Genetic variability and taxonomic revision of the genus Auxenochlorella (Shihira et Krauss) Kalina et Puncocharova (Trebouxiophyceae,Chlorophyta)

The monotypic genus Auxenochlorella with its type species A. protothecoides is so far only known from specific habitats such as the sap of several tree species. Several varieties were described according to physiological performances in culture on different organic substrates. However, two strains designated as Auxenochlorella were isolated from other habitats (an endosymbiont of Hydra viridis and an aquatic strain from an acidic volcano stream). We studied those isolates and compared them with six strains of Auxenochlorella belonging to different varieties. The integrative approach used in this study revealed that all strains showed similar morphology but differed in their SSU and ITS rDNA sequences. The Hydra endosymbiont formed a sister taxon to A. protothecoides, which included the varieties protothecoides, galactophila, and communis. The variety acidicola is not closely related to Auxenochlorella and represented its own lineage within the Trebouxiophyceae. In view of these results, we propose a new species of Auxenochlorella, A. symbiontica, for the Hydra symbiont, and a new genus Pumiliosphaera, with its type species, P. acidophila, for acidophilic strain. These results are supported by several compensatory base changes in the conserved region of ITS‐2 and ITS‐2 DNA barcodes.     

New species of Teratosphaeria associated with leaf diseases on Corymbia calophylla (Marri)

Corymbia calophylla is an important eucalypt species endemic to the south-west of Western Australia. It is currently in decline across its native habitat for unknown reasons but perhaps due to climate change or to the presence of a recently described canker disease caused by Quambalaria coyrecup. Few fungal leaf pathogens have been described from C. calophylla. The following paper describes two new species of Teratosphaeria from C. calophylla, Teratosphaeria calophylla sp. nov and T. rubidae sp. nov. In addition, a new epitype was designated for T. australiensis.     

Growth profile of ectomycorrhizal fungal mycelium: emphasis on substrate pH influence

A knowledge of behaviour of ectomycorrhizal fungal isolates to substrate pH would help in identifying candidate fungi for plantation programs. Ectomycorrhizal fungus isolates were studied to determine the pH optima for growth, substrate acidification by the culture and the effect of substrate acidification on culture growth. Of the isolates tested, the members of Agaricales (except Laccaria laccata) and Aphyllophorales favored neutral to near neutral pH, while members of the order Sclerodermatales strictly favored acidic pH. The change in substrate pH (ΔpH) was maximum at the optimum growth pH for acidophilic isolates but minimum for neutro/basophilic isolates. The experiments indicate that the substrate pH would not only determine the growth rate of the fungus but also limits further proliferation of the fungus in medium. This gives vital information for determining the subculture frequency and for designing substrate parameters for nursery/plantation programs. This revised version was published online in June 2006 with corrections to the Cover Date.     

Bioleaching in brackish waters—effect of chloride ions on the acidophile population and proteomes of model species

High concentrations of chloride ions inhibit the growth of acidophilic microorganisms used in biomining, a problem particularly relevant to Western Australian and Chilean biomining operations. Despite this, little is known about the mechanisms acidophiles adopt in order to tolerate high chloride ion concentrations. This study aimed to investigate the impact of increasing concentrations of chloride ions on the population dynamics of a mixed culture during pyrite bioleaching and apply proteomics to elucidate how two species from this mixed culture alter their proteomes under chloride stress. A mixture consisting of well-known biomining microorganisms and an enrichment culture obtained from an acidic saline drain were tested for their ability to bioleach pyrite in the presence of 0, 3.5, 7, and 20 g L⁻¹ NaCl. Microorganisms from the enrichment culture were found to out-compete the known biomining microorganisms, independent of the chloride ion concentration. The proteomes of the Gram-positive acidophile Acidimicrobium ferrooxidans and the Gram-negative acidophile Acidithiobacillus caldus grown in the presence or absence of chloride ions were investigated. Analysis of differential expression showed that acidophilic microorganisms adopted several changes in their proteomes in the presence of chloride ions, suggesting the following strategies to combat the NaCl stress: adaptation of the cell membrane, the accumulation of amino acids possibly as a form of osmoprotectant, and the expression of a YceI family protein involved in acid and osmotic-related stress.     

