Ectomycorrhiza formation between Pseudotsuga menziesii seedling roots and monokaryotic and dikaryotic isolates of Laccaria bicolor

Seedling roots of Pseudotsuga menziesii were colonized with three monokaryotic isolates and one dikaryotic isolate of Laccaria bicolor to assess the effect of fungal genotype on ectomycorrhiza formation. Ectomycorrhizas resulting from colonization by the dikaryotic isolate had a multilayered mantle and a cortical Hartig net. One monokaryotic isolate (ss7) formed ectomycorrhizas comparable in anatomy to those induced by the dikaryotic isolate. Two other monokaryotic isolates (ss5, ss1) failed to form mantles or Hartig nets. Roots colonized by these isolates developed characteristics indicating an incompatible reaction.     

Characterization of protoplasts prepared from the edible fungus, Stropharia rugoso-annulata

Protoplast preparation and regeneration conditions of the edible fungus, Stropharia rugoso-annulata Farlow apud Murrill were studied, and the regenerated progenies were characterized in this study. The optimal condition for protoplast preparation was incubation of young mycelia with gentle shaking in 1.5%(w/v) Lywallzyme at 30 °C for 3 h. PGPM (potato/glucose/peptone/mannitol) was the most suitable regeneration medium. Served as osmotic stabilizer, sugars (mannitol and sucrose) were better than inorganic salts (MgSO₄) for clone development and growth. Pre-incubation of protoplasts in liquid regeneration medium resulted in a significantly decreased regeneration rate. Both dikaryotic isolates and monokaryotic isolates could be identified from protoplast-regenerated progenies, with a much higher frequency of monokaryotic isolates identified from the early-developed and fast-growing regenerated clones. Two parental mating types were also identified from protoplasted monokaryotic isolates, but not segregated by 1:1. The mycelial growth rate of protoplasted monokaryotic isolates showed a mating type-dependent model when cultured at different incubation temperatures and pH values, with A2B2 mating type monokaryotic isolates growing faster than those of A1B1 mating type monokaryotic isolates.     

<Emphasis Type="Italic">Coprinopsis austrophlyctidospora</Emphasis> sp. nov., an agaric ammonia fungus from Southern Hemisphere plantations and natural forests

A new ammonia fungus, Coprinopsis austrophlyctidospora, is described from Nothofagus and Pinus forests in New Zealand and from Eucalyptus forest in Australia. In ecology and macro-morphology, this species is similar to the Northern Hemisphere species C. phlyctidospora, but the new species differs in morphological characters of the basidiospore, i.e., in having a plage, more minute surface warts, and the smaller size of the basidiospore.     

Nuclear selection in oidium formation from dikaryotic mycelia ofFlammulina velutipes

The effect of nuclear dominance in monokaryotic oidium formation from dikaryotic mycelia in a tetrapolar basidiomycete,Flammulina velutipes, was examined. A total of 46 monokaryotic stocks were used to produce 194 hybrid dikaryotic stocks by crossing. The proportion of homokaryons among the oidium isolates from dikaryotic mycelia was over 95%. The staining of nuclei of oidia with propidium iodide showed that over 90% of oidia were monokaryotic and suggested that these oidia had single haploid nuclei at the G1 stage. The monokaryotic oidium isolates from hybrid dikaryons were backrossed to parental monokaryotic stocks. Although most of the monokaryotic oidium isolates (except for those from 17 hybrid dikaryons from a total of 194 test stocks) showed nuclear types similar to only one of the parental stocks, the process seems to produce essentially the split nuclear type composition. Therefore, the monokaryotization in oidium formation from dikaryotic mycelia essentially involves the process of nuclear selection. The two separate results of hierarchies of relative dominance among two nuclei of the parental dikaryons in the monokaryotic oidium formation by grouping with incompatibility factor compositions were determined. Only a few discrepancies were found in the hierarchies between the two specific nuclear compositions of hybrid dikaryons.     

