Mycologia
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Services
Right arrow Similar articles in this journal
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Hietala, A. M.
Right arrow Articles by Sen, R.
Right arrow Search for Related Content
PubMed
Right arrow Articles by Hietala, A. M.
Right arrow Articles by Sen, R.
Agricola
Right arrow Articles by Hietala, A. M.
Right arrow Articles by Sen, R.
Mycologia, 95(2), 2003, pp. 239-250.
© 2003 by The Mycological Society of America

An unknown mechanism promotes somatic incompatibility in Ceratobasidium bicorne


Ari M. Hietala 1

     Skogforsk, Bioteknologi, Avd. 1, Høgskoleveien 12, 1432 Ås, Norway

Kari Korhonen

     Finnish Forest Research Institute, P.O. Box 18 (Jokiniemenkuja 1), FIN-01301 Vantaa, Finland

Robin Sen

     Division of General Microbiology, Department of Biosciences, P.O. Box 56 (Viikinkaari 9), FIN-00014 University of Helsinki, Finland

   ABSTRACT
TOP
 ABSTRACT
INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 LITERATURE CITED
 

Strains of Ceratobasidium bicorne (anamorph uninucleate Rhizoctonia), causing root dieback in nursery-grown conifer seedlings, were fruited in the laboratory and the pairing interactions among sibling, single-basidiospore progeny were investigated. No mating reactions were observed. Instead, a high frequency of somatic incompatibility was observed in progeny pairings, indicated by a killing reaction in hyphal anastomosis and by formation of a demarcation line. The F1 progeny also could be fruited, and the level of somatic incompatibility within the F2 progeny remained high, even if lower than in the F1 progeny. The interaction types in pairings within a family of progeny were similar in all respects to those between field isolates, indicating that the species is homothallic. The uninucleate condition of vegetative cells and the basidial characteristics would indicate homokaryotic fruiting, but the possibility of pseudohomothallism remains. We currently are not able to provide an explanation for the mechanism promoting somatic incompatibility in this species, but it seems likely that the classic heterogenic model of somatic incompatibility recognized in basidiomycetes is not applicable here. Alternative mechanisms are discussed.

Key words: basidiomycetes, homothallism


   INTRODUCTION
TOP
 ABSTRACT
 INTRODUCTION
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 LITERATURE CITED
 
The anamorph genus Rhizoctonia contains soil-inhabiting fungi with diverse taxonomic relationships. Vegetative mycelia of several basidiomycete genera, e.g., Thanatephorus Donk, Waitea Warcup and Talbot, Tulasnella J. Schröt. and Ceratobasidium Rogers, possess characteristics that fit the criteria described for Rhizoctonia (Stalpers and Andersen 1996Citation). Because laboratory induction of fruiting is needed to determine teleomorph affinity, identification of these fungi has been problematic and traditionally based on anamorphic characteristics.

The common phytopathogen Rhizoctonia solani Kühn [teleomorph Thanatephorus cucumeris (Frank) Donk] is by far the most studied species in the genus. Field isolates of R. solani have multinucleate vegetative cells and are regarded as heterokaryotic. Isolates are subdivided into anastomosis groups (AGs) based on hyphal anastomosis behavior. With few exceptions, anastomosis takes place only between members within a single AG. Currently, R. solani is known to consist of 13 anastomosis groups (AGs) (Carling 1996Citation, Carling et al 1999Citation). Numerous studies using a variety of genetic and biochemical markers have shown that these AGs represent genetically isolated populations (e.g., Vilgalys and Cubeta 1994Citation, Kuninaga et al 1997Citation). Therefore, R. solani is regarded as a species complex.

The genetic basis of hyphal anastomosis in Rhizoctonia has not been studied, but a somatic incompatibility system has been offered as an explanation for the phenomenon (e.g., Anderson 1982Citation, Adams 1996Citation). Somatically compatible isolates produce a hyphal interaction termed perfect fusion (C3 reaction; Carling 1996Citation). This involves the fusion of cell walls and mixing of cytoplasm between the paired isolates, which results in the formation of living hyphal bridges between the colonies. Such a reaction is typical in self pairings. Excluding pairings between isolates originating from the same field or plant tissues, perfect fusion is a rare event in non-self pairings. Anastomosis between somatically incompatible isolates results in a killing reaction (C2; Carling 1996Citation), in which anastomosing and adjacent cells die after cell-wall fusion, thus preventing development of cytoplasmic connections. This is a typical interaction type in non-self pairings within an anastomosis group.

