Hymenoscyphus Ericae Research Articles

Fungi in hair roots of Vaccinium spp. (Ericaceae) growing on decomposing wood: colonization patterns, identity, and in vitro symbiotic potential

Most of our knowledge on the ericoid mycorrhizal (ErM) symbiosis comes from temperate heathlands characterized by acidic peaty soils and many experiments with a few ascomycetous fungi. However, ericaceous plants thrive in many other ecosystems and in temperate coniferous forests, their seedlings often prosper on decomposing wood. While wood is typically exploited by basidiomycetous ectomycorrhizal (EcM) and saprobic fungi, the role of ErM fungi (ErMF) is much less clear. We explored the cultivable mycobiota of surface sterilized hair roots of Vaccinium spp. growing on decomposing wood in two coniferous forests in Mid-Norway (Scandinavia) and Northern Bohemia (Central Europe). Obtained isolates were identified using molecular tools and their symbiotic potential was tested in vitro. While the detected community lacked the archetypal ErMF Hyaloscypha hepaticicola and the incidence of dark septate endophytes and EcM fungi was negligible, it comprised other frequent asexual ascomycetous ErMF, namely H. variabilis and Oidiodendron maius, together with several isolates displaying affinities to sexual saprobic H. daedaleae and H. fuckelii. Ascomycete-suppressing media revealed representatives of the saprobic basidiomycetous genera Coprinellus, Gymnopilus, Mycena (Agaricales), and Hypochnicium (Polyporales). In the resyntheses, the tested basidiomycetes occasionally penetrated the rhizodermal cells of their hosts but never formed ericoid mycorrhizae and in many cases overgrew and killed the inoculated seedlings. In contrast, a representative of the H. daedaleae/H. fuckelii-related isolates repeatedly formed what morphologically appears as the ErM symbiosis and supported host's growth. In conclusion, while basidiomycetous saprobic fungi have a potential to colonize healthy-looking ericaceous hair roots, the mode(-s) of their functioning remain obscure. For the first time, a lineage in Hyaloscypha s. str. (corresponding to the former Hymenoscyphus ericae aggregate) where sexual saprobes are intermingled with root symbionts has been revealed, shedding new light on the ecology and evolution of these prominent ascomycetous ErMF.

Hyaloscypha gabretae and Hyaloscypha gryndleri spp. nov. (Hyaloscyphaceae, Helotiales), two new mycobionts colonizing conifer, ericaceous and orchid roots.

Historically, Hyaloscypha s. lat. (Hyaloscyphaceae, Helotiales) included various saprobes with small apothecia formed on decaying plant matter, usually wood, that were defined by chemical and (ultra)structural aspects. However, recent molecular phylogenetic and resynthesis studies have narrowed the concept of the genus and shown that it contains several widely distributed species with unknown sexual morphs that form ectomycorrhizae, ericoid mycorrhizae, and mycothalli and also grow endophytically in plant roots and hypogeous ectomycorrhizal (EcM) fruitbodies (i.e., the historical Hymenoscyphus ericae aggregate). Hence, some of the sexually reproducing saprobic Hyaloscypha s. lat. and the symbionts belong to the monophyletic Hyaloscypha s. str. Here, we introduce two new root-symbiotic Hyaloscypha s. str. species, i.e., H. gabretae and H. gryndleri spp. nov. While the former was isolated only from ericaceous hosts (Vaccinium myrtillus from Southern Bohemia, Czechia and Calluna vulgaris from England, UK), the latter was obtained from a basidiomycetous EcM root tip of Picea abies (Pinaceae), roots of Pseudorchis albida (Orchidaceae), and hair roots of V. myrtillus from Southern Bohemia and C. vulgaris from England. Hyaloscypha gryndleri comprises two closely related lineages, suggesting ongoing speciation, possibly connected with the root-symbiotic life-style. Fungal isolates from ericaceous roots with sequences similar to H. gabretae and H. gryndleri have been obtained in Japan and in Canada and Norway, respectively, suggesting a wide and scattered distribution across the Northern Hemisphere. In a series of in vitro experiments, both new species failed to form orchid mycorrhizal structures in roots of P. albida and H. gryndleri repeatedly formed what morphologically corresponds to the ericoid mycorrhizal (ErM) symbiosis in hair roots of V. myrtillus, whereas the ErM potential of H. gabretae remained unresolved. Our results highlight the symbiotic plasticity of root-associated hyaloscyphoid mycobionts as well as our limited knowledge of their diversity and distribution, warranting further ecophysiological and taxonomic research of these important and widespread fungi.

