First Report of Root Rot Caused by Plectosphaerella cucumerina on Cabbage in China

Severe root rot was observed in fields of cabbages (Brassica oleracea L.) in 2015 in China. Cardinal symptoms of this disease included root rot and wilting leaves. A fungus was isolated from diseased tissues consistently. Based on the morphological features and molecular analysis of the ITS-5.8S rDNA and D1/D2 domain of the 28S rRNA gene, it was identified as Plectosphaerella cucumerina. This is the first report of P. cucumerina causing cabbage root rot in China and the world.

rDNA-ITS and 28 S rDNA D1/D2 sequence analysis of downy mildew pathogen from Aconitum carmichaeli / 中草药

Objective: To define sequences about rDNA-ITS and 28 S rDNA D1/D2 of Peronospora aconiti separated from cultivated Aconitum carmichaeli in Jiangyou area of Sichuan province and provide a theoretical basis for the diagnosis and prevention of downy mildew disease. Methods: Spores and hyphae of P. aconiti from diseased plants were collected and total genomic DNA of pathogen were extracted and then rDNA-ITS and 28 S rDNA D1/D2 fragment were amplified and sequenced. According to the above results, the abutment (Neighbor-joining, NJ) phylogenetic tree of pathogen was constructed and analyzed. Results: The rDNA-ITS and 28 S rDNA D1/D2 sequences of P. aconiti were sequenced and compared according to the database from NCBI. Compared that with P. pulveracea and P. aparines, the similarity of rDNA-ITS sequences of P. aconiti was 94%. The similarity of 28 S rDNA D1/D2 sequences of P. aconiti was 97% compared that with P. pulveracea, P. ficariae and P. bulbocapni. Conclusion: The results of morphological identification of downy mildew pathogen separated from A. carmichaeli are consistent with those from molecular identification (rDNA-ITS and 28 S rDNA D1/D2 sequences) and the pathogen of Aconitum downy mildew should be P. aconiti. Therefore, rDNA-ITS and 28 S rDNA D1/D2 sequences constructed in this paper can be used to identify downy mildew pathogen from Aconitum carmichaeli Debx.

Characterization and amylolytic activity of yeast and mold strains from Thai sweet rice

Aims: The aim of this study was to isolate and identify the yeast and mold strains from the starter and to investigate their amylolytic activity. Methodology and results: Thirty-two yeasts were isolated from ten samples of look-paeng khao-mak, a traditional starter culture in the production of khao-mak (sweet rice) in several places in Thailand. All isolates were identified based on their morphological and biochemical characteristics including the sequencing of D1/D2 domain of large subunit (26S) ribosomal DNA analyses. They were identified as Saccharomycopsis fibuligera (9 isolates), Candida rugosa (2 isolates), C. tropicalis (1 isolate), Clavispora lusitaniae (1 isolate), Wickerhamomyces anomalus (15 isolates) and Meyerozyma guilliermondii (4 isolates). All isolates of S. fibuligera showed high amylolytic activity. One-hundred isolates of mold were isolated from twenty-one samples of look-paeng khao-mak. They were belonged to Amylomyces sp. (42 isolates), Rhizopus sp. (30 isolates), Mucor sp. (12 isolates) and Penicillium sp. (16 isolates) based on their morphological characteristics. Four isolates, LK4-1, LK8-2, LK12-5 and LK17-1 showed amylase activity ranged from 32.24 to 39.74 unit/mL by dinitrosalicylic acid (DNSA) method. The isolates LK4-1, LK8-2 and LK12-5 were identified as Amylomyces rouxii and LK17-1 was Rhizopus microsporus based on their morphological characteristics and the ribosomal RNAencoding DNA (rDNA) internal transcribed spacer (ITS) sequences. Conclusion, significance and impact study: The characterization and evaluation of yeast and mold species based on their phenotypic and genetic characteristics including their amylase activity will be useful for the diversity and sweet rice production.

Identification of Clinical Mold Isolates by Sequence Analysis of the Internal Transcribed Spacer Region, Ribosomal Large-Subunit D1/D2, and beta-Tubulin

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.

Laboratory Experience in Phenotypic and Molecular Identification of Blastomyces dermatitidis First Isolated in Korea / 대한임상미생물학회지

Blastomycosis, endemic in North America, has been hardly reported in Korea. We describe laboratory experience in phenotypic and molecular identification of Blastomyces dermatitidis first isolated in Korea. The patient was a 45-year-old male with pulmonary blastomycosis mimicking pulmonary tuberculosis. Diagnosis was based on culture and dimorphism combined with DNA target sequencing of internal transcribed spacers (ITS) and D1/D2 regions.

