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Department of Plant Biology, University of Illinois, 265 Morrill Hall, 505 South Goodwin Avenue, Urbana, Illinois 61801
J. L. Crane
Center for Biodiversity, Illinois Natural History Survey, 172 Natural Resources Building, 607 E. Peabody Drive, Champaign, Illinois 61820
P. M. Fallah
Environmental Microbiology Laboratory, Inc., 1800 Sullivan Avenue Suite 209, Daly City, California 94044
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ABSTRACT |
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Ceriospora caudae-suis and Submersisphaeria aquatica, two freshwater pyrenomycetes reported infrequently since their original description, occur commonly on submerged woody debris in the USA. Based on analyses of 28S rDNA sequence data and morphology, both species belong in the Annulatascaceae. Ceriospora caudae-suis is transferred to Pseudoproboscispora, a genus in the Annulatascaceae with similar overall morphology and ecology. Submersisphaeria aquatica is redescribed and illustrated based on additional collections.
Key words: Annulatascaceae, aquatic, fungi, systematics, 28S rDNA
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSTAXONOMYDISCUSSIONLITERATURE CITED |
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from submerged twigs of Fraxinus sp. collected from Lake Windermere, England. Although Ingold referred this species to the genus Ceriospora, he commented that it differed from all other known species of the genus in the length and coiling of the ascospore appendages. Ceriospora Niessl was established in 1876 for fungi that have immersed to superficial, ostiolate ascomata, septate filamentous paraphyses, asci that are unitunicate, cylindric to clavate, with a J+ plug-like apical ring, and ascospores that are two-celled with bipolar spine-like appendages (Müller and von Arx 1962, Hyde 1993). It is currently placed in the Amphisphaeriaceae, Xylariales (Eriksson et al 2001).
Ceriospora caudae-suis was reported recently from North America for the first time from lakes in northern Wisconsin (Fallah and Shearer 2001). The authors found that the ascus apical ring of C. caudae-suis was J- and was not plug-like in morphology, characteristics in disagreement with those of the type species of Ceriospora, C. dubyi Niessl (Müller and von Arx 1962, Hyde 1993). Also, the asci of C. caudae-suis are cylindrical with overlapping uniseriate ascospores, while those of C. dubyi are cylindric-clavate with overlapping bi- and triseriate ascospores. The ascospores also differ: those of C. caudae-suis are ellipsoidal and broadly tapered at the apices while those of C. dubyi are fusiform and narrowly tapered (Hyde 1993). Thus the placement of C. caudae-suis in Ceriospora is questionable.
Wong and Hyde (1999) described a fungus from north Queensland, Australia, that they considered related to C. caudae-suis but which had three- rather than one-septate ascospores. They considered that Ceriospora was not a suitable genus for their taxon since Ceriospora has asci with J+ apical rings (although they did not give the staining reaction for their taxon) and uniseptate ascospores with spine-like appendages. They therefore established a new genus and species, Proboscispora aquatica Wong & K.D Hyde (Lasiosphaeriaceae, Sordariales). The name Proboscispora had been used previously for a coelomycetous fungus, Proboscispora manihotis Punith., and was therefore illegitimate according to Article 53.1 of the International Code of Botanical Nomenclature (Greuter 1994) and a new name, Pseudoproboscispora Punith., was hence established for the genus (Punithalingam 1999). Although Wong and Hyde (1999) suggested that C. caudae-suis might belong in Pseudoproboscispora, they did not have any type material of C. caudae-suis available for study and so considered it premature to make any changes. There are morphological similarities between C. caudae-suis and P. aquatica with respect to their cylindrical, pedicellate asci with an apical ring, and ellipsoidal ascospores with bipolar unfurling appendages. The differences between them are in ascomal, ascus, and ascospore size and in ascospore septation.
We were able to make single ascospore cultures from collections of C. caudae-suis and a similar fungus with three septate ascospores, isolate A336–2 (Figs. 4, 5). Molecular sequence data were obtained to determine the relationship between these two taxa and to examine their ordinal relationships with other morphologically similar pyrenomycetes.