Root-associated ectomycorrhizal fungi shared by various boreal forest seedlings naturally regenerating after a fire in interior alaska and correlation of different fungi with host growth responses.

The role of common mycorrhizal networks (CMNs) in postfire boreal forest successional trajectories is unknown. We investigated this issue by sampling a 50-m by 40-m area of naturally regenerating black spruce (Picea mariana), trembling aspen (Populus tremuloides), and paper birch (Betula papyrifera) seedlings at various distances from alder (Alnus viridis subsp. crispa), a nitrogen-fixing shrub, 5 years after wildfire in an Alaskan interior boreal forest. Shoot biomasses and stem diameters of 4-year-old seedlings were recorded, and the fungal community associated with ectomycorrhizal (ECM) root tips from each seedling was profiled using molecular techniques. We found distinct assemblages of fungi associated with alder compared with those associated with the other tree species, making the formation of CMNs between them unlikely. However, among the spruce, aspen, and birch seedlings, there were many shared fungi (including members of the Pezoloma ericae [Hymenoscyphus ericae] species aggregate, Thelephora terrestris, and Russula spp.), raising the possibility that these regenerating seedlings may form interspecies CMNs. Distance between samples did not influence how similar ECM root tip-associated fungal communities were, and of the fungal groups identified, only one of them was more likely to be shared between seedlings that were closer together, suggesting that the majority of fungi surveyed did not have a clumped distribution across the small scale of this study. The presence of some fungal ribotypes was associated with larger or smaller seedlings, suggesting that these fungi may play a role in the promotion or inhibition of seedling growth. The fungal ribotypes associated with larger seedlings were different between spruce, aspen, and birch, suggesting differential impacts of some host-fungus combinations. One may speculate that wildfire-induced shifts in a given soil fungal community could result in variation in the growth response of different plant species after fire and a shift in regenerating vegetation.

Chapter Six: The Ascomycete Rhizoscyphus ericae Elicits a Range of Host Responses in the Rhizoids of Leafy Liverworts: An Experimental and Cytological Analysis

Abstract Ascomycete fungi, identified cytologically from their simple septa and Woronin bodies and genetically diagnosed as belonging to the Rhizoscyphus ericae (D. J. Read) W. Y. Zhuang & Korf (Hymenoscyphus ericae (D. J. Read) Korf & Kernan) aggregate form a range of specialized mutualistic associations with the rhizoids in several families of leafy liverworts. To test the hypothesis that this fungus can produce these associations between liverworts from widely separate geographical regions, we carried out cross-infection experiments between mycobionts that induce branching and septation in the rhizoids in the Schistochilaceae from the southern hemisphere and those from a range of disparate taxa (Bazzania, Calypogeia, Cephalozia, Kurzia, Lepidozia, Odontoschisma) from the northern hemisphere and from Calluna in the Ericaceae. All the fungal isolates produced rhizoid infections identical to those found in nature. The same fungal isolates also invaded the rhizoids of axenically cultured Nardia scalaris, w...

Interactions between iron availability, aluminium toxicity and fungal siderophores

The influence of iron, aluminium and of the combined application of both metals on microbial biomass and production of siderophores by three fungi (Aspergillus nidulans, Neurospora crassa and Hymenoscyphus ericae) were investigated. All three species showed a strong iron regulation and Al-sensitivity of siderophore biosynthesis although several differences remained species dependent. Inhibitory effects of Fe and Al on siderophore-production were additive and the higher binding capacity of siderophores towards iron could be compensated by a higher Al-availability. Although pH itself is also important for regulation of siderophore biosynthesis, an indirect effect of Al on siderophore production via an Al-induced pH decrease could be outlined. The toxic effects of Al resulting in a reduced biomass production were compensated by high Fe-availability, whereas the addition of DFAM, a bacterial siderophore, enhanced Al-toxicity.