Somatic incompatibility maintains individuality of confronting mycelia and usually prevents genetic exchange between them. In basidiomycetes, it typically occurs between secondary (heterokaryotic) mycelia and is regarded as a heterogenic system primarily controlled by nuclear genes; a difference at any of the corresponding loci leads to an incompatibility reaction (Worrall 1997Citation). An approach that has been used in several studies is the construction of pedigreed heterokaryons, in which one nucleus is kept constant in all mycelia. Such an approach permits the analysis of variation due to one set of nuclei in secondary mycelia in the absence of confounding variation from the conjugate nucleus. Detailed information concerning the somatic incompatibility system is lacking for most basidiomycetes, but several multiallelic loci have been suggested in Heterobasidion annosum (Fr.) Bref. (Hansen et al 1993Citation), a single locus in Phellinus gilvus (Schwein) Fr. (Rizzo et al 1995Citation), two loci in Armillaria ostoyae (Romag.) Herink. (Guillaumin 1998Citation), and three to four in Collybia fusipes (Bull. : Fr.) Quel. (Marçais et al 2000Citation). The genetic basis of somatic incompatibility has been well established in some ascomycetes; it is a heterogenic mechanism involving many genes. Eleven genes, including those at the mating type locus, control somatic incompatibility in Neurospora crassa Shear & Dodge (Perkins 1988Citation), nine in Podospora anserina (Cesati) Niessl (Bégueret et al 1994Citation) and eight in Aspergillus nidulans (Eidam) G. Winter (Croft and Dales 1984Citation). In contrast to the somatic incompatibility reactions that occur between secondary mycelia in basidiomycetes, those in the ascomycetes take place between homokaryons, the system preventing formation of heterokaryons between different individuals. Fully compatible reactions occur only between isolates with the same allele at each locus.

In R. solani, typical diagnostic markers enabling separation of homokaryons from heterokaryons are absent; both possess multinucleate cells with no clamp connections. The genetics of several R. solani AGs has been studied primarily by analysis of heterokaryotic tuft formation, which arise at the contact zone between the paired homokaryons. Currently, the genetics of only two AGs (AG 1 and AG 4) are understood to some extent; data have been presented showing that field isolates of these AGs are heterokaryotic and possess a distinctive bipolar mating system with multiple alleles (Adams and Butler 1982Citation, Anderson 1982Citation, Adams 1996Citation, Julian et al 1996Citation). Very little is known about the mating system of the remaining AGs. Evidence has been presented that mating processes (indicated by formation of heterokaryotic mycelial tufts between paired homokaryons) occur independently of somatic incompatibility processes in AG 1-IC (Julian et al 1996Citation).

A uninucleate Rhizoctonia species causing root dieback of conifer seedlings has been identified from forest nurseries in Finland and Norway (e.g., Venn et al 1986Citation, Hietala 1995Citation). On the basis of basidial characteristics and sequence analysis of the ITS region of ribosomal DNA, it was identified as Ceratobasidium bicorne J. Erikss. & Ryvarden, a species originally described as a parasite on mosses (Hietala et al 1994Citation, 2001Citation). Before these findings, the genus Ceratobasidium was considered binucleate in the vegetative stage. Almost nothing is known about the genetics of binucleate Ceratobasidium spp., which are divided into at least 21 AGs (see e.g., Sneh et al 1991Citation). Based on anastomosis reactions, field isolates of the uninucleate Ceratobasidium form a single AG, typically presenting a somatic incompatibility reaction when paired against each other (Hietala et al 1994Citation, Hietala 1995Citation, Lilja et al 1992Citation). Isolates of this fungus can be fruited readily under laboratory conditions (Hietala et al 1994Citation, Hietala 1997Citation). Attempts to investigate the sexual system indicated that the fungus is homothallic but a high degree of somatic incompatibility unexpectedly was observed in pairings between single-spore isolates. Here we present results from pairings between single-basidiospore isolates of the F1 and F2 progeny of this fungus.


   MATERIALS AND METHODS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
 LITERATURE CITED
 
Fungal strains and pairings – The field isolates of Ceratobasidium bicorne were obtained from conifer seedlings suffering from root dieback in forest nurseries. Source details are presented in Table I. Isolates were inoculated on potato dextrose agar (PDA) approx 5 cm apart and incubated in the dark at room temperature for two weeks until visual examination of the confrontation zone. For anastomosis observations, the paired isolates were inoculated 5 cm apart on 0.1% malt-extract agar (MEA). Pairings were incubated in the dark at room temperature until the hyphae of opposing colonies began to overlap. The confrontation zone was examined under a light microscope using magnifications 100–400x.


View this table:
[in this window]
[in a new window]
  		TABLE I. The field isolates of C. bicorne (uninucleate Rhizoctonia sp.) investigated

 
Fruiting – For fruiting, mycelial plugs from the margins of PDA grown colonies were inoculated into a Petri dish containing aseptically grown Scotch pine seedlings in distilled water (Hietala et al 1994Citation). The dishes were incubated on a laboratory bench under natural indirect light. After the development of hymenium, determined at 40x magnification, samples were taken for spore isolation and for nuclear observations. For single-spore isolations, water samples were spread on 1% malt-extract agar and incubated at room temperature in darkness. Germinating basidiospores were isolated under a microscope, using a modified Pasteur pipette (Korhonen and Hintikka 1980Citation), and transferred to PDA.

