IB-371 – GENERAL MYCOLOGY LABORATORY
PYRENOMYCETES
XYLARIALES, SORDARIALES, HALOSPHAERIALES
DIAPORTHALES, HYPOCREALES
Thursday, November 12, 2003
I. XYLARIALES
Stromata usually well developed, mostly consisting only of fungal tissue. Ascomata
perithecial, rarely cleistothecial, +/- globose, superficial or immersed in the stroma. Peridium is
usually black and thick walled; ostiole is papillate with periphyses. Interascal tissue well
developed, of narrow paraphyses. Asci are cylindrical, persistent, relatively thick-walled but
without separable layers, with an often complex J+ apical ring. Asci contain eight ascospores.
Ascospores are usually pigmented, sometimes transversely septate, with germ pores or slits,
sometimes with a gelatinous sheath. (See pictures in your handouts.)
Representative genera: Xylaria, Daldinia, Hypoxylon
Examine several types of stromata and look for embedded perithecia. Observe the
prepared slides and make drawings of the position of perithecia in the stroma. Can you see the
beaks (papillae) of the ascomata? Look for the paraphyses and periphyses. Provide sketches
of Xylaria below.
II. SORDARIALES
Members of this order rarely form stromata. The ascomata are perithecial or
cleistothecial, with a thin or thick peridium. Interascal tissue is inconspicuous or lacking at
maturity. Asci are unitunicate, cylindrical or clavate, persistent or evanescent. Ascospores
usually have at least one dark cell with a germ pore and often have a gelatinous sheath or
appendages. Anamorphs are mostly absent or spermatial. There are about eight families (still
controversial) but you will be responsible for only four genera representative of the order. (See
pictures of the representative genera in your handouts.)
Representative genera: Sordaria, Gelasinospora, Chaetomium, Microthecium
Also look at Annulatascus triseptatus (For apical ring)
A. Set up the mating experiment for the black and tan spored isolates of Sordaria
fimicola as directed.
For each species provided in culture, remove 2 or 3 perithecia with a sterile needle and
place the perithecia on a slide in a drop of distilled water. Examine with the dissecting
microscope using light from above. Sketch the ascomata. Using your needles crush the
perithecia and watch for the asci and ascospores to emerge. Cover the material with a
coverslip and examine the material with the compound microscope. Below sketch the asci and
ascospores of each fungus.
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III. HALOSPHAERIALES
This order is considered polyphyletic at present and most likely contains several
evolutionary lines. The fungi are grouped together mainly because they have membranous or
brittle perithecia, usually with beaks, but some are cleistothecial. The asci are unitunicate, thinwalled, usually lack an apical apparatus and are evanescent. The asci deliquesce to release
ascospores to the centrum and the ascospores are then released through the beak. Sterile
tissues within the ascomata consist of chains of hyaline thin-walled cells called catenophyses.
Most of the species occur in the marine habitat and the ascospores are equipped with
appendages and sheaths that keep them suspended in the water and assist with attachment to
substrates.
Laboratory Example: Halosphaeria mediosetigera, Nais inornata
Using the dissecting microscope, locate some ascomata in the culture tube. Remove a
few with a flamed needle and crush them in a drop of water on a slide. Locate asci, ascospores
and catenophyses. Draw all three structures below.
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IV. DIAPORTHALES - The centrum at early stages of development is composed of
pseudoparenchymatous tissue that disintegrates as the asci grow into it. Paraphyses are
absent in mature ascomata. The asci are evanescent or the base of the ascus gelatinizes,
releasing intact asci into the venter of the fruit body.
Family – GNOMONIACEAE - Numerous important plant pathogens occur in this family. The
perithecia may or may not be produced in stromata. Asci are unitunicate and have a refractive
apical ring. They are free-floating in the centrum at maturity. The ascospores are one- or twocelled and hyaline or yellowish.
Laboratory Example: Cryphonectria (Endothia) parasitica (Causal agent of chestnut blight).
Examine prepared slides of the teleomorph and locate the: (a) stroma, (b) asci, and (c)
ascospores. Examine prepared slides of the anamorph (Cytospora) and locate the: (a)
pycnidia, (b) conidia, and (c) conidiogenous cells.
Examine some of the dried specimens and describe the appearance of the fungus on the host.
Illustrate all stages of the life cycle.
Laboratory Example: Gnomonia ulmea (Causal agent of leaf spot of elm).
Examine the prepared slides and locate the: (a) long necks, (b) periphyses in the neck, (c) asci
with refractive apical rings. Illustrate the structure of the asci and ascospores.
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V. HYPOCREALES - This order is characterized by perithecia (and stromata, if present) that
are typically soft and fleshy or membranous and white to brightly colored. The centrum is
typically composed of apical paraphyses originating from the perithecial apex just below the
periphyses, growing down as a palisade layer and finally disintegrating as the asci grow up
among them. The ascus wall is uniformly thin-walled, sometimes thickened at the apex, with or
without a pore; asci are unitunicate and form a single layer lining the base and sides of the
perithecial cavity and growing upward among the apical paraphyses.
Family - HYPOCREACEAE - The ascomata are pale to brightly colored (pink, red, blue,
dark purple, rarely brown). The peridium is of large, pseudoparenchymatous cells. The asci
are cylindrical, oblong or inflated and are in a basal hymenium. Paraphyses are apical and
usually deliquescent and therefore not present at maturity. Ascospores are hyaline, yellowish,
pinkish to greenish and only occasionally brown, 0 to several septate.
Laboratory Examples: Prepared slides of Nectria cinnabarina and Gibberella saubinetii and
cultures of the anamorphs of Nectria haematococca (Fusarium solani ) and Gibberella zea
(Fusarium graminearum) (We have pictures of the Fusarium anamorphs). From the prepared
slides, note the structure and position of the apical paraphyses and asci. Note the relationship
between the perithecia and their substrates. Examine material from culture. Find the
conidiogenous cells and conidia. Illustrate the anamorphic and and teleomorphic states of
Nectria cinnabarina. Draw the conidiogenous cell and conidia of Fusarium solani. Examine the
fresh material of Nectria and note the color and position of the ascomata. Crush an ascoma in a
drop of water and cover with a coverslip. Press lightly on the coverslip to flatten the peridium of
the ascoma. Describe the cells making up the peridium. What type of fungal cells are they?
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Family – CLAVICIPITACEAE – Members of this group are distinguished by the morphology of
their asci and ascospores. The ascus has a thick cap, perforated by a central narrow canal
through which needle-shaped ascospore are successively ejected. Members of this group are
parasitic on plants, insects or on other fungi.
Laboratory Examples: Claviceps purpurea and Cordyceps
1. Claviceps purpurea.
a. Conidial stage: Study prepared slides showing section of mycelial mats with layers of
conidiophores. Observe the large numbers of conidia produced. The conidiosporus mycelial
mats develop eventually into the sclerotia of the fungus.
b. Sclerotial stage: Examine heads of rye infected with Claviceps purpurea. Note the
size, shape and color of the sclerotia. Note the pseoudparenchymatous tissue composing the
rind and the interior of the sclerotium. Test the hardness of a sclerotium by cutting it with a
scalpel.
c. Ascigerous stage: Observe germinated sclerotia bearing stromata. Study stained
longitudinal sections through a stroma. Note the position of the perithecia. Find the perithecial
wall as distinguished from the stromatic tissue. Search your slide for a perithecium that has
been sectioned exactly through the center; note the ostiole, periphyses, the asci and the
ascospores.
2. Cordyceps.
Examine an insect parasitized by Cordyceps. Where are the asci located?
Illustrate a, b, and c above.
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