorph production of A. gallicaa Fungus

Number of rhizomorphs

Number of rhizomorphs apices

Length of rhizomorphs (mm)

Weight of rhizomorph (mg)

A. gallica — control A. kanagawaensis C. merdarium C. pannorum C. destructans 1 C. destructans 2 C. destructans 3 C. didymum 1 C. didymum 2 C. didymum 3 C. didymum 4 M. gracilis M. hygrophila M. microspora var. macrocystis P. adametzii 1 P. adametzii 2 P. adametzii 3 P. adametzii 4 P. adametzii 5 P. adametzii 6 P. chrysogenum P. daleae 2 P. daleae 3 P. janczewskii 2 P. janczewskii 3 P. roseus S. candelabrum

3.75 1.0 2.5 0.25 2.75 1.5 3.75 5.5 0 1.75 0.25 2.5 5.25 3.5 3.25 4.0 4.75 3.75 3.5 5.25 2.0 3.0 2.25 0 1.25 3.0 0

9.5 1.0 25.75∗ 0.25 53.75∗∗ 11.25 59.25∗∗ 57.5∗∗ 0 37.0∗∗ 0.25 19.5 70.0∗∗ 14.5 55.75∗∗ 22.75∗ 68.75∗∗ 47.0∗∗ 32.5∗∗ 37.5∗∗ 5.75 38.75∗∗ 47.5∗∗ 0 5.5 16.25 0

16.75 1.0 48.0∗∗ 0.125 91.25∗∗ 33.0∗∗ 101.25∗∗ 150.0∗∗ 0 44.5∗ 0.5 39.5∗ 187.5∗∗ 43.5∗∗ 105.5∗∗ 52.5∗∗ 105.0∗∗ 74.25∗∗ 73.5∗∗ 75.0∗∗ 20.0 125.0∗∗ 135.0∗∗ 0 14.5 45.0∗∗ 0

36.25 3.0 47.75 0.25 190.0∗∗ 42.5 147.5∗∗ 325.0∗∗ 0 48.5 0.5 43.75 410.0∗∗ 36.25 162.5∗∗ 60.0∗ 232.5∗∗ 135.0∗∗ 112.5∗∗ 80.0∗∗ 12.5 215.0∗∗ 187.5∗∗ 0 18.75 52.5 0

Mean, n = 4. Significantly different at P ≤ 0.05. ∗∗ Significantly different at P ≤ 0.001. a

∗

stimulated the formation and growth of Armillaria rhizomorphs. The greatest stimulating effect was caused by species whose density on stump roots increased moderately. Other species with locally high density on stump roots, e.g. A. kanagawaensis and S. candelabrum, were able to inhibit rhizomorph growth entirely. Since the ‘test’ fungi are also often isolated from inside tree roots (author, unpublished), it is presumed that in the test with oak sections they grew not only on the surface but also penetrated the cylinders. Reisolation of fungi from the oak sections, however, was not carried out. A few ‘test’ fungi caused significant increases in length and weight of A. ostoyae rhizomorphs. Significant increases in the numbers of rhizomorph apices were observed rarely and increases in the numbers of

rhizomorphs were exceptional. In axenic cultures, in the control treatment, A. ostoyae either did not produce rhizomorphs or produced them very rarely. They were single, very short and unbranched. The inhibition of rhizomorph formation by A. ostoyae in excised roots and shoots as well as in soil, in sterile and unsterile conditions, was also observed by Rishbeth (1984; 1985a,b; 1988). Rishbeth observed also that the growth of A. ostoyae in excised oak roots and shoots could be erratic; the fungus often grew well in some lengths and did not grow in others. The ‘test’ fungi had significantly greater influence on rhizomorph production by A. gallica than A. ostoyae. In axenic cultures, in the control treatments, A. gallica produced rhizomorphs easily. They were monopodially branched, and usually consisted of one