The biotransformation of mercury in pH-stat cultures of microfungi

The biotransformation of mercury was monitored in Hymenoscyphus ericae (Read) Korf & Kernan, Neocosmospora vasinfecta E. F. Sm., and Verticillium terrestre (Link) Lindau following the exposure of these fungi to environmentally relevant doses of HgII(HgCl2) in aerated pH-controlled cultures. Mercury applied at 120 and 300 μg·L–1quickly associated with cells of N. vasinfecta. Within 3 h, approximately 80% of the remaining Hg in both treatments had been converted to β-HgS, and at 8 h, 15% and 53% of the original doses had been converted into volatile Hg0. Hymenoscyphus ericae was less efficient at converting HgIIto β-HgS, but it volatilized larger amounts of Hg0. A mercury-tolerant isolate of V. terrestre withstood levels of Hg in the parts per million. When exposed to 270, 1000, or 2000 μg·L–1HgCl2, 93% ± 3% of the amounts remaining in cultures of V. terrestre after 2 h was β-HgS. At 2 mg·L–1, 72% of the Hg was reduced to Hg0in 2 h. Volatilization accounts for the tolerance of V. terrestre to high doses of Hg, whereas the conversion of Hg to β-HgS was the major detoxifying mechanism at lower doses. The formation of β-HgS occurs preferentially with volatilization operating as the default pathway, when the former biotransformation mechanism is saturated.

SENSITIVITY OF ERICOID MYCORRHIZAL FUNGI AND MYCORRHIZAL CALLUNA VULGARIS TO COPPER MINE SPOIL FROM AVOCA, COUNTY WICKLOW

Ericoid mycorrhizal fungi were isolated from roots of Calluna vulgaris collected from mine spoil at abandoned copper mines at Avoca, County Wicklow, Ireland, and Parys Mountain, Anglesey, Wales. Two isolates were morphologically identical to Hymenoscyphus ericae and formed typical ericoid mycorrhizal associations. Four fungal isolates— the two mine-site isolates and two H. ericae isolates (obtained from uncontaminated sites) were grown in liquid media containing elevated levels of Cu and Zn. All isolates were determined to have a low sensitivity to these metals, with EC₅₀ values (concentration of metal causing 50% reduction in mycelial dry mass) in the range 0.3-0.9mM Cu and 1.2-5.5mM Zn, but there were differences in metal sensitivity between isolates. There was a significant decline in the tolerance of each isolate to increasing concentrations of mine spoil (up to 10% in the agar medium). Mycorrhizal amelioration of heavy-metal toxicity was determined by growing mycorrhizal (M) and non-mycorrhizal (NM) rooted cuttings of C. vulgaris on mine spoil. Rooted cuttings obtained from an uncontaminated site showed no response to mycorrhizal colonisation. However, C. vulgaris obtained from the Avoca site and inoculated with H. ericae performed better than NM plants. The improved growth of these plants was associated with a reduction in Cu within the host's shoot. Inoculation of C. vulgaris with an ericoid mycorrhizal fungus isolated from the Avoca mine site also reduced Cu accumulation within shoots but there was no change in the growth and Zn accumulation of C. vulgaris on mine spoil. Results are discussed in relation to the potential benefit of the ericoid mycorrhizal association to C. vulgaris at metal contaminated sites.

Sensitivity of Ericoid Mycorrhizal Fungi and Mycorrhizal <i>Calluna vulgaris</i> to Copper Mine Spoil from Avoca, County Wicklow

Ericoid mycorrhizal fungi were isolated from roots of Calluna vulgaris collected from mine spoil at abandoned copper mines at Avoca, County Wicklow, Ireland, and Parys Mountain, Anglesey, Wales. Two isolates were morphologically identical to Hymenoscyphus ericae and formed typical ericoid mycorrhizal associations. Four fungal isolates? the two mine-site isolates and two H. ericae isolates (obtained from uncontaminated sites) were grown in liquid media containing elevated levels of Cu and Zn. All isolates were determined to have a low sensitivity to these metals, with EC50 values (concentration of metal causing 50% reduction in mycelial dry mass) in the range 0.3-0.9mM Cu and 1.2-5.5mM Zn, but there were differences in metal sensitivity between isolates. There was a significant decline in the tolerance of each isolate to increasing concentrations of mine spoil (up to 10% in the agar medium). Mycorrhizal amelioration of heavy-metal toxicity was determined by growing mycorrhizal (M) and non-mycorrhizal (NM) rooted cuttings of C. vulgaris on mine spoil. Rooted cuttings obtained from an uncontaminated site showed no response to mycorrhizal colonisation. However, C. vulgaris obtained from the Avoca site and inoculated with H. ericae performed better than NM plants. The improved growth of these plants was associated with a reduction in Cu within the host's shoot. Inoculation of C. vulgaris with an ericoid mycorrhizal fungus isolated from the Avoca mine site also reduced Cu accumulation within shoots but there was no change in the growth and Zn accumulation of C. vulgaris on mine spoil. Results are discussed in relation to the potential benefit of the ericoid mycorrhizal association to C. vulgaris at metal contaminated sites. Brian R. Gibson, Division of Food Sciences, School of Biosciences,

