• Journal of Microbiology and Biotechnology
• /
• v.18 no.4
• /
• pp.624-630
• /
• 2008

Terminal restriction fragment length polymorphism (T-RFLP) is a method that has been frequently used to survey the microbial diversity of environmental samples and to monitor changes in microbial communities. T-RFLP is a highly sensitive and reproducible procedure that combines a PCR with a labeled primer, restriction digestion of the amplified DNA, and separation of the terminal restriction fragment (T-RF). The reliable identification of T-RF requires the information of nucleotide sequences as well as the size of T-RF. However, it is difficult to obtain the information of nucleotide sequences because the T-RFs are fragmented and lack a priming site of 3'-end for efficient cloning and sequence analysis. Here, we improved on the T-RFLP method in order to analyze the nucleotide sequences of the distinct T-RFs. The first method is to selectively amplify the portion of T-RF ligated with specific oligonucleotide adapters. In the second method, the termini of T-RFs were tailed with deoxynucleotides using terminal deoxynucleotidyl transferase (TdT) and amplified by a second round of PCR. The major T-RFs generated from reference strains and from T-RFLP profiles of activated sludge samples were efficiently isolated and identified by using two modified T-RFLP methods. These methods are less time consuming and labor-intensive when compared with other methods. The T-RFLP method using TdT has the advantages of being a simple process and having no limit of restriction enzymes. Our results suggest that these methods could be useful tools for the taxonomic interpretation of T-RFs.

• Korean Journal of Microbiology
• /
• v.39 no.1
• /
• pp.16-20
• /
• 2003

A phylogenetic analysis of bacterial populations inhabiting soil derived from serpentine was conducted. The samples were collected from adjacent metamorphic rocks and serpentinite soil at Kwangcheon. The pH of the serpentine areas ranged from 8.5 to 9.2. The number of bacteria on the DAL medium which was diluted with $10^{-2}$ of AL medium was 10~100 fold higher than that from the full strength of AL medium, and which indicates that oligotrophs are distributed in the serpentinite soil. Of a total of 76 isolates, 42 isolates were oligotrophic bacteria, which grew only on the DAL medium. Based on a phylogenetic analysis using 16S rDNA sequences, these isolates are found to fall within five major phylogenetic groups: proteobacteria $\alpha$-subdivision (3 strains), $\alpha$-subdivision (7 strains), $\gamma$-subdivision (2 trains); high G+C gram-positive bacteria (19 strains); low G+C grampositive bacteria (14 strains). Bacteria of the genus Streptomyces (high G+C division) and Bacillus (low G+C division) have been considered to form a numerically important fraction of serpentinite soil. Oligotrophic strains categorized as Afipia ($\alpha$-subdivision), Ralstonia, Variovorax ($\beta$-subdivision), Pseudomonas ($\gamma$ -subdivision), Arthrobacter (high G+C division), and Streptomyces (low G+C division).

• Journal of the Korean Society of Food Science and Nutrition
• /
• v.43 no.5
• /
• pp.749-755
• /
• 2014