226

H. Kwa´sna / Applied Soil Ecology 17 (2001) 215–227

long, central, thicker cord and many shorter, thinner branches developing along the central one. In the forest, A. gallica also produces rhizomorphs more easily and abundantly than A. ostoyae (Redfern, 1975; Rishbeth, 1984; Gregory, 1985). The observations of a stimulating effect of oak fungi on Armillaria rhizomorph production was not new, having been made earlier by Rishbeth (1972) who found very extensively ramifying rhizomorphs of Armillaria sp. (in the formerly named A. mellea complex) growing through oak tissue occupied by other fungi. Apart, however, from this general observation, Rishbeth (1972) did not study the effect of individual fungi on rhizomorph formation. It should be mentioned that, although the experiment gave positive results, they were based on yields of rhizomorphs produced mainly through the cut ends of sections of wood, rather than through the bark as occurs naturally. In the forest, rhizomorphs in their subcortical and subterranean forms grow in soil, on the surface of roots and trunks and beneath the bark from where the pathogen colonizes the wood. The stimulating effect of rhizosphere fungi on A. ostoyae rhizomorph formation was smaller compared to A. gallica. Considering, however, the very large increase in density of rhizosphere fungi on stump roots, it might be predicted that decaying oak stumps favour the growth of both pathogen. The situation is different when broadleaved tree stumps are still alive. Armillaria grows in such stumps very slowly. When the regrowth dies, often due to the increased shading by surrounding trees, Armillaria spreads easily and fast. The abundance of Armillaria rhizomorphs when they grow (i) in the presence of fungal secretions and (ii) on sterile laboratory media, rich in nutrients, e.g. malt extract agar, seems to suggest two mechanisms involved in the stimulation of rhizomorph production. A direct effect of fungi may be exerted by fungal liquid or volatile metabolites secreted to the habitat and acting as a growth-promoting substances. This thesis is supported by earlier findings, e.g. tryptophol (an indole-3-ethanol analogue), which is a major secondary metabolite produced by Z. moelleri, stimulated the growth of A. ostoyae (Kwa´sna and Łakomy, 1998) and abundant volatiles produced by Ceratocystis virescens (Davids) C. Moreau stimulated growth of Armillaria in vitro (Wargo and Harrington, 1991).

An indirect effect may be elicited by the participation of microfungi in the degradation of wood and in the supply of nutrients. The latter suggestion is supported by the abundance of rhizomorphs in substrates rich in organic matter, e.g. peat, pine bark and compost (Redfern, 1973; Rykowski, 1984). Rhizosphere fungi are usually studied as potential antagonists of Armillaria species (Peno et al., 1975). The basic concept of this biological control is that fungi incapable of causing disease occupy roots which otherwise would be invaded by Armillaria. The general opinion is that the extent to which Armillaria species become established in a stump is influenced by the amount of competition from other fungi (mainly Basidiomycotina) and, perhaps, especially by the rate at which this competition develops. Studies on the soil, root and rhizosphere microfungi as stimulants of the growth of pathogens, development of infection and increase of disease risk are exceptional and, so far, are not advanced but are continued (Kwa´sna and Łakomy, unpublished).

Acknowledgements I am grateful to Dr Piotr Łakomy (Department of Forest Pathology, Agricultural University, Pozna´n, Poland) for providing both Armillaria isolates and to Dr G.L. Bateman (Rothamsted Experimental Station, Harpenden, United Kingdom) for critically reviewing the manuscript and correcting its English. I gratefully appreciate the helpful comments of the referees. References Childs, L., Zeller, S.M., 1929. Observations on Armillaria root rot of orchard trees. Phytopathology 19, 869–873. Domsch, K.H., Gams, W., Anderson, T.H., 1980. Compendium of Soil Fungi. Academic Press, New York. Gregory, S.C., 1985. The use of potato tubers in pathogenicity studies of Armillaria isolates. Plant Pathol. 34, 41–48. Guillaumin, J.J., Mohammed, C., Anselmi, N., Courtecuisse, R., Gregory, S.C., Holdenrieder, O., Intini, M., Lung, B., Marxmüller, H., Morrison, D., Rishbeth, J., Termorshuizen, A.J., Tirro, A., Van Dam, B., 1993. Geographical distribution and ecology of the Armillaria species in western Europe. Eur. J. For. Pathol. 23, 321–341. Kwa´sna, H., 1996a. Mycobionta of birch roots and birch stump roots and their possible effect on the infection by Armillaria spp. (Romagn) Herink growth. Part I. Acta Mycol. 31, 101–110.

