<Emphasis Type="Italic">Rhizoglomus venetianum</Emphasis>, a new arbuscular mycorrhizal fungal species from a heavy metal-contaminated site,downtown Venice in Italy

Rhizoglomus venetianum, a new arbuscular mycorrhizal fungal species, has been isolated and propagated from a heavy metal-contaminated site in Sacca San Biagio island, downtown Venice, Italy. Interestingly, under the high levels of heavy metals occurring in the site, the new fungus was able to grow only intraradically. In greenhouse trap and single species cultures under low heavy metal levels, the fungus produced innumerous spores, clusters, and sporocarps extraradically, which were formed terminally on subtending hyphae either singly, in small spore clusters, or, preferably, in loose to compact non-organized sporocarps up to 2500?×?2000?×?2000 μm. Spores are golden-yellow to bright yellow brown, globose to subglobose to rarely oblong, 75–145?×?72–140 μm in diameter, and have four spore wall layers. Morphologically, the new fungus is similar to R. intraradices, and phylogenetically, it forms a monophyletic clade next to R. irregulare, which generally forms irregular spores and lacks, like R. intraradices, the flexible innermost wall layer beneath the structural/persistent third wall layer. A key for the species identification is presented comprising all 18 Rhizoglomus species, so far described or newly combined.     

THE SPORE PATTERN IN SOME SPECIES OF CHEILANTHES

Of the 32 taxa examined, 13 contained 32 spores in each sporangium and are considered apo-gamous, 14 were sexual species with 64 spores per sporangium, and 5 had 32 spores in some sporangia and 64 in others. When considered as a whole, the spores ranged in size from 29.9 to 74.88μ. Most species had oval or globose spores but several had tetrahedral spores. The spores of all were radially symmetrical. Almost all of the species possessed a crassimarginate type of laesura and all except C. cooperae and C. viscida had a perispore. The ornamentation of the perispore showed the following patterns: napate, granulate, psilate, lobate, foveate, and echinate. The exine pattern was predominantly psilate but foveate, rugulate, napate, and granulate conditions were observed. Seventeen taxa were found to have some degree of spore abortion.     

Taxonomic studies on Asplenium sect. Thamnopteris (Aspleniaceae) II: spore morphology

Asplenium sect. Thamnopteris is a morphologically distinct group within Asplenium, however, the species delimitation within the section is difficult. To evaluate the utility of spore morphology in the taxonomy of Asplenium sect. Thamnopteris, we examined 64 samples representing ca 25 species using scanning electron microscopy. The results show that the spores are monolete and ellipsoidal in shape, and range between 35.7–50.1 × 24.1–32.4 µm in size. The perispore is morphologically very diverse but relatively stable within species. Based on the morphology of spores, we discuss the species concept of A. nidus L. and A. phyllitidis D. Don, propose a new combination (A. latibasis) for Neottopteris latibasis Ching, and treat A. phyllitidis subsp. malesicum Holttum, N. latibasis, N. latipes Ching ex S. H. Wu, and N. subantiqua Ching ex S. H. Wu as new synonyms. A key to all known species of sect. Thamnopteris is provided using characters of spore morphology.     

Analysis of broth-cultured Bacillus atrophaeus and Bacillus cereus spores

Aims: To compare physical properties of spores that were produced in broth sporulation media at greater than 10⁸ spores ml⁻¹. Methods and Results: Bacillus atrophaeus reproducibly sporulated in nutrient broth (NB) and sporulation salts. Microscopy measurements showed that the spores were 0·68 ± 0·11 μm wide and 1·21 ± 0·18 μm long. Coulter Multisizer (CM3) measurements revealed the spore volumes and volume-equivalent spherical diameters, which were 0·48 ± 0·38 μm³ and 0·97 ± 0·07 μm, respectively. Bacillus cereus reproducibly sporulated in NB, sporulation salts, 200 mmol l⁻¹ glutamate and antifoam. Spores were 0·95 ± 0·11 μm wide and 1·31 ± 0·17 μm long. Spore volumes were 0·78 ± 0·61 μm³ and volume-equivalent spherical diameters were 1·14 ± 0·11 μm. Bacillus atrophaeus spores were hydrophilic and B. cereus spores were hydrophobic. However, spore hydrophobicity was significantly altered after treatment with pH-adjusted bleach. Conclusions: The utility of a CM3 for both quantifying Bacillus spores and measuring spore sizes was demonstrated, although the volume between spore exosporium and spore coat was not measured. This study showed fundamental differences between spores from a Bacillus subtilis- and B. cereus-group species. Significance and Impact of the Study: This is useful for developing standard methods for broth spore production and physical characterization of both living and decontaminated spores.     