Bikaverin production and applications

Bikaverin is a reddish pigment produced by different fungal species, most of them from the genus Fusarium, with antibiotic properties against certain protozoa and fungi. Chemically, bikaverin is a polyketide with a tetracyclic benzoxanthone structure, resulting from the activity of a specific class I multifunctional polyketide synthase and subsequent group modifications introduced by a monooxygenase and an O-methyltransferase. In some fungi, bikaverin is found with smaller amounts of a precursor molecule, called norbikaverin. Production of these metabolites by different fungal species depends on culture conditions, but it is mainly affected by nitrogen availability and pH. Regulation of the pathway has been investigated in special detail in the gibberellin-producing fungus Fusarium fujikuroi, whose genes and enzymes responsible for bikaverin production have been recently characterized. In this fungus, the synthesis is induced by nitrogen starvation and acidic pH, and it is favored by other factors, such as aeration, sulfate and phosphate starvation, or sucrose availability. Some of these inducing agents increase mRNA levels of the enzymatic genes, organized in a coregulated cluster. The biological properties of bikaverin include antitumoral activity against different cancer cell lines. The diverse biological activities and the increasing information on the biochemical and genetic basis of its production make bikaverin a metabolite of increasing biotechnological interest.     

Biodegradation of polycyclic aromatic hydrocarbons by an acidophilic Stenotrophomonas maltophilia strain AJH1 isolated from a mineral mining site in Saudi Arabia

The present study aims at analyzing the degradation of polycyclic aromatic hydrocarbons (PAHs) at acidic conditions (pH = 2) by acidophilic Stenotrophomonas maltophilia strain AJH1 (KU664513). The strain AJH1 was obtained from an enrichment culture obtained from soil samples of mining area in the presence of PAH as sole sources of carbon and energy. Strain AJH1was able to degrade low (anthracene, phenanthrene, naphthalene, fluorene) and high (pyrene, benzo(e)pyrene and benzo(k)fluoranthene) molecular weight PAHs in acidophilic mineral salt medium at pH 2, with removal rates of up to 95% (LMW PAH) and 80% (HMW PAH), respectively. In addition, strain AJH1 treated petroleum wastewater with 89 ± 1.1% COD removal under acidic condition (pH 2) in a continuously stirred reactor. Acidophilic S. maltophilia strain AJH1, hence holds the promise as an effective degrader for biological treatment of PAHs contaminated wastewater at acidic pH.

     

Identification of deoxynivalenol, 3-acetyldeoxynivalenol and zearalenone in the galactose oxidase-producing fungus Dactylium dendroides

The galactose oxidase-producing fungus Dactylium dendroides was re-identified as a Fusarium species. Fungi of this genus are well known for the production of mycotoxins. Verification of growth of this fungus on rice, corn and liquid medium described for the production of galactose oxidase is provided to determine whether the fungus could produce Fusarium toxins, namely, moniliformin, fusaric acid, fumonisin, zearalenone and the trichothecenes, deoxynivalenol, 3-acetyldeoxynivalenol, fusarenone, nivalenol, diacetoxyscirpenol, neosolaniol, and toxin T-2. Under the culture conditions used, deoxynivalenol, 3-acetyldeoxynivalenol and zearalenone were detected in the fungal culture medium. The finding is consistent with the hypothesis that the fungus is in fact a Fusarium species. This revised version was published online in July 2006 with corrections to the Cover Date.     

The macroinvertebrate fauna of an acid-stressed headwater stream system in the Adirondack Mountains, New York

SUMMARY.

• 1 The Silver Run system in the Adirondack Mountains of New York includes three first-order streams with very different pH regimes: one moderately acidic (pH 5.8-7.2), the others much more so (pH generally 4.4-5.0)
• 2 The less acidic (acidophilic) site yielded a relatively diverse, well balanced fauna, with at least 10% each of midges, mayflies, stoneflies and elmid beetles. Ephemerella funcralis (Ephemeroptera) and Oulimnius latiusculus (Coleoptera) were dominant in this community. They appear to be among the most acid-tolerant species of their respective families and may become abundant under moderately acidic conditions. Their absence or low abundance in the more acidic tributaries of the Silver Run system suggests that they arc eliminated as acidification intensifies, These species therefore may he useful indicators of advancing acidification.
• 3 The more acidic (acidobiontic) communities contained fewer Hum half as many taxa as the acidophilic communities, contained few elmid beetles or mayflies, and were heavily dominated In stoneflies. Leuctra feiruinea and Isopecrla sp. were dominant, comprising.56-86% of the individuals. Other common constituents included black flies (Simuliumn gouldingi and S. venustum) and the caddisfly Rhyacvphila fuscula.