H. Kwa´sna / Applied Soil Ecology 17 (2001) 215–227 Kwa´sna, H., 1996b. Mycobionta of birch roots and birch stump roots and their possible effect on the infection by Armillaria spp. (Romagn) Herink growth. Part II. Acta Mycol. 31, 111– 122. Kwa´sna, H., 1997. Fungi on the surface of roots of Scots pine and its stumps and their effect on Heterobasidion annosum (Fr.) Bref and Armillaria ostoyae (Romagn.) Herink growth. Pol. Agric. Ann., S. E. 28 (1/2), 109–123. Kwa´sna, H., Łakomy, P., 1998. Stimulation of Armillaria ostoyae vegetative growth by tryptophol and rhizomorph formation by Zygorhynchus moelleri. Eur. J. For. Pathol. 28, 53–61. Łakomy, P., 1998. Monitoring huby korzeni i opie´nkowej zgnilizny korzeni w wybranych uprawach sosnowych Krainy Wielkopolsko-Pomorskiej (Monitoring of Heterobasidion annosum and Armillaria root and butt rot in a few Scots pine plantations in central and northern Poland). Roczn. A.R. w Poznaniu, Rozp. Nauk. 283, 1–82. Mohammed, C., Guillaumin, J.J., 1989. Competition phenomena between European species of Armillaria. In: Morrison, D.J., (Ed.), Proceedings of the 7th International Conference on Root and Butt Rots, I.U.F.R.O. 9–16 August 1988, B.C. Canada, pp. 447–457. Nirenberg, H., 1976. Untersuchungen uber die morphologische und biologische Differenzierung in der Fusarium-Section Liseola. Mitteilungen aus der Biologische Bundesanstalt für Land- und Forstwirtschafi. Berlin-Dahlem 169, 1–117. Peace, T.R., 1962. Pathology of Trees and Shrubs. Oxford University Press, Oxford. Peno, M., Veselinovi´c, N., Plavši´c, V., 1975. Prilog ispitivanju mogu´cnosti primeno bioloske borbe protiv Armillaria mellea pri rekonstrukciji degradiranih liˆsc´ arskih sˆuma (Possibilities of biological control of Armillaria mellea in the conversion of degraded hardwood stands). Zaš. Bil. 26, 187–197. Redfern, D.B., 1968. The ecology of Armillaria mellea in Britain. Biological Control Ann. Bot. 32, 293–300. Redfern, D.B., 1973. Growth and behaviour of Armillaria mellea rhizomorphs in soil. Trans. Br. Mycol. Soc. 61, 569–581. Redfern, D.B., 1975. The influence of food base on rhizomorph growth and pathogenicity of Armillaria mellea isolates. In: Bruehl, G.W. (Ed.), Biology and Control of Soil-Borne Plant Pathogens. St. Paul, The American Phytopathological Society, MN, pp. 69–73. Redfern D.B., Filip G.M., 1991. Inoculum and infection. In: Shaw C.G., Kile G.A. (Eds.), Armillaria root disease. Agricultural Handbook No. 691, Forest Service US Department of Agriculture,Washington, D.C., pp. 48–62. Rishbeth, J., 1972. The production of rhizomorphs by Armillaria mellea from stumps. Eur. J. For. Pathol. 2, 193–205.

227

Rishbeth, J., 1982. Species of Armillaria in southern England. Plant Pathol. 31, 9–17. Rishbeth, J., 1984. Pathogenicity tests for Armillaria. In: Kile, G.A., (Ed.), Proceedings of the 6th International Conference on Root and Butt Rot of Forest Trees 25–31 August 1983, International Union of Forestry Research Organizations, Melbourne, Victoria and Gympie, Australia, pp. 131–139. Rishbeth, J., 1985a. Infection cycle of Armillaria and host response. Eur. J. For. Pathol. 15, 332–341. Rishbeth, J., 1985b. Armillaria: resources and hosts. In: Moore, D., Casselton, L.A., Wood, D.A., et al. (Eds.), Developmental biology of higher fungi, Cambridge University Press, Cambridge, pp. 87–101. Rishbeth, J., 1988. Stump infection by Armillaria in first rotation conifers. Eur. J. For. Pathol. 18, 401–408. Rovira, A.D., 1965. Plant root exudates and their influence upon soil microorganisms. In: Baker, K.F., Snyder, W.C. (Eds.), Ecology of Soil-Borne Plant Pathogens. University of California Press, Berkeley, CA, pp. 107–186. Rykowski, K., 1984. Niektóre trofficzne uwarunkowania patogeniczno´sci Armillaria mellea (Vahl) Quel. w uprawach sosnowych (Some trophic factors in the pathogenicity of Armillaria mellea in Scots pine plantations), Vol. 640, Prace Instytutu Badawczego Le´snictwa (Papers of Polish Forest Research Institute, Warsaw), pp. 1–140. Shaw, III, C.G., Kile, G.A., 1991. Armillaria Rot Disease, Agriculture Handbook No. 691, USDA Forest Service, USDA, Washington, DC. Smith, W.H., 1976. Character and significance of forest tree root exudates. Ecology 57, 324–331. Wargo, P.M., Harrington, T.C., 1991. Host stress and susceptibility. In: Shaw, III, C.G., Kile, G.A., (Eds.), Armillaria root disease, Agriculture Handbook No. 691, USDA Forest Service, Washington, DC, pp. 88–101. Wicklow, D.T., Whittingham, W.F., 1974. Soil microfungal changes among the profiles of disturbed conifer–hardwood forests. Ecology 55, 3–16. ˙ Zółciak, A., 1999a. Identyfikacja gatunków grzybów z rodzaju Armillaria (Fr.:Fr.) Staude w Polsce (Identification of species of the Armillaria (Fr.:Fr.) Staube genus in Poland), Prace Instytutu Badawczego Le´snictwa (Papers of Polish Forest Research Institute, Warsaw), 888, 3–19. ˙ Zółciak, A., 1999b. Wystepowanie grzybów z rodzaju Armillaria (Fr.:Fr.) Staude w kompleksach le´snych w Polsce (The occurrence of species from the Armillaria (Fr.:Fr.) Staube genus in forests in Poland), Prace Instytutu Badawczego (Le´snictwa Papers of Polish Forest Research Institute, Warsaw), 890, 29–40.