Spore morphology and wall ultrastructure of Hymenophyllaceae Link (Pteridophyta) from north-west Argentina

The family Hymenophyllaceae is represented in the study area by six species in two genera, Hymenophyllum J. E. Smith and Trichomanes L. The study was based on herbarium material and spores were studied under light microscope (LM), scanning electron microscope (SEM) and transmission electron microscope (TEM). Both genera have trilete spores, 23 to 45 μm in equatorial diameter, with an ornamentation of echinulae and cones in Hymenophyllum and of verrucae, gemmae and granules in Trichomanes. Mature spores have a sporoderm composed of a perispore, an exospore and a fibrillar endospore; the exospore is 0.5 to 2.5 μm thick, compact and with an irregular margin. In some cases radial channels and other channels associated with the middle and inner parts of the laesurae were evident. A series of cavities filled with an opaque content line the inner margin of the exospore. The perispore is 20 to 400 nm thick and unevenly differentiated along the surface of a same spore. Under TEM, two main differentially contrasted portions could be distinguished: a dark massive portion with structural components could not be distinguished, and a light portion with several plates arranged in piles. The inner surface of the perispore exhibit short scales. Globules are immersed within the perispore at some depth from the perispore surface and others connected to it by structural threads. The spore characters observed including shape, ornamentation, laesurae length and wall structure are useful in distinguishing the two genera studied, but less useful in differentiation at the species level.     

Emission rate of charged spores in basidiomycetous fungi and the relaxation time of their electric charges

Basidiospores are one of the main components of coarse fraction of atmospheric aerosol. Majority of them, the ballistospores of 20,000 species of Basidiomycotina, carry electrostatic charges when getting airborne. To study the polarity and magnitude of primary charges and the hymenial emission rate of charged spores, 128 spore samples of 31 species of Agaricomycetes were collected in natural conditions. A portable device was placed under the fruiting body and the freely falling charged spores were extracted from the air by a horizontal homogeneous electrostatic field. The charge polarity distribution was the same in all intraspecies spore samples; it was unipolar-positive, unipolar-negative, or bipolar, depending on the species. The mean spore charge magnitude was 21–981 e, and it was not related to the emission rate of charged spores. The hymenial emission rate was fluctuating, and the maximum value was 715 charged spores cm^?2 s^?1. To estimate the territorial emission rate of charged spores, area of the hymenial surface per hectare of forest was calculated for three species and the maximum values were 11 m² ha^?1 and 8.6 × 10⁷ charged spores ha^?1 s^?1. Calculations showed that a spore charge diminished sevenfold within 47 min. Ecologists, health and agricultural scientists could be interested in this information. It could be useful by investigating the role of microorganisms in meteorological phenomena and in atmospheric processes in general.     

Ortholinea alata n. sp. (Myxosporea: Ortholineidae) in the Northern Butterfly Fish Chaetodon rainfordi

Ortholinea alata n. sp. is described from the northern butterfly fish, Chaetodon rainfordi collected at Heron Island, Great Barrier Reef, Australia. Spherical, disporous trophozoites (10–15 μ) and spores were observed in the lumina of kidney tubules and collecting ducts. Spores are broadly triangular with two short, broad processes that extend dorsoventrad from the posterior end of each of the two spore valves. Valves are bisected by a suture in the plane of the polar capsules. Spores are 12.6 μ (length) × 9.6 μ (width) × 9.9 μ (length), and at the anterior end contain two spherical, divergent polar capsules measuring 4.6 ( 4.1 –5.1) μ. Sporogenesis is similar to that of renal Sphaerospora spp.: the intraluminal trophozoites of O. alata n. sp. correspond to pseudoplasmodia described for Sphaerospora spp. and no large, multinucleate plasmodia are formed. No significant histopathological changes were observed in the kidneys of infected fish.     