Interactions between field isolates and within F1 and F2 progeny – Field isolates were paired in all combinations on PDA and on 0.1% MEA as described above. Pairings were replicated (n = 3) on both media. Selected field isolates were fruited and single-spore isolations performed as described above. Randomly selected single-spore isolates from each progeny were paired with each other and the parent on PDA and on 0.1% MEA in all combinations. For control, hyphal-tip cell isolations (Korhonen and Hintikka 1980Citation) were made from 2-day-old mycelium of field isolates (250, 263, 264 and T8A) and paired in all combinations. As a further control, one single-cell isolate from 250 and 263 were fruited and the progeny were examined for somatic incompatibility reactions on PDA. To investigate the F2 progeny, randomly selected single-spore isolates from field isolates 263 and T8A were fruited and the progeny were screened for variation in pairings on PDA. In Isolate 263, the parent and selected F1 and F2 single-spore isolates also were paired on PDA.

The behavior of nuclei in vegetative hyphae growing in Petri dishes on 0.1% MEA was investigated with phase-contrast microscopy. Young colonies growing on MEA-coated slides also were stained with HCl-Giemsa (Wilson 1992Citation). To observe nuclear behavior in basidia, hymenial samples were taken from Isolate 263 and stained with HCl-Giemsa or DAPI, following the procedure described by Cooke et al (1987)Citation.


   RESULTS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
DISCUSSION
 LITERATURE CITED
 
Somatic incompatibility between field isolates – In all self pairings, the colonies merged on PDA medium and showed a perfect hyphal fusion in microscopic studies on malt-extract agar, indicating somatic compatibility (Table II). With two exceptions, all the pairings between field isolates originating from different nurseries showed a somatic incompatibility reaction. This reaction was characterized visually by the presence of a 1–2-mm wide demarcation line with sparse aerial hyphae between the confronting colonies (Fig. 1) and microscopically by the presence of the killing reaction. The number of dying cells was 1–3 on either side of anastomosing cells. Formation of mycelial tufts at the interaction zone was not observed in any pairing. With few exceptions, identical anastomosis reaction categories were detected in the three replicate pairings performed on both media.


View this table:
[in this window]
[in a new window]
  		TABLE II. Pairing reactions between field isolates of C. bicorne (uninucleate Rhizoctonia sp.) on potato dextrose agar and 0.1% malt extract agar (n = 3)

 


View larger version (81K):
[in this window]
[in a new window]
  		 FIG. 1. Somatic incompatibility reactions in representative parental and F1 progeny pairings of C. bicorne (Strain 263). The upper row shows parental and F1 progeny self pairings and the lower row two parental x F1 pairings and two F1 progeny non-self pairings. Note the presence of a demarcation line (arrowed) in somatically incompatible pairings

 
Genetic variation within F1 progeny – All of the field isolates included in the fruiting experiments produced basidia and basidiospores. However, the vast majority of basidia lacked sterigmata or, when sterigmata were produced, spore formation was inhibited. The number of sterigmata ranged from one to four, two being the most common case.

Mycelial interactions in pairings between single-basidiospore isolates were similar to the pairings between field isolates. Somatic incompatibility, expressed by the formation of a demarcation line (Fig. 1) and the presence of a killing reaction, was frequent in pairings between progeny members. Its occurrence showed no distinct pattern. Within the F1 progeny of the field isolate 260, 81 and 83 non-self pairings out of 90 showed a demarcation line on PDA and a killing reaction on 0.1% MEA, respectively (Table III). The corresponding frequencies within the F1 progeny of isolate 264 were 77 and 84 out of 90 (Table IV); within the F1 progeny of isolate 250, they were 24 and 22 out of 28 (data not shown); within the F1 progeny of 83-111/1N, they were 23 and 18 out of 28 (Table V). In nine non-self pairings within the F1 progeny of 83-111/1N, both killing reaction and perfect fusion were observed in the same replicate. Within the F1 progeny of field isolates T8A and 263, the rate of incompatibility in non-self pairings on PDA was 29/36 and 31/45, respectively (data not shown). With one exception (isolates 260 vs 260/7; Table III), a demarcation line was produced in pairings between the parental isolate and its progeny member.


View this table:
[in this window]
[in a new window]
  		TABLE III. Pairing reactions within the progeny of parent isolate 260 on potato dextrose agar and on 0.1% malt extract agar (n = 2)

 

View this table:
[in this window]
[in a new window]
  		TABLE IV. Pairing reactions within the progeny of parent isolate 264 on potato dextrose agar and on 0.1% malt extract agar (n = 2)

 

View this table:
[in this window]
[in a new window]
  		TABLE V. Pairing reactions within the F1 progeny of parent isolate 83/111/1N on potato dextrose agar and on 0.1% malt extract agar (n = 1)

 
With two exceptions, the single-cell cultures isolated from hyphal tips of a colony were identical genetically, thus differing from single-basidiospore isolates. Paired single-cell isolates from the strains 250, 263 and 264 fused (10 from each strain) without formation of a demarcation line. However, two pairings out of the 45 non-self pairings between single-cell isolates from Strain T8A formed a demarcation line indicating somatic incompatibility (data not shown). Uninucleate single-cell isolates obtained from 250 and 263 were fruited; in these progeny, 23 and 28 non-self pairings out of 28, respectively, resulted in the formation of a demarcation line on PDA (data not shown).