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described another new genus and species, Submersisphaeria aquatica, from wood submerged in a small stream in north Queensland, Australia. Although this fungus has not been reported from other geographical areas, we have encountered it numerous times in North America (Fig. 2). Based on ascospore pigmentation and appendages, this species bears little resemblance to C. caudae-suis and P. aquatica. There are, however, similarities in ascomal anatomy, hamathecium, ascus and ascospore morphology, and habitat. Although originally placed in the Lasiosphaeriaceae, Sordariales (Hyde 1996), S. aquatica was moved to the Annulatascaceae, Sordariales by Wong et al (1998). In this study, the morphology of S. aquatica was studied in further detail and molecular sequence data were obtained and analyzed to provide information about the phylogeny of this species. |
MATERIALS AND METHODS |
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. Abbreviations for frequent collectors are: Jinx Campbell (JC), J. Leland Crane (JLC), Walter L. Hurley (WLH), Payam M. Fallah (PMF), Kevin M. Robertson (KMR), Carol A. Shearer (CAS), and Mark J. Wetzel (MJW). Specimens are deposited at ILL unless otherwise indicated. Cultures are deposited at ATCC and the Department of Plant Biology, University of Illinois Fungus Collection.
Fungal isolates –
Cultures of the eight species used in this study (Table I) were obtained from single ascospores or asci according to the procedures of Shearer (1993) and Fallah and Shearer (2001). To confirm the identities of cultures, isolates were grown on peptone-yeast-glucose (PYG) agar slants (0.125% peptone, 0.125% yeast extract, 0.6% glucose, 1.8% agar) containing strips of balsa wood to stimulate reproduction. For DNA extraction, isolates were grown for 6–8 wk on PYG agar plates in the dark at 24 C. Sequences for additional species were obtained from GenBank (Table II). Species were selected for the molecular analyses based on morphological similarity to C. caudae-suis and S. aquatica and the current taxonomic placement of these two taxa. For example, several species with unfurling apical ascospore appendages similar to those of C. caudae-suis (Ceriosporopsis halima, Haligena salina, Halosarpheia fibrosa and H. viscosa) were included in the analyses. Since Ceriospora has been included provisionally within the Amphisphaeriaceae, Xylariales (Eriksson et al 2001), two representatives of the family were included. Both S. aquatica and P. aquatica are currently included in the Annulatascaceae, thus three representatives of this family were included. Ceriospora caudae-suis and S. aquatica both have asci that separate from the hymenium and lie free in the ascomal venter, a morphological character state of the Diaporthales, hence, four representatives of the Diaporthales were included.
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). DNA was amplified with Taq PCR Master Mix Kit (Qiagen 1999c) using the polymerase chain reaction (PCR) in a PTC-100 model 60 (MJ Research INC, Watertown, Massachusetts) thermal cyder. Universal fungal primers LROR, LR7 (Bunyard et al 1994), and LR6 (Vilgalys and Hester 1990) were used to amplify 1400-bp and 1100-bp regions of the 28S ribosomal gene. PCR reactions were carried out in 0.5 mL micro-centrifuge tubes in 50 µL volume containing 50 ng template DNA, 1 xPCR buffer, 1.5 mM MgCl2, 2 mM dNTP, 0.5 µM each primer, and 1.25 units Taq polymerase. Cycling parameters were as follows: LROR–LR7: an initial step of 96 C for 2 min, followed by 35 cycles of 96 C for 1 min, 55 C for 1 min, and 72 C for 1 min, and completed with a final step of 72 C for 10 min; LROR–LR6: 30 cycles of 94 C for 1 min, 50 C for 45 s, 50–72 C ramp for 1 min, 72 C for 1 min, and a final step of 72 C for 7 min. The PCR products were purified to remove excess primers, dNTPs and non-specific amplification products using a Qiaquick PCR Purification Kit (Qiagen 1999b). PCR products were sequenced directly using primers LROR (Bunyard et al 1994), and either LR7 (Bunyard et al 1994) or LR6 (Vilgalys and Hester 1990), at the University of Illinois Biotechnology Center, using Prizm fluorescent dye terminators on ABI 373A automated sequencers. The sequences were aligned with published sequence data (Table II) in Clustal X (Thompson et al 1997), then refined manually in Se-Al (Rambaut 1996), with ambiguous regions of alignment excluded from the analyses.
