• Restorative Dentistry and Endodontics
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• 제39권4호
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• pp.288-295
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• 2014

Objectives: This study was designed to evaluate the synergistic antibacterial effect of xylitol and ursolic acid (UA) against oral biofilms in vitro. Materials and Methods: S. mutans UA 159 (wild type), S. mutans KCOM 1207, KCOM 1128 and S. sobrinus ATCC 33478 were used. The susceptibility of S. mutans to UA and xylitol was evaluated using a broth microdilution method. Based on the results, combined susceptibility was evaluated using optimal inhibitory combinations (OIC), optimal bactericidal combinations (OBC), and fractional inhibitory concentrations (FIC). The anti-biofilm activity of xylitol and UA on Streptococcus spp. was evaluated by growing cells in 24-well polystyrene microtiter plates for the biofilm assay. Significant mean differences among experimental groups were determined by Fisher's Least Significant Difference (p < 0.05). Results: The synergistic interactions between xylitol and UA were observed against all tested strains, showing the FICs < 1. The combined treatment of xylitol and UA inhibited the biofilm formation significantly and also prevented pH decline to critical value of 5.5 effectively. The biofilm disassembly was substantially influenced by different age of biofilm when exposed to the combined treatment of xylitol and UA. Comparing to the single strain, relatively higher concentration of xylitol and UA was needed for inhibiting and disassembling biofilm formed by a mixed culture of S. mutans 159 and S. sobrinus 33478. Conclusions: This study demonstrated that xylitol and UA, synergistic inhibitors, can be a potential agent for enhancing the antimicrobial and anti-biofilm efficacy against S. mutans and S. sobrinus in the oral environment.

• Journal of Microbiology and Biotechnology
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• 제31권9호
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• pp.1191-1199
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• 2021

Enteropathogenic Escherichia coli (EPEC), which belongs to the attaching and effacing diarrheagenic E. coli strains, is a major causative agent of life-threatening diarrhea in infants in developing countries. Most EPEC isolates correspond to certain O serotypes; however, many strains are non-typeable. Two EPEC strains, EPEC001 and EPEC080, which could not be serotyped during routine detection, were isolated. In this study, we conducted an in-depth characterization of their putative O-antigen gene clusters (O-AGCs) and also performed constructed mutagenesis of the O-AGCs for functional analysis of O-antigen (OAg) synthesis. Sequence analysis revealed that the occurrence of O-AGCs in EPEC001 and E. coli O132 may be mediated by recombination between them, and EPEC080 and E. coli O2/O50 might acquire each O-AGC from uncommon ancestors. We also indicated that OAg-knockout bacteria were highly adhesive in vitro, except for the EPEC001 wzy derivative, whose adherent capability was less than that of its wild-type strain, providing direct evidence that OAg plays a key role in EPEC pathogenesis. Together, we identified two EPEC O serotypes in silico and experimentally, and we also studied the adherent capabilities of their OAgs, which highlighted the fundamental and pathogenic role of OAg in EPEC.

• Parasites, Hosts and Diseases
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• 제57권4호
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• pp.369-377
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• 2019

Artemisinin-based combination therapy (ACT) resistance is widespread throughout the Greater Mekong Subregion. This raises concern over the antimalarial treatment in Thailand since it shares borders with Cambodia, Laos, and Myanmar where high ACT failure rates were reported. It is crucial to have information about the spread of ACT resistance for efficient planning and treatment. This study was to identify the molecular markers for antimalarial drug resistance: Pfkelch13 and Pfmdr1 mutations from 5 provinces of southern Thailand, from 2012 to 2017, of which 2 provinces on the Thai- Myanmar border (Chumphon and Ranong), one on Thai-Malaysia border (Yala) and 2 from non-border provinces (Phang Nga and Surat Thani). The results showed that C580Y mutation of Pfkelch13 was found mainly in the province on the Thai-Myanmar border. No mutations in the PfKelch13 gene were found in Surat Thani and Yala. The Pfmdr1 gene isolated from the Thai-Malaysia border was a different pattern from those found in other areas (100% N86Y) whereas wild type strain was present in Phang Nga. Our study indicated that the molecular markers of artemisinin resistance were spread in the provinces bordering along the Thai-Myanmar, and the pattern of Pfmdr1 mutations from the areas along the international border of Thailand differed from those of the non-border provinces. The information of the molecular markers from this study highlighted the recent spread of artemisinin resistant parasites from the endemic area, and the data will be useful for optimizing antimalarial treatment based on regional differences.