A point of interest was the occasional detection of bi- and trinucleate progeny. These were observed in most progeny, the proportion being usually less than 10%. In cultural appearance, these differed from the uninucleate progeny by having fluffy aerial hyphae and a slow growth rate (0.01–0.15 mm h-1 versus the normal 0.63–0.34 mm h-1). In preliminary pairings, no interactions (e.g., formation of mycelial tufts or a demarcation line) were observed, even when they were paired with the normal uninucleate offspring. Attempts to isolate uninucleate colonies from these, using the protoplasting technique (Phillips 1993Citation), failed. For these reasons, the bi- and trinucleate colonies were excluded from further tests.

Genetic variation within F2 progeny – All of the F1 single-spore isolates included in the fruiting test produced basidia. As in parental fruiting, the vast majority of basidia lacked sterigmata or spores. Varying degrees of somatic incompatibility, expressed as the formation of a demarcation line in non-self pairings, was observed also in the F2 progeny pairings. However, the frequency of somatic incompatibility generally was lower than within the F1 progeny. In three F2 progeny (parents T8A/8, T8A/10, and T8A/13), a demarcation line was formed in 19/28 (data not shown), 15/36 (data not shown), and 11/28 (Table VI) of the non-self pairings, respectively. In these series, a demarcation line between the F1 parent and a progeny member was formed in 3/8, 8/9 and 7/8 cases, respectively. In the F2 progeny of the single-spore isolate 263/5, a demarcation line was formed in 17 out of 45 non-self pairings (data not shown). When field Isolate 263 was paired with randomly selected members of its F1 and F2 progeny, the result was always an incompatible reaction (Table VII). In contrast, F1 Isolate 263/5 produced an incompatible reaction in only four cases out of eight when it was paired with its progeny.


View this table:
[in this window]
[in a new window]
  		TABLE VI. Somatic incompatibility reactions within the progeny of F1 single-spore isolate T8A/13 on potato dextrose agar (n = 1)

 

View this table:
[in this window]
[in a new window]
  		TABLE VII. Somatic incompatibility reactions within and between F1 and F2 progeny of the field isolate 263 on potato dextrose agar (n = 1)

 
Summary of the pairing results – The pairing data are summarized in Table VIII. In pairings between the field isolate and F1 and F2 progeny, the frequency of somatic incompatibility ranged between 90–100%. Within the F1 and F2 progeny, the levels of somatic incompatibility ranged between 64–92% and 39–78%, respectively.


View this table:
[in this window]
[in a new window]
  		TABLE VIII. Summary of the frequenciesa of somatic incompatibility in non-self pairings

 
Nuclear behavior in hyphae and basidia – Although most cells in vegetative hyphae were uninucleate, cells with two nuclei were not rare and occasionally cells with three or four nuclei were observed. A striking phenomenon was the rarity of nuclear divisions in mycelia stained with HCl-Giemsa. Among thousands of cells checked in actively growing colonies, no distinct division figure was seen.

A vast majority of hymenial cells were uninucleate, but binucleate cells also were observed. The cells from which basidia developed were invariably uninucleate. In basidia, the nuclear number was highly variable and no distinct pattern could be observed (Table IX). No clear meiotic divisions were observed in any case. In basidia where basidiospores had not developed, the nuclear numbers ranged from one to seven. In basidia with basidiospores, the nuclear numbers ranged from two to seven, when counting also the nuclei in the basidiospores. The most common nuclear number in basidia was four in both cases (Fig. 2). In basidiospores attached to sterigmata, the nuclear number ranged from one to three.


View this table:
[in this window]
[in a new window]
  		TABLE IX. Observations on the nuclear number in basidia and basidiospores in strain 263

 


View larger version (114K):
[in this window]
[in a new window]
  		 FIG. 2. A developing, four-nucleate basidium of Strain 263 stained with HCl-Giemsa. Note the uninucleate condition of the cell, giving rise to basidia (scalebar = 15 µm)

 

   DISCUSSION
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
LITERATURE CITED
 
All the single-spore isolates tested were self-fertile, indicating that Ceratobasidium bicorne is homothallic. No mating reactions, e.g., tuft formation, were observed in this study. However, tuft formation does not always take place in compatible matings within R. solani (Thanatephorus cucumeris) complex. A good example is R. solani AG 4, where homokaryons generally do not form tufts with their parental isolates or other heterokaryotic field isolates. Synthesized heterokaryons usually give barrage or neutral reactions with the contributing homokaryons (e.g., Garza-Chapa and Anderson 1966Citation, Vest and Anderson 1968Citation). In this study, mycelial interactions in F1 and F2 progeny pairings in all respects were similar to those between field isolates. Together with the self-fertility of the basidiospores, this would imply that the progeny share the same karyotic stage as field isolates.

