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Trichomycetes from China and the description of three new Smittium species

     Department of Biology, Saint Mary’s University, Halifax, Nova Scotia, B3H 3C3 Canada

Shengquan Xu

     College of Life Science, Shaanxi Normal University, Xi’an, PR China

	ABSTRACT

TOP ABSTRACT INTRODUCTION MATERIAL AND METHODS RESULTS TAXONOMY DISCUSSION LITERATURE CITED

 

Trichomycetes were recovered from the guts of aquatic insect larvae collected from a stream in the Qinling Mountains in western China. These collections included Smittium hecatei, known only from Spain, as well as Smittium simulii and Stachylina penetralis, which appear to be more widely distributed. Caudomyces japonicus, previously recorded only from Japan, also is reported from crane fly larvae (Antocha sp.) from China. We describe three new species, Smittium chinliense from a tipulid host, as well as Smittium naiadis and Smittium nodifixum, both from chironomid larvae. A probable new species of Gauthieromyces was collected in mayfly nymphs and is illustrated but not described here.

Key words: aquatic insects, Caudomyces, Gauthieromyces, gut fungi, Harpellales

	INTRODUCTION

TOP ABSTRACT INTRODUCTION MATERIAL AND METHODS RESULTS TAXONOMY DISCUSSION LITERATURE CITED

 
Trichomycetes living in the guts of arthropods, mostly immature stages of insects in aquatic systems, are widely distributed. The best documented occurrences of these fungi are from the United States and western Europe (France, England and Spain) (Manier 1970, Moss et al 1975, Lichtwardt 1986, Santamaria 1997, Lichtwardt et al 2001, Valle and Santamaria 2004). Fewer published records exist for South America (Lichtwardt and Arenas 1996, Lichtwardt et al 2000), Central America (Lichtwardt 1994, 1997), Australia (Lichtwardt and Williams 1990) and New Zealand (Williams and Lichtwardt 1990). Although the Trichomycete survey to date has covered many widely separated continents, large areas with prime habitat and high arthropod diversity have yet to be examined in a systematic way (e.g. in Canada, Africa and China). The abundance and frequency of occurrence of Trichomycetes in hosts from areas that already have been surveyed suggest that sampling in new geographic areas will yield many undescribed species and may help better elucidate the ecology and biogeography of this fungal group.

Trichomycetes are poorly known from Asia. One publication reported species recovered in Japan (Lichtwardt et al 1987) and another describing parasites of blackflies from China noted the presence of a Trichomycete, Harpella melusinae Leger & Dubosq, in these hosts (Adler et al 1996). Chien and Hsieh (2001) reported Trichomycetes in the orders Eccrinales and Asellariales from aquatic and terrestrial crustaceans as well as millipedes in Taiwan, including two new species of eccrinids from crustacean hosts.

This paper reports Trichomycetes from aquatic insects collected in the Niu Bei Liang nature protection area, Qinling Mountains approximately 40 km from Xi’an, in western China. We describe three new species of Trichomycetes in the genus Smittium from these collections as well as six other taxa of trichomycetes.

	MATERIAL AND METHODS

TOP ABSTRACT INTRODUCTION MATERIAL AND METHODS RESULTS TAXONOMY DISCUSSION LITERATURE CITED

 
Insects were collected in a D-net by kick-sampling (Lichtwardt et al 2001) in a fast-flowing, shallow stream (Hao River) in the Niu Bei Liang nature protection area, western China (33° 55.21N 108° 56.49E) 26 and 27 Apr 2005. The stream had a sandy and pebbly bottom with little organic material and flowed through an area of large boulders. The stream yielded immature stages of mayflies (Ephemeroptera), stoneflies (Plecoptera), larval flies (Diptera) and beetles (Coleoptera). Samples of tipulid (Diptera) larvae also were hand-picked from rocks in the splash zone of a small waterfall near the sampling area.

The hindgut and peritrophic matrix (in the dipterans) were dissected out of the hosts and examined microscopically for the presence of trichomycetes and other gut inhabitants. Fungal identifications were made with the Lucid keys available at the University of Kansas Trichomycete Website (www.nhm.ku.edu/fungi/). Semipermanent slides, stained with lactophenol cotton blue were prepared as described in Lichtwardt et al (2001). Digital photomicrographs of the diagnostic features were taken at the Saint Mary’s University Taxonomy Laboratory, Halifax, Nova Scotia, Canada. Voucher specimens of new species and Caudomyces japonicus Lichtw. Kobayasi & Indoh were deposited in the herbarium (SANU), College of Life Science, Shaanxi Normal University, Xi’an, 710062, Shaanxi, People’s Republic of China. Other voucher specimens are in the collection of DBS, Saint Mary’s University.