Chitinolytic activities of ericoid mycorrhizal and other root-associated fungi from Epacris pulchella ( Ericaceae)

Ericoid mycorrhizal endophytes and other root-associated fungi from Epacris pulchella (Ericaceae) in an eastern Australian sclerophyll forest, along with Hymenoscyphus ericae, were tested for their abilities to produce extracellular chitinolytic activities during growth in axenic culture. Two root-associated fungi produced activities that were active against only a monomeric 4-methylumbelliferyl (4-MU) glycoside of N-acetylglucosamine, suggesting exo-acting beta-N-acetylhexosaminidase (EC 3.2.1.52) activity. All ericoid mycorrhizal fungi and two root-associated fungi produced activities against dimeric and trimeric 4-MU glycosides of N-acetylglucosamine, suggesting production of chitobiosidase and endo-acting chitinase (EC 3.2.1.14) respectively in addition to beta-N-acetylhexosaminidase. Specific activities for all ericoid mycorrhizal fungi, including H. ericae, were of the same order of magnitude, suggesting that their chitinolytic potential is broadly similar. Chitinase activities were only produced by an ericoid mycorrhizal fungus when chitin was included in the medium, however, no activity was produced if glucose was also present in the medium.

Oribatid mite (Acari, Oribatida) feeding on ectomycorrhizal fungi

The coexistence of a large number of soil animals without extensive niche differentiation is one of the great riddles in soil biology. The main aim of this study was to explore the importance of partitioning of food resources for the high diversity of micro-arthropods in soil. In addition, we investigated if ectomycorrhizal fungi are preferentially consumed compared to saprotrophic fungi. Until today, ectomycorrhizal fungi have never been tested as potential food resource for oribatid mites. We offered six ectomycorrhizal fungi [Amanita muscaria (L.) Hook., Boletus badius (Fr.) Fr., Cenococcum geophilum Fr., Laccaria laccata (Scop.) Fr., Paxillus involutus (Batsch) Fr. and Piloderma croceum J. Erikss. & Hjortstam], one ericoid mycorrhizal fungus [Hymenoscyphus ericae (D.J. Read) Korf & Kernan] and three saprotrophic fungi [Agrocybe gibberosa (Fr.) Fayod, Alternaria alternata (Fr.) Keissl. and Mortierella ramanniana (A. Møller) Linnem.] simultaneously to each of the mainly mycophagous oribatid mite species Carabodes femoralis (Nicolet), Nothrus silvestris Nicolet and Oribatula tibialis Nicolet. The ericoid mycorrhizal fungus H. ericae and the ectomycorrhizal fungus B. badius were preferentially consumed by each oribatid mite species. However, feeding preferences differed significantly between the three species, with O. tibialis being most selective. This study for the first time documented that oribatid mites feed on certain ectomycorrhizal fungi.