Regarding the applicability of the classic heterogenic model of somatic incompatibility here, the key question is the ploidy level of the species. In this study, unexpectedly, no nuclear division figures were observed in HCl-Giemsa-stained vegetative hyphae, despite a relatively large sampling. This is surprising when compared with studies on other basidiomycetes, e.g., Armillaria (Korhonen and Hintikka 1974Citation) and suggests that the nuclear division takes place rapidly. In accordance, no meiotic divisions were observed in the basidia. However, the most common number of nuclei in basidia was four, so the possibility of meiosis remains, due to limited observations. In any case, there is an essential difference between nuclear divisions that take place in the vegetative hyphae and in the basidium: The former produced somatically compatible cells, the latter mostly incompatible ones. Commonly only one or two sterigmata were formed per basidium in this study. The basidia retained features of homokaryotic fruiting; basidial development was interrupted in most cases, and these cells did not produce sterigmata or basidiospores. The low production of basidiospores also has been noted in other studies on homokaryotic fruiting of R. solani (e.g., Adams and Butler 1982Citation, Julian et al 1997Citation). Julian et al (1997)Citation also recorded the nuclear behavior in the basidia; in some homothallic isolates, meiotic divisions were observed, whereas in others meiosis was interrupted. Regarding the karyotic stage of the now-studied cultures, the data are limited and do not exclude the possibility of diploidy. The basidial characteristics would suggest haploid homokaryotic fruiting, but in this case it is not possible to relate the progeny pairing data to the classic heterogenic model of somatic incompatibility.

The heterogenic model of somatic incompatibility would be applicable to C. bicorne only if the species was diploid. In this case, the species would be pseudohomothallic (syn. secondarily homothallic). Among pseudohomothallic basidiomycetes, Agaricus bisporus (Lange) Imbach var. bisporus is the best known. In this species, two nuclei with compatible mating types typically migrate into each basidiospore; the mycelium originating from a single-basidiospore is thus heterokaryotic and able to produce fruit bodies without mating. Royse and May (1982)Citation and Summerbell et al (1989)Citation compared the level of heterozygosity in the parents and offspring of A. bisporus using isoenzymes and RFLP markers, respectively. In the first study, the loss of heterozygosity in the progeny was at maximum 6.6%, depending on the loci studied, and in the second 1.9%, indicating that in general the frequency of crossing-over is relatively low in this species. Since crossing-over is restricted, the level of heterozygosity in the parent is efficiently maintained in the offspring. Construction of pedigreed heterokaryons is complicated in secondarily homothallic species, and the mechanism controlling somatic incompatibility has not been studied in A. bisporus. However, roughly 10–20% of sibling pairings in this species, based on incidental occasions, might show behavior consistent with some form of somatic incompatibility (Richard W. Kerrigan, pers comm). This would fit well with a somatic incompatibility system controlled by 3–5 loci if applying the crossing-over frequencies mentioned above.

If C. bicorne represents a pseudohomothallic species, it would not be haploid, and meiotic divisions followed by nuclear fusion in the basidia would be expected to occur. The results indicate that the degree of somatic incompatibility is higher in F1 than in F2 progeny. The heterogenic model of somatic incompatibility could explain the high degree of variation observed in F1 and the reduction in F2 in case the parent isolates were heterozygous at several loci controlling somatic incompatibility, and the frequency of crossing-over were sufficiently high to allow efficient fixation of alleles. In F1 progeny pairings, the highest frequency of somatic incompatibility was now 92%. The complexities of differing unknown recombination rates (crossing-over) for an unknown number of loci argue against presenting a model here. However, if the parental isolate was heterozygous at 10 loci controlling somatic incompatibility, each having a crossing-over frequency of 20%, somatic incompatibility would reach a similar level (89%) in the progeny. If this was indeed the case, C. bicorne would differ considerably from A. bisporus var. bisporus in recombination rate and, regarding the number of loci involved, it would show more similarity to ascomycetes. In this kind of a system, a puzzling question also would be how a high potential for incompatibility could be maintained in wild isolates in the absence of mating.

Flentje and Stretton (1964)Citation examined anastomosis in the progeny of one homothallic R. solani isolate; perfect fusion was observed in all pairing combinations. Unfortunately Julian et al (1997)Citation did not include anastomosis testing in their study on homokaryotic fruiting in R. solani. In homothallic populations of the basidiomycete Stereum sanguinolentum (Albertini & Schwein. : Fr.) Fr., normal meiosis takes place in the basidia but all the siblings within an offspring are somatically compatible, whereas non-sibling interactions typically result in an incompatibility reaction (Ainsworth 1987Citation). There are several other examples of homothallic basidiomycetes that produce genetically identical progeny. If C. bicorne is haploid, it is not a typical homothallic species.