Nuclear selection in monokaryotic oidium formation from dikaryotic mycelia in a basidiomycete,Pholiota nameko

The effect of nuclear dominance in monokaryotic oidium formation from dikaryotic mycelia inPholiota nameko was examined. Over 90% of oidium isolates from dikaryotic mycelia were monokaryotic. Although only one parental nuclear type was recovered from an average of about 80% in these isolates, the nuclear selection process in oidium formation seems essentially to produce split nuclear type composition in oidium products. The hierarchy of relative dominance among the nuclear types of the parental dikaryons in monokaryotic oidium formation was determined. The two hierarchies in nuclear selection between monokaryotic oidium formation and monokaryotic mycelium formation coincided at a level of at least 75%.     

Phylogeographic analysis of the brown alga Cutleria multifida (Tilopteridales,Phaeophyceae) suggests a complicated introduction history

In depth genetic comparisons of populations of Cutleria multifida (Tilopteridales, Phaeophyceae) collected from Europe, the northwestern Pacific Ocean, Australia and New Zealand using the DNA sequences of four gene regions (the mitochondrial cox2 and cox3 genes, the intergeneric spacer region adjacent to cox3, and the open reading frame) suggested that the northwestern European and Japanese populations were considerably greater in terms of their genetic divergence than Mediterranean, Australian or New Zealand populations. The haplotypes of the populations in northwestern European (distribution range including the type locality, seven haplotypes) and Japanese populations (seven haplotypes) were unique except for one shared haplotype. There were weak but positive correlations between the geographical distance and the genetic divergence among northwestern European and Japanese populations. Moreover, both female and male gametophytes occurred in eight of the nine Japanese localities, suggesting Japanese populations showed normal sexual heteromorphic life history of the species. In light of these results, it appears that Japanese populations were native to the area despite earlier hypothesis. In contrast, Australian and New Zealand populations were composed of only one haplotype that is very close to those found in northwestern Europe and Japan, suggesting a recent introduction history from Europe (or from northeastern Asia via Europe) by ship transport to Australia and New Zealand. The Mediterranean populations included two haplotypes identical to those found in northwestern Europe and Japan, and it is suggestive of transoceanic introductions of some populations between Mediterranean and Japanese coasts.     

Alternation of nuclear phase in the filamentous basidiomycete,Helicobasidium mompa

Variation in the number of nuclei and cellular ploidy were observed in eight strains ofHelicobasidium mompa. The basidiospores, single-spore isolates and field-isolated strains were all dikaryons. The cellular ploidy, which was assessed by analyzing the fluorescence emitted by DAPI-stained nuclei, was unstable: monokaryotic strains derived from the original dikaryotic strains by successive subcultures were mainly tetraploid, although the original dikaryon was in most cases diploid. On the other hand, a dikaryotic strain derived by treatment with benomyl was haploid. These results suggest that diploid dikaryon is a normal nuclear phase ofH. mompa in nature, and the alternation of ploidy may be due to a feature of the mating system of this fungus.     

BIOGEOGRAPHY OF ASEXUAL AND SEXUAL REPRODUCTION IN CALOGLOSSA (DELESSERIACEAE,RHODOPHYTA) FROM AUSTRALIA AND NEW ZEALAND

Caloglossa species are widely distributed in mangroves and salt marshes around the world and their life history patterns are being investigated in laboratory culture. In Australia all isolates of C. monosticha, C. postiae and C. ogasawaraensis have Polysiphonia‐type (P‐type) sexual life histories. Among the 70 C. leprieurii isolates from Australia and New Zealand P‐type sexual reproduction also is dominant. However, ten isolates of C. leprieurii from the Spencer Gulf and the Gulf of St. Vincent in South Australia give rise to successive tetrasporphyte generations without gametophytes. Moreover, one isolate from Queensland is asexual. Only one South Australia isolate, obtained from Lake Alexandrina at the mouth of the Murray River, is sexual. South Australia and Pacific Mexico are two regions in which asexual reproduction is dominant. In another mangrove dwelling red alga Bostrychia moritiziana (Rhodomelaceae) non‐sexual reproduction also is frequent in Australia, New Caledonia and Bali (Indonesia). This asexual reproductive pattern of tetrasporophytic recycling appears to have arisen independently among individual populations of various red algal species in different regions. Investigations are underway on the molecular phylogeny of the Caloglossa leprieurii isolates.     

