Epitypification can solve many taxonomic problems and stabilize the understanding of species, genera, families or orders. The aim of this paper is to illustrate how to epitypify. A few examples where taxa have been epitypified are considered and the benefits and disadvantages of epitypification are discussed. We also outline some examples of taxa which need to be epitypified with reasons.
Colletotrichum ; classification ; genetics ; DNA, Fungal ; genetics ; isolation & purification ; Fungi ; classification ; genetics ; Mycological Typing Techniques ; methods ; Phylogeny

Appressorium formation is an important event in establishing a successful interaction between the rice blast fungus, Magnaporthe oryzae, and its host plant, rice. An understanding of molecular events occurring in appressorium differentiation will give new strategies to control rice blast. A quick and reliable method to extract total RNA from appressorium is essential for studying gene expression during appressorium formation and its mechanism. We found that duplicate film is an efficient substratum for appressorium formation, even when inoculated with high density conidia. When inoculated with conidia at 1x10(6) ml(-1), the percentages of conidium germination and appressorium formation were (97.98+/-0.67)% and (97.88+/-0.45)%, respectively. We applied Trizol before appressorium collection for total RNA isolation, and as much as 113.6 microg total RNA was isolated from the mature appressoria at 24 h after inoculation. Functional analysis of two genes, MNH6 and MgATG1, isolated from the cDNA subtractive library, revealed that the quantity of RNA was good enough to construct a cDNA (complementary DNA) library or a cDNA subtractive library. This method may be also applicable for the appressorium RNA isolation of other pathogenic fungi in which conidia differentiate into appressoria in the early stages of host infection.
DNA, Complementary ; genetics ; isolation & purification ; DNA, Fungal ; genetics ; isolation & purification ; Gene Library ; Genes, Fungal ; Magnaporthe ; genetics ; growth & development ; pathogenicity ; Oryza ; microbiology ; Plant Diseases ; microbiology ; RNA, Fungal ; genetics ; isolation & purification

To establish a simple, sensitive and effective technique for the identification of six common dermatophytes, polymerase chain reaction (PCR) and PCR-restriction fragment length polymorphism (RFLP) targeting Topoisomerase II gene were used. The DNA of 6 common dermatophytes was amplified by primer dPsD1 and then primers dPsD2. The products generated by dPsD2 were digested with restriction enzyme Hinc II and Hinf I separately. A DNA fragment of about 3390 bp was amplified by using primer dPsD1 from the genomic DNA of each dermatophyte species. The product of dPsD2 was 2380 bp and the restriction profiles of Hinc II and Hinf I were between 58-1670 bp. By using PCR-RFLP, all of the 6 dermatophytoses were diagnosed to species level and no obvious difference identification between Hinc II and Hinf I. It is concluded that the PCR-RFLP identification of dermatophytes by Hinc II or Hinf I is efficient and rapid in clinical practice.
Arthrodermataceae/*classification ; Arthrodermataceae/genetics ; Arthrodermataceae/*isolation & purification ; DNA Topoisomerases, Type II/genetics ; DNA, Fungal/analysis ; DNA, Fungal/genetics ; Dermatomycoses/*microbiology ; Polymerase Chain Reaction ; Polymorphism, Restriction Fragment Length

The swollen root of Rehmannia glutinosa is used as one kind of important Chinese traditional medicine. The root of R. glutinosa usually swelled in rotational cropping but not in continuous cropping. The rhizosphere microorganisms of R. glutinosa under different farming condition were thought related to that. In this study, the endophytic fungi in the root of R. glutinosa growing in various soil conditions were isolated for the study of the relationship between the microorganisms and the root enlargement of their host plants. The dominant endophytes, Verticillium spp., Fusarium oxysporum, F. redolens and Ceratobasidium spp. were identified by morphological observation and 18S rDNA and ITS sequence analysis. The preliminary investigation showed that the excessive growth of Verticillium and Fusarium genus fungi is unfavorable for the R. glutinosa root swelling, but Ceratobasidium fungi has no effects on the root enlargement.
DNA, Fungal ; genetics ; DNA, Ribosomal ; genetics ; Fungi ; classification ; genetics ; isolation & purification ; Molecular Sequence Data ; Plant Roots ; microbiology ; Rehmannia ; microbiology

OBJECTIVETo identify the endophytic fungal strain PR35 separated from Paeonia delavayi and study chemical constituents of its secondary metabolites.