Phylogenetic analysis –
Cladistic analyses were performed in PAUP* 4.0b10 (Swofford 2002) using maximum parsimony, weighted parsimony, distance methods, and maximum likelihood criteria on 24 taxa. Outgroup comparison was used to polarize the character states and hence root the trees (Stevens 1980, Watrous and Wheeler 1981, Farris 1982, Darlu and Tassey 1987), employing the strategies of Maddison et al (1984) and Campbell (1999). These strategies demonstrate that global parsimony is best achieved when the pleisomorphic states are indicated in a more distant ancestor than the most recent common ancestor of the ingroup taxa. Several basal ascomycetes were explored as outgroup taxa (data not shown) and Gyromitra esculenta, Morchella esculenta, and Helvella lacunosa were chosen on the basis of reduced homoplasy and increased congruence in the phylogenetic reconstructions.
Maximum parsimony analyses were performed with heuristic searches employing random starting trees, random stepwise addition on 100 replicates, gaps treated as missing and a tree-bisection-reconnection branch-swapping algorithm. Parsimony tree scores for the consistency index (CI), retention index (RI) and rescaled consistency index (RC) were calculated for each tree generated. Weighted parsimony analyses were performed using a step matrix to weight nucleotide transformations based on the reciprocal of the observed transition: transversion (ti/tv) ratio (Spatafora et al 1998), which was calculated using MacClade (Maddison and Maddison 1992). Distance analyses were performed using Jukes-Cantor (1969), HKY85 (Hasegawa et al 1985), and uncorrected "P" distance methods. The maximum likelihood model was selected using Modeltest (Posada and Crandall 1998), which selects the appropriate evolutionary model for the dataset. Analyses were then performed in PAUP* with heuristic searches, and the evolutionary model set to GTR (Rodriguez et al 1990) using the base frequencies and among site rate variation that was calculated in Modeltest, and a tree-bisection-reconnection branch-swapping algorithm. The alternative topologies were tested using the Kishino-Hasegawa maximum likelihood (K-H) test (Kishino and Hasegawa 1989). Where more than one tree was generated in an analysis, the K-H test was applied to those trees first and the best tree from that analysis used in the K-H test on the alternative models. Bootstrap analyses (Felsenstein 1985) were performed on 1000 replicates using the same criteria for each evolutionary model as above. Decay indices (Bremer 1988, 1994) were calculated in AutoDecay (Eriksson 1998).
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RESULTS |
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TAXONOMY |
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), P. aquatica appears to have two spore forms. One spore form is ellipsoidal and similar in shape to those of C. caudae-suis, while the other form is cylindrical with distinct constrictions at the septa; C. caudae-suis has only ellipsoidal ascospores. We were unable to obtain the type specimen of P. aquatica to confirm the presence of two spore forms. Ceriospora caudae-suis has been described and illustrated in detail recently (Fallah and Shearer 2001), hence only ascus and ascospore measurements based on collections cited herein are provided.
Basionym: Ceriospora caudae-suis Ingold. Trans. Brit. Mycol. Soc. 34 (2): 210. 1951.
Asci 190–343 x 15–21 µm (range of means for asci in six collections = 210–270 x 17–20 µm, n = 10–30); ascospores 18–36 x 9–18 µm (range of means for ascospores in 34 collections = 26–31 x 10.5–15 µm, n = 7–50).