Mating type of isolates derived from the spermogonial state ofPuccinia coronata var.coronata

Hyphal confrontation between two haploid cultures originating from single basidiospores was used to determine the mating type ofPuccinia coronata var.coronata. Pairs of 15 single-basidiospore cultures were placed approximately 1 mm apart on the medium in all possible combinations. Hyphae of the pairs of colonies came into contact with each other in all combinations approximately two weeks after confrontations. When the nuclear number of hyphal cells in a contact zone was investigated one month after confrontation, monokaryotic hyphae were observed in selfing combination. On the other hand, dikaryotic hyphae were observed in 90.5% of crossing combinations between different cultures. Two isolates are considered compatible if dikaryotic hyphae are present in the contact zone but incompatible if they are absent. The mating type of the fungus was found to be characterized by multiple-allelomorphic tetrapolar incompatibility controlled by the “A” and “B” incompatible factors. Contribution No. 116, Laboratories of Plant Pathology and Mycology, Institute of Agriculture and Forestry, University of Tsukuba     

Nuclear selection in monokaryotization of dikaryotic mycelia ofPholiota nameko as described by leading and following nuclei

To examine monokaryotization of dikaryotic mycelia ofPholiota nameko, 18 monokaryotic stocks were used to produce a total of 130 dikaryotic stocks by reciprocal crossing. Monokaryotized mycelium was raised from dikaryotic mycelium in the peripheral zone of the growing colony. The stocks mated with a particular group of monokaryons produced wide-range monokaryotization at higher rates than the other combinations of hybridization. The growth rates of the monokaryotized mycelia exceeded from those of the corresponding parental dikaryons. The monokaryotized mycelium was isolated and back-crossed to parental monokaryotic stocks. Most of the isolates had nuclear types similar to only one of the parental stocks, while the replicates of isolates from two dikaryotic hybrids showed split nuclear type compositions. It is suggested that a relative dominance is active in the selection of one of the two nuclei of the dikaryotic cells in monokaryotization. The hierarchy of relative dominance among nuclei of 18 parental monokaryotic stocks in the monokaryotization of their reciprocal crossing products was estimated. We propose the involvement of a cascade process in dikaryotic cell division, in which the first dividing nucleus (to be found in the monokaryotized cell) may act as the leading nucleus and the other one as the following nucleus.     

Genetic analysis of nuclear ribosomal DNA of Lentinula edodes

Genetic analysis of nuclear ribosomal DNA (rDNA) of Lentinula edodes was carried out using rDNA restriction fragment length polymorphisms (RFLPs) as genetic markers. Two compatible monokaryotic strains that differed in the endonuclease digestion patterns of their rDNA were used. The dikaryotic strain established by crossing them produced mixed RFLP patterns. Single-spore isolates derived from the dikaryotic strain showed three types of rDNA RFLP patterns: either one of the two parental types or a mixed type. From the frequency of the mixed type, the recombination value of rDNA tandem repeats was calculated to be 31.4%. Linkage analysis between rDNA and two incompatibility factors (A and B) revealed that rDNA was not linked to either factor. The rDNA genotypes did not affect mycelial growth among the single-spore isolates.     

The first records of Stylodrilus heringianus (Oligochaeta: Lumbriculidae) from the Southern Hemisphere

Abstract

The oligochaete family Lumbriculidae is well represented in the Northern Hemisphere, but for the Southern Hemisphere only Lumbriculus variegatus (Müller) is recorded, from Africa, Australia, and New Zealand; no species are known from South America (Brinkhurst & Jamieson 1971). According to Brinkhurst (1971), L. variegatus may be a recent introduction to New Zealand, where it is now widely distributed in a range of inland waters.     