Spore wall ultrastructure of Polypodiaceae from north-western Argentina

The spore wall ultrastructure of Campyloneurum, Microgramma, Pecluma, Phlebodium, Pleopeltis and Serpocaulum (Polypodiaceae) from north-western Argentina has been studied using transmission electron microscopy (TEM). The exospore is 0.4–3 μm thick, two-layered and variously ornamented in all taxa. The exospore surface is distinctive, but in general ultrastructure the exospore is similar in all species studied. The structural elements of the exospore consist of cavities in the inner part as well as channels with a radial orientation and channels at both sides of the laesura. Variation in the exospore surface was observed in spores at different stages of maturation. The perispore is darkly contrasted and 0.04–2 μm thick. Three different structure types were recognised, including fibrillar, multilamellar and lacunose. Scattered globules and spherules were always present on the perispore surface. The structural variability of the perispore was surveyed within complete sporangia. We concluded that the observed variability may be related to the stage in spore maturation and, consequently, to the stages in perispore differentiation. As the exospore ultrastructure is similar and interpreted as related to functional activity in the studied material, it cannot be used for systematic delimitations at this generic or specific level.     

Spore morphology of pteridophytes from China Ⅱ. Sinopteridaceae

Spores of 61 species and 6 varieties in 9 genera of the Sinopteridaceae were examined under scanning electron microscope (SEM). Based on surface ornamentation and other features, the spores of the Sinopteridaceae are divided into three types. In type Ⅰ , the exospore is smooth and the surface ornamentation, which is reticulate, cristate, echinate or rugate, is formed by the perispore. All the other genera of this family, except for Onychium and Cryptogramma, have this pattern of spores. In type Ⅱ, the surface ornamentation is formed by both perispore and exospore. This pattern is found only in Cryptogramma. In type Ⅲ, the perispore is thin and the surface ornamentation is formed by the exospore. Onychium is characterized by this type of spores. Those genera with spores of type Ⅰ of the Sinopteridaceae seem to be closely related to each other and should be natural members of this family. The systematic position of Cryptogramma and Onychium, with spores of type Ⅱ and type Ⅲ respectively, however, should be reconsidered. Aleuritopteris might be the mostprimitive member of the Sinopteridaceae from the evidence of spore morph     

狗脊孢壁发育超微结构的研究

鳞毛蕨型孢子类型众多,初步研究表明形态相似的孢子类型其孢壁发育特征存在差异,因此有必要对各代表类群的孢壁发育进行深入地研究。该文利用透射电镜对乌毛蕨科(Blechnaceae)狗脊(Woodwardia japonica)孢壁结构和发育的超微结构进行研究。结果表明:(1)狗脊孢子囊的结构由外向内分别为孢子囊壁细胞、两层绒毡层细胞和孢子母细胞;(2)狗脊孢子具乌毛蕨型(Blechnoid type)外壁,表面光滑,由两层构成,裂缝区域具辐射状的槽;(3)周壁属于空心型(cavity type),由四层构成,从内向外分别为P1、P2、P3和P4层,前三层叠合在一起,层间有不同程度的空隙,P4层与前三层之间具有明显而连续的空腔,并隆起形成片状褶皱纹饰;(4)有小球体和小杆共同参与孢子周壁的形成,周壁部分或全部来源于孢子囊壁细胞。综上所述,狗脊孢子与同属于鳞毛蕨型的贯众(Cyrtomium fortunei)和朝鲜介蕨(Dryoathyrium coreanum)孢壁的发育在周壁结构、周壁各层的发育顺序、周壁来源和参与成壁的特征物质等方面存在差异。该研究有利于进一步理解蕨类植物孢壁所蕴含的分类和演化上的科学意义和价值。     