Exopolysaccharides (EPSs) have been widely used in the food industry as viscofying, stabilizing, and emulsifying agents as well as in the pharmaceutical industry for their immunomodulatory, anti-tumor, and anti-inflammatory effects. A total of 458 lactic acid bacteria (LAB) strains isolated from several kinds of soybean pastes were screened for the production of homo-EPS (HoPS). LAB isolates were primarily screened using thin layer chromatography (TLC) and further screened polymerase chain reaction (PCR) targeting genes involved in HoPS production. Six LAB isolates producing high amounts of HoPS were identified by TLC. Among these isolates, glucansucrase gene was amplified in two strains (JSA57, JSB22), whereas the fructansucrase gene was detected in three strains (JSA57, JSB22, JSB66). After isolating the strains, their morphological characteristics and 16S rDNA sequences were determined. Six species were identified as L. alimentarius HSB15, L. plantarum JSA22, L. pentosus JSA57, L. brevis JSB22, L. alimentarius JSB66, and L. parabrevis JSB89. To evaluate the potential probiotic properties of these LAB, their survival rates against a simulated intestinal environment were determined. After 2 hr of incubation in artificial gastric juice, survival rates of JSA57, JSB90, JSB22, and JSB66 were all greater than 50%. After 2 hr of incubation in bile juice, viable cell count of JSB22 was similar with initial vial cell counts. Growth of the six LAB was screened in arabino-oligosaccharide (AOS)-containing MRS broth. Results showed that growth of the isolates selectively increased after culture in AOS-containing media. Strain JSB22 (6 hr), JSB66 (6 hr), HSB15 (20 hr), and JSA22 (29 hr) showed maximum growth rate. Especially, JSB22 showed the highest growth rate. These results suggest that EPS-producing LAB isolated from Deonjang could be applied as synbiotics.

• The Plant Pathology Journal
• /
• v.32 no.6
• /
• pp.575-579
• /
• 2016

Burkholderia glumae (bacterial grain rot), Xanthomonas oryzae pv. oryzae (bacterial leaf blight), and Acidovorax avenae subsp. avenae (bacterial brown stripe) are major seedborne pathogens of rice. Based on the 16S and 23S rDNA sequences for A. avenae subsp. avenae and B. glumae, and transposase A gene sequence for X. oryzae pv. oryzae, three sets of primers had been designed to produce 402 bp for B. glumae, 490 bp for X. oryzae, and 290 bp for A. avenae subsp. avenae with the $63^{\circ}C$ as an optimum annealing temperature. Samples collected from naturally infected fields were detected with two bacteria, B. glumae and A. avenae subsp. avenae but X. oryzae pv. oryzae was not detected. This assay can be used to identify pathogens directly from infected seeds, and will be an effective tool for the identification of the three pathogens in rice plants.

• Korean Journal of Fisheries and Aquatic Sciences
• /
• v.54 no.4
• /
• pp.568-573
• /
• 2021

A single leptocephalus specimen of Albula glossodonta belonging to the family Albulidae under the order Albuliformes, was collected from the southern sea of Korea in May 21, 2019. This species is distinguished from A. vulpes in the number of melanophores at the base of the caudal fin. In addition, our specimen was readily distinguished from A. argentea in having no melanophores on the operculum, at the base of the dorsal fin and in front of the pectoral fin. Also it was well distinguished from A. gilberti in having no melanophores on the operculum, in front of the pectoral fin and on the lateral body. A molecular analysis based on 467 bp sequences of the mitochondrial DNA 16S rRNA shows that the leptocephalus is well matched to A. glossodonta adult (K2P distance=0.002). We report the first occurrence of A. glossodonta leptocephalus in the Korean waters, and propose it's new Korean name, "Nam-bang-yeoeul-myeol".

• Research in Plant Disease
• /
• v.29 no.3
• /
• pp.304-309
• /
• 2023

Wilted and water-soaked lesion symptoms were observed on cucumbers in greenhouses located in Daejeon, Chungcheongnam-do, Korea, in June 2021. A bacterial strain, designated KNUB-04-21, was isolated from the cucumbers, which was subsequently identified as Pectobacterium brasiliense through a phylogenetic analysis based on sequences of the 16S rRNA region, dnaX, leuS, and recA genes. The biochemical characteristics of KNUB-04-21 were also similar to those of P. brasiliense through investigation using the API ID 32 GN system. The pathogenicity of KNUB-04-21 was confirmed by inoculating it into healthy cucumber plants. The reisolated strains were also found to be same to the original strain. To our knowledge, this is the first report of P. brasiliense being identified as the causative agent of cucumber soft rot in Korea.

