Caecomyces sympodialis sp. nov., a new rumen fungus isolated from Bos indicus

A new anaerobic rumen fungus was isolated from the rumen fluid of a yellow cow (Bos indicus). This fungus appears to be a previously undescribed species of the genus Caecomyces, it possessing uniflagellate zoospores, a spherical holdfast, tubular sporangiophores and bulbous rhizoids. This new fungus also features distinctive multisporangiate thallus sympodially distributed on sporangiophores. The fungus resembles Caecomyces communis and C. equi in that it characterizes bulbous rhizoids and uniflagellate zoospores but differs from C. communis and C. equi in that it possesses multisporangiate and sympodial sporangia. This new fungus and Cyllamyces aberensis both reveal similar morphology during early thallus development in having a spherical holdfast, but they vary from unbranched sporangiophores and additional bulbous rhizoids. In addition, the molecular phylogenetic analyses ITS1 (internal transcribed spacer 1) also conform to the results of the morphological examinations of Caecomyces. For the mentioned reasons, this new species of fungus is described as Caecomyces sympodialis sp. nov. The genera of Neocallimasticaceae and species of Caecomyces are also keyed out.     

Ultrastructural observations of mature and encysting zoospores ofPythium proliferum de Bary

Summary The process of zoospore maturation and encystment inP. proliferum was studied by electron microscopy. General ultrastructural features of the mature, swimming zoospore were found to be similar to those previously described for other oomycetes in both the attachment and ultrastructure of the flagella as well as the type and distribution of cellular organelles. Associated with extensive areas of RER in the mature zoospores were unusual, electrondense, bar-like structures. These structures were found in the groove region of young zoospores and at the periphery of encysting zoospores. Their possible function is discussed. The five main types of vesicles observed during encystment, as seen grouped in this study, along with the vesicles described in previous studies of oomycete encystment, were in table form and individually discussed. Interesting correlations appear to exist in the types of vesicles that are present within the oomycetes studied thusfar.     

Two New Kremastochrysopsis species,K. austriaca sp. nov. and K. americana sp. nov. (Chrysophyceae)1

Melting summer snow in the Austrian Alps exhibited a yellowish bloom that was mainly comprised of an unidentified unicellular chrysophyte. Molecular data (18S rRNA and rbcL genes) showed a close relationship to published sequences from an American pond alga formerly identified as Kremastochrysis sp. The genera Kremastochrysis and Kremastochrysopsis are morphologically distinguished by the number of flagella observed with the light microscope, and therefore we assigned the Austrian snow alga and an American pond alga to the genus Kremastochrysopsis. Transmission and scanning electron microscopy revealed that swimming cells had two flagella oriented in opposite directions, typical for the Hibberdiales. Molecular phylogenetic analyses showed that both new species were closely related to Hibberdia. Kremastochrysopsis ocellata, the type species and only known species, has two chloroplasts per cell and the zoospores have red eyespots. Our two organisms had only a single chloroplast and no zoospore eyespot, but their gene sequences differed substantially. Therefore, we described two new species, Kremastochrysopsis austriaca sp. nov and Kremstochrysopsis americana sp. nov. When grown in culture, both taxa showed a characteristic hyponeustonic growth (hanging below the water surface), whereas older immotile cells grew at the bottom of the culture vessel. Ecologically, Kremastochrysopsis austriaca sp. nov., which caused snow discolorations, had no close phylogenetic relationships to other psychrophilic chrysophytes, for example, Chromulina chionophilia, Hydrurus sp., and Ochromonas-like flagellates.     