Dikaryotic fruiting body development in a single dikaryon of <Emphasis Type="Italic">Agrocybe aegerita</Emphasis> and the spectrum of monokaryotic fruiting types in its monokaryotic progeny

Using monokaryotic offspring from several dikaryotic parental strains, the phenomenon of monokaryotic fruiting has been previously analysed in the commercially cultivated high-quality edible mushroom Agrocybe aegerita, revealing a variety of monokaryotic fruiting types. Here, we report a single dikaryotic A. aegerita strain, A. aegerita AAE-3, and 40 monokaryons derived from it, which exhibit a wide spectrum of monokaryotic fruiting types, including a rare, previously unknown type. Advantageously, the selected parental strain A. aegerita AAE-3 completes its life cycle within three weeks by the formation of dikaryotic fruiting bodies of typical agaric morphology on malt extract agar plates. In order to morphologically compare normal dikaryotic fruiting to monokaryotic fruiting, histology was performed from all dikaryotic fruiting body development stages and all fruiting types of monokaryotic origin. No clamp connections or dikaryotic hyphae were observed within the plectenchyma of monokaryotic fruiting stages. Among the monokaryotic fruiting types of the A. aegerita AAE-3-derived monokaryons, we also characterised the rare ‘stipe type’ here described as ‘elongated initials type’ as no differentiation into a future cap and stipe was seen. The two mating-compatible monokaryotic strains representing the extremes of the fruiting type spectrum observed, A. aegerita AAE-3-13 (‘mycelium type’) and A. aegerita AAE-3-32 (‘abortive?+?true homokaryotic fruiting fruiter type, AHF?+?THF fruiter type’), were also found to readily produce oidia (arthrospores). In order to obtain a set of mating-compatible monokaryons covering the whole observed spectrum of monokaryotic fruiting, the two monokaryons A. aegerita AAE-3-40 (‘initials type’) and A. aegerita AAE-3-37 (‘elongated initials type’) have been selected for their mating compatibility with A. aegerita AAE-3-32 and A. aegerita AAE-3-13, respectively. Together with the parental dikaryotic strain A. aegerita AAE-3, this set of standard monokaryons could prove useful for studies exploring the factors regulating monokaryotic fruiting in comparison to dikaryotic mushroom formation.     

Mating system and DNA polymorphism of monokaryons with different mating type of Stropharia rugoso-annulata

The mating system of Stropharia rugoso-annulata Farlow apud Murrill was studied by pairing single spore isolates from the same fruitbody, and the genetic diversity of monokaryotic strains with different mating types was evaluated by the RAPD technique. Basidiospores could germinate normally on PGP (potato/glucose/peptone) medium at 30 °C. Analysis of self- and cross-pairings revealed that Stropharia rugoso-annulata was heterothallic and tetrapolar. RAPD analysis detected polymorphism among monokaryotic strains, with more genetic variation within monokaryotic strains with non-parental mating type compared to monokaryotic strains with parental mating type. These results were in general agreement with the existing knowledge, confirming the validity and usefulness of the RAPD technique. Therefore, the RAPD technique will provide an exciting and valuable tool for a large-scale study on identification and genetic resources of monokaryotic strains, and should lead to a more efficient understanding and utilization of genetic diversity of monokaryotic strains in cross breeding by breeders.     

A new biological function of Shiitake mushroom,Lentinula edodes, in a myco-heterotrophic orchid,Erythrorchis ochobiensis

The biological function ofLentinula edodes in a myco-heterotrophic orchid,Erythrorchis ochobiensis was examined, using one local variant each from Japan (JPN), Papua New Guinea (PNG) and New Zealand (NZ). All variants induced seed germination: PNG and NZ isolates were effective at 25°C and JPN isolate showed the highest germination rate at 30°C. Germinated seeds developed into plants and formed normal endomycorrhizas. Hence, it is concluded thatL. edodes has a perfect symbiotic potential withE. ochobiensis, though it has not been observed in the root of the orchid in the field.     

