• 한국동물위생학회지
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• 제39권1호
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• pp.1-6
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• 2016

In this study, Bacillus licheniformis which has been used as probiotics was isolated from Korean traditional fermented soybean. A total of 69 strains were presumptively identified as B. licheniformis by phenotypic methods. Based on PCR amplification and 16S rRNA gene sequencing, the multilocus sequence typing of gyrA and rpoB, followed by phylogenetic analysis was performed. The isolates were distinctly differentiated and found to be closely related to B. amyloliquefaciens, B. subtilis, and B. aerius. The partial 16S rRNA gene sequences of those strains matched those of B. sonorensis (97%) and B. aerius (98%) in the phylogenetic tree. In contrast, multilocus phylogenetic analysis (MLPA) showed that only 61 (86.9%) out of 69 strains were B. licheniformis. The rest of those strains were found to be B. subtilis (5.8%), B. amyloliquefaciens (2.9%), and B. sonorensis (2.9%), respectively. Therefore, our results suggested that since the 16S rRNA gene sequencing alone was not sufficient to compare and discriminate closely related lineages of Bacillus spp., it was required to analyze the MLPA simultaneously to avoid any misleading phenotype-based grouping of these closely related species.

• Genomics & Informatics
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• 제11권4호
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• pp.272-276
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• 2013

Sequence analysis of the 16S rRNA gene has been widely used for the classification of microorganisms. However, we have been unable to clearly identify five Flavobacterium species isolated from a freshwater by using the gene as a single marker, because the evolutionary history is incomplete and the pace of DNA substitutions is relatively rapid in the bacteria. In this study, we tried to classify Flavobacterium species through multilocus sequence analysis (MLSA), which is a practical and reliable technique for the identification or classification of bacteria. The five Flavobacterium species isolated from freshwater and 37 other strains were classified based on six housekeeping genes: gyrB, dnaK, tuf, murG, atpA, and glyA. The genes were amplified by PCR and subjected to DNA sequencing. Based on the combined DNA sequence (4,412 bp) of the six housekeeping genes, we analyzed the phylogenetic relationship among the Flavobacterium species. The results indicated that MLSA, based on the six housekeeping genes, is a trustworthy method for the identification of closely related Flavobacterium species.

• Food Science and Biotechnology
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• 제27권6호
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• pp.1755-1760
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• 2018

The objective of this study was to perform genetic diversity analysis of 13 strains isolated from South Korean foods by multilocus sequence typing (MLST). For typing, seven housekeeping loci (atpA, dnaA, dnaK, gyrB, pheS, pyrG, and rpoA) were selected, amplified and analyzed. Fifty-one polymorphic sites varying from 1 to 22 in each species were identified. Thirteen sequence types were generated with allele numbers ranged from 2 to 10. The overall relationship between strains was assessed by unweighted pair group method with arithmetic mean dendrogram and minimum spanning tree. In addition, combined spits tree analysis revealed intragenic recombination. No clear relationship was observed between the isolation sources and strains. The developed MLST scheme enhanced our knowledge of the population diversity of Leu. citreum strains and will be used further for the selection of industrially important strain.

• 한국어병학회지
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• 제32권2호
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• pp.69-80
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• 2019