The cassette system first found in yeasts could offer one alternative explanation for the now reported variation. In Saccharomyces cerevisiae Hansen, each haploid genome contains both the a and {alpha} mating type gene, and normally one of them is transcribed with its interaction with the MAT locus while the other is silent (e.g., Perkins 1987Citation). Occasionally, a new copy of the silent-mating type gene is made, the other one is removed from the MAT locus and the new type is transcribed. Mating type switching also has been shown in another ascomycetous genus, Ceratocystis (Witthuhn et al 2000Citation). In addition, Labarère and Noël (1992)Citation reported mating type switching in the tetrapolar basidiomycete Agrocybe aegerita (Brig.) Singer. This study showed that some haploid homokaryotic strains spontaneously can switch their mating specificities at the two unlinked A and B mating type factors. This event caused the dikaryotization of homokaryotic mycelia without plasmogamy and led to "pseudo-homokaryotic fruiting", in which parental and switched mating types segregated meiotically as Mendelian markers. As far as we know, there are no reports on a cassette system controlling somatic incompatibility in any filamentous fungus. If a cassette system was involved in Ceratobasidium bicorne, several factors affecting somatic incompatibility should be expected to have a cassette system and the switching would occur, as in A. aegerita, in relation to fruiting. If this is the case, it should be possible to recover parental factors in the F2 progeny as shown by Labarère and Noël (1992)Citation. Further pairings (parent x F2 and F1parent x F3) will be needed to test this hypothesis.

Recently it was shown that prions might be involved in controlling individualism in fungi. One of the loci controlling vegetative incompatibility in the ascomycete Podospora anserina produces a protein that behaves as a prion analog (Coustou et al 1997Citation). This protein can catalyze its own alteration to an abnormal form (prion form) and this post-translational alteration changes the vegetative incompatibility group of the strain in question. Of interest in this study, two single-cell isolates from parental Strain T8A showed somatic parental incompatibility. There were also cases in which both perfect fusion and killing reaction were observed in the same replicate. The latter observation was noted also, e.g., by Flentje and Stretton (1964)Citation and by Julian et al (1996)Citation, when studying anastomosis behavior of R. solani. These observations show that there is plasticity in the occurrence of incompatibility reaction; it could result from events analogous to the post-translational alteration case described above.

In addition to nuclear genes, mitochondria have been implicated in somatic compatibility: In Coprinus cinereus (Schaeff. : Fr.) S.F. Gray sensu Konr. different mitochondria in the same nuclear background led to strong incompatibility reaction in one of eight cases tested (May 1988Citation). McCabe et al (1999a)Citation performed single hyphal isolations from AG 4 strains of R. solani and a low percentage of these were somatically incompatible with the parent. The mechanism behind this was not revealed, but the authors speculated that the parents might have contained more than two nuclear types, or they might be a mosaic in regard to mitochondria. Regarding this study, this hypothesis seems the least likely because the mitochondrial DNA type should be expected to be uniform within each of the family lineages.

The presence of a strongly promoted somatic incompatibility system within C. bicorne also raises a question of its ecological implications. Hyphal fusion with compatible strains provides larger nutrient resources and faster reproduction; in fact, this is regarded as a great benefit of homothallism. The size structure of some fungal populations suggests that survival is low at small ramets (Holmer and Stenlid 1991Citation, Worrall 1994Citation). In this respect, high levels of somatic incompatibility within the progeny of this species cannot be regarded as beneficial. On the other hand, fungal viruses are transmitted through cytoplasmic bridges and somatic incompatibility systems create a significant barrier to this movement (e.g., McCabe et al 1999bCitation). dsRNA viruses have been reported in R. solani, both in association with reduced virulence and with increased virulence (reviewed by Rubio et al 1996Citation). The occurrence of viruses has not been studied in the C. bicorne.

In conclusion, the uninucleate condition and the basidial characteristics would imply that C. bicorne is homothallic. The element that does not fit this picture is the high frequency of somatic incompatibility among progeny. We currently are not able to provide a satisfactory explanation for this phenomenon, but as discussed above, several mechanisms could account for at least some of the features now reported. In further work, molecular tools should be employed to characterize nuclear DNA and cytoplasmic elements and events separately. The genetic basis of somatic incompatibility is still known only for a handful of basidiomycetes and generalizations should be avoided. Due to the uninucleate nature of this species, it should be a convenient candidate to study the factors controlling somatic incompatibility in Fungi.


   ACKNOWLEDGMENTS
 
The first author is grateful to the Department of Applied Biology, University of Helsinki, for providing facilities for the study. This work was financially supported by the Natural Resources Research Foundation of Finland (Suomen Luonnonvarain Tutkimussäätiö) and the Academy of Finland (FIBRE). Carl Gunnar Fossdal, Richard W. Kerrigan and one anonymous reviewer are thanked for comments improving the manuscript.