• 한국미생물·생명공학회지
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• 제47권3호
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• pp.381-389
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• 2019

Chlorella vulgaris was isolated from the Nile River, Qalubia Governorate, Egypt, for possible use in biodiesel production. BG-II nutrient growth media was used for isolation and laboratory growth. Identification was performed via 18S rRNA gene amplification, followed by sequencing. The alga was exposed to UV-C (254 nm) for 15, 30, and 45 s to improve dry weight accumulation and to increase the oil production. Daily measurements of dry weight ($g{\cdot}l^{-1}$) were performed; oil content and volumetric lipid productivity were also determined. UV-C exposure led to an increase in the volumetric lipid productivity by 27, 27.3, and $32.4mg{\cdot}l^{-1}{\cdot}d^{-1}$ with 15, 30, and 45 s, respectively, as compared with the control, which resulted in $18mg{\cdot}l^{-1}{\cdot}d^{-1}$. Of the examined mutants, the one with the highest productivity was re-irradiated by UV-C (254 nm) for 15, 30, 45, and 60 s. For 15 s of exposure time, the oil content increased to 34%, while it was 31% at 30 s; further, it decreased to 22% at 45 and 60 s exposures. The fatty acid methyl ester profile was 82.22% in the first mutant at 45 s, compared with the wild strain that contained a total of 66.01% of FAs. Furthermore, the highest levels of polyunsaturated fatty acid methyl ester were observed in the mutant exposed for 45 s, and it reached 11.41%, which reduced the cetane number to 71.3.

• Journal of Microbiology and Biotechnology
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• 제29권3호
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• pp.357-366
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• 2019

We first confirmed the involvement of MalQ (4-${\alpha}$-glucanotransferase) in Escherichia coli glycogen breakdown by both in vitro and in vivo assays. In vivo tests of the knock-out mutant, ${\Delta}malQ$, showed that glycogen slowly decreased after the stationary phase compared to the wild-type strain, indicating the involvement of MalQ in glycogen degradation. In vitro assays incubated glycogen-mimic substrate, branched cyclodextrin (maltotetraosyl-${\beta}$-CD: G4-${\beta}$-CD) and glycogen phosphorylase (GlgP)-limit dextrin with a set of variable combinations of E. coli enzymes, including GlgX (debranching enzyme), MalP (maltodextrin phosphorylase), GlgP and MalQ. In the absence of GlgP, the reaction of MalP, GlgX and MalQ on substrates produced glucose-1-P (glc-1-P) 3-fold faster than without MalQ. The results revealed that MalQ led to disproportionate G4 released from GlgP-limit dextrin to another acceptor, G4, which is phosphorylated by MalP. In contrast, in the absence of MalP, the reaction of GlgX, GlgP and MalQ resulted in a 1.6-fold increased production of glc-1-P than without MalQ. The result indicated that the G4-branch chains of GlgP-limit dextrin are released by GlgX hydrolysis, and then MalQ transfers the resultant G4 either to another branch chain or another G4 that can immediately be phosphorylated into glc-1-P by GlgP. Thus, we propose a model of two possible MalQ-involved pathways in glycogen degradation. The operon structure of MalP-defecting enterobacteria strongly supports the involvement of MalQ and GlgP as alternative pathways in glycogen degradation.