Specimens examined. ENGLAND. Anglia: Westmorland, Lake Windermere, on submerged, decorticated twigs of Fraxinus, 14 Oct 1949, C.T. Ingold (IMI 38506) Holotype. USA. ARKANSAS: Lee County, St. Francis River at Phillip's Bayou, St. Francis National Forest, 34°38'55''N, 84°38'12''W, UTM Zone 15, 716600mE, 3836425mN, water temperature 13.5 C, pH 7.5, on submerged, corticated wood, 14 Nov 1992, JLC, A40-04. FLORIDA: Hamilton County, Suwanee River State Park, sandy edge of the Suwanee River, water temperature 25 C, pH 6.5, on submerged, corticated wood (possibly Betula nigra), 16 June 1997, KMR, A40-25. ILLINOIS: Johnson County, Cache River at foot bridge to Heron Pond, 37°21'07''N, 88°55'27''W, on submerged, decorticated wood, 25 June 1996, CAS & WLH, A40-14. LOUISIANA: Caldwell Parish, bank of the Ouachita River where Hwy. 165 crosses it at Columbia, 30°01'51''N, 92°00'01''W, UTM Zone 15, 594399mE, 3544290mN, water temperature 32 C, pH 6.0, on submerged, corticated wood, 15 July 1997, KMR, A40-24; Washington Parish, east of Enon, on property of Weyerhauser Forest Industries, Hwy 437, depression in Nyssa sylvatica forest, 30°43'38''N, 90°05'03''E, water temperature 26 C, pH 6.5, on submerged, decorticated wood, 27 August 1997, KMR, A40-23; spring fed creek, water temperature 22 C, water pH 6.5; on submerged, corticated wood, 27 Aug 1997, KMR, A40-26; Bogue Chitto River, 30°43'38''N, 90°05'03''W, water temperature 28 C, pH 6.5, on submerged, partially decorticated wood, 27 August 1997, KMR, A40-21, on submerged wood, 27 Aug 1997, KMR A336-1. MAINE: York County, Waterboro, Buganut Pond, 43°30'39''N, 70°42'42''W, UTM Zone 19, 361640mE, 4818720mN, water temperature 26 C, pH 5.5, on submerged, decorticated wood, 03 July 1994, JLC, A40-09, A40-15. MISSISSIPPI: Franklin County, near Bude, edge of a sandbar of the Hamochitto River near intersection with Hwy 98, on submerged, corticated wood, 14 July 1997, KMR, A40-29; Holmes County, Big Black River at jct. with Rt. 19, east of West, 33°11'38''N, 89°46'14''W, UTM Zone 16, 241690mE, 3676040mN, water temperature 7 C, pH 5.5, on submerged wood, 27 Dec 1993, JLC, A40-06. NEW HAMPSHIRE: Coos County, Androscoggin River, 4.3 miles north of Milan village church, 44°36'53''N, 71°14'5''W, water temperature 13 C, water pH 4.5, on submerged, corticated wood, 24 May 1993, JLC, A40-10; White Mountain National Forest, Saco River at Davis Path off Rt. 302, 44°07'12''N, 71°21'38''W, UTM Zone 19, 311650mE, 4887700mN, water temperature 15 C, water pH 5.4, on submerged, partially decorticated wood, 04 Sep 1992, JLC, A40-03. NEW YORK: Adirondack Park, Raquette River at Tupper Lake Rod and Gun Club Landing, 44°11'78''N, 74°27'74''W, on submerged, decorticated wood, 05 Jul 1995, JLC, A40-12; South Bay of Raquette Lake at jct. with South Inlet and bridge on Rt. 28, 43°48'26''N, 86°36'36''W, UTM Zone 18, 531360mE, 4850320mN, water temperature 22 C, water pH 6.4, on submerged, decorticated wood, 08 Sep 1992, JLC, A40-02. NORTH CAROLINA: Cheoah River, 35°23'56'' N, 83°52'07'' W, altitude 1666 ft., water temperature 27 C, pH 5.5, on submerged, decorticated wood, 18 July 2000, JC, A40-32, Oconaluftee River, 35°28'47'' N, 83°19'12'' W, altitude 1900 ft, water temperature 19.5 C, pH 5, on submerged, decorticated wood, 18 July 