METHODThe fungal strain PR35 was identified by morphological observation and ITS rDNA sequence analysis. Various chromatographic methods were adopted to separate and purify its secondary metabolites, and their structures were identified by physiochemical properties and spectral data

RESULTThe fungal strain PR35 was identified as Trichoderma longibrachiatum. Five compounds were separated from fermentation products of fungal strain PR35 and identified as 1-(2,6-dihydroxyphenyl)-3-hydroxybutan-1-one (1), 1-(2,6-dihydroxypheny) propan-1-one (2), 1-(2,4,6-trihydroxyphenyl) butan-1-one (3), 4-methoxy-1-naphthol (4), and cerevisterol (5). Among them, compounds 1-3 showed notable antifungal activities against Botrytis cinerea, Fusarium avenaceum and Hormodendrum compactum.

CONCLUSIONThe endophytic fungus T. longibrachiatum was separated from the plant P. delavayi for the first time. Five compounds were first separated from endophytic fungus of P. delavayi. Among them, compound 4 was separated from microbial fermentation products for the first time.

DNA, Fungal ; genetics ; DNA, Intergenic ; genetics ; Endophytes ; classification ; genetics ; isolation & purification ; metabolism ; Paeonia ; microbiology ; Phylogeny ; Trichoderma ; classification ; genetics ; isolation & purification ; metabolism

OBJECTIVETo promote development and utilization of Ophiocordyceps gracilis in xinjiang and provide basic data for researching and sustainable developing medicine fungus related to O. gracilis.

METHODA white strain SFYT002 isolated from the sclerotium of O. gracilis in Xinjiang was researched by morphological observation, ITS and 18SrDNA sequencing. The ITS and 18SrDNA sequences of the strain were determined, BLAST was compared with the other sequences of Tolypocladium in GenBank. The phylogenetic trees of ITS and 18SrDNA sequences were analyzed in Tolypocladium. In addition, the filter paper method was used to study the antibacterial effects.

RESULTThe main morphological characters of this strain were white cotton-like colonies, phialide with inflated base, drastically sharping with partially bending tips, small and transparent budding spores with being always assemble to spearhead and globular, subglobular or ellipse conidiospores. The phylogenetic trees of ITS and 18SrDNA sequences were constructed and analyzed in Tolypocladium. It was resulted that Tolypocladium was confirmed to be monophyletic, and the strain SFYT002 was the same as the systematic position of others of T. inflatum. Meanwhile, the antibacterial test was performed against the 4 common pathogenic bacteria. It was showed that both fermentation and its extracts of different polar from this strain possessed good anti-bacteria capacities.

CONCLUSIONThe strain SFYT02 was identified as T. inflatum, and inhibited effectively growth of bacteria.

Anti-Bacterial Agents ; isolation & purification ; pharmacology ; China ; DNA, Fungal ; genetics ; DNA, Intergenic ; genetics ; Hypocreales ; genetics ; isolation & purification ; physiology ; Medicine, Chinese Traditional ; methods ; Mycelium ; Phylogeny