• 대한수의학회지
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• 제61권2호
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• pp.13.1-13.8
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• 2021

Mammalian reovirus (MRV) causes respiratory and intestinal disease in mammals. Although MRV isolates have been reported to circulate in several animals, there are no reports on Korean MRV isolates from wildlife. We investigated the biological and molecular characteristics of Korean MRV isolates based on the nucleotide sequence of the segment 1 gene. In total, 144 swabs from wild animals were prepared for virus isolation. Based on virus isolation with specific cytopathic effects, indirect fluorescence assays, electron microscopy, and reverse transcription-polymerase chain reaction, only one isolate was confirmed to be MRV from a Korean roe deer (Capreolus pygargus). The isolate exhibited a hemagglutination activity level of 16 units with pig erythrocytes and had a maximum viral titer of 10^5.7 50% tissue culture infectious dose (TCID₅₀)/mL in Vero cells at 5 days after inoculation. The nucleotide and amino-acid sequences of the partial segment S1 of the MReo2045 isolate were determined and compared with those of other MRV strains. The MReo2045 isolate had nucleotide sequences similar to MRV-3 and was most similar (96.1%) to the T3/Bat/Germany/342/08 strain, which was isolated in Germany in 2008. The MReo2045 isolate will be useful as an antigen for sero-epidemiological studies and developing diagnostic tools.

• Journal of Microbiology and Biotechnology
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• 제32권9호
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• pp.1134-1145
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• 2022

SCO6993 (606 amino acids) in Streptomyces coelicolor belongs to the large ATP-binding regulators of the LuxR family regulators having one DNA-binding motif. Our previous findings predicted that SCO6993 may suppress the production of pigmented antibiotics, actinorhodin, and undecylprodigiosin, in S. coelicolor, resulting in the characterization of its properties at the molecular level. SCO6993-disruptant, S. coelicolor ΔSCO6993 produced excess pigments in R2YE plates as early as the third day of culture and showed 9.0-fold and 1.8-fold increased production of actinorhodin and undecylprodigiosin in R2YE broth, respectively, compared with that by the wild strain and S. coelicolor ΔSCO6993/SCO6993⁺. Real-time polymerase chain reaction analysis showed that the transcription of actA and actII-ORF4 in the actinorhodin biosynthetic gene cluster and that of redD and redQ in the undecylprodigiosin biosynthetic gene cluster were significantly increased by SCO6993-disruptant. Electrophoretic mobility shift assay and DNase footprinting analysis confirmed that SCO6993 protein could bind only to the promoters of pathway-specific transcriptional activator genes, actII-ORF4 and redD, and a specific palindromic sequence is essential for SCO6993 binding. Moreover, SCO6993 bound to two palindromic sequences on its promoter region. These results indicate that SCO6993 suppresses the expression of other biosynthetic genes in the cluster by repressing the transcription of actII-ORF4 and redD and consequently negatively regulating antibiotic production.

• Journal of Microbiology and Biotechnology
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• 제31권7호
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• pp.912-920
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• 2021

SOS response is a conserved response to DNA damage in prokaryotes and is negatively regulated by LexA protein, which recognizes specifically an "SOS-box" motif present in the promoter region of SOS genes. Myxococcus xanthus DK1622 possesses a lexA gene, and while the deletion of lexA had no significant effect on either bacterial morphology, UV-C resistance, or sporulation, it did delay growth. UV-C radiation resulted in 651 upregulated genes in M. xanthus, including the typical SOS genes lexA, recA, uvrA, recN and so on, mostly enriched in the pathways of DNA replication and repair, secondary metabolism, and signal transduction. The UV-irradiated lexA mutant also showed the induced expression of SOS genes and these SOS genes enriched into a similar pathway profile to that of wild-type strain. Without irradiation treatment, the absence of LexA enhanced the expression of 122 genes that were not enriched in any pathway. Further analysis of the promoter sequence revealed that in the 122 genes, only the promoters of recA2, lexA and an operon composed of three genes (pafB, pafC and cyaA) had SOS box sequence to which the LexA protein is bound directly. These results update our current understanding of SOS response in M. xanthus and show that UV induces more genes involved in secondary metabolism and signal transduction in addition to DNA replication and repair; and while the canonical LexA-dependent regulation on SOS response has shrunk, only 5 SOS genes are directly repressed by LexA.