   FOOTNOTES
 
1 Corresponding author. ari.hietala{at}skogforsk.no Back

Accepted for publication April 11, 2002.


   LITERATURE CITED
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 LITERATURE CITED
 
Adams GC., 1996 Genetics of Rhizoctonia species. In: Sneh B, Jabaji-Hare S, Neate S, Dijst G, eds. Rhizoctonia species: taxonomy, molecular biology, ecology, pathology and disease control. Dordrecht, The Netherlands: Kluwer Academic Publishers. p 101–116

———, Butler EE., 1982 A reinterpretation of the sexuality of Thanatephorus cucumeris anastomosis group four. Mycologia 74:793-800

Ainsworth AM., 1987 Occurrence and interactions of outcrossing and non-outcrossing populations in Stereum, Phanerochate and Coniophora. In: Brasier ADM, Brasier CM, Moore D, eds. Evolutionary biology of the fungi. Cambridge, U.K: Cambridge University Press. p 285–299

Anderson NA., 1982 The genetics and pathology of Rhizoctonia solani. Ann Rev Phytop 20:329-347

Bégueret J, Turcq B, Clavé C., 1994 Vegetative incompatibility in filamentous fungi: het genes begin to talk. Trends in Genetics 10:441-446[Medline]

Carling DE., 1996 Grouping in Rhizoctonia solani by hyphal anastomosis reaction. In: Sneh B, Jabaji-Hare S, Neate S, Dijst G, eds. Rhizoctonia species: taxonomy, molecular biology, ecology, pathology and disease control. Dordrecht, The Netherlands: Kluwer Academic Publishers. p 37–47

———, Pope EJ, Brainard KA, Carter DA., 1999 Characterization of mycorrhizal isolates of Rhizoctonia solani from an orchid, including AG-12, a new anastomosis group. Phytopathology 89:942-946

Cooke JC, Gemma JN, Koske RE., 1987 Observations of nuclei in vesicular-arbuscular mycorrhizal fungi. Mycologia 79:331-333

Coustou V, Deleu C, Saupe S, Bégueret J., 1997 The protein product of the het-s heterokaryon incompatibility gene of the fungus Podospora anserina behaves as a prion analog. Proc Natl Acad Sci 94:9773-9778[Abstract/Free Full Text]

Croft JH, Dales RBG., 1984 Mycelial interactions and mitochondrial inheritance in Aspergillus. In: Jennings DH, Rayner ADM, eds. The ecology and physiology of the fungal mycelium. London, New York: Cambridge Univ. Press. p 433–450

Flentje NT, Stretton HM., 1964 Mechanisms of variation in Thanatephorus cucumeris and T. praticolus. Aust J Biol Sci 17:686-704

Garza-Chapa R, Anderson NA., 1966 Behavior of single-basidiospore isolates and heterokaryons of Rhizoctonia solani from flax. Phytopathology 56:1260-1268

Guillaumin JJ., 1998 Genetic control of somatic incompatibility in Armillaria ostoyae—a preliminary analysis. In: Delatour C, Guillaumin JJ, Lung-Escarmant B, Marçais B, eds. Proceedings of the ninth International Conference on Root and Butt Rots of Forest Trees. France, Colloques de l'INRA no. 89. p 31–38

Hansen EM, Stenlid J, Johansson M., 1993 Genetic control of somatic incompatibility in the root-rotting basidiomycete Heterobasidion annosum. Mycol Res 97:1229-1233

Hietala AM, Sen R, Lilja A., 1994 Anamorphic and teleomorphic characteristics of a uninucleate Rhizoctonia sp. isolated from the roots of nursery grown conifer seedlings. Mycol Res 98:1044-1050

———. 1995 Uni- and binucleate Rhizoctonia spp. coexisting in the roots of Norway spruce seedlings suffering from root dieback. Eur J For Path 25:136-144

———. 1997 The mode of infection of a pathogenic uninucleate Rhizoctonia sp. in conifer seedling roots. Can J For Res 27:471-480

———, Vahala J, Hantula J., 2001 Molecular evidence suggests that Ceratobasidium bicorne has an anamorph known as a conifer pathogen. Mycol Res 105:555-562

Holmer L, Stenlid J., 1991 Population structure and mating system in Marasmius androsaceus Fr. New Phytol 119:307-314

Julian MC, Debets F, Keijer J., 1996 Independence of sexual and vegetative compatibility mechanisms of Thanatephorus cucumeris (Rhizoctonia solani) anastomosis Group 1. Phytopathology 86:566-574

———, Dullemans AM, van Silfhout CH, Keijer J., 1997 Nuclear behavior in homokaryotic and heterokaryotic fruiting of Thanatephorus cucumeris (Rhizoctonia solani) anastomosis Group 1, Subgroup IC. Mycologia 89:361-374

Korhonen K, Hintikka V., 1974 Cytological evidence for somatic diploidization in dikaryotic cells of Armillariella mellea. Arch Microbiol 95:187-192