Culture studies of Acrochaete leptochaete comb. nov. and A. wittrockii comb. nov. (Chaetophoraceae, Chlorophyceae)

A culture study of the plants known as Phaeophila leptochaete (Huber) Nielsen and P. wittrockii (Wille) Nielsen proved that these are distinct species. Both have the Acrochaete type of hairs, and therefore, the new combinations Acrochaete leptochaete and A. wittrockii are introduced. Several isolates of both species were started from plants collected at different places in Europe.
A. leptochaete in culture was mainly characterised by having two or more (up to six) pyrenoids in many of its cells, while in A. wittrockii there invariably was only one. The swarmers formed by A. leptochaete had two or four flagella. Several hairs from one basal swelling were sometimes observed in one of the isolates. Another isolate often formed a hair on the germinated zoospore. The isolates of A. wittrockii had a variable morphology, from unicellular plants to large pseudoparenchymatous cell masses. Different kinds of swarmers were observed; zoospores with three flagella seemed typical, but ones with two or four flagella were also observed. Small, pale biflagellate swarmers were assumed to be gametes. One of the isolates of A. wittrockii differed from the rest as the only swarmer type observed was zoospores with two flagella. Chlorophilum ephemerum is considered identical to A. wittrockii. Observations on hair formation in A. repens suggested that the hair structure represents a separate cell.     

Zoosporulation of a new Perkinsus species isolated from the gills of the softshell clam Mya arenaria

A gill-associated Perkinsus sp. isolated from the softshell clam (Mya arenaria) is described as a new species, P. chesapeaki sp. nov. Examination of the parasite in seawater cultures revealed life cycle stages and zoosporulation processes similar to those described for other species of the genus Perkinsus. Prezoosporangia developed thickened cell walls upon contraction of the cytoplasm and development of a distinctive clear area between the cell wall and the protoplast. Successive bipartition of the protoplast led to the formation of hundred's of zoospores within mature sporangia. Zoospores were released into seawater through one or more discharge tubes. Ultrastructural studies revealed an oblong zoospore possessing two flagella that arose from a concave side located in the upper third of the zoospore body. The anterior flagellum possessed a unilateral array of hair-like structures. A large anterior vacuole and basolateral nucleus dominated the cytoplasm of the zoospore body. The presence of a rudimentary apical complex including an open-sided conoid, rhoptries, micronemes, and subpellicular microtubules were also discerned. Differences in zoospore morphology, and sequence analyses of two genes previously reported, support the designation of the gill-associated Perkinsus from the softshell clam as a new species.     

ULTRASTRUCTURE OF CEPHALEUROS VIRESCENS (CHROOLEPIDACEAE; CHLOROPHYTA). III. ZOOSPORES

Transmission electron microscopic examination of Cephaleuros virescens Kunze growing on leaves of Camellia spp. and Magnolia grandiflora L. indicates that unreleased zoospores in mature zoosporangia are similar to those produced by the related genus Phycopeltis epiphyton Millardet and unlike the quadriflagellate motile cells produced by taxa in other families of Chlorophyta. The zoospores bear four smooth isokont bilaterally “keeled” flagella containing typical “9 + 2” axonemes and lacking scales. Flagellar insertion is apical and the parallel basal bodies overlap laterally at two levels. A cross section through the four basal bodies shows a trapezoidal arrangement wherein the two upper (anterior) basal bodies are closer together than are the lower (posterior) two. Serial sections indicate that diagonally opposing upper and lower basal bodies anchor flagella which emerge from the same side of the apical papilla. Each of the four basal bodies is associated with a microtubular spline which extends beneath the plasmalemma to the posterior end of the zoospore. A distinct multilayered structure is associated with each of the lower basal bodies. A nucleus, mitochondria (two of which are closely associated with the nucleus and spline microtubules), a chloroplast, and cytoplasmic haematochrome droplets are present in each zoospore. Pyrenoids and eyespots are absent. Flagellar insertion is characterized by “reversed bilateral symmetry”; and zoospores with both right-handed and left-handed arrangements are produced. The ultrastructure of the zoospores clearly indicates that: 1) the mode of flagellar insertion: 2) morphology, number, and arrangement of multilayered structures, and 3) bilaterally keeled flagella are characteristic of the Chroolepidaceae.     