Electrophoretic karyotype of <Emphasis Type="Italic">Flammulina velutipes</Emphasis> and its variation among monokaryotic progenies

 The karyotype of Flammulina velutipes (Curt. : Fr.) Sing. was investigated using contour-clamped homogeneous electric fields (CHEF) gel electrophoresis. A parental dikaryotic stock, JA, was resolved into at least eight chromosomal DNA bands ranging from 1.4- to 4.9-megabase (Mb) pairs. Overall, little size variation was found among monokaryotic strains with a few major exceptions. Among 13 monokaryotic progenies examined, 11 strains were resolved into at least eight chromosomal DNA bands in a manner similar to the parent dikaryon, whereas the other 2 were resolved into at least seven chromosomes lacking the 2.1-Mb chromosome possessed in the former. A slightly larger size variation was found in a chromosome carrying ribosomal DNA. An estimated haploid genome size of this stock was 24.0 Mb or more. Received: October 11, 2001 / Accepted: November 11, 2002 Acknowledgments We thank Professor T. Morinaga, Hiroshima Prefectural University, and Dr. T. Arima for their technical advice regarding CHEF gel electrophoresis. Correspondence to:E. Tanesaka     

Reproductive patterns of Caloglossa species (Delesseriaceae, Rhodophyta) from Australia and New Zealand: multiple origins of asexuality in C. leprieurii. Literature review on apomixis, mixed-phase, bisexuality and sexual compatibility

Reproduction and life history patterns in culture of five Caloglossa speaes from Australia and New Zealand are compared. Caloglossa adhaerens King et Puttock and Caloglossa bengalensis (Martens) King et Puttock have a Polyslphonla‐type sexual life history (P‐type, isomorphic alternation of generations). Caloglossa monosticha Kamiya occurs only in Western Australia (WA) and is a P‐type. Caloglossa ogasawaraensis Okamura occurs in WA, Northern Territory (NT), Queensland (QLD), New South Wales (NSW), Victoria (VIC) and South Australia (SA) and is for the most part a P‐type in culture. A few isolates have bisexual gametophytes that are self‐compatible, while most are unisexual. Caloglossa ogasawaraensis from Adelaide, SA and from Wilsons Promontory, VIC are new records for these States. In Australia, Caloglossa postiae (King et Puttock) Kamiya et King occurs in NSW, NT and QLD. All nine isolates are P‐type. Isolates of Caloglossa leprieurii (Montagne) G. Martens from NSW, NT, QLD, Tasmania, VIC and New Zealand are P‐type except for the freshwater isolates in which tetraspore germlings do not reproduce. In some isolates mixed‐phase reproduction is seen with male gametophytes producing both viable spermatia and tetrasporangia and female gametophytes producing procarps and sori with non‐dividing sporangia. All isolates of C. leprieurii irom Spencer Gulf and Gulf of St Vincent, SA and one isolate from QLD give rise to successive asexual generations of tetrasporophytes. Based on RuBisCO spacer DNA data an asexual life history has arisen several times in the C. leprieurii complex. The literature on apomixis, mixed‐phase reproduction, bisexuality and sexual compatibility in red algae is surveyed.     

Gondwanan radiation of the Southern Hemisphere crayfishes (Decapoda: Parastacidae): evidence from fossils and molecules

