Hormographiella verticillata and an Ozonium stage as anamorphs of Coprinellus domesticus

Living cultures of Hormographiella verticillata and Coprinellus domesticus, as well as herbarium specimens of Ozonium spp. have been examined and compared. Based on morphological and molecular data, we present for the first time C. domesticus as a teleomorphic stage of H. verticillata and a first record of H. verticillata isolated from human skin in America (Panama). In addition to Hormographiella, species of Ozonium have been described as anamorphic stages of species of Coprinus s.l. Based on extensive review of the literature and own observations, we propose to apply the names Hormographiella spp. to conidiogenous stages of species of Coprinus s.l. and Ozonium stage to the sterile brown hyphae which can be observed in nature and in older cultures of species of Hormographiella. Because species of Ozonium are not distinguishable morphologically, we do not use species names in this genus. The two imperfect stages represent synanamorphs which are designated explicitly for a species of Coprinus s.l. for the first time.     

Separate and interactive effects of competition and herbivory on the growth, expansion, and tuber formation of Hydrilla verticillata

This study examined the interaction and main-effect impacts of herbivory by the leaf-mining fly Hydrellia pakistanae and plant competition from Vallisneria americana on the growth, expansion and tuber formation of Hydrilla verticillata in a 2 × 2 factorial design experiment. The study was conducted in 14,000-L tanks, over two growing seasons. Each tank represented a single experimental unit and contained 32 1-L pots. At the beginning of the experiment half of these were planted with H. verticillata while the other half were either left empty or planted with V. americana (the competitor). H. pakistanae fly larvae (the herbivore) were added to tanks as appropriate. No significant interactions were identified between herbivory and competition on any parameter of H. verticillata growth analyzed (i.e., total tank biomass accumulation, total number of rooting stems, total tuber number, total tuber mass, and tuber size), indicating that the factors were operating independently and neither antagonism nor synergism was occurring. Both competition and herbivory impacted the growth of H. verticillata. H. verticillata plants grown in the presence of V. americana developed less total biomass, had fewer total basal stems, had fewer tubers and less tuber mass per tank, and produced significantly smaller tubers relative to control plants. Herbivory also suppressed H. verticillata biomass accumulation and tended to suppress the number and total mass of tubers produced in each tank. Both factors showed 30–40% reduction of total H. verticillata biomass, although the mechanism of impact was different. Competition suppressed expansion of H. verticillata into adjoining pots but had little impact on its growth in pots where it was originally planted. Herbivory resulted in a general suppression of growth of H. verticillata in all pots. Although herbivory significantly impacted H. verticillata biomass, it did not result in competitive release for V. americana under the current experimental conditions. We conclude that management activities that promote competition or herbivory will impact the growth and expansion of H. verticillata. Furthermore, since these factors operated independently, the combined use of both factors should be beneficial for suppression of H. verticillata dominance.     

Virulence of the entomopathogenic nematodes Heterorhabditis bacteriophora, Heterorhabditis zealandica, and Steinernema scarabaei against five white grub species (Coleoptera: Scarabaeidae) of economic importance in turfgrass in North America

We compared the virulence of the entomopathogenic nematodes Steinernema scarabaei, Heterorhabditis zealandica, and Heterorhabditis bacteriophora (GPS11 and TF strains) against third instars of the Japanese beetle, Popillia japonica, the oriental beetle, Anomala (=Exomala) orientalis, the northern masked chafer, Cyclocephala borealis, the European chafer, Rhizotrogus majalis, and the Asiatic garden beetle, Maladera castanea, in laboratory and greenhouse experiments. The virulence of the nematode species relative to each other differed greatly among white grub species. H. bacteriophora and H. zealandica had similar modest virulence to P. japonica, A. orientalis, C. borealis, and M. castanea. But against R. majalis, H. zealandica showed low virulence with a clear concentration response whereas H. bacteriophora caused only erratic and very low mortality. In contrast, S. scarabaei had modest virulence against C. borealis, but was highly virulent against R. majalis, P. japonica, A. orientalis, and M. castanea with R. majalis being the most susceptible and M. castanea the least susceptible.     