BACKGROUND: The identification of molds in clinical laboratories is largely on the basis of phenotypic criteria, the classification of which can be subjective. Recently, molecular methods have been introduced for identification of pathogenic molds in clinical settings. Here, we employed comparative sequence analysis to identify molds. METHODS: A total of 47 clinical mold isolates were used in this study, including Aspergillus and Trichophyton. All isolates were identified by phenotypic properties, such as growth rate, colony morphology, and reproductive structures. PCR and direct sequencing, targeting the internal transcribed spacer (ITS) region, the D1/D2 region of the 28S subunit, and the beta-tubulin gene, were performed using primers described previously. Comparative sequence analysis by using the GenBank database was performed with the basic local alignment search tool (BLAST) algorithm. RESULTS: For Aspergillus, 56% and 67% of the isolates were identified to the species level by using ITS and beta-tubulin analysis, respectively. Only D1/D2 analysis was useful for Trichophyton identification, with 100% of isolates being identified to the species level. Performances of ITS and D1/D2 analyses were comparable for species-level identification of molds other than Aspergillus and Trichophyton. In contrast, the efficacy of beta-tubulin analysis was limited to genus identification because of the paucity of database information for this gene. CONCLUSIONS: The molecular methods employed in this study were valuable for mold identification, although the different loci used had variable usefulness, according to mold genus. Thus, a tailored approach is recommended when selecting amplification targets for molecular identification of molds.
Aspergillus/genetics/isolation & purification ; DNA, Fungal/analysis/isolation & purification ; Databases, Genetic ; Fungi/genetics/*isolation & purification ; Humans ; Polymerase Chain Reaction ; RNA, Ribosomal, 28S/*genetics ; Sequence Analysis, DNA ; Trichophyton/genetics/isolation & purification ; Tubulin/*genetics

OBJECTIVETo isolate and identify the anamorph of Shiraia bambusicola.

METHODFungus strains were isolated from mature ascospores and stroma. They were identified by means of morphological identification including colony and microscope characteristic, the molecular identification was done by 5.8S-ITS rDNA.

RESULTThe strains GZDXIFR-171 and GZDXIFR-181 were isolated and obtained with the separation of different methods, which had the same colony morphology. With the universal primers of the ITS1-5.8S rDNA-ITS2, the 5.8S-ITS rDNA sequence of GZDXIFR-171 and GZDXIFR-181 were obtained by the PCR amplification and sequencing. Compared with the published nucleotide sequence of 5.8S-ITS rDNA in NCBI (National Center for Biotechnology Information), GZDXIFR-171 and GZDXIFR-181 were highly identical with S. bambusicola.

CONCLUSIONThe isolated strains GZDXIFR-171 and GZDXIFR-181 were confirmed to be the true anamorph of S. bambusicola.

Ascomycota ; classification ; genetics ; isolation & purification ; DNA, Fungal ; genetics ; DNA, Ribosomal Spacer ; genetics ; Molecular Sequence Data ; Mycological Typing Techniques ; Phylogeny ; RNA, Ribosomal, 5.8S ; genetics

A new stem rot disease is found to occur naturally on Arabidopsis plants in greenhouses of Fuzhou, China. In order to identify its pathogen, we conducted a series of fungal isolation and purification, plant reinoculation, and ascus and ascospore induction from the sclerotia. The isolate caused typical water-soaked lesions after reinoculation and produced sclerotia both on Arabidopsis plants and culture medium plates, and the sclerotia could be induced to produce discal apothecia and 8 binucleate ascospores per ascus. These disease symptom and fungal morphology data revealed that the fungus Sclerotinia sclerotiorum (Lib.) de Bary was the pathogen for Arabidopsis stem rot. To confirm this, we further amplified its large subunit ribosomal DNA (LSU rDNA) by polymerase chain reaction (PCR), and compared the sequence with the known LSU rDNA sequences in GenBank. The results show that the sequence shares the highest identities with the LSU rDNAs of different S. sclerotiorum strains. Taking all these data together, we concluded that the fungus that caused the Arabidopsis stem rot is S. sclerotiorum (Lib.) de Bary. This is the first report that Arabidopsis is naturally infected by S. sclerotiorum.
Arabidopsis ; microbiology ; Ascomycota ; classification ; genetics ; isolation & purification ; pathogenicity ; Base Sequence ; China ; DNA, Fungal ; genetics ; DNA, Ribosomal ; genetics ; Phylogeny ; Plant Diseases ; microbiology ; Plant Stems ; microbiology

Aspergillosis ; diagnosis ; microbiology ; Aspergillus fumigatus ; genetics ; isolation & purification ; DNA, Fungal ; genetics ; isolation & purification ; Dermatomycoses ; diagnosis ; microbiology ; Eye Diseases ; diagnosis ; microbiology ; Female ; Humans ; Middle Aged ; Polymerase Chain Reaction