The mycorrhizal community in a forest chronosequence of Sitka spruce [Picea sitchensis (Bong.) Carr.] in Northern England

Demography and fungal diversity of the belowground ectomycorrhizal community in a chronosequence of Sitka spruce [Picea sitchensis (Bong.) Carr.] in Northumberland, Northern England, were analysed; mycorrhizal root samples were taken from 6-, 12-, 30- and 40-year-old stands, and fungal fruiting bodies were collected in autumn to complement the survey. Naturally germinated seedlings less than 1 year of age (taken from the 30-year-old stand) were also examined. A total of 118,000 mycorrhizal root tips were extracted from 40 soil cores (ten per age class) and from the complete root systems of 25 seedlings and separated into active and senescent root tips according to their morphology and anatomy. Active tips were distinguished according to their mycobionts which were characterised and identified microscopically. Although almost 100% of all fine roots were mycorrhizal, EM fungal diversity throughout the chronosequence was low, consisting of a total of 16 species of which three were only found as fruiting bodies. Of the six mycobionts found most regularly below ground, Tylospora fibrillosa was the most common, colonising about 70% of all root tips and more than 90% of those of seedlings and young trees. Root density and mycorrhizal diversity increased, but percentage of vital root tips decreased with increasing tree age, levelling off in the 30- and 40-year-old stand. Among the five subdominant fungal species, Dermocybe crocea was found to have its peak of distribution in the 12-year-old stand and Russula emetica, Lactarius rufus, Hymenoscyphus ericae agg. and the unidentified Piceirhiza sulfo-incrustata in the 30- and 40-year-old stands. The possible correlations between the mycorrhizal community structure and biotic and abiotic factors are discussed.

Phosphatases of ericoid mycorrhizal fungi: kinetic properties and the effect of copper on activity

Ericoid endomycorrhizal fungi (two isolates of Hymenoscyphus ericae obtained from unpolluted heathlands and two H. ericae-type endophytes isolated from Calluna vulgaris growing on Cu-contaminated mine spoil) were grown for 14 d on 10% Rorison's solution containing sodium phytate as the sole P source and either trace (0.16 microM) or elevated (0.25 mM) concentrations of Cu. The elevated levels of Cu in the medium had no effect on the growth of the two H. ericae-type endophytes from mine spoil sites but caused a significant reduction in growth of the two H. ericae isolates from unpolluted sites. Wall, cytoplasmic and extracellular fractions were assayed for phosphomonoesterase (PMEase) and phosphodiesterase (PDEase) activity. K(m) and V(max) values varied between the different endophytes and both were highest in the wall fractions. Wall-bound phosphatase activity, excluding PDEase of one H. ericae-type endophyte, was generally unaffected after the isolates had been grown on medium containing 0.25 mM Cu. Extracellular PDEase of the two H. ericae-type endophytes from mine spoil sites was stimulated by 0.25 mM Cu in the growth medium. Cu concentrations up to 5.0 mM in the assay medium did not inhibit wall-bound phosphatase activity whereas three of the isolates showed a stimulation of extracellular activity with increasing Cu. The results are discussed in relation to phosphatase activity of ericoid endophytes on Cu-contaminated substrates.

Influence of pH on copper and zinc sensitivity of ericoid mycobionts in vitro

The effect of pH on growth, metal uptake and toxicity in four isolates of ericoid mycobionts (two Hymenoscyphus ericae from unpolluted heathland sites and two H. ericae-type mycobionts from metal-contaminated mine spoil) was assessed in vitro. These isolates were incubated in liquid medium (10% Rorison's medium, glucose at 10 g l(-1)) containing either 0.25 mM Cu or 2.0 mM Zn and adjusted to pH 2, 3, 4, 5 or 6. After 30 days incubation, dry mass and mycelial metal content were determined and growth was expressed as tolerance index, i.e. dry mass in the presence of metal as a percentage of dry mass in the absence of metal. Initial medium pH had a significant effect on both tolerance index and metal accumulation. Tolerance indices were highest at pH 2, with several isolates showing a stimulation of growth (i.e. tolerance index >100%) at this pH. Tolerance index decreased at higher initial pH values and growth of two mycobionts was completely inhibited (tolerance index=0) in the Cu-supplemented media at pH 6. Reduction in tolerance index coincided with an increase in mycelial accumulation of Cu and Zn. Practical and environmental implications of these results are discussed.

Are liverworts imitating mycorrhizas?

Are liverworts imitating mycorrhizas?