Discrimination of species and strains of basidiomycete genusCoprinus by random amplified polymorphic DNA (RAPD) analysis

All five examined strains ofCoprinus cinereus could be clearly discriminated from the strains of five otherCoprinus species by RAPD patterns with 12 of 13 primers. Also one specimen of unknownCoprinus strain was identified to beC. cinereus by this method. The RAPD patterns were similar among the strains in the same species; many common DNA fragments were recognized as well as some strain-specific DNA fragments. Thus all seven strains ofC. cinereus and all four strains ofC. angulatus examined could be distinguished individually. Diakryotic strains showed the combined RAPD patterns of the two monokaryotic strains constituting the dikaryon. The combined RAPD markers observed in the dikaryons were segregated in their basidiospore progeny. All 18 randomly picked progeny showed different combinations of RAPD markers from the parental strains.     

Mapping a new nematode resistance locus in Lycopersicon peruvianum

Accessions of the wild tomato species L. peruvianum were screened with a root-knot nematode population (557R) which infects tomato plants carrying the nematode resistance gene Mi. Several accessions were found to carry resistance to 557R. A L. peruvianum backcross population segregating for resistance to 557R was produced. The segregation ratio of resistant to susceptible plants suggested that a single, dominant gene was a major factor in the new resistance. This gene, which we have designated Mi-3, confers resistance against nematode strains that can infect plants carrying Mi. Mi-3, or a closely linked gene, also confers resistance to nematodes at 32°C, a temperature at which Mi is not effective. Bulked-segregant analysis with resistant and susceptible DNA pools was employed to identify RAPD markers linked to this gene. Five-hundred-and-twenty oligonucleotide primers were screened and two markers linked to the new resistance gene were identified. One of the linked markers (NR14) was mapped to chromosome 12 of tomato in an L. esculentum/L. pennellii mapping population. Linkage of NR14 and Mi-3 with RFLP markers known to map on the short arm of chromosome 12 was confirmed by Southern analysis in the population segregating for Mi-3. We have positioned Mi-3 near RFLP marker TG180 which maps to the telomeric region of the short arm of chromosome 12 in tomato.     

Differences in susceptibility of introduced and native white grub species to entomopathogenic nematodes from various geographic localities

Invasive, non-native, white grubs (Coleoptera: Scarabaeidae) cause significant damage in urban landscapes. Although the lack of natural enemies in their new home is often suggested as an important factor in the establishment and spread of invasive species, the potential of incumbent generalist parasites and pathogens to delay their establishment and spread has not been explored. We compared the susceptibility of the introduced Popillia japonica and the native Cyclocephala borealis to 16 species and strains of entomopathogenic nematodes isolated from within or outside the geographic ranges of the two scarabs. We found large variation in the virulence of the species/strains of nematodes with over 50% mortality of P. japonica produced by Heterorhabditis zealandica strain X1 and H. bacteriophora strain GPS11 and of C. borealis by H. zealandica and H. bacteriophora strains KMD10 and NC1. Heterorhabditis indica and H. marelatus caused less than 20% mortality of both scarab species. When considered as a group the nematode species and strains from within and outside the geographic ranges of either P. japonica or C. borealis did not differ in virulence towards either scarab species. Dose response studies with selected nematode species and strains against P. japonica and two additional non-native species Anomala (Exomala) orientalis and Rhizotrogus majalis and the native C. borealis indicated that R. majalis was the least susceptible and P. japonica and A. orientalis were as susceptible as the native C. borealis. Heterorhabditis zealandica was significantly more virulent than any other species or strain against P. japonica with a LC₅₀ of 272 IJs/grub. The LC₃₀ and LC₅₀ values for H. zealandica were also the lowest among the four nematode species/strains tested against A. orientalis and C. borealis. The LC₅₀ values for H. zealandica and H. megidis (UK strain) were significantly lower for the native C. borealis than the introduced A. orientalis. H. zealandica also showed the highest penetration efficiency and the lowest encapsulation in P. japonica and C. borealis grubs. Results suggest that the introduction of the exotic H. zealandica into the front-line states with respect to the movement of P. japonica and A. orientalis should be explored as a tactic to delay their establishment and spread. The results also suggest that the manipulation of the indigenous H. bacteriophora populations may help in delaying spread and mitigating losses caused by the invasive grub species.     

Ribosomal DNA repeat unit polymorphism in 25 Hordeum species

Summary Tandemly repeated DNA sequences containing structural genes encoding ribosomal RNA (rDNA) were investigated in 25 species of Hordeum using the wheat rDNA probe pTA71. The rDNA repeat unit lengths were shown to vary between 8.5 and 10.7 kb. The number of length classes (1–3) per accession generally corresponded to the number of nucleolar organizing regions (NORs). Intraspecific variation was found in H. parodii, H. spontaneum and H. leporinum, but not in H. bulbosum. Restriction analysis showed that the positions of EcoRI, SacI and certain BamHI cleavage sites in the rRNA structural genes were highly conserved, and that repeat unit length variation was generally attributable to the intergenic spacer region. Five rDNA BamHI restriction site maps corresponded to the following groups of species: Map A — H. murinum, H. glaucum, H. leporinum, H. bulbosum, H. marinum, H. geniculatum; Map B — H. leporinum; Map C — H. vulgare, H. spontaneum, H. agriocrithon; Map D — H. chilense, H. bogdanii; and Map E — remaining 14 Hordeum species. The repeat unit of H. bulbosum differed from all other species by the presence of a HindIII site. The closer relationship of H. bulbosum to H. leporinum, H. murinum and H. glaucum than to H. vulgare was indicated by their BamHI restriction maps.Contribution No. 1169, Plant Research Centre     

Salt Tolerance in Aquatic Macrophytes: Ionic Relation and Interaction

Effects of seawater salinity (SWS) and pure NaCl on the intracellular contents of Na⁺, K⁺, Mg²⁺, Ca²⁺, chlorophylls (Chl) and carotenoids (Car) were studied in three submerged aquatic macrophytes, Hydrilla verticillata, Najas indica and Najas gramenia, which differed in their tolerance to salinity. NaCl resulted in significant increase in Chl/Car ratio in the salt-sensitive H. verticillata and moderately salt-tolerant N. indica, but not in the salt-tolerant N. gramenia. SWS treatment did not result in any significant change in the ratio. The intracellular content of Na⁺ increased significantly in all the test plants upon exposure to both NaCl and SWS. The content of K⁺ decreased significantly in these plants upon salinity treatment, except in N. gramenia. The contents of Ca²⁺ and Mg²⁺ decreased significantly upon NaCl treatment and remained unchanged or increased upon SWS treatment. No relationship between salt tolerance and K⁺/Na⁺ ratio was observed. The maintenance of a minimal level of K⁺ was observed to be the most probable requirement of salt tolerance in aquatic macrophytes.     

An expanded family of fungalysin extracellular metallopeptidases of Coprinopsis cinerea

Proteolytic enzymes, particularly secreted proteases of fungal origin, are among the most important of industrial enzymes, yet the biochemical properties and substrate specificities of these proteins have been difficult to characterize. Genomic sequencing offers a powerful tool to identify potentially novel proteases. The genome of the model basidiomycete Coprinopsis cinereus was found to have an unusually high number of metalloproteases that closely match the M36 peptidase family known as fungalysins. The eight predicted C. cinereus fungalysins divide into two groups upon comparison with fungalysins from other fungi. One member, CcMEP1, is most similar to the single representative fungalysins from the basidiomycetes Phanerochaete chrysosporium, Cryptococcus neoformans, and Ustilago maydis, and to the fungalysin type-protein from Aspergillus fumigatus. The remaining seven C. cinereus predicted fungalysins form a group with similarity to three predicted M36 peptidases of Laccaria bicolor. All eight of the C. cinereus enzymes contain both the signature M36 Pfam domain and the FTP propeptide domain. All contain large propeptides with considerable sequence conservation near a proposed cleavage site. The predicted mature enzymes range in size from 37–46 kDa and have isoelectric points that are mildly acidic to neutral. The proximity of these genes to telomeres and/or to transposable elements may have contributed to the expansion of this gene family in C. cinereus.     

香菇印gpd-Le和ras-Le启动子的功能分析

利用从香菇菌丝体中克隆的启动子片段gpd-Le(613bp)和ras-Le(715bp)分别连接于报告基因gfp(绿色荧光蛋白基因)的上游,构建了启动子功能活性检测表达质粒pLg-gfp和pLr-gfp。采用PEG介导法把表达质粒pLg-gfp和pLr-gfp分别与辅助质粒pCc1001(含有trp1基因)共转化进色氨酸营养缺陷型的灰盖鬼伞粉孢子的原生质体中。经过选择培养基筛选、假定转化子的分子鉴定以及GFP荧光检测。结果表明:香菇gpd-Le启动子在灰盖鬼伞的菌丝中具有较强驱动外源gfp基因表达的活性,在荧光显微镜和共聚焦显微镜下观察到gfp基因表达的绿色荧光。而香菇ras-Le启动子没有检测到有驱动外源gfp基因表达的活性。     

香菇印gpd-Le和ras-Le启动子的功能分析

利用从香菇菌丝体中克隆的启动子片段gpd-Le(613bp)和ras-Le(715bp)分别连接于报告基因gfp(绿色荧光蛋白基因)的上游,构建了启动子功能活性检测表达质粒pLg-gfp和pLr-gfp。采用PEG介导法把表达质粒pLg-gfp和pLr-gfp分别与辅助质粒pCc1001(含有trp1基因)共转化进色氨酸营养缺陷型的灰盖鬼伞粉孢子的原生质体中。经过选择培养基筛选、假定转化子的分子鉴定以及GFP荧光检测。结果表明:香菇gpd-Le启动子在灰盖鬼伞的菌丝中具有较强驱动外源gfp基因表达的活性,在荧光显微镜和共聚焦显微镜下观察到gfp基因表达的绿色荧光。而香菇ras-Le启动子没有检测到有驱动外源gfp基因表达的活性。     

Structural organization of the nuclear ribosomal RNA genes in Cannabis and Humulus (Cannabaceae)

The structural organization of the nuclear ribosomal DNA (rDNA) of Humulus lupulus, H. japonicus and Cannabis sativa was determined by restriction site mapping. A high degree of DNA sequence similarity was evident in the coding regions of the rDNA repeats of the taxa and supports the placement of Cannabis and Humulus in one family, Cannabaceae. However, the presence of a BstEII site, an additional SacI site, absence of the SpeI site and positional differences of the SspI sites in the 25 S gene distinguished H. japonicus from H. lupulus. Humulus lupulus has an additional EcoRV site in the IGS region. A XhoI site in the 18S region of C. sativa distinguishes it from the two hop species. The diagnostic differences in the IGS of C. sativa include the EcoRI, HindIII and XhoI sites. These sites were not detected in the IGS of the two hop species.     

Molecular characterization and identification ofSaprolegnia by restriction analysis of genes coding for ribosomal RNA

Restriction fragment length polymorphisms (RFLPs) in two regions of the ribosomal DNA (rDNA) repeat unit were examined in 33 strains representing 18 species ofSaprolegnia. The Polymerase Chain Reaction (PCR) was used to separately amplify the 18S rDNA and the region spanning the two internal transcribed spacers (ITS) and the 5.8S ribosomal RNA gene. Amplified products were subjected to a battery of restriction endonucleases to generate various fingerprints. The internal transcribed spacer region exhibited more variability than the 18S rDNA and yielded distinctive profiles for most of the species examined. Most of the species showing 100% similarity for the 18S rDNA could be distinguished by 5.8S + ITS restriction polymorphisms except forS. hypogyna, S. delica, S. lapponica, andS. mixta. The rDNA data indicate thatS. lapponica andS. mixta are conspecific withS. ferax, whereas there is no support for the proposed synonymies ofS. diclina withS. delica and ofS. mixta withS. monoica. Results from cluster analysis of the two data sets were very consistent and tree topologies were the same, regardless of the clustering method used. A further examination of multiple strains in theS. diclina-S. parasitica complex showed that restriction profiles are conserved across different strains ofS. parasitica originating from the U.K. and Japan.HhaI andBsaI restriction polymorphisms were observed in isolates from the U.S. and India. The endonucleaseBstUI was diagnostic forS. parasitica, generating identical fingerprints for all strains regardless of host and geographic origin. Except for the atypical strain ATCC 36144, restriction patterns were also largely conserved inS. diclina. Correlation of the rDNA data with morphological and ultrastructural features showed thatS. diclina andS. parasitica are not conspecific. Restriction polymorphisms in PCR-amplified rDNA provide a molecular basis for the classification ofSaprolegnia and will be useful for the identification of strains that fail to produce antheridia and oogonia.     