	RESULTS

TOP ABSTRACT INTRODUCTION MATERIAL AND METHODS RESULTS TAXONOMY DISCUSSION LITERATURE CITED

 
The insects contained several species of Trichomycetes in the order Harpellales as well as protists in the genus Paramoebidium, which is currently classified within the Trichomycetes in the order Amoebidiales (Lichtwardt et al 2001). Harpellids recovered from midges (Diptera: Chironomidae) included Smittium hecatei L.G.Valle & Santam. and Sm. simulii Lichtw. from the hindgut. Stachylina penetralis Lichtw. was observed on the peritrophic matrix of these hosts. In addition two species of Smittium collected in chironomids had unique morphological features that warrant description as new species, Sm. naiadis (FIGS. 1–4) and Sm. nodifixum (FIGS. 5–8). Thalli of Paramoebidium spp. were common in the hindgut of these dipterans but insufficient characters were available to identify them to species.

View larger version (158K): [in this window] [in a new window]   FIGS. 1–4. Smittium naiadis. 1. Thallus branching and holdfast characteristics. 2. Details of the holdfast (arrow). 3, 4. Trichospores. The arrows indicate apparent thickenings in collar. Bars = 20 µm.

View larger version (146K): [in this window] [in a new window]   FIGS. 5–8. Smittium nodifixum. 5. Thallus arising from bulbous holdfast (arrow). 6. Details of knobby holdfast and individual branches. 7. Trichospore showing campanulate collar. 8. Trichospore with single appendage. Bars = 20 µm.

View larger version (131K): [in this window] [in a new window]   FIGS. 9–11. Smittium chinliense. 9. Thallus with foot-like holdfast (arrow). 10, 11. Released trichospores. Bars = 20 µm.

View larger version (90K): [in this window] [in a new window]   FIGS. 12–13. Caudomyces japonicus. 12. Trichospores with long collars attached to fertile branch. 13. A single trichospore with long collar typical of the species. Bars = 20 µm.

View larger version (94K): [in this window] [in a new window]   FIGS. 14–15. Gauthieromyces sp. 14. Immature thallus showing sparse branching from swollen basal cell at arrow. 15. Terminal fertile branch with many horseshoe-shaped trichospores. Bars = 20 µm.

	TAXONOMY

TOP ABSTRACT INTRODUCTION MATERIAL AND METHODS RESULTS TAXONOMY DISCUSSION LITERATURE CITED

 
Smittium naiadis Strongman et Shengquan Xu, sp. nov. FIGS. 1–4

Thallus diffusus, ramis primariis e fasciculo simplici cellularum basalium muco velatarum orientibus; ramificatio secundaria per longitudinem et in extremis ramorum principalium valde verticillata. Trichosporae cylindricae longae angustaeque (34–36 x 2.5–3.5 µm), cum collo conspicuo breve (3 µm) quod ut videtur 1–2 tumores plerumque habet. Trichosporae appendice singula tenui instructae. Zygosporae incognita. In proctodaeo larvarum Chironomidarum.

Thallus diffuse, primary branches arise from a simple cluster of basal cells covered with mucilage (FIGS. 1, 2), secondary branching strongly verticilliate along the length and at the end of the primary branches (FIG. 1). Trichospores cylindrical, long and narrow (34–36 x 2.5–3.5 µm) with a conspicuous, short (3 µm) collar that commonly appears to have 1 or 2 thickenings (FIGS. 3, 4). Trichospores with a single, thin appendage. Zygospores not found. In hindgut of bloodworms (Chironomidae).

Etymology. – from the Latin naias meaning from a nymph, referring to the source being the immature (larval) stage of the host.

Specimens examined. – CHINA. SHAANXI PROVINCE: Hao River, Niu Bei Liang nature protection area, 33° 55.21N 108° 56.49E. Slide CHI-2 prepared from the hindgut of a chironomid larva collected 27 Apr 2005 (HOLOTYPE SANU).

Note. – This slide also contains thalli and trichospores of Sm. nodifixum.

Commentary. – The trichospores in Sm. naiadis are long and narrow compared to most other species of Smittium (www.nhm.ku.edu/fungi/). The trichospore collar has distinctive thickenings and this feature seems to be consistent, therefore diagnostic. Smittium gravimetallum Lichtw. Ferrington & Hayford (Ferrington et al 2000) has narrow spores but in this species they are shorter on average and have a distinct swelling in the midregion, which is lacking in Sm. naiadis. Another species with long spores is Sm. elongatum Lichtw. but they are wider than in Sm. naiadis, the holdfast in Sm. elongatum is inconspicuous (Lichtwardt 1972) whereas it is prominent and embedded in mucilage in this new species. There was no apparent coiling in the appendage as is described for Sm. elongatum. Smittium angustum M.C. Williams & Lichtw. has narrow spores but they are shorter and the thallus features differ greatly from Sm. naiadis (Lichtwardt and Williams 1992).