2000, JC, A40-31. OREGON: Florence County, Mercer Lake at boat launch 4092, 44°03'03'' N, 124°04'07''W, on submerged wood, 28 Aug 1999, WLH & CW Hurley, A336-2, water temperature 19.5 C, water pH 5, on submerged, decorticated wood, 26 Jun 2001, CAS & WLH, A40-36; Mill Creek on Rd. to Loon Lake North of bridge at Camp Creek Rd. cut-off, 43°35'58''N, 123°50'48''W, water temperature 17 C, water pH 5.3, on submerged, decorticated wood, 26 Jun 2001, CAS & WLH, A40-36. PENNSYLVANIA: Columbia County, Catawwiia Creek at jct. with Rt. 924, 40°52'56''N, 76°09'15''W, on submerged, decorticated wood, 06 Jul 1995, JLC, A40-13. TENNESSEE: Great Smoky Mountains National Park, near Gatlinburg, Mill Creek, Cades Cove Loop Road, 39°16'19''N, 75°29'25''W, water temperature 11 C, pH 4.5–5.0, on submerged, decorticated wood, 10 Dec 1999, CAS & WLH, A40-30. WEST VIRGINIA: Pocahontas County, Williams River at jct. with Rt. 150, 38°19'85''N, 80°13'30''W, on submerged, decorticated wood, 07 Jul 1995, JLC, A40-11; Knapps Creek west of Marlington, 38°12'69''N, 80°02'91''W, on submerged, decorticated wood, 07 Jul 1995, JLC, A40-16. WISCONSIN: Adams County, Lemonweir River at jct. with Rt. HH south of Rt. 82, 43°46'16''N, 89°53'10''W, water temperature 20 C, water pH 5.5, on submerged, decorticated wood, 31 Jul 1992, JLC, WLH & CAS, A40-01; Iron County, Layman Creek at jct. with Rt. 51, 46°19'55''N, 90°09'43''W, on submerged, decorticated wood, 29 Nov 1995, PMF & JLC, A40-17; Iron County, Manitowish River at jct. with Rt. 51, Manitowish Waters, 46°08'14''N, 89°54'42''W, on submerged, decorticated wood, 02 May 1997, PMF, A40-22; Oneida County, Tomahawk River, 45°50'06''N, 89°48'23''W, UTM Zone 16, 282017mE, 5079591mN, on submerged, decorticated wood, 19 Sep 1996, PMF, A40-27; on submerged, corticated wood, 02 May 1997, PMF, A40-28; Vilas County, Grassy Creek, 46°09'07''N, 89°35'56''W, UTM Zone 16, 299326mE, 5114215mN, on submerged, decorticated wood, 18 June 1996, PMF & JLC, A40-19; on submerged, decorticated wood, 02 May 1997, PMF, A40-20; Mishonogam Creek, 45°54'43''N, 89°45'33''W, UTM Zone 16, 286011mE, 5087969mN, on submerged, decorticated wood, 28 June 1995, PMF, A40-18, Rice Creek at jct. with Rt. K, 1.8 miles north of Newcomb Lane, 46°08'23''N, 89°44'58''W, UTM Zone 16, 287724mE, 5113272mN, on submerged, decorticated wood, 02 May 1997, PMF, A40-35; Washburn County, St. Croix National Scenic Riverway, Fritz Landing, Namekagon River, 46°2'41''N, 92°2'0''W, UTM Zone 15, 574790mE, 5099270mN, water temperature 9 C, water pH 5.0, on submerged, decorticated wood, 25 Oct 1993, JLC, A40-08.
Although unreported for more than fifty years since its original description from the Lake District by Ingold (1951), Pseudoproboscispora caudae-suis is one of the most commonly occurring species in temperate North America (Fig. 1). It is often found fruiting on twigs examined immediately after collection, indicating that it is fruiting in situ. It occurs on submerged corticated or decorticated woody debris in both lentic (Fallah and Shearer 2001) and lotic habitats. We have collected it at water temperatures from 7–32 C and at pH (4.5–) 5–6.5 (–7.5). This species occurs at latitudes 30°–55°N and longitudes 3°–124°W; it has not yet been reported from the eastern or southern hemisphere.