———, ———. 1980 Simple isolation and inoculation methods for fungal cultures. Karstenia 20:19-22

Kuninaga S, Natsuaki T, Takeuchi T, Yokosawa R., 1997 Sequence variation of the rDNA ITS regions within and between anastomosis groups in Rhizoctonia solani. Curr Genet 32:237-243[Medline]

Labarère J, Noël T., 1992 Mating type switching in the tetrapolar basidiomycete Agrocybe aegerita. Genetics 131:307-319[Abstract]

Lilja A, Lilja S, Poteri M, Ziren L., 1992 Conifer seedling root fungi and root dieback in Finnish nurseries. Scand J For Res 7:547-556

Marçais B, Caël O, Delatour C., 2000 Genetics of somatic incompatibility in Collybia fusipes. Mycol Res 104:304-310

May G., 1988 Somatic incompatibility and individualism in the coprophilous basidiomycete Coprinus cinereus. Trans Br Myc Soc 91:443-451

McCabe PM, Gallacher MP, Deacon JW., 1999a Evidence for segregation of somatic incompatibility during hyphal tip subculture of Rhizoctonia solani AG 4. Mycol Res 103:1323-1331

———, Pfeiffer P, van Alfen NK., 1999b The influence of dsRNA viruses on the biology of plant pathogenic fungi. Trends in Microbiology 9:377-381

Perkins DD., 1987 Mating type switching in filamentous ascomycetes. Genetics 115:215-216[Free Full Text]

———. 1988 Main features of vegetative incompatibility in Neurospora. Fungal Genetics Newsletter 35:44-46

Phillips AJL., 1993 The use of protoplasts for the preparation of homokaryons from heterokaryotic isolates of R. solani. Mycol Res 97:456-460

Rizzo DM, Rentmeester RM, Burdsall HH., 1995 Sexuality and somatic incompatibility in Phellinus gilvus. Mycologia 87:805-820

Royse DJ, May B., 1982 Use of isozyme variation to identify genotypic classes of Agaricus brunnescens. Mycologia 74:93-102

Rubio V, Tavantzis SM, Lakshman DK., 1996 Extrachromosomal elements and degree of pathogenicity in Rhizoctonia solani. In: Sneh B, Jabaji-Hare S, Neate S, Dijst G, eds. Rhizoctonia species: taxonomy, molecular biology, ecology, pathology and disease control. Dordrecht, The Netherlands: Kluwer Academic Publishers. p 127–138

Sneh B, Burbee L, Ogoshi A., 1991 Identification of Rhizoctonia species. St. Paul, Minnesota: The American Phytopathological Society Press. 133 p

Stalpers JA, Andersen TF., 1996 A synopsis of the taxonomy of teleomorphs connected with Rhizoctonia s.l. In: Sneh B, Jabaji-Hare S, Neate S, Dijst G, eds. Rhizoctonia species: taxonomy, molecular biology, ecology, pathology and disease control. Dordrecht, The Netherlands: Kluwer Academic Publishers. p 49–63

Summerbell RC, Castle AJ, Horgen PA, Anderson JB., 1989 Inheritance of restriction fragment length polymorphisms in Agaricus brunnescens. Genetics 123:293-300[Abstract/Free Full Text]

Venn K, Sandvik M, Langerud BR., 1986 Nursery routines, growth media and pathogens affect growth and root dieback in Norway spruce seedlings. Meddel Norsk Inst Skogforsk 39:313-328

Vest G, Anderson NA., 1968 Studies on heterokaryosis and virulence of Rhizoctonia solani from flax. Phytopathology 58:802-807

Vilgalys R, Cubeta MA., 1994 Molecular systematics and population biology of Rhizoctonia. Ann Rev Phytop 32:135-155

Wilson AD., 1992 A versatile giemsa protocol for permanent nuclear staining of fungi. Mycologia 84:585-588

Witthuhn RC, Harrington TC, Wingfield BD, Steimel JP, Wingfield MJ., 2000 Deletion of the MAT-2 mating-type gene during unidirectional mating-type swithing in Ceratocystis. Curr Genet 38:48-52[Medline]

Worrall JJ., 1994 Population structure of Armillaria species in several forest types. Mycologia 86:401-407

———. 1997 Somatic incompatibility in basidiomycetes. Mycologia 89:24-36




This article has been cited by other articles:


Home page
 MycologiaHome page
T. Toda and M. Hyakumachi
Heterokaryon formation in Thanatephorus cucumeris anastomosis group 2-2 IV.
Mycologia, September 1, 2006; 98(5): 726 - 736.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Services
Right arrow Similar articles in this journal
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Hietala, A. M.
Right arrow Articles by Sen, R.
Right arrow Search for Related Content
PubMed
Right arrow Articles by Hietala, A. M.
Right arrow Articles by Sen, R.
Agricola
Right arrow Articles by Hietala, A. M.
Right arrow Articles by Sen, R.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS