• Journal of Microbiology and Biotechnology
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• v.31 no.12
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• pp.1701-1708
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• 2021

Food safety is the most important global health issue due to foodborne pathogens after consumption of contaminated food. Foodborne bacteria such as Escherichia coli, Salmonella enterica, Staphylococcus aureus, Campylobacter spp., Bacillus cereus, Vibrio spp., Yersinia enterocolitica and Clostridium perfringens are leading causes of the majority of foodborne illnesses and deaths. These foodborne pathogens often come from the livestock feces, thus, we analyzed fecal microbial communities of three different livestock species to investigate the prevalence of foodborne pathogens in livestock feces using metagenomics analysis. Our data showed that alpha diversities of microbial communities were different according to livestock species. The microbial diversity of cattle feces was higher than that of chicken or pig feces. Moreover, microbial communities were significantly different among these three livestock species (cattle, chicken, and pig). At the genus level, Staphylococcus and Clostridium were found in all livestock feces, with chicken feces having higher relative abundances of Staphylococcus and Clostridium than cattle and pig feces. Genera Bacillus, Campylobacter, and Vibrio were detected in cattle feces. Chicken samples contained Bacillus, Listeria, and Salmonella with low relative abundance. Other genera such as Corynebacterium, Streptococcus, Neisseria, Helicobacter, Enterobacter, Klebsiella, and Pseudomonas known to be opportunistic pathogens were also detected in cattle, chicken, and pig feces. Results of this study might be useful for controlling the spread of foodborne pathogens in farm environments known to provide natural sources of these microorganisms.

• Journal of Microbiology and Biotechnology
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• v.31 no.11
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• pp.1552-1558
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• 2021

The diverse microbial communities in kimchi are dependent on fermentation period and temperature. Here, we investigated the effect of enterotoxigenic Escherichia coli (ETEC) during the fermentation of kimchi at two temperatures using high-throughput sequencing. There were no differences in pH between the control group, samples not inoculated with ETEC, and the ETEC group, samples inoculated with ETEC MFDS 1009477. The pH of the two groups, which were fermented at 10 and 25℃, decreased rapidly at the beginning of fermentation and then reached pH 3.96 and pH 3.62. In both groups, the genera Lactobacillus, Leuconostoc, and Weissella were predominant. Our result suggests that microbial communities during kimchi fermentation may be affected by the fermentation parameters, such as temperature and period, and not enterotoxigenic E. coli (ETEC).

• Journal of Veterinary Science
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• v.23 no.3
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• pp.41.1-41.15
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• 2022

Background: Proliferative enteritis caused by Lawsonia intracellularis undermines the economic stability of the swine industry worldwide. The development of cost-effective animal models to study the pathophysiology of the disease will help develop strategies to counter this bacterium. Objectives: This study focused on establishing a model of gastrointestinal (GI) infection of L. intracellularis in C57BL/6 mice to evaluate the disease progression and lesions of proliferative enteropathy (PE) in murine GI tissue. Methods: We assessed the murine mucosal and cell-mediated immune responses generated in response to inoculation with L. intracellularis. Results: The mice developed characteristic lesions of the disease and shed L. intracellularis in the feces following oral inoculation with 5 × 10⁷ bacteria. An increase in L. intracellularis 16s rRNA and groEL copies in the intestine of infected mice indicated intestinal dissemination of the bacteria. The C57BL/6 mice appeared capable of modulating humoral and cell-mediated immune responses to L. intracellularis infection. Notably, the expression of genes for the vitamin B12 receptor and for secreted and membrane-bound mucins were downregulated in L. intracellularis -infected mice. Furthermore, L. intracellularis colonization of the mouse intestine was confirmed by the immunohistochemistry and western blot analyses. Conclusions: This is the first study demonstrating the contributions of bacterial chaperonin and host nutrient genes to PE using an immunocompetent mouse model. This mouse infection model may serve as a platform from which to study L. intracellularis infection and develop potential vaccination and therapeutic strategies to treat PE.