• Mycobiology
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• 제49권6호
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• pp.582-588
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• 2021

The interaction of mating pheromone and pheromone receptor from the B mating-type locus is the first step in the activation of the mushroom mating signal transduction pathway. The B mating-type locus of Lentinula edodes is composed of Bα and Bβ subloci, each of which contains genes for mating pheromone and pheromone receptor. Allelic variations in both subloci generate multiple B mating-types through which L. edodes maintains genetic diversity. In addition to the B mating-type locus, our genomic sequence analysis revealed the presence of a novel chromosomal locus 43.3 kb away from the B mating-type locus, containing genes for a pair of mating pheromones (PHBN1 and PHBN2) and a pheromone receptor (RCBN). The new locus (Bα-N) was homologous to the Bα sublocus, but unlike the multiallelic Bα sublocus, it was highly conserved across the wild and cultivated strains. The interactions of RcbN with various mating pheromones from the B and Bα-N mating-type loci were investigated using yeast model that replaced endogenous yeast mating pheromone receptor STE2 with RCBN. The yeast mating signal transduction pathway was only activated in the presence of PHBN1 or PHBN2 in the RcbN producing yeast, indicating that RcbN interacts with self-pheromones (PHBN1 and PHBN2), not with pheromones from the B mating-type locus. The biological function of the Bα-N locus was suggested to control the expression of A mating-type genes, as evidenced by the increased expression of two A-genes HD1 and HD2 upon the treatment of synthetic PHBN1 and PHBN2 peptides to the monokaryotic strain of L. edodes.

• 생명과학회지
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• 제26권10호
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• pp.1121-1129
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• 2016

밀의 고분자 글루테닌 서브유닛[high molecular-weight glutenin subunit (HMW-GS)]은 밀가루의 성질을 결정하는데 가장 중요한 요소이며 가공적성을 나타내는데 중요한 역할을 수행한다. 우리는 Agrobacterium 동시 형질전환법을 이용하여 한국 밀 품종인 ‘조경’으로부터 밀 HMW-GS을 암호화하는 TaGlu-Ax1 유전자를 가지는 marker-free 형질전환 벼를 생산하였다. TaGlu-Ax1 유전자의 종자 특이적 발현을 위하여 밀에서 존재하는 TaGlu-Bx7 유전자의 자체 프로모터를 벡터 내에 삽입하였다. 동시 접종을 위해서 오직 TaGlu-Ax1 유전자와 hygromycin phosphotransferase II (HPTII) 저항성 유전자만으로 구성된 두 종류의 발현 카세트를 독립적으로 Agrobacterium EHA105에 도입하였고, TaGlu-Ax1와 HPTII가 도입된 각각의 EHA105 Agrobacterium을 3:1 비율로 혼합하여 벼 캘러스에 접종하였다. 210개의 HPTII 저항성 형질전환체 중에서 벼 게놈에 TaGlu-Ax1과 HPTII가 모두 삽입된 20개의 형질전환 라인을 획득하였다. TaGlu-Ax1와 HPTII가 벼 게놈에 도입된 것을 Southern blot을 통해서 다시 확인하였다. 형질전환 벼 T₁ 세대의 종자에서 밀 TaGlu-Ax1 유전자가 전사와 번역되어 오직 TaGlu-Ax1만을 가지는 marker-free 식물체를 T₁세대에서 성공적으로 선발할 수 있었다. TaGlu-Ax1 유전자가 발현되는 marker-free 형질전환 식물체는 야생형(wild type)과의 표현형 차이는 없었다. 형질전환 벼의 쌀가루의 제빵적성을 비교하였을 때 TaGlu-Ax1 유전자만이 발현되어서는 제빵적성이 더 나아지지 않았다. 그러므로 더 많은 밀 고분자 및 저분자 글루테닌, 글리아딘의 유전자의 집적과 조합이 쌀가루 가공적성을 증진시키는데 필요하다. 결론적으로 TaGlu-Ax1 marker-free 형질전환 벼는 쌀가루 가공적성을 증진시키는데 좋은 재료로 사용될 것이다.