中国蕨类植物孢子形态的研究 Ⅱ. 中国蕨科

利用扫描电镜对国产中国蕨科Sinopteridaceae植物9属61种6变种的孢子进行了观察。结果表明,该科植物的孢子可分为3种类型：(1)孢子球形,三裂缝;周壁较厚,疏松地包在孢子之外;外壁光滑,表面纹饰由周壁形成,呈网状、嵴状、刺状或皱状。除金粉蕨属Onychium和珠蕨属Cryptogramma外,该科其他属的植物都具此类型孢子。(2)孢子钝三角形,三裂缝;周壁较薄,由周壁和外壁共同形成表面轮廓,表面具疣状或颗粒状纹饰。具此类型孢子的只有珠蕨属。(3)孢子钝三角形,三裂缝,沿裂缝两侧各有一脊状隆起或瘤状纹饰;周壁薄,由外壁形成表面纹饰的基本轮廓;具赤道环、近极脊和远极脊。具此类型孢子的只有金粉蕨属。另外,从孢粉学的角度对该科的分类和系统演化进行了探讨。     

A DEVELOPMENTAL STUDY OF GUTTULINOPSIS VULGARIS (ACRASIALES)

The cellular slime mold, Guttulinopsis vulgaris E. W. Olive, is a common and widely distributed species. Its myxamoebae have lobose pseudopodia and aggregate, not by streaming, but by movement singly or in small groups into the aggregation center. The resulting shield-shaped pseudoplasmodium gives rise to one (or possibly more on occasion) whitish, simple or compound sorocarp. Sorocarps are 95–465 μ in height, typically with relatively short, stout stalks and with mucilaginous spore heads that measure 75–350 μ broad. The spores are mostly irregular, with flattened or concave sides, and are surrounded by a thin spore wall. They measure 3.8–9.2 μ diam., or 2.6–7.8 × 3.9–9.2 μ. During sorocarp development the myxamoebae become grouped within membranous compartments both in the sorocarp and pseudoplasmodium. In the upper part of the sorocarp they develop almost entirely into spores, while in the lower stalk region of the sorocarp and in the pseudoplasmodium, some myxamoebae develop into spores while others degenerate. The phylogenetic position of Guttulinopsis is unclear, but the genus is probably not closely related to the Dictyosteliaceae. This investigation has been supported by National Science Foundation Grants G–44263 and GB-501. The writer is grateful to Miss Carmen Stoianovitch for her assistance.     

华中铁角蕨孢子纹饰形成的超微结构观察

利用透射电子显微镜对铁角蕨科(Aspleniaceae)华中铁角蕨(Asplenium sarelii Hook.)孢子及其纹饰的形成过程进行观察。结果表明:①华中铁角蕨孢子囊发育为薄囊蕨型;②孢子外壁表面光滑,远极面的外壁厚约0.8~1.1μm,近极面的外壁厚约1.4~1.8μm;③孢子周壁厚度约4~5μm,染色较外壁深,分为内层和外层;内层紧帖外壁表面,其上具柱状、瘤状或疣状突起;外层向外隆起形成脊状纹饰的轮廓,脊的下方具空腔,脊的顶端具翅;④铁角蕨型与鳞毛蕨型孢子外壁和周壁纹饰的形成过程具有相似性;⑤孢子的成熟度对于孢子形态的研究是至关重要的,只有完全成熟的孢子的表面纹饰才是稳定的。     

Wet and dry density of Bacillus anthracis and other Bacillus species

Aims: To determine the wet and dry density of spores of Bacillus anthracis and compare these values with the densities of other Bacillus species grown and sporulated under similar conditions. Methods and Results: We prepared and studied spores from several Bacillus species, including four virulent and three attenuated strains of B. anthracis, two Bacillus species commonly used to simulate B. anthracis (Bacillus atrophaeus and Bacillus subtilis) and four close neighbours (Bacillus cereus, Bacillus megaterium, Bacillus thuringiensis and Bacillus stearothermophilus), using identical media, protocols and instruments. We determined the wet densities of all spores by measuring their buoyant density in gradients of Percoll and their dry density in gradients of two organic solvents, one of high and the other of low chemical density. The wet density of different strains of B. anthracis fell into two different groups. One group comprised strains of B. anthracis producing spores with densities between 1·162 and 1·165 g ml^?1 and the other group included strains whose spores showed higher density values between 1·174 and 1·186 g ml^?1. Both Bacillus atrophaeus and B. subtilis were denser than all the B. anthracis spores studied. Interestingly and in spite of the significant differences in wet density, the dry densities of all spore species and strains were similar. In addition, we correlated the spore density with spore volume derived from measurements made by electron microscopy analysis. There was a strong correlation (R² = 0·95) between density and volume for the spores of all strains and species studied. Conclusions: The data presented here indicate that the two commonly used simulants of B. anthracis, B. atrophaeus and B. subtilis were considerably denser and smaller than all B. anthracis spores studied and hence, these simulants could behave aerodynamically different than B. anthracis. Bacillus thuringiensis had spore density and volume within the range observed for the various strains of B. anthracis. The clear correlation between wet density and volume of the B. anthracis spores suggest that mass differences among spore strains may be because of different amounts of water contained within wet dormant spores. Significance and Impact of the Study: Spores of nonvirulent Bacillus species are often used as simulants in the development and testing of countermeasures for biodefense against B. anthracis. The similarities and difference in density and volume that we found should assist in the selection of simulants that better resemble properties of B. anthracis and, thus more accurately represent the performance of countermeasures against this threat agent where spore density, size, volume, mass or related properties are relevant.     