Aim The sequential break‐up of Gondwana is thought to be a dominant process in the establishment of shared biota across landmasses of the Southern Hemisphere. Yet similar distributions are shared by taxa whose radiations clearly post‐date the Gondwanan break‐up. Thus, determining the contribution of vicariance versus dispersal to seemingly Gondwanan biota is complex. The southern freshwater crayfishes (family Parastacidae) are distributed on Australia and New Guinea, South America, Madagascar and New Zealand and are unlikely to have dispersed via oceans, owing to strict freshwater limitations. We test the hypotheses that the break‐up of Gondwana has led to (1) a predominately east–west (((Australia, New Zealand: 80 Ma) Madagascar: 160–121 Ma) South America: 165–140 Ma), or (2) a southern (((Australia, South America: 52–35 Ma) New Zealand: 80 Ma) Madagascar: 160–121 Ma) pattern for parastacid crayfish. Further, we examine the evidence for a complete drowning of New Zealand and subsequent colonization by freshwater crayfish. Location Southern Hemisphere. Methods The evolutionary relationships among the 15 genera of Parastacidae were reconstructed using mitochondrial [16S, cytochrome c oxidase subunit I (COI)] and nuclear (18S, 28S) sequence data and maximum likelihood and Bayesian methods of phylogenetic reconstruction. A Bayesian (multidivtime ) molecular dating method using six fossil calibrations and phylogenetic inference was used to estimate divergence time among crayfish clades on Gondwanan landmasses. Results The South American crayfish are monophyletic and a sister group to all other southern crayfish. Australian crayfish are not monophyletic, with two Tasmanian genera, Spinastacoides and Ombrastacoides, forming a clade with New Zealand and Malagasy crayfish (both monophyletic). Divergence of crayfish among southern landmasses is estimated to have occurred around the Late Jurassic to Early Cretaceous (109–178 Ma). Main conclusions The estimated phylogenetic relationships and time of divergence among the Southern Hemisphere crayfishes were consistent with an east–west pattern of Gondwanan divergence. The divergence between Australia and New Zealand (109–160 Ma) pre‐dated the rifting at around 80 Ma, suggesting that these lineages were established prior to the break‐up. Owing to the age of the New Zealand crayfish, we reject the hypothesis that there was a complete drowning of New Zealand crayfish habitat.     

Establishment of monokaryotic and dikaryotic isolates of Hedgehog mushrooms (Hydnum repandum and related species) from basidiospores

Hydnum repandum and its relatives are gourmet edible ectomycorrhizal mushrooms. However, no reliable pure cultures have been reported in this genus. Here, we report for the first time the successful isolation of mycelial strains from basidiospores in the genus Hydnum. Basidiospores obtained from basidioma samples were aseptically inoculated onto modified Norkran's C (MNC) medium, MNC containing n-butyric acid (n-MNC), or MNC with gellan gum instead of agar (G-MNC). Although basidiospore germination was observed in most samples, the isolation rate was higher from MNC (91.7%) and G-MNC (93.8%) than from n-MNC (36.4%). Most established isolates were monokaryotic and lacked a clamp connection, but three were dikaryotic and had clamp connections. Established isolates were identified by molecular phylogenetic analysis of the internal transcribed spacer region of the rRNA gene. These results suggest that basidiospores can be used to establish monokaryotic and dikaryotic isolates of Hydnum species.     

The constitution of incompatibility factor and mating characteristics of spore isolates in a bipolar mushroom, <Emphasis Type="Italic">Pholiota nameko</Emphasis>

Pholiota nameko is a wood-rotting edible mushroom that carries a bipolar A incompatibility factor gene. The linkage analysis of the multiple allelomorphic A factor gene demonstrated that sexual reproduction produced a monospore isolate carrying a new A factor gene in addition to two parental mating types of isolates. However, 10%–30% of the modified monospore isolates could not produce a dikaryon with both of the parental monokaryons by crossing. It is concluded that the bipolar A incompatibility factor gene of P. nameko is constituted of two functional subunits, Aα and Aβ, which might be successively located beside each other with an apparent genetic distance of 0.3 centi-Morgan between them on the same chromosome. Further, some monospore isolates that did not conjugate with both parental monokaryons could produce dikaryons with different monokaryotic stocks with either one of the parental mating types. This result suggests that the crossing capability of these isolates were essentially those for one of the mating types of the parental monokaryons, but that their function for mating activity was made partially by unequal crossing-over in the process of sexual recombination. Received: May 1, 2001 / Accepted: December 5, 2001