Ultrastructure of the scale-covered zoospores of the green alga Chaetosphaeridium, a possible ancestor of the higher plants and bryophytes

The zoospores of the green alga Chaetosphaeridium globosum are covered on all surfaces with tiny diamond-shaped scales similar to those of the prasinophycean flagellates and the Charales. The flagella also bear striated hairs (hair scales) so far considered to be a characteristic of the Prasinophyceae. The flagellar apparatus differs from that observed in the Prasinophyceae, shows many similarities to that of the Charales, and is identical with the "Vierergruppe" of the pteridophytes, cycads and bryophytes.
The zoospores are opisthokont, with two flagella inserted subapically. There is a lateral chloroplast containing typical grana and intergranal lamellae, but no eyespot. The very complicated Golgi body/contractile vacuole system comprises 10–20 contractile vacuoles. A microbody occupies a characteristic position in the cell, and in a young germling contains a crystalline inclusion.
The ultrastructure of the zoospore supports the old theory that the ancestors of the higher plants may well be found among Coleochaete and its relatives, past and present.     

Taxonomic studies on the Chlorarachniophyta. II. Generic delimitation of the chlorarachniophytes and description of Gymnochlora stellata gen. et sp. nov. and Lotharella gen. nov.

A new chlorarachniophytan alga, Gymnochlora stellata Ishida et Y. Hara gen. et sp. nov., has been isolated from Anae Island in Guam. It is a green, star-shaped, unicellular, amoeboid organism with several filopodia that do not form a reticulopodial network. Neither zoospores nor walled coccoid cells have been observed throughout the life cycle. The chloroplast ultrastructure is similar to those of described species; however, the pyrenoid matrix, which is invaded by many tubular structures originating from the inner membrane of the chloroplast envelope, is unique. A classification system is proposed for the Chlorarachniophyta. In this system, the ultrastructural features of the pyrenoid and the location of the nucleomorph in the periplastidial compartment are used as generic criteria, while the morphological features of the vegetative cells and life cycle patterns are used for species criteria. The described species, except for Cryptochlora perforans Calderon-Saenz et Schnetter, are also reassessed under the new system, and consequent nomenclatural requirements for the genus Chlorarachnion are dealt with in this paper. The taxonomic rank of a previously described species, Chlorarachnion globosum Ishida et Y. Hara, is elevated and Lotharella globosa (Ishida et Y. Hara) Ishida et Y. Hara gen. nov. et comb. nov. is proposed.     

Ultrastructural study of two rumen fungi:Piromonas communis andSphaeromonas communis

Two species of strictly anaerobic rumen fungi,Sphaeromonas communis andPiromonas communis, were examined by light and electron microscopy (scanning and transmission). Although morphologically different (vegetative body, number of flagella per zoospore), the ultrastructure of these two microorganisms was rather similar to that ofNeocallimastix frontalis andN. patriciarum. Two types of organelles were regularly found, i.e., isolated or associated ribosomes in the form of aggregates and hydrogenosome-like organelles with an amorphous content that may be involved in energy generation for these mitochondria-free strictly anaerobic fungi. UnlikeN. frontalis, the distribution of organelles was homogenous.     

On the induction of zoosporogenesis in the rumen phycomycetes Neocallimastix frontalis, Piromonas communis and Sphaeromonas communis.

The plant components inducing zoosporogenesis in the rumen phycomycetes Neocallimastix frontalis, Sphaeromonas communis and Piromonas communis were widely distributed in the plant kingdom with no apparent taxonomic relationship. In Lolium perenne L. (perennial rye-grass) and Hordeum distichon (barley), the components were principally present in the leaves and aerial tissues. Sufficient inducer was present in the normal diet of the host animal to trigger the differentiation and release of the zoospores from all the sporangia of each phycomycete species present in the rumen fluid tested. The inducers were unstable to oxygen, especially at elevated temperatures, and were destroyed by rumen micro-organisms. They may be similar compounds for each species.     

Degradation of maize stem by two rumen fungal species, Piromyces communis and Caecomyces communis, in pure cultures or in association with cellulolytic bacteria.

Two species of rumen fungi, Piromyces (Piromonas) communis FL and Caecomyces (Sphaeromonas) communis FG10, were cultured alone or in association with the cellulolytic bacteria Ruminococcus flavefaciens or Fibrobacter succinogenes on maize stem. A kinetic study of the degradation of the substrate was then made. After 48 h of culture, all non-lignified tissues observed by scanning electron microscopy disappeared with P communis and degradation was as complete as that observed in the rumen. In contrast, C communis degraded little of the plant cell walls. The ability of P communis to more rapidly degrade maize stem was probably due to the presence of filamentous rhizoids. The extent of dry matter loss after 8 days of incubation was practically the same in all the monocultures and in the 4 cocultures. However, the rate of degradation was faster in the bacterial than in the fungal monocultures and the co-cultures. No metabolic interaction was observed.     