Kuraishia MolischianaSp. Nov., the Teleomorph of Candida Molischiana

Thirty-two strains, many of them isolated from wood-associated habitats, and designated as Kuraishia (Pichia) capsulata and Candida molischiana according to their phenotype, exhibited two types of HaeIII restriction fragment patterns of their small subunit rDNA with the neighboring ITS. One fragment pattern corresponded to that of the type strain of K. capsulata, whereas the other pattern was unique to the typestrain of C. molischiana. Sequencing of the D1/D2 domain of the large subunit rDNA confirmed that the different HaeIII restriction fragment patterns of small subunit rDNA with the neighboring ITS reliably distinguished K. capsulata from C. molischiana. Ascospore formation was observed in several C. molischiana strains and K. molischiana (type strain: NCAIM Y.01725, CBS 9993) is proposed as the teleomorphic state of Candida molischiana.     

凤眼莲入侵程度对金鱼藻和黑藻生长及种间关系的影响

湿地生态系统中凤眼莲（Eichhornia crassipes）入侵造成湿地植物群落结构退化及功能崩溃,直接影响沉水植物的生长繁殖及初级生产力。目前关于凤眼莲的入侵机制有一定的研究,而关于凤眼莲入侵程度对沉水植物金鱼藻（Ceratophyllum demersum）和黑藻（Hydrilla verticillate）生长及种间关系的影响相对缺乏。以外来入侵植物凤眼莲,沉水植物金鱼藻和黑藻为研究对象,设计凤眼莲入侵程度（无入侵,轻度入侵对应盖度25%,重度入侵对应盖度75%）交叉定植方式（黑藻单种模式、金鱼藻单种模式,金鱼藻和黑藻混种模式）的控制实验,探究凤眼莲入侵强度对沉水植物金鱼藻和黑藻生长及种间关系的影响。结果表明,凤眼莲入侵程度显著降低了金鱼藻的生物量、分枝数;黑藻的株高、分枝数和分节数。无凤眼莲入侵时,两种沉水植物生物量均最大,两者种间竞争关系较强;随凤眼莲入侵盖度增加,两种沉水植物的生物量先急剧降低后略微增加,种间关系经过微弱促进后又变为竞争作用,其中黑藻表现出明显的竞争优势。此外,凤眼莲入侵略微降低了水体中的总氮、总磷含量。结构方程模型分析结果表明凤眼莲入侵以及水体总氮、总磷等水体理化性质对沉水植物生长均有显著负向影响（P<0.05）,且水体理化性质对沉水植物生长的影响强于凤眼莲入侵。总之,凤眼莲入侵显著降低了金鱼藻和黑藻生长繁殖,随着凤眼莲入侵程度增加,两种沉水植物种间关系由竞争转变为促进再转变为竞争。研究结果为凤眼莲入侵有效控制及湿地沉水植被的恢复与重建提供了一定的理论依据和技术支撑。     

Inheritance of chromosome length polymorphisms inCoprinus cinereus

The sexually compatible strains ofCoprinus cinereus 5302 and Dd 13 revealed chromosome length polymorphisms in their electrophoretic karyotypes. The dikaryon derived from two monokaryons contained a mixture of the two electrophoretic patterns. F₁ progenies were isolated by crossingC. cinereus 5302 and Dd 13 strains and it showed unique karyotypes. Chromosome length polymorphisms of both parental strains were inherited at random in the F₁ progenies. As a result, several novel electrophoretic karyotypes which had not been observed in either parental strains were found in the F₁ progeny. The rDNA probe hybridized with one chromosome in both parental strains, with two chromosomes in the hybridization pattern of both parental strains in the dikaryon, and with one to two chromosomes in the F₁ progenies. The relation between mating type and hybridization pattern has thus not been made clear in the case of F₁ progeny.     