• Research in Plant Disease
• /
• v.29 no.1
• /
• pp.72-78
• /
• 2023

In September 2021, gray-to-brown discoloration and expanding water-soaked lesions were observed on the outer and inner layers and the core of kimchi cabbage (Brassica rapa subsp. pekinensis) in fields located in Samcheok, Gangwondo, Korea. A bacterial strain designated as KNUB-02-21 was isolated from infected cabbage samples. Phylogenetic analysis based on the sequences of the 16S rRNA region and the dnaX, leuS, and recA genes confirmed that the strain was affiliated with Pectobacterium versatile. Additionally, the biochemical and morphological profiles of the isolate were similar to those of P. versatile. Based on these results, the isolate was identified as a novel strain of P. versatile. Healthy kimchi cabbage slices developed soft rot upon inoculation with P. versatile KNUB-02-21 and exhibited symptoms similar to those observed in the diseased plants in fields. The re-isolated strains were similar to those of P. versatile. Prior to our study, P. versatile as the causative pathogen of kimchi cabbage soft rot had not been reported in Korea.

• Research in Plant Disease
• /
• v.29 no.4
• /
• pp.445-451
• /
• 2023

Zamioculcas zamiifolia is a popular indoor ornamental plant in Korea. In August 2021, a severe outbreak of soft rot disease affected Z. zamiifolia in Emseong, Chungcheongbuk-do, Korea. Infected plants displayed wilting, water-soaked lesions, stem collapse, and green-brown discoloration. The bacterial strain KNUB-05-21 was isolated from infected stems and identified as Pectobacterium aroidearum using 16S rRNA nucleotide sequencing and multilocus sequence analysis based on partial sequences of dnaX, leuS, and recA genes. Confirmation of its affiliation with P. aroidearum was also obtained through biochemical and morphological characterization. To confirm the pathogenicity of strain KNUB-05-21, its suspension was injected into Z. zamiifolia stems. Within a week, soft rot developed on the stems, exhibiting symptoms similar to those observed in field-infected plants. The reisolated strain was identical to those of P. aroidearum. Before this study, P. aroidearum was not reported as a causative pathogen of Z. zamiifolia soft rot in Korea.

• Research in Plant Disease
• /
• v.29 no.4
• /
• pp.452-458
• /
• 2023

In the Goesan region, located in Chungcheongbuk-do, Korea, a significant outbreak of soft rot infections was documented in August 2021, affecting fruits of Momordica charantia, commonly known as bitter melon or bitter gourd. The symptoms included a noticeable transition to yellowing in the affected fruits, eventually leading to their collapse. The bacterial strain KNUB-09-21 was isolated from the diseased fruits. Molecular analysis, using the sequences of the 16S rRNA region and three housekeeping genes (dnaX, recA, and leuS), along with the results of compound utilization in the API ID 32 GN system, provide strong evidence for the identification of the isolate KNUB-09-21 as Pectobacterium brasiliense. The pathogenicity of strain KNUB-09-21 on M. charantia was confirmed through a controlled inoculation test. Within two days, inoculated fruits displayed soft rot symptoms closely resembling those observed in naturally affected fruits. This is the first report of soft rot on M. charantia in Korea.

• Journal of Life Science
• /
• v.14 no.2
• /
• pp.229-234
• /
• 2004

Feathers, which are almost pure keratin protein, are produced in large amounts as a waste by-product at poultry-processing plants. Keratinolytic enzymes may have important uses in biotechnological processes involving keratin-containing wastes from poultry and leather processes. In this study, screening and identification of keratin-degrading bacteria were investigated. Five keratin-degrading bacterial strains (F3-1, F3-4, F7-1, C1-1, C1-2) were isolated from compost and decayed chicken feather. On the basis of morphological, physiological studies, and Biolog system, all isolates were identified as the genus Bacillus. Among them, the strain F7-1 had the highest feather-degrading activity and was selected for further taxonomical study. Phylogenetic analysis of strain F7-1 based on comparison of 165 rDNA sequences revealed that this strain is closely related to Bacillus megaterium.