Identification of Trichosporon spp. Strains by Sequencing D1/D2 Region and Sub-typing by Sequencing Ribosomal Intergenic Spacer Region of Ribosomal DNA / 华中科技大学学报（医学）（英德文版）

To re-identify and further group 25 isolates of Trichosporon spp. identified morphologically previously, sequences of D1/D2 region of large subunit (LSU) of ribosomal DNA (rDNA) of 25 tested strains for identification and those of ribosomal intergenic space 1 (IGS1) region of 11 strains for sub-grouping were detected. The identifications of tested strains were changed except 6 strains. According to the alignment of the IGS1 region, 6 T. asahii isolates tested fell into 4 groups and 5 T. faecale isolates into 3 groups. Polymorphism of 2 T.japonicum isolates was found in 10 positions. With the alignments obtained in this research compared with the relative GenBank entries, it was found that T. asahii, T.faecale and T.japonicum species were divided into 7, 3 and 2 subtypes respectively. Morphological and biophysical methods are not sufficient for Trichosporon spp. identification. Sequencing becomes neces-sary for Trichosporon diagnosis. There is obvious diversity within a species.

The Determination of the Partial 28S Ribosomal DNA Sequences and Rapid Detection of Phellinus linteus and Related species

Species of Phellinus were known to harmful fungi causing white pocket rot and severe plant disease such as canker or heart-rot in living trees in the West, but some species have been used to traditional medicines in the Orient for a long time. In this study the partial D1-D2 nucleotide sequences of 28S ribosomal DNA from 13 Phellinus strains were determined and compared with the sequences of 21 strains obtained from GenBank database. According to the neighbor-joining (NJ) method comparing the sequence data the phylogenetic tree was constructed. The phylogenetic tree displayed the presence of four groups. Group I includes P. ferreus, P. gilvus and P. johnsonianus, Group II contains P. laevigatus, P. conchatus and P. tremulae, Group III possesses P. linteus, P. weirianus, P. baumii, P. rhabarbarinus and P. igniarius, and Group IV comprises P. pini, P. chrysoloma. P. linteus and P. baumii, which were used mainly in traditional medicine, belong to the same group, but exactly speaking both were split into two different subgroups. To detect P. linteus only, we developed the PCR primer, D12HR. The primer showed the specific amplification of P. linteus, which is permitted to medicinal mushroom in the East. The results make a potential to be incorporated in a PCR identification system that could be used for the rapid identification of this species from its related species, P. linteus especially.

Prognosis for Patients with Early Gastric Cancer Comparison of D1 vs D2 lymphadenectomy

PURPOSE: Early gastric cancer is now considered to be a curable disease, and its traditional treatment is a D2 lymphadenectomy. However, the low rate of lymph node metastasis, the recent developments of endoscopic and laparoscopic surgery, and concerns for postoperative quality of life have led to less invasive therapeutic options. The D1 lymphadenectomy is one such option, so we investigated its adequacy as a substitute for a D2 lymphadenectomy as a treatment modality for early gastric cancer by comparing the prognoses of the two approaches. METHODS: A retrospective analysis of the case histories of 332 patients who had received an operation for early gastric cancer at Korea University Guro Hospital from 1984 to 1997 was performed. These cases were divided into D1 and D2 groups, and the groups were compared on the basis of clinicopathologic features, operative procedures, and 5-year survival rates. RESULTS: The D1 group included 160 cases, and the D2 group had 172 cases. The D2 group included more distal one-third cancer (66.3% vs 51.9%), more submucosal tumors (51.2% vs 38.7%), and more dissected lymph nodes (31.1+/-12.8 vs 23.0+/-11.3) than the D1 group (p0.05). When the tumor depth was considered, the 5-year survival rates of the D1 and the D2 groups were not significantly different for mucosal and submucosal tumors (p>0.05). CONCLUSION: A D2 lymphadenec tomy for early gastric cancer can harvest more lymph nodes, but it has no survival benefit over a D1 lymphadenectomy. The result of this retrospective study suggests that a D1 lymphadenectomy may be used as a replacement for a D2 lymphadectomy in early gastric-cancer surgery, although prospective randomized studies are needed.