Canaleparolina darwiniensis, gen. nov., sp. nov., and other pillotinaceous spirochetes from insects.

We describe two new pillotinaceous spirochetes (Canaleparolina darwiniensis, Diplocalyx cryptotermitidis) and identify for the first time Hollandina pterotermitidis from both the subterranean termite Cryptotermes cavifrons and the wood-eating cockroach Cryptocercus punctulatus based on morphometric analysis of transmission electron micrographic thin sections. C. darwiniensis, gen. nov., sp. nov., limited to near Darwin, Australia, invariably is present on the surface of the treponeme-studded trichomonad Mixotricha paradoxa, a consistent inhabitant of the hindgut of healthy termite Mastotermes darwiniensis. The spirochete both attached to the surface of protists and free-swimming in the paunch (hindgut) lumen of the insect has 16 periplasmic flagella (16:32:16) and imbricated wall structures that resemble flattened crenulations of Pillotina. The flagella surround half the protoplasmic cylinder. C. darwiniensis is the largest (0.5 microm diameter x 25 microm length) of the three epibiotic bacteria (two spirochetes, one rod) that comprise the complex cortex of its host Mixotricha paradoxa. Several criteria distinguish Diplocalyx cryptotermitidis sp. nov. isolated from Cryptotermes cavifrons intestine: smaller diameter, fewer flagella, absence of inner and outer coats of the outer membrane, wider angle subtended by its flagella and, most notably, cytoplasmic tubule-associated centers, which are periodic electron dense spheres within the protoplasmic cylinder from which emanate cytoplasmic tubules up to 24 nm in diameter. This is also the first report of abundant populations of Hollandina in Cryptotermes cavifrons (those populations belong to the species H. pterotermitidis). Morphometric analysis of the first thin sections of any spirochetes (published nearly 40 years ago by A.V. Grimstone) permits us to identify the large (0.9 microm diameter) free-swimming intestinal symbiont of Cryptocercus punctulatus also as Hollandina pterotermitidis.     

Studies on British chytrids XXVIII. Rhizophydium nobile sp. nov., parasitic on the resting spore of Ceratium hirundinella O.F.Müll. from the plankton

The life history of a new species of the Chytridiales Rhizophydium nobile is described. It occurs in the autumn on resting spores of the alga Ceratium hirundinella O.F.M. in Blelham Tarn, Windermere and Esthwaite Water, lakes in the English Lake District. The sporangium develops from the zoospore and possesses a branched rhizoidal system. The zoospores are fully formed in the sporangium. On dehiscence part of the sporangium content flows out surrounded by a vesicle which eventually bursts and liberates the zoospores. Only a few resting spores were seen associated with the sporangial stage. They were small spherical thick-walled bodies containing several globules. Further observations need to be made upon this stage.     

Life cycle,behavior and morphology of a new tide pool dinoflagellate,Gymnodinium pyrenoidosum sp. nov. (Gymnodiniales,Pyrrhophyta)

A new tide pool dinoflagellate,Gymnodinium pyrenoidosum Horiguchi et Chihara sp. nov. is described from central Japan. It was found to form dense blooms with a characteristic greenish color from April to November. The species exhibits a characteristic diurnal vertical migration and an alternation of a motile with a nonmotile phase, which are dependent on light intensity and tidal movement. Cells of the motile phase are unarmored and relatively small. They have a single, reticulate chloroplast, orange stigma situated near the sulcus and conspicuous pyrenoid in epicone. The alga reproduces itself by means of zoospores which are produced by the bipartition of protoplasm within the parent cell wall during the nonmotile stage which occurs at night. The occurrence of another type of motile cell, termed a macroswarmer, which differs from normal zoospore in size and shape has also been demonstrated.     