Smittium nodifixum Strongman et Shengquan Xu, sp. nov. FIGS. 5–8

Thallus compactus, ramis primariis e haptero conspicuo nodoso orientibus; ramificatio secundaria sparsa. Trichosporae subcylindricae, 26–29.5 x 5–6 µm, cum collo campanulato 5–6 µm longo. Zygosporae incognitae. In proctodaeo larvarum Chironomidarum.

Thallus compact, primary branches arising from a conspicuous knobby holdfast, secondary branching sparse (FIGS. 5, 6). Trichospores subcylindrical 26–29.5 x 5–6 µm with a campanulate collar 5–6 µm long (FIGS. 7, 8). Zygospores not found. In hindgut of bloodworms and other midge species (Chironomidae).

Etymology. – from the Latin nodos and fixum, meaning fasten with a knot, in reference to the conspicuous, knobby holdfast in this species.

Specimens examined. – CHINA. SHAANXI PROVINCE: Hao River, Niu Bei Liang nature protection area, 33° 55.21N 108° 56.49E. Slide CHI-4 prepared from the hindgut of a chironomid larva collected 27 Apr 2005 (HOLOTYPE SANU).

Commentary. – The spore dimensions in Sm. nodifixum overlap with nine described species in this genus (www.nhm.ku.edu/fungi/). However, when thallus features are considered along with spore shape and collar characteristics, the species can be separated. Smittium acutum Lichtw. & Grigg and Sm. annulatum Lichtw. share some features with this new species but the holdfast structures differ; Sm. acutum has a flared collar (Lichtwardt and Grigg 1998), compared to a campanulate collar in Sm. nodifixum, and the spores of Sm. annulatum are smaller on average (Lichtwardt 1997). Smittium commune Lichtw. & Grigg has variable spores (Lichtwardt and Grigg 1998) so the dimensions overlap substantially with Sm. nodifixum. However the spores in Sm. commune are ellipsoidal and shorter, on average, with a shorter collar so it can be differentiated from Sm. nodifixum on this basis. The new species resembles Sm. cylindrosporum Lichtw. & Arenas but trichospores in that species have a conspicuous bulge in the middle and the branching pattern in the thallus differs (Lichtwardt and Arenas 1996). There is no mention of a distinctive holdfast in the description of Sm. cylindrosporum, which is a conspicuous feature in Sm. nodifixum.

Smittium chinliense Strongman et Shengquan Xu, sp. nov. FIGS. 9–11

Thallus e haptero conspicuo pediformi oriens; ramificatio sparsa, trichosporis 3–4 in terminiis ramorum fertilium typice formantibus. Trichosporae cylindricae vel subcylindricae, 28.5–36 x 5–6 µm, cum collo tubulari 5–6 x 4 µm saepe sporam latitudine aequanti, appendice singula instructae. Zygosporae incognitae. In proctodaeo larvarum Tipulidarum.

Thallus arising from a conspicuous foot-like holdfast, branching sparse, typically with 2–4 trichospores forming at the tips of fertile branches (FIG. 9). Trichospores cylindrical to subcylindrical, 28.5–36 x 5–6 µm with a tubular collar 5–6 x 4 µm, often as wide as the spore, with a single appendage (FIGS. 10, 11). Zygospores not found. In hindgut of crane fly larvae (Tipulidae).

Etymology. – from Qinling (pronounced chinlee) Mountains of China where the collections were made.

Specimens examined. – CHINA. SHAANXI PROVINCE: Hao River, Niu Bei Liang nature protection area, 33° 55.21N 108° 56.49E. Slide CHI-13 prepared from the hindgut of a tipulid (Antocha sp.) larva collected 27 Apr 2005 (HOLOTYPE SANU).

Note. – This slide also contains thalli and trichospores of Caudomyces japonicus.

Commentary. – This fungus does not resemble any of the three species known from tipulid larvae, Sm. simulatum Lichtw. & Arenas (Lichtwardt and Arenas 1996), Sm. simulii (Lichtwardt 1964) and Sm. tipulidarum M.C. Williams & Lichtw. (Williams and Lichtwardt 1987) (www.nhm.ku.edu/fungi/). The long, cylindrical trichospores of Sm. chinliense are similar in dimensions to Sm. elongatum and Sm. cylindrosporum, both from dipteran larvae (chironomids), but the collar in these species is narrower than the spore (Lichtwardt 1972, Lichtwardt and Arenas 1996) whereas the collar in Sm. chinliense is as wide as the spore. Also the thallus in both known species has verticilliate branching whereas Sm. chinliense does not.