• Research in Plant Disease
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• v.28 no.3
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• pp.152-161
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• 2022

Several types of chemical bactericides have been used to control fire blight. However, their excessive usage leads to environmental deterioration. Therefore, several researchers have analyzed antagonistic microorganisms as promising, effective, and safe biological control agents (BCAs). The primary aim of this study was to screen for potential antagonistic bacteria that suppress Erwinia amylovora. Among the 45 isolates studied, 5 strains showed the largest inhibition zone against E. amylovora. 16S rRNA gene sequencing identified them as Bacillus amyloliquefaciens (KPB 15), B. stratosphericus (KPB 21), B. altitudinis (KPB 25), B. safensis (KPB 31), and B. subtilis (KPB 39). KPB 25 and 31 reduced the lesion size of fire blight by 50% in immature apple fruits, and did not show antagonism against each other. Therefore, KPB 25 and 31 were selected to develop an antagonistic mixture against fire blight. Although the mixture with KPB 25 and 31 showed a slightly increased ability to reduce lesion size on immature fruits, they did not exhibit a synergistic effect in reducing E. amylovora population compared to each strain alone. Nevertheless, we have identified these two strains as useful and novel BCAs against fire blight with additional benefits safety and potential in developing a mixture without loss of their activity, owing to the absence of antagonism against each other.

• Journal of Microbiology and Biotechnology
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• v.33 no.2
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• pp.195-202
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• 2023

Protease is a widely used enzyme particularly in the detergent industry. In this research, we aimed to isolate alkaline protease-producing bacteria for characterization as a laundry detergent additive. The screening of alkaline protease production was investigated on basal medium agar plus 1% skim milk at pH 11, with incubation at 30℃. The highest alkaline protease-producing bacterium was 6BS15-4 strain, identified as Bacillus gibsonii by 16S rRNA gene sequencing. While the optimum pH was 12.0, the strain was stable at pH range 7.0-12.0 when incubated at 45℃ for 60 min. The alkaline protease produced by B. gibsonii 6BS15-4 using dairy effluent was characterized. The optimum temperature was 60℃ and the enzyme was stable at 55℃ when incubated at pH 11.0 for 60 min. Metal ions K⁺, Mg²⁺, Cu²⁺, Na⁺, and Zn²⁺ exhibited a slightly stimulatory effect on enzyme activity. The enzyme retained over 80% of its activity in the presence of Ca²⁺, Ba²⁺, and Mn²⁺. Thiol reagent and ethylenediaminetetraacetic acid did not inhibit the enzyme activity, whereas phenylmethylsulfonyl fluoride significantly inhibited the protease activity. The alkaline protease from B. gibsonii 6BS15-4 demonstrated efficiency in blood stain removal and could therefore be used as a detergent additive, with potential for various other industrial applications.

• Journal of Microbiology and Biotechnology
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• v.33 no.1
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• pp.61-74
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• 2023

The global increase in multidrug-resistant (MDR) bacteria has inspired researchers to develop new strategies to overcome this problem. In this study, 23 morphologically different, soil-isolated actinomycete cultures were screened for their antibacterial ability against MDR isolates of ESKAPE pathogens. Among them, isolate BOGE18 exhibited a broad antibacterial spectrum, so it was selected and identified based on cultural, morphological, physiological, and biochemical characteristics. Chemotaxonomic analysis was also performed together with nucleotide sequencing of the 16S rRNA gene, which showed this strain to have identity with Streptomyces lienomycini. The ethyl acetate extract of the cell-free filtrate (CFF) of strain BOGE18 was evaluated for its antibacterial spectrum, and the minimum inhibitory concentration (MIC) ranged from 62.5 to 250 ㎍/ml. The recorded results from the in vitro anti-biofilm microtiter assay and confocal laser scanning microscopy (CLSM) of sub-MIC concentrations revealed a significant reduction in biofilm formation in a concentration-dependent manner. The extract also displayed significant scavenging activity, reaching 91.61 ± 4.1% and 85.06 ± 3.14% of 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), respectively. A promising cytotoxic ability against breast (MCF-7) and hepatocellular (HePG2) cancer cell lines was obtained from the extract with IC₅₀ values of 47.15 ± 13.10 and 122.69 ± 9.12 ㎍/ml, respectively. Moreover, based on gas chromatography-mass spectrometry (GC-MS) analysis, nine known compounds were detected in the BOGE18 extract, suggesting their contribution to the multitude of biological activities recorded in this study. Overall, Streptomyces lienomycini BOGE18-derived extract is a good candidate for use in a natural combating strategy to prevent bacterial infection, especially by MDR pathogens.