We performed MLSA (multilocus sequence analysis) and phenotypic characterization of Aliivibrio species isolated from walleye pollock (Gadus chalcogrammus) maintained in 3 different facilities of Gangwon Province, the east coast of Korea. Of 38 Aliivibrio species identified by 16S rDNA sequences, 12 strains were randomly selected and MLSA was conducted with 5 house-keeping genes (gapA, gyrB, pyrH, recA and rpoA) and 16S rDNA gene. Phylogenetic analysis and homology of the concatenated sequences (4,580 bp) with other Vibrionaceae genera revealed that 4 strains (GNGc16.1, YYGc16.1, YYGc16.2, GSGc18.1) were identified as Aliivibrio logei and one strain (GSGc16.1) as A. wodanis. One strain (GSGc17.1) was tentatively identified as A. logei, but needs further analysis because it did not belong to the same clade with A. logei type strain. 6 strains (GSGc17.2, GNGc16.2, GSGc16.2, GSGc17.3, GSGc18.2, GSGc17.4) need further investigation as potential novel species. Either phenotypic characterization or 16S rDNA sequence alone did not provide enough information for identification of Aliivibrio strains at the species level. A. logei and A. wodanis are generally known as non-pathogenic bacteria, but also known as opportunistic or secondary pathogens of cold water fishes. Cares should be taken to prevent potential outbreaks due to these bacteria, although there was no outbreaks during the sampling period.

• Mycobiology
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• 제49권4호
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• pp.308-345
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• 2021

Lactifluus (Pers.) Roussel is an ectomycorrhizal genus that was recently recognized to be distinct from the genus Lactarius. To date, 226 Lactifluus species have been reported worldwide. Misidentification of Lactifluus species is common because of intraspecific morphological variation, cryptic diversity, and the limited number of taxonomic keys available. Molecular data are indispensable for species delimitation; a multilocus phylogenetic analysis showed that most Asian Lactifluus species are not conspecific with morphologically similar species present on other continents. In particular, Korea has misused European and North American Lactifluus names. In this study, we evaluated the taxonomy of Lactifluus in Korea using both morphological and multilocus molecular (ITS, nrLSU, rpb1, and rpb2) data. We examined 199 Lactifluus specimens collected between 1980 and 2016, and a total of 24 species across the four Lactifluus subgenera were identified. All Korean species are distinct and clearly separated from European and North American species. Five taxa corresponded to previously described species from Asia and the remaining 19 taxa are confirmed as new species. Herein, we provide keys to the Korean Lactifluus species within their subgenera, molecular phylogenies, a summary of diversity, and detailed description of the new species.

• 한국미생물·생명공학회지
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• 제46권2호
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• pp.162-170
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• 2018

Non-typhoidal Salmonella is one of the main pathogens causing food-borne illness in humans, with up to 20% of cases resulting from consumption of pork products. Over the gastroenteritis signs, multidrug resistant Salmonella has arisen. In this study, pan-susceptible phenotypic strains of Salmonella enterica serotype Weltevreden recovered from pig production chain in Chiang Mai, Thailand during 2012-2014 were chosen for analysis. The aim of this study was to use whole genome sequencing (WGS) data with an emphasis on antimicrobial resistance gene investigation to assess their pathogenic potential and genetic diversity determination based on whole genome Multilocus Sequence Typing (wgMLST) to expand epidemiological knowledge and to provide additional guidance for disease control. Analyis using ResFinder 3.0 for WGS database tracing found that one of pan-susceptible phenotypic strain carried five classes of resistance genes: aminoglycoside, beta-lactam, phenicol, sulfonamide, and tetracycline associated genes. Twenty four and 36 loci differences were detected by core genome Multilocus Sequence Typing (cgMLST) and pan genome Multilocus Sequence Typing (pgMLST), respectively, in two matching strains (44/13 vs A543057 and A543056 vs 204/13) initially assigned by conventional MLST and Pulsed-field Gel Electrophoresis (PFGE). One hundread percent discriminant ability can be achieved using the wgMLST technique. WGS is currently the ultimate molecular technique for various in-depth studies. As the findings stated above, a new of "gold standard typing method era" for routine works in genome study is being set.

• Mycobiology
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• 제43권3호
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• pp.360-365
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• 2015

Multilocus sequence typing analysis was applied to determine the genotypes of 147 (137 clinical and 10 environmental) Cryptococcus neoformans and three clinical Cryptococcus gattii isolates from 1993 to 2014 in Korea. Among the 137 clinical isolates of C. neoformans, the most prevalent genotype was ST5 (n = 131), followed by ST31 (n = 5) and ST127 (n = 1). Three C. gattii strains were identified as ST57, ST7, and ST113. All environmental isolates were identified as C. neoformans with two genotypes, ST5 (n = 7) and ST31 (n = 3). Our results show that C. neoformans isolates in Korea are genetically homogeneous, and represent a close genetic relationship between clinical and environmental isolates.