Ingold (1951), in his original description of this species, accurately characterized the morphology and behavior of the asci and ascospore appendages. There are, however, two notable features of this fungus that have not yet been reported. At first, the appendages appear to be a single filament surrounded by gelatinous material that stains in aqueous nigrosin (Fig. 6). As the gelatinous material dissolves in water, the appendage, which initially is tightly coiled, begins to uncoil (Fig. 7) and the distal ends of the appendages separate into many fine, long filaments (Fig. 8); staining with aqueous nigrosin enhances the resolution of the filaments. These fine filaments appear to be compacted together near the ascospore apices to form the stout base of the appendage (Figs. 7, 8). A second observation is that the pedicels of the asci separate from the hymenium (deciduous asci) when the ascoma is crushed in water (Figs. 44, 45 in Fallah and Shearer 2001). As pointed out by Ingold (1951), the ascus breaks in half along a transverse line of weakness to release ascospores, and the lower part of the ascus disintegrates (a sort of back door discharge). Deciduous asci and back door discharge occur in species of Annulatascus velatisporus, the type of the genus and family, Submersisphaeria, Ophioceras Sacc. (Shearer et al 1999), and Pseudohalonectria Minoura & T. Muroi (Shearer 1989), all pyrenomycete genera with species that occur commonly in freshwater habitats.
Two collections of P. caudae-suis, A336-1 and A336-2, had mostly 3-septate ascospores (Figs. 4, 5). Isolates of A336-2 produced both one- and three-septate ascospores in culture (cultures of A336-1 were not made). Although older discharged ascospores of P. caudae-suis with two or three septa were found in other collections, ascospores in the A336 collections were septate prior to discharge from the ascus. Initially, we thought these collections might be a new species of Pseudoproboscispora but careful morphological comparisons did not reveal differences between the A336 collections and P. caudae-suis, other than septation, sufficient to support the establishment of a new species. Wong and Hyde (1999) report the ascospores of P. caudae-suis as being pale brown. We did not find pale brown ascospores except for discharged older spores that often had more than one septum. Mature, freshly discharged ascospores have hyaline walls but appear pale yellow because they are finely multiguttulate with pale-yellow lipid material. Ingold (1951) reports the ascospores as being pale yellow due to presence of yellow oil globules.
Pseudoproboscispora caudae-suis differs from P. aquatica in ascus and ascospore size and ascospore septation. In addition, the ascospores of P. aquatica appear to be somewhat dimorphic, i.e., fusiform and not constricted at the septa or cylindrical and constricted at the septa (as seen from the illustrations but unreported in Wong and Hyde 1999), while those of P. caudae-suis are consistently broadly fusiform (Figs. 4–8).
Ascomata scattered or aggregated, immersed, partially immersed or superficial, black, ostiolate, with a short, central neck (Fig. 12). Venter subglobose or globose, 300–520 x240–530 µm. Peridium 31–43 µm thick, composed of 5 to 7 layers of rather large brown pseudoparenchymatic cells, 6–17 x 3–4 µm (Fig. 15), outer cells darker, inner cells subhyaline, textura angularis in face view and median longitudinal section (Fig. 14). Neck cylindrical, 210–1120 x 90–140 µm, tapering to 60–70 µm, black, lighter at the apex, periphysate (Figs. 12, 13). Paraphyses simple, septate, hyaline, 125–180 x 7–8 µm at base, tapering to 2 µm at the apex (Fig. 16). Asci deciduous from the hymenial layer (Figs. 17, 18, 20, 21), cylindrical, with rounded or truncated apex, narrowing to an attenuated stipe, 160–245 x 8–12 µm (Figs. 17, 18), extending in water to 280 µm (Fig. 20) and rupturing near the midpoint of the ascus to release ascospores, with a prominent refractive apical ring, 4–6 x 5–6 µm (Figs. 10, 19), J-, staining light blue with a weak solution of aqueous cotton blue containing 2% lactic acid, with 8 uniseriate or overlapping uniseriate ascospores (Figs. 17, 18). Ascospores 18–26 x 7–9 µm, ellipsoidal to oval, brown, with a single septum at the midpoint or above, multiguttulate, with very small, hyaline, cap-like appendages about <2 µm wide (Figs. 22, 23).