     

Acidobacterium capsulatum gen. nov., sp. nov.: An acidophilic chemoorganotrophic bacterium containing menaquinone from acidic mineral environment

Acidobacterium is proposed as a new genus for the acidophilic, chemoorganotrophic bacteria containing menaquinone isolated from acidic mineral environments.Acidobacterium capsulatum is proposed for the singleAcidobacterium species which consists of eight strains (Biogroup 5). The members of this species are gram-negative, aerobic, mesophilic, non-spore-forming, capsulated, saccharolytic, and rod-shaped bacteria. They are motile by peritrichous flagella. They can grow between pH 3.0 and 6.0, but not at pH 6.5. They give positive results in tests for esculin hydrolysis, catalase, and -galactosidase. Oxidase and urease are negative. They can use glucose, cellobiose, starch, maltose, or -gentiobiose as a sole carbon source, but cannot use elemental sulfur and ferrous iron as an energy source. The DNA base composition is 59.7–60.8 guanine plus cytosine (G+C) mol%. The major isoprenoid quinone is menaquinone with eight isoprene units (MK-8). The major fatty acid is 13-methyltetradecanoic acid. DNA relatedness between this species and the species ofAcidiphilium, Acidomonas, andDeinobacter was 18 to 2%. From phenotypic and chemotaxonomic characters, these member do not belong to any known taxa of gram-negative bacteria. A culture of the type strain (strain 161) has been deposited in the Japan Collection of Microorganisms as JCM 7670.     

Fungi associated with cellulose decomposition in the tidal mud-flats of Kuwait

Fungi potentially able to decompose cellulose, in the tidal mud-flats of Kuwait, have been investigated. More than 58 species were isolated and their cellulytic ability were tested. Many of the species recorded are well known cellulose decomposers, while for many others the strong cellulytic activity is a new record e.g. Arachniotus dankaliensis, Lasiobolidium orbiculoides, Corynascus sepedonium, and Pesotum sp. Comparison of the frequency of occurrence and cellulytic ability, in all species reported, revealed that there is no coincidence between the two parameters. The high frequency of any fungus does not imply that this fungus is an active cellulose decomposer in the soil and similarly low frequency fungi are not necessarily weak decomposers. It is likely possible, that the competitive saprophytic ability of the fungus which determine its role in the process of cellulose decomposition in the soil rather than its cellulytic ability in pure culture.     

<Emphasis Type="Italic">Ancylobacter abiegnus</Emphasis> sp. nov., an oligotrophic member of the xylotrophic mycobacterial community

A new species of the genus Ancylobacter, exemplified by strain Z-0056, was isolated from dystrophic humified waters formed by xylotrophic fungi grown on decaying spruce wood. The cells of strain Z-0056 (0.65–0.9 μm) are coccoid, gram-negative, with fimbriae, and nonmotile. The strain is pleomorphic and reproduces by nonuniform division. Strain Z-0056 is an aerobic organoheterotroph and a mesophilic and moderately acidophilic oligotrophic microorganism. As an inhabitant of dystrophic ultrafresh waters, strain Z-0056 is sensitive to NaCl. The bacterium utilizes organic acids (acetate, pyruvate, oxalate, gluconate, malate, succinate, and citrate), as well as xylose and xylan. The microorganism grows in a pH range of 4.0–8.0, with an optimum at pH 5.5. The temperature range for growth is 15–25°C, with an optimum at 20°C. According to its ecophysiological properties, strain Z-0056 belongs to the group of ombrophilic dissipotrophs. The DNA G+C base content is 66.8 mol %. According to phylogenetic analysis, strain Z-0056 belongs to the genus Ancylobacter. Strain Z-0056 showed the highest similarity (98.3%) with the type strain of the species A. oerskovii. The phenotypic and phylogenetic properties of strain Z-0056 support classification of this microorganism within the genus Ancylobacter as the novel species Ancylobacter abiegnus sp. nov.     

A new species of Melampsora on Populus yunnanensis from China

Melampsora nujiangensis, a rust fungus found on Populus yunnanensis in China, is described as a new species. Light and scanning electron microscopy with herbarium specimens of the rust fungus show that the shape of its urediniospores differs from that of other known species of Melampsora, and its urediniospore walls are thinner than the other species. Furthermore, in phylogenetic trees based on the DNA sequences (28S and ITS) the rust fungus is phylogenetically separated by high bootstrap values. These results indicate that the fungus is an isolated species among the genus Melampsora. Contribution no. 202 from the Laboratory of Plant Parasitic Mycology, Graduate School of Life and Environmental Sciences, University of Tsukuba, Japan     