• The Plant Pathology Journal
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• 제37권3호
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• pp.243-257
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• 2021

Bacillus pumilus is the causal agent of trunk bulges disease affecting rubber and rubberwood quality and yield production. In this study, B. pumilus and other closely related species were included in B. pumilus group, as they shared over 99.5% similarity from 16S rRNA analysis. Multilocus sequence analysis (MLSA) of five housekeeping genes and repetitive elements-based polymerase chain reaction (rep-PCR) using REP, ERIC, and BOX primers conducted to analyze the diversity and systematic relationships of 20 isolates of B. pumilus group from four rubber tree plantations in Peninsular Malaysia (Serdang, Tanah Merah, Baling, and Rawang). Multi rep-PCR results revealed the genetic profiling among the B. pumilus group isolates, while MLSA results showed 98-100% similarity across the 20 isolates of B. pumilus group species. These 20 isolates, formerly established as B. pumilus, were found not to be grouped with B. pumilus. However, being distributed within distinctive groups of the B. pumilus group comprising of two clusters, A and B. Cluster A contained of 17 isolates close to B. altitudinis, whereas Cluster B consisted of three isolates attributed to B. safensis. This is the first MLSA and rep-PCR study on B. pumilus group, which provides an in-depth understanding of the diversity of these rubber-pathogenic isolates in Malaysia.

• Mycobiology
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• 제42권1호
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• pp.73-78
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• 2014

Cryptococcal infection is primarily caused by two species, Cryptococcus neoformans and C. gattii. Between the two species, C. neoformans var. grubii is the major causative agent of cryptococcosis in Asia. We investigated the molecular characteristics of 46 isolates of C. neoformans from patients with cryptococcosis between 2008 and 2012 in Seoul, Korea. All the isolates were determined to be C. neoformans var. grubii (serotype A), mating type $MAT{\alpha}$, and molecular type VNI by PCR-restriction fragment length polymorphism of the URA5 gene. Multilocus sequencing type (MLST) analysis using the International Society of Human and Animal Mycoses (ISHAM) consensus MLST scheme identified two sequence types (ST). Out of the 46 strains, 44 (95.7%) were identified as ST5, and remaining 2 were identified as ST31. Our study revealed that the clinical strains of C. neoformans in Korea are genetically homogeneous with the VNI/ST5 genotypes, and new appearance of VNI/ST31 genotype may serve as an important indicator of global genetic analysis.

• The Plant Pathology Journal
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• 제33권1호
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• pp.21-33
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• 2017

Citrus blast caused by bacterium Pseudomonas syringae is a very important disease of citrus occuring in many areas of the world, but with few data about genetic structure of the pathogen involved. Considering the above fact, this study reports genetic characterization of 43 P. syringae isolates obtained from plant tissue displaying citrus blast symptoms on mandarin (Citrus reticulata) in Montenegro, using multilocus sequence analysis of gyrB, rpoD, and gap1 gene sequences. Gene sequences from a collection of 54 reference pathotype strains of P. syringae from the Plant Associated and Environmental Microbes Database (PAMDB) was used to establish a genetic relationship with our isolates obtained from mandarin. Phylogenetic analyses of gyrB, rpoD, and gap1 gene sequences showed that P. syringae pv. syringae causes citrus blast in mandarin in Montenegro, and belongs to genomospecies 1. Genetic homogeneity of isolates suggested that the Montenegrian population might be clonal which indicates a possible common source of infection. These findings may assist in further epidemiological studies of this pathogen and for determining mandarin breeding strategies for P. syringae control.