	DISCUSSION

TOP ABSTRACT INTRODUCTION MATERIAL AND METHODS RESULTS TAXONOMY DISCUSSION LITERATURE CITED

 
This paper expands the known geographic range of several Trichomycete species to another continent, China, and describes three new species, Sm. naiadis, Sm. nodifixum and Sm. chinliense. Smittium hecatei is a recently described species from Spain (Valle and Santamaria 2004) and the Chinese collection fits the trichospore size ranges and characteristics for both spores and spores ß as described by these authors. We did not see the "elephant leg" basal cell that is typical for this species but the thallus characteristics match otherwise. Smittium simulii is a species that is widely distributed and common in chironomid larvae (Lichtwardt et al 2001) so it is not surprising that we collected it in these hosts from China as well. Stachylina penetralis has been reported from Japan as well as Europe (Lichtwardt et al 2001) suggesting it might have broad distribution.

Smittium is the largest, most widely distributed genus of Trichomycetes with more than 70 species described to date from a variety of insect hosts. Therefore typical morphological characteristics used to define species (e.g. spore dimensions) overlap greatly, which makes species identification challenging. This problem is further complicated by the occurrence of several dimorphic species that exhibit two distinct spore morphologies arising from the same thallus (Valle and Santamaria 2004, White and Lichtwardt 2004). Thallus characteristics such as the holdfast and a more standardized description of the branching patterns may help delineate taxa. More molecular studies like those described in Gottlieb and Lichtwardt (2001) will provide data on genetic relatedness that might help refine our interpretation of the significance of certain morphological characters and variation among species.

Caudomyces japonicus specimens recovered from Antocha sp. larvae in the Qinling Mountains matched the type species description well and are from the same genus of host (Lichtwardt et al 1987). This is an extension of the geographic range for this fungus and although we, like the authors on the original description, were unable to identify the host to species it might be an indication that the host and fungus are widespread in Asia. The original description of C. japonicus included an illustration but this second collection from China supplements the illustration by providing photomicrographs (FIGS. 12, 13), which might help other collectors recognize the species and give us a better understanding of the morphological variation within the species.

The discovery of another species of Gauthieromyces is significant in that this monotypic genus has not been reported since its description from France in 1960 (Lichtwardt 1983). No type material exists for G. microsporus and subsequent attempts to collect the species at the type locality were unsuccessful (Lichtwardt 1983). The characteristics of the Gauthieromyces sp. from China differ from the type in that the swollen basal cell is shorter, tapers to the central axis more abruptly (FIG. 14) and the branching is more profuse with typically many spores (20+) per fertile branch (FIG. 15). The structure of the thallus is similar to species of Graminella, also known from baetid mayflies (www.nhm.ku.edu/fungi/). However the trichospores are diagnostic in that no other genus contains species with small, horseshoe-shaped spores. The spores in the Chinese collection are the same length around the curve (10–12[–14] µm) as the type and about 2 µm wide; no width is given in the original description (Lichtwardt 1983). The independent discovery of another species in both India and China provides reference material that will be useful when other species are found. The results from this limited survey of two habitats along a short stretch of one river suggest that China and other unexplored areas in Asia are rich in Trichomycetes and no doubt more extensive surveys will add many more new taxa.

	ACKNOWLEDGMENTS

 
We thank Dr Zhezhi Wang, dean of the College of Life Science, Shaanxi Normal University (SNNU), for financing the field trip to the Qinling Mountains, and both SNNU and Saint Mary’s University for financing travel for DBS. We thank Dr Zhongmin Dong, Biology Department, Saint Mary’s University, for arranging the visit (by DBS) to China and for aiding the communication that helped to produce this manuscript. Mr Yi Bai and Ms Deng Sufang ably assisted us in the field. Carolyn Bird provided the Latin diagnosis. We also thank Dr R.W. Lichtwardt and Dr M.M. White for their help and discussions surrounding Smittium species and for editorial suggestions on the paper.

	FOOTNOTES

 
Accepted for publication March 1, 2006.

¹ Corresponding author. E-mail: doug.strongman{at}smu.ca

	LITERATURE CITED

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This Article Abstract Full Text (PDF) Services Similar articles in this journal Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Strongman, D.B. Articles by Xu, S. Search for Related Content PubMed Articles by Strongman, D.B. Articles by Xu, S. Agricola Articles by Strongman, D.B. Articles by Xu, S.