地黄内生放线菌的分离及地黄轮纹病拮抗菌Streptomyces folium leaf-16的新种鉴定

药用植物内生放线菌具有合成天然活性化合物的潜力,放线菌新种是寻找新型抗生素先导化合物的一个重要来源。【目的】挖掘药用植物地黄内生放线菌资源,并对地黄轮纹病拮抗菌株leaf-16进行新种鉴定。【方法】本研究采用五步消毒法分离河南道地药材地黄的内生放线菌,以地黄轮纹病原真菌草茎点霉(Phoma herbarum)为指示菌,采用平板对峙法筛选对该病菌有抑制作用的菌株,16S rRNA基因测序发现一株抗地黄轮纹病的放线菌新种leaf-16。通过形态、生理生化、细胞壁化学组分和分子生物学等特征对菌株leaf-16进行多相分类学鉴定。【结果】经平板对峙实验得到8株抗地黄轮纹病的放线菌,其中菌株leaf-16经16S rRNA基因测序、形态比较、生理生化、化学组分和分子生物学以及DNA-DNA杂交分析,确定菌株leaf-16为1株链霉菌新种,并命名为Streptomyces folium。【结论】菌株leaf-16为1株链霉菌新种,具有抑制地黄轮纹病原真菌的活性,为进一步分离新型抗地黄轮纹病的生物制剂奠定物质基础。     

Purification,Characterization, and Molecular Cloning of Acidophilic Xylanase from Penicillium sp.40

Penicillum sp. 40, which can grow in an extremely acidic medium at pH 2.0 was screened from an acidic soil. This fungus produces xylanases when grown in a medium containing xylan as a sole carbon source. A major xylanase was purified from the culture supernatant of Penicillium sp. 40 and designated XynA. The molecular mass of XynA was estimated to be 25,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. XynA has an optimum pH at 2.0 and is stable in pH 2.0-5.0. Western blot analysis using anit-XynA antibody showed that XynA was induced by xylan and repressed by glucose. Also, its production was increased by an acidic medium. The gene encoding XynA (xynA) was isolated from the genomic library of Penicillium sp. 40. The structural part of xynA was found to be 721 bp. The nucleotide sequence of cDNA amplified by RT-PCR showed that the open reading frame of xynA was interrupted by a single intron which was 58 bp in size and encoded 221 amino acids. Direct N-terminal amino acid sequencing showed that the precursor of XynA had a signal peptide composed of 31 amino acids. The molecular mass caliculated from the deduced amino acid sequence of XynA is 20,713. This is lower than that estimated by gel electrophoresis, suggesting that XynA is a glycoprotein. The predicted amino acid sequence of XynA has strong similarity to other family11 xylanases from fungi.     

Large-Scale Cultivation of Acidophilic Hyperthermophiles for Recovery of Secreted Proteins

An electric water heater was modified for large-scale cultivation of aerobic acidophilic hyperthermophiles to enable recovery of secreted proteins. Critical changes included thermostat replacement, redesign of the temperature control circuit, and removal of the cathodic anticorrosion system. These alterations provided accurate temperature and pH control. The bioreactor was used to cultivate selected strains of the archaeon Sulfolobus solfataricus and other species within this genus. Reformulation of a basal salts medium facilitated preparation of large culture volumes and eliminated sterilization-induced precipitation of medium components. Substrate induction of synthesis of the S. solfataricus-secreted alpha-amylase during growth in a defined medium supported the utility of the bioreactor for studies of physiologically regulated processes. An improved purification strategy was developed by using strong cation-exchange chromatography for recovery of the alpha-amylase and the processing of large sample volumes of acidic culture supernatant. These findings should simplify efforts to study acidophilic hyperthermophilic microbes and their secreted proteins.     

Aecidium dispori is the aecial anamorph ofPuccinia albispora, sp. nov. (Uredinales)

Aecidium dispori forms spermogonium and aecium onDisporum sessile andD. smilacinum, which are distributed in East Asia. TheAecidium species is found to be an aecial anamorph of aPuccinia fungus, with its uredinial-telial stage being formed onCarex conica, C. dolichostachya subsp.multifolia, C. pisiformis subsp.alterniflora andC. rugata. Urediniospores of this fungus are large, colorless, thick-walled with 4–5 equatorial germ pores. The morphological characteristics of urediniospores and the spermogonial-aecial host do not fit to any set of circumscribing characters of previously described species. We consider the fungus to be a new species and propose a new name,Puccinia albispora, for the fungus.     