Piromyces citronii sp. nov., a strictly anaerobic fungus from the equine caecum: a morphological, metabolic, and ultrastructural study

Abstract Piromyces citronii sp. nov. was isolated from the caecum of one pony and three donkeys. It differed from other anaerobic fungal species in that it had a filamentous monocentric thallus composed of a globular or elliptic sporangium, which was occasionally bifid or trifid (monocarpic thallus), or several sporangia (polycarpic thallus), with a short sporangiophore. P. citronii also differed from other species in that it did not grow with starch or maltose as carbon source and it did not produce d-lactate. The uniflagellate zoospores presented a standard ultrastructure.     

禾谷多粘菌游动孢子超微结构观察

本文描述了寄生在大麦根部的禾谷多粘菌Polymyxa graminis Led.的次生游动孢子的超微结构,包括核、内质网、高尔基体、线粒体、脂质粒、排泄泡、小囊、具膜小囊、核糖体以及鞭毛基体(Kinetosome)和鞭毛杆等细胞器。游动孢子中未见微体。同时也在电镜下观察了游动孢子接触和穿透根细胞时所形成的管腔(Rohr)和棘杆(Stachel)以及游动孢子穿透细胞壁的详细过程。     

Molecular phylogenetic and zoospore ultrastructural analyses of Chytridium olla establish the limits of a monophyletic Chytridiales

Chytridium olla A. Braun, the first described chytrid and an obligate algal parasite, is the type for the genus and thus the foundation of family Chytridiaceae, order Chytridiales, class Chytridiomycetes and phylum Chytridiomycota. Chytridium olla was isolated in coculture with its host, Oedogonium capilliforme. DNA was extracted from the coculture, and 18S, 28S and ITS1-5.8S-ITS2 rDNA were amplified with universal fungal primers. Free swimming zoospores and zoospores in mature sporangia were examined with electron microscopy. Molecular analyses placed C. olla in a clade in Chytridiales with isolates of Chytridium lagenaria and Phlyctochytrium planicorne. Ultrastructural analysis revealed C. olla to have a Group II-type zoospore, previously described for Chytridium lagenaria and Phlyctochytrium planicorne. On the basis of zoospore ultrastructure, family Chytridiaceae is emended to include the type of Chytridium and other species with a Group II-type zoospore, and the new family Chytriomycetaceae is delineated to include members of Chytridiales with a Group I-type zoospore.     

Structure and reproduction of Hormidiella bharatiansis sp. nov.

The structure and reproduction of Hormidiella bharatiansis sp. nov. is described. Asexual reproduction is by zoospores: sexual reproduction is by morphologically similar gametes, some of which, after a brief period of swarming, settle down, round off, enlarge and act as females gametes, the others remaining active and functioning as male gametes. Fertilisation takes place about six to eight hours after liberation: one to two percent of the zygotes are abortive.     

Zoospore ultrastructure and phylogenetic position of Phlyctochytrium aureliae Ajello is revealed (Chytridiaceae, Chytridiales, Chytridiomycota)

From forest soils in Scotland Phlyctochytrium aureliae was observed and brought into pure culture. Previously included in a molecular phylogenetic study of Chytridiales as Phlyctochytrium sp. KP 061, the organism groups with Phlyctochytrium planicorne, P. bullatum, Chytridium olla and C. lagenaria in the family Chytridiaceae. Thallus morphology and development as well as zoospore ultrastructure are detailed herein. The sporangium is epibiotic, spherical or subspherical, apophysate or non-apophysate, and ornamented with dentate enations. The overall zoospore ultrastructural features are consistent with the Group II type zoospore that characterizes family Chytridiaceae in the Chytridiales, although the zoospore also has two character states unique to this taxon: the MLC cisterna fenestrations are one-third to one-half the diameter of fenestrations in other Chytridiaceae zoospores and an accumulation of electron-dense material (a kinetosome-associated structure, or KAS) proximal to the kinetosome and non-flagellated centriole is extensive and unique. This study verifies that zoospore ultrastructure of P. aureliae zoospores places this species in the Chytridiales and Chytridiaceae, as indicated in a previous molecular phylogenetic study.