CO-REQUIREMENT FOR CALCIUM AND POTASSIUM IN THE GERMINATION OF SPORES OF THE FERN ONOCLEA SENSIBILIS

Spores of Onoclea sensibilis L. do not germinate on distilled H₂O if they are pretreated for sufficient time with dilute NaClO solution. However, spores will germinate, after NaClO pretreatment, on a simple mineral medium containing the major and trace elements. Complete germination after pretreatment also is obtained on a solution containing only Ca²⁺ and K⁺ as the cations, but neither ion by itself is sufficient. Rb⁺, but not Li⁺ or Na⁺, can replace K⁺. Hypochlorite-treated spores do not require the continuous presence of Ca²⁺ and K⁺ to germinate; exposure during the first 4 hr of culture, with the remainder of the time on distilled H₂O, is sufficient. Extraction of spores with ethylene glycol bis(aminoethyl ether) tetraacetic acid [EGTA] makes their germination dependent on Ca²⁺, as reported by other workers, but it does not produce a co-requirement for K⁺. Colorimetric analysis with arsenazo III confirms that Ca²⁺ is extracted from Onoclea spores by NaClO. Extractable Ca²⁺ amounts to about 78 nmol/mg spore dry wt. Of this amount, 31% is contained in the perispore. The perispore comprises 13% of the total spore dry wt.     

A novel milky disease organism from Australian scarabaeids: Field occurrence,isolation, and infectivity

A novel milky disease organism has been found causing disease in Aphodius tasmaniae and other scarabaeid larvae in the field in Australia. The sporangium is exceptionally long, measuring 10.5 × 1.5 μm, with a small central spore, measuring 1.0 × 0.6 μm. The vegetative cell is about half the size of the sporangium. The disease was easily transmitted by injection of spores into the hemocoel, with typically milky symptoms developing in 2–4 weeks. Spores will form in vivo at temperatures down to 12°C. For A. tasmaniae third-instar larvae, the ID₅₀ by injection was 3 × 10² spores/larva, yet no infection resulted when larvae were reared in peat containing up to 10⁸ spores/g, i.e., the disease was not successfully transmitted per os. All 10 species of scarabaeids tested were susceptible to the disease when spores were injected; however, all attempts to infect larvae per os were unsuccessful. In vitro culture was also unsuccessful.     

THE ULTRASTRUCTURE OF PALEOZOIC FERN SPORES: I. BOTRYOPTERIS

The spores of four species of the Paleozoic filicalean fern Botryopteris are examined at the ultrastructural level. Spores of B. cratis, B. forensis, B. globosa, and an unnamed species from the Lower Pennsylvanian, are compared on the basis of sporoderm stratification and the presence or absence of a sculptine layer. The species examined differ widely as to the type of reproductive unit in which they are borne and include forms that range throughout the Pennsylvanian. In all species the exine is homogeneous, lacking cavities and lamellae. A thin nexine is present in the Middle and Upper Pennsylvanian taxa, but is absent in the Lower Pennsylvanian spores. Only one spore type (B. cratis) possesses a clearly defined sculptine layer. Features of the sporoderm are compared with those of extant, homosporous pteridophyte spores.     