Molecular and morphological studies of Paecilomyces sinensis reveal a new clade in clavicipitaceous fungi and its new systematic position

The type materials of Paecilomyces sinensis, including herbarium specimen and ex-type strain, were re-examined to clarify its relationships with other species. Morphological observations on the strain grown in various culture media revealed that the fungus was morphologically related to Polycephalomyces, since it produced conidial mass and lanceolate or narrowly lageniform phialides. Six genes, including ITS, nrSSU, nrLSU, tef1, rpb1 and rpb2, were amplified from the type materials and used in phylogenetic analyses to determine the systematic position of the fungus in the framework of clavicipitaceous fungi. The results place P. sinensis with Polycephalomyces formosus, the type species of Polycephalomyces, and Cordyceps ramosopulvinata forming a clade unaffiliated with the known families of clavicipitaceous fungi. Based on both morphological and molecular data, a new combination, Polycephalomyces sinensis, is proposed for Paecilomyces sinensis. The new clade found in this study is designated as Polycephalomyces clade and expands the phylogenetic diversity for clavicipitaceous fungi. The teleomorph–anamorph connection between Berkelella and Polycephalomyces previously conceived cannot be retained as the type species of Polycephalomyces, P. formosus, is closely linked to species of Cordyceps s.l. in the new clade.     

In vitro screening of taxol,an anticancer drug produced by the fungus,Colletotrichum capsici

The fungus Colletotrichum capsici was isolated from the diseased fruits of Chilli plant, Capsicum annuum. The isolated test fungus was identified by its morphological and molecular characteristic features. For the first time, the fungus was screened for the production of taxol on modified liquid medium. The presence of taxol was confirmed by the spectroscopic and chromatographic methods of analyses. The amount of taxol produced by this fungus was quantified by HPLC. The maximum amount of fungal taxol production was recorded as 687 μg/L. The production rate was 13 740‐fold higher than that, previously reported for the fungus Taxomyces andreanae. The extracted fungal taxol showed a strong cytotoxic activity in an in vitro culture of human cancer cells indicating that the increase in taxol concentration induces increased cell death. A PCR‐based screening for taxadiene synthase (ts), a unique gene in the formation of the taxane skeleton, confirmed the molecular blueprint for taxol biosynthesis. The results show that the fungus C. capsici is an excellent candidate for an alternate source of taxol supply and can serve as a potential species for genetic engineering to enhance the production of taxol to a higher level.     

Micro‐ and macrospatial scale analyses illustrates mixed mating strategies and extensive geneflow in populations of an invasive haploid pathogen

Sexual reproduction in fungi involves either a single individual (selfing) or two individuals (outcrossing). To investigate the roles that these two strategies play in the establishment of an invasive alien pathogen, the Eucalyptus leaf‐infecting fungus, Teratosphaeria (Mycosphaerella) nubilosa was studied. Specifically, the genetic diversity of the pathogen was investigated at micro and macrospatial scales. Interestingly, while data obtained at microspatial scales show clearly that selfing is the main reproductive strategy, at macrospatial scales the population genetic structure was consistent with a genetically outcrossing organism. Additional analyses were performed to explore these apparently discordant results at different spatial scales and to quantify the contribution of selfing vs. outcrossing to the genotypic diversity. The results clearly show that the fungus has a mixed mating strategy. While selfing is the predominant form of mating, outcrosses must have occurred in the pathogen that increased the genotypic diversity of the fungus over time. This mating strategy, coupled with the high levels of geneflow between distant populations of the pathogen, has created an even distribution of maximum diversity from the smallest (leaf) to largest scales (>500 km), which will make breeding for resistance difficult. These data illustrate the evolutionary potential and danger of the introduction of multiple genotypes of a potentially outcrossing pathogen, especially when it has a high dispersal potential.     

New acidic chitinase isoforms induced in tobacco roots by vesicular-arbuscular mycorrhizal fungi

Summary Chitinase activities have been compared in tobacco roots (Nicotiana tabacum cv. Xanthi nc) infected by the pathogenic fungus Chalara elegans or three species of vesicular arbuscular mycorrhizal (VAM) fungi: Glomus versiforme, G. intraradix and G. fasciculatum, using native polyacrylamide gel electrophoresis (PAGE). All previously known acidic chitinase isoforms were stimulated in roots by the pathogenic fungus and by the VAM fungi, while two new acidic chitinase isoforms were specifically induced in response to the endomycorrhizal association. After separation in sodium dodecyl sulphate polyacrylamide denaturing gels (SDS-PAGE) under non-reducing conditions, the estimated apparent molecular mass for these additional acidic chitinase isoforms from VAM-colonized samples was 33 kDa, compared to 30 kDa for the main activity stimulated in C. elegans-infected root extracts.