Nutritional influences on the solubilization of metal phosphate by ericoid mycorrhizal fungi

Four ericoid mycobionts (two isolates of Hymenoscyphus ericae, and two dark, sterile ericoid mycobionts isolated from metal-contaminated mine sites) were grown on solid agar plates supplemented with zinc phosphate (0.25 %) containing different forms of nitrogen (nitrate, ammonium or alanine) and different concentrations of carbon (glucose) and phosphorus (K2HPO4). The influence of nutrient variation on solubilizing ability of the fungi was assessed by measuring the zones of solubilization appearing beneath the growing colonies. All four mycobionts were capable of zinc phosphate solubilization in the presence of all three nitrogen sources and in media containing no nitrogen. No solubilization was observed at 0 mM glucose-C but was observed with increasing glucose concentration from 300 to 600 mM C. Increasing phosphorus concentration (0-5 mM P) had no effect on the solubilizing ability of the isolates. All but one of the mycobionts were capable of solubilizing calcium phosphate (CaHPO4), while no solubilization was observed in media containing aluminium phosphate (AlPO4), iron phosphate (FePO4 x 4H2O) or copper phosphate (Cu3O8P2 x 2H2O) under conditions which were found to be optimal for zinc phosphate solubilization. Under conditions of glucose at 300 mM C and alanine as the N source in the zinc phosphate-amended agar medium, one of the mycobionts produced new crystals, which were morphologically distinct from the original zinc phosphate crystals. It is concluded that medium composition influences the metal-phosphate solubilizing ability of ericoid mycobionts. The results are discussed in relation to the possible mechanisms involved in solubilization and the potential benefits of metal-phosphate solubilization to ericoid mycobionts and their host plants.

Zinc Phosphate and Pyromorphite Solubilization by Soil Plant-Symbiotic Fungi

Acidolysis, complexolysis and metal accumulation were involved in solubilization of zinc phosphate and pyromorphite by a selection of soil fungi representing ericoid and ectomycorrhizal plant symbionts and an endophytic/entomopathogenic fungus, Beauveria caledonica. Zinc phosphate was much more readily solubilized than pyromorphite. According to the relationship between metal mobilization and pH, acidolysis (protonation) was found to be the major mechanism of both zinc phosphate and pyromorphite dissolution for most of the fungi examined. In general, the more metal tolerant fungal strains yielded more biomass, acidified the medium more and dissolved more of the metal mineral than less tolerant strains. However, B. caledonica 4 excreted a substantial amount of oxalic acid (∼ 0.8 mM) in the presence of pyromorphite that coincided with a dramatic increase in lead mobilization providing a clear example of complexolysis. Organic acid excretion by fungi was inter- and intraspecific and was strongly influenced by the presence of the toxic metal minerals. When grown on zinc phosphate or pyromorphite, Hymenoscyphus ericae DGC3(UZ) accumulated the lowest metal concentration, but Thelephora terrestris accumulated the highest metal concentration in the biomass. The ability to accumulate water-soluble lead species, representing mainly cytosolic and vacuolar pools, seemed to be connected with pyromorphite-solubilizing ability. B. caledonica 4, which demonstrated the highest ability to dissolve pyromorphite, accumulated the highest water-soluble fraction and total lead concentration in the mycelium. Generally, isolates with a higher zinc-tolerance accumulated significantly less total zinc from zinc phosphate (including the sum of water-soluble and NaCl-extractable zinc) than non-tolerant strains.

Graffenrieda emarginata (Melastomataceae) forms mycorrhizas with Glomeromycota and with a member of the Hymenoscyphus ericae aggregate in the organic soil of a neotropical mountain rain forest

Graffenrieda emarginata (Ruiz & Pav.) Triana (Melastomataceae) was found to be the most frequent tree on the nutrient-poor organic soil in a species-rich tropical mountain rain forest of southern Ecuador. Our studies revealed that the rootlets are symbiotically associated with arbuscular mycorrhizal fungi and simultaneously form ectomy corrhizas displaying a superficial Hartig net but no hyphal mantle. Molecular phylogeny revealed a multitude of previously unknown glomeromycete sequence types. Ultrastructure of the ectomycorrhiza-forming fungus indicated an ascomycete, and molecular phylogenetic studies identified the fungal symbiont as a new taxon within the Hymenoscy phus ericae (Read) Korf & Kernan aggregate (Ascomycota). The mycorrhizal state of G. emarginata is considered crucial for its competitiveness and abundance in these sites.Key words: arbuscular mycorrhiza, ectomycorrhiza, Glomeromycota, Hymenoscyphus ericae aggregate, neotropical mountain rain forest, Ecuador.