Consecutive hydrolysis of creatinine using creatininase and creatinase encapsulated in <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis> spores

Objectives

To achieve consecutive conversion from creatinine to urea and sarcosine using creatininase and creatinase encapsulated in spores of Saccharomyces cerevisiae.

Results

Creatininase encapsulated into the spore wall was produced and its specific activity was 3.4 ± 0.4 U/mg. By deletion of OSW2 gene, which causes a mild spore wall defect, the activity was increased to 10.9 ± 0.5 U/mg. Compared with soluble enzymes, spore-encapsulated creatininase was tolerant to environmental stresses; creatininase encapsulated in osw2? spores retained more than 90 % of the activity after treatment by SDS or proteinase K. Creatinase capsules could also be produced through spore encapsulation. The mixture of spores containing either creatininase or creatinase could mediate a two-step reaction to produce urea from creatinine; 5 mg spores produced 19 µmol urea in 10 min. Spores co-expressing creatininase and creatinase could also mediate the reactions more efficiently than the mixture of spores individually expressing each enzyme; the yield in 10 min was 38 µmol.

Conclusions

Yeast spores can hold creatininase and creatinase simultaneously and catalyze the consecutive reactions.

     

Glomus drummondii and G. walkeri,two new species of arbuscular mycorrhizal fungi (Glomeromycota)

Two new ectocarpic arbuscular mycorrhizal fungal species, Glomus drummondii and G. walkeri (Glomeromycota), found in maritime sand dunes of northern Poland and those adjacent to the Mediterranean Sea are described and illustrated. Mature spores of G. drummondii are pastel yellow to maize yellow, globose to subglobose, (58–)71(–85) μm diam, or ovoid, 50–80 × 63–98 μm. Their wall consists of three layers: an evanescent, hyaline, short-lived outermost layer, a laminate, smooth, pastel yellow to maize yellow middle layer, and a flexible, smooth, hyaline innermost layer. Spores of G. walkeri are white to pale yellow, globose to subglobose, (55–)81(–95) μm diam, or ovoid, 60–90 × 75–115 μm, and have a spore wall composed of three layers: a semi-permanent, hyaline outermost layer, a laminate, smooth, white to pale yellow middle layer, and a flexible, smooth, hyaline innermost layer. In Melzer's reagent, only the inner- and outermost layers stain reddish white to greyish rose in G. drummondii and G. walkeri, respectively. Both species form vesicular–arbuscular mycorrhizae in one-species cultures with Plantago lanceolata as the host plant. Phylogenetic analyses of the ITS and parts of the LSU of the nrDNA of spores placed both species in Glomus Group B sensu Schüßler et al. [Schüßler A, Schwarzott D, Walker C, 2001. A new fungal phylum, the Glomeromycota: phylogeny and evolution. Mycolological Research 105: 1413-1421.]     

Fine structure of Chloromyxum menticirrhi n. sp. (Myxozoa) infecting the urinary bladder of the marine teleost Menticirrhus americanus (Sciaenidae) in Southern Brazil

A myxosporidian was found in the urinary bladder of the teleost Menticirrhus americanus Linnaeus, 1758 (Sciaenidae) collected from the South Atlantic coast of Brazil. Polysporic amoeboid plasmodia containing sporoblasts, developing pansporoblasts and spores were free in the bladder lumen. The prevalence of infection was 17.64% (15/85). Unfixed spores were spherical to subspherical, on average 10.5 μm long, 9.8 μm wide and 10.1 μm thick (n=25), and fixed spores measured 10.1×9.5×9.7 μm. The two spore valves were of equal size and each possessed prominent sutural lines and about 41 (37–45) surface ridges aligned parallel with the suture line. These ridges gave transverse sections a cog-wheel-like outline. The spores contained four pyriform polar capsules of equal size (3.20×2.0 μm) (n=25) (fixed), each with a polar filament having 3–4 (rarely 5) coils. The binucleate sporoplasm was irregular in shape, with granular matrix and randomly distributed dense bodies. The shape and dimensions of the spore, as well as the number, position and arrangement of the surface ridges, polar capsules and polar filament indicate that this is a new species, herein designated Chloromyxum menticirrhi. The gill, liver, gall bladder and intestine of the host showed no abnormalities.