Saccharomyces barnetti andSaccharomyces spencerorum: two new species ofSaccharomyces sensu lato (van der Walt)

In the course of a study of DNA base sequence homology, 19 strains labelled asSaccharomyces exiguus, its imperfect state,Candida holmii, orC. milleri were examined. Results confirmed the separation ofC. milleri as a separate species. The remaining strains can be divided into three distinct groups of genomic relatedness. The type cultures ofS. exiguus andC. holmii form a cluster with 10 other strains showing variable reassociation values ranging from 100 to 40%. The remaining four strains comprise two separate species showing no nucleotide relatedness between themselves nor to either theS. exiguus complex or toC. milleri. Physiological analyses demonstrate that it is possible to separate these four taxa on the basis of a few simple tests. The two new species are described respectively asSaccharomyces barnetti honoring James Barnett in recognition of his invaluable work in the field of yeast taxonomy andSaccharomyces spencerorum in honor of J.F.T. and Dorothy M. Spencer who have made innumerable contributions to the genetics and biotechnological application of yeasts.     

Adsorption of lead(II) from aqueous solution onto Hydrilla verticillata

The adsorption of Pb(II) onto Hydrilla verticillata was examined in aqueous solution with parameters of pH, adsorbent dosage, contact time and temperature. The linear Langmuir and Freundlich models were applied to describe equilibrium isotherms, and both models fitted well. The monolayer adsorption capacity of Pb(II) was found as 104.2 mg/g at pH 4 and 25°C. Dubinin–Radushkevich (D–R) isotherm model was also applied to the equilibrium data. The mean free energy of adsorption (15.81 kJ/mol) indicated that the adsorption of Pb(II) onto H. verticillata may be carried out via chemical ion-exchange mechanism. Thermodynamic parameters, free energy (ΔG ⁰), enthalpy (ΔH ⁰) and entropy (ΔS ⁰) of adsorption were also calculated. These parameters showed that the adsorption of Pb(II) onto H. verticillata was a feasible, spontaneous and exothermic process in nature. The influence of Cd²⁺, Cu²⁺ and Ni²⁺ on adsorption of Pb²⁺ onto H. verticillata was studied, too. In the investigated range of operating conditions, it was found that the existence of Cd ²⁺, Cu ²⁺ and Ni ²⁺ had no impact on the adsorption of Pb²⁺.     

Mitochondrial DNA analysis of Exophiala jeanselmei var. lecanii-corni and Exophiala castellanii

A Japanese clinical isolate (KU-A-0094) which was identified by de Hoog et al. as Exophiala jeanselmei var. lecanii-corni with difficulty, was compared with 5 strains including the type cultures of E. jeanselmei var. lecanii-corni, var. jeanselmei and E. castellanii using RFLP (restriction fragment length polymorphism) patterns of mtDNA (mitochondrial DNA). RFLP patterns of KUA-0094 were identical with those of E. jeanselmei var. lecanii-corni and different from those of E. castellanii with restriction enzymes of HaeIII, MspI and hindIII. Therefore, de Hoog et al.'s identification of KU-A-0094 was confirmed. Additionally, mtDNA-RFLP patterns of E. jeanselmei var. lecanii-corni and E. jeanselmei var. jeanselmei were also different from each other. Consequently E. jeanselmei var. lecanii-corni seem to be a species in its own right rather than a variant of E. jeanselmei. This revised version was published online in June 2006 with corrections to the Cover Date.     

Ultrastructure of chitin in hyphae ofCandida albicans and other dimorphic and mycelial fungi

Summary Chitin microfibrils exposed by chemical extraction of hyphal walls ofCandida albicans, Histoplasma capsulatum, Blastomyces dermatitidis, Paracoccidiodes brasiliensis, Coprinus cinereus andMucor mucedo were of variable morphology but gave identical infrared spectra and behaved as pure chitin in chromatographic analyses. The microfibrils of the four dimorphic fungi studied were shorter than those in the mouldsC. cinereus andM. mucedo but were similar to those reported for the yeastSaccharomyces cerevisiae. InC. albicans the microfibrils in the septal plates of hyphae were predominantly tangentially orientated and were longer than those in the lateral walls. Microfibrils produced by chitin synthasein vitro were very much longer than any observed from hyphal preparations.