Host Range of a Select Isolate of the Ericoid Mycorrhizal Fungus Hymenoscyphus ericae

Studies were conducted to examine the host range of a select isolate of the ericoid mycorrhizal fungus Hymenoscyphus ericae (Read) Korf and Kernan [American Type Culture Collection (ATCC) #32985]. Host status was tested for 15 ericaceous species, including: Calluna vulgaris (L.) Hull, Enkianthus campanulatus (Miq.) Nichols, Gaultheria procumbens L., Kalmia latifolia L., Leucothoe fontanesiana Sleum., Oxydendrum arboreum (L.) DC.,Pierisfloribunda (Pursh) Benth. & Hook.,Rhododendron calendulaceum (Michx.) Torr.,Rhododendron carolinianum Rehd., Rhododendron catawbiense Michx., Rhododendron maximum L., Rhododendron mucronulatumTurcz., Vaccinium corymbosum L., and Vaccinium macrocarpon Ait. Arbutus unedo L., an ericaceous species that forms arbutoid, not ericoid, mycorrhizae, was used as a negative control. All of the species were colonized by the ericoid isolate with the exception of Enkianthus campanulatus and the negative control. Inoculation with this isolate of H. ericae resulted in a significant increase in shoot growth. However, intensity of root colonization was not correlated to amount of shoot growth. In fact, an increase in growth was observed in the two species that lacked fungal colonization.

Filamentous ascomycetes inhabiting the rhizoid environment of the liverwort Cephaloziella varians in Antarctica are assessed by direct PCR and cloning

We molecularly assesed the ascomycetous fungal communities inhabiting the rhizoid environment of Cephaloziella varians, collected at Rothera Point on the western Antarctic Peninsula. The RFLP-phenotyped and cloned PCR products of a partial small subunit of the ribosomal RNA gene were sequenced and analyzed with neighbor joining (NJ) and maximum parsimony (MP). Both analyses identified four bootstrap-supported groups: (i) a sister group to Onygenales, (ii) a well-supported clade with Phialocephala fortinii, (iii) a large group of clones nested within Chaetothyriales, and (iv) a group nested within Eurotiales with a likely affinity to the genus Aspergillus. An additional marginally supported clade, including helotialean Hymenoscyphus fructigenus, was detected in the NJ analysis. Placement of one clone (possibly helotialean) was not supported by either analysis. We included Hymenoscyphus ericae (Helotiales) in our analyses to test for its presence in clone libraries. None of our clones showed strongly supported affinity to H. ericae. The culture-independent technique proved useful for assessing the composition of rhizoid fungal communities, although it remains unknown whether any of these fungi colonize C. varians tissues. Direct-community assays of this kind might be best combined with traditional isolation techniques to get a more holistic view of fungi occupying plant tissues.

Filamentous Ascomycetes Inhabiting the Rhizoid Environment of the Liverwort Cephaloziella varians in Antarctica Are Assessed by Direct PCR and Cloning

We molecularly assesed the ascomycetous fungal communities inhabiting the rhizoid environment of Cephaloziella varians, collected at Rothera Point on the western Antarctic Peninsula. The RFLP-phenotyped and cloned PCR products of a partial small subunit of the ribosomal RNA gene were sequenced and analyzed with neighbor joining (NJ) and maximum parsimony (MP). Both analyses identified four bootstrap-supported groups: (i) a sister group to Onygenales, (ii) a well-supported clade with Phialocephala fortinii, (iii) a large group of clones nested within Chaetothyriales, and (iv) a group nested within Eurotiales with a likely affinity to the genus Aspergillus. An additional marginally supported clade, including helotialean Hymenoscyphus fructigenus, was detected in the NJ analysis. Placement of one clone (possibly helotialean) was not supported by either analysis. We included Hymenoscyphus ericae (Helotiales) in our analyses to test for its presence in clone libraries. None of our clones showed strongly supported affinity to H. ericae. The culture-independent technique proved useful for assessing the composition of rhizoid fungal communities, although it remains unknown whether any of these fungi colonize C. varians tissues. Direct-community assays of this kind might be best combined with traditional isolation techniques to get a more holistic view of fungi occupying plant tissues.