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Our collections agree in general with the protologue of S. aquatica (Hyde 1996), however, we were unable to obtain the type specimen for a direct comparison. Ascomal venters were longer and wider, and necks longer in our collections. The neck of the holotype was reported as 560 µm in diam, but our measurements never exceeded 140 µm in diam. The range in ascus dimensions was slightly greater in our collections than that reported in the type description but this likely reflects within species variation revealed by the greater number of collections examined. Although ascospore measurements were very similar to those reported in the protologue (Hyde 1996), our interpretation of ascospore morphology differs. Hyde described the ascospores as having apical germ pores. The definition of a germ pore is "a hole or opening in the wall of a spore" (Ulloa and Hanlin 2000). In our collections, the ascospores have small hyaline, cap-like appendages at both ends (Figs. 22, 23). In early stages of ascospore formation, the ascospore wall is hyaline throughout (Fig. 17). Pigment deposition occurs throughout the entire ascospore wall (Figs. 10, 17, 18) and holes at the apices could not be seen. The apical appendages are only visible when the ascospores are mounted in water or glycerin but disappear in lactic acid. Thus, spores mounted in lactic acid might appear to have biapiculate germ pores at the point of attachment of the appendages. Similar hyaline cap-like apical appendages are illustrated for S. aquatica (Hyde 1996) and S. bambusicola Zhou & Hyde (Zhou and Hyde 2000) but they are labeled as germ pores. We were unable to find germ pores with the light microscope at 1000x. EM and/or SEM studies of the ascospores may be required to resolve the questionable presence of germ pores and the origin of the apical appendages, which is also unknown.
Submersisphaeria aquatica was originally described from Queensland, Australia, and we report it from six states in the USA (Fig. 2). We have collected it on corticated and decorticated woody debris submerged in lentic and lotic habitats at temperatures ranging from 11–30 C and pH from 3.7–6.5.
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DISCUSSION |
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) that is bipartite at the EM level (Wong et al 1999). Réblová and Winka (2001) used molecular data to demonstrate that the presence of such a ring did not unite otherwise phenotypically diverse fungi in a clade. Our molecular results support the inclusion of P. caudae-suis in the Annulatascaceae even though it does not have a massive apical ring (Fig. 9). Although Réblová and Winka (2001) lacked molecular data for some genera of Annulatascaceae, they speculated based on morphological data that Pseudoproboscispora (sub Proboscispora) and Submersisphaeria were difficult to assign to any of the ascomycete families and indicated that S. aquatica shows an affinity to Pseudovalsaria Spooner, Clypeosphaeriaceae, Xylariales. Our 28S rDNA data (Fig. 3) shows very strong support for the inclusion of Pseudoproboscispora and Submersisphaeria in Annulatascaceae. Although the apical ring in P. caudae-suis is not massive, it is J- and refractive (Fig. 9). Other shared phenotypic characters that support placement of P. caudae-suis and S. aquatica in Annulatascaceae include: occurrence on submerged wood; globose, ostiolate, dark-pigmented ascomata; peridium of thin-walled cells darkened to the outside; periphysate neck; hamathecium of long, tapering, septate paraphyses; long cylindrical asci that are deciduous from the hymenium, extend in length in water, and exhibit back door dehiscence; and uniseriate to overlapping uniseriate, ellipsoid ascospores. In addition to the shared characters listed above, P. caudae-suis, and P. aquatica have bipolar unfurling apical appendages with a stout region next to the ascospore apices. These characteristic appendages appear to be apomorphic and a defining character for the genus Pseudoproboscispora.
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ACKNOWLEDGMENTS |
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FOOTNOTES |
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Accepted for publication May 14, 2002.
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LITERATURE CITED |
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