• Journal of Environmental Science International
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• v.31 no.3
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• pp.265-274
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• 2022

Lipoxygenase is an enzyme, mainly produced by plants, capable of converting unsaturated fatty acids to fatty acids. It has vast application potential in the food, pharmaceutical and agricultural industries. The aim of this study was to isolate novel lipoxygenase-producing bacteria from the environment and to investigate the lipoxygenase enzymatic properties for industrial production. The strain, NC1, isolated from cultivation soils, was identified as Bacillus subtilis based on the phenotypic characteristics and 16S rRNA gene sequencing. This strain formed a pink color around the colony when cultured on indamine dye formation plates. The production of lipoxygenase by B. subtilis NC1 was influenced by the composition of the medium and linoleic acid concentrations. The optimum temperature and pH for lipoxygenase activity was determined to be 40 ℃ and pH 6, respectively. The enzyme showed relatively high stability at temperatures ranging from 20-50 ℃ and acid-neutral regions. In addition, the lipoxygenase produced by B. subtilis NC1 was able to degrade commercially available oils including sunflower seed oil and Perilla oil. In this study, a useful indigenous bacterium was isolated, and the fundamental physicochemical data of bacterial lipoxygenase giving it industrial potential are presented.

• Journal of Microbiology and Biotechnology
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• v.33 no.5
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• pp.607-620
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• 2023

The biocontrol approach using beneficial microorganisms to control crop diseases is becoming an essential alternative to chemical fungicides. Therefore, new and efficient biocontrol agents (BCA) are needed. In this study, a rhizospheric actinomycete isolate showed unique and promising antagonistic activity against three of the most common phytopathogenic fungi, Fusarium oxysporum MH105, Rhizoctonia solani To18, and Alternaria brassicicola CBS107. Identification of the antagonistic strain, which was performed according to spore morphology and cell wall chemotype, suggested that it belongs to the Nocardiopsaceae. Furthermore, cultural, physiological, and biochemical characteristics, together with phylogenetic analysis of the 16S rRNA gene (OP869859.1), indicated the identity of this strain to Nocardiopsis alba. The cell-free filtrate (CFF) of the strain was evaluated for its antifungal potency, and the resultant inhibition zone diameters ranged from 17.0 ± 0.92 to 19.5 ± 0.28 mm for the tested fungal species. Additionally, the CFF was evaluated in vitro to control Fusarium wilt disease in Vicia faba using the spraying method under greenhouse conditions, and the results showed marked differences in virulence between the control and treatment plants, indicating the biocontrol efficacy of this actinomycete. A promising plant-growth promoting (PGP) ability in seed germination and seedling growth of V. faba was also recorded in vitro for the CFF, which displayed PGP traits of phosphate solubilization (48 mg/100 ml) as well as production of indole acetic acid (34 ㎍/ml) and ammonia (20 ㎍/ml). This study provided scientific validation that the new rhizobacterium Nocardiopsis alba strain BH35 could be further utilized in bioformulation and possesses biocontrol and plant growth-promoting capabilities.

• Animal Bioscience
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• v.36 no.9
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• pp.1453-1464
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• 2023

Objective: This study investigated the changes in bacterial communities within decomposing swine microcosms, comparing soil with or without intact microbial communities, and under aerobic and anaerobic conditions. Methods: The experimental microcosms consisted of four conditions: UA, unsterilized soil-aerobic condition; SA, sterilized soil-aerobic condition; UAn, unsterilized soil-anaerobic condition; and San, sterilized soil-anaerobic condition. The microcosms were prepared by mixing 112.5 g of soil and 37.5 g of ground carcass, which were then placed in sterile containers. The carcass-soil mixture was sampled at day 0, 5, 10, 30, and 60 of decomposition, and the bacterial communities that formed during carcass decomposition were assessed using Illumina MiSeq sequencing of the 16S rRNA gene. Results: A total of 1,687 amplicon sequence variants representing 22 phyla and 805 genera were identified in the microcosms. The Chao1 and Shannon diversity indices varied in between microcosms at each period (p<0.05). Metagenomic analysis showed variation in the taxa composition across the burial microcosms during decomposition, with Firmicutes being the dominant phylum, followed by Proteobacteria. At the genus level, Bacillus and Clostridium were the main genera within Firmicutes. Functional prediction revealed that the most abundant Kyoto encyclopedia of genes and genomes metabolic functions were carbohydrate and amino acid metabolisms. Conclusion: This study demonstrated a higher bacteria diversity in UA and UAn microcosms than in SA and SAn microcosms. In addition, the taxonomic composition of the microbial community also exhibited changes, highlighting the impact of soil sterilization and oxygen on carcass decomposition. Furthermore, this study provided insights into the microbial communities associated with decomposing swine carcasses in microcosm.

• Microbiology and Biotechnology Letters
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• v.51 no.2
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• pp.191-202
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• 2023

Elevated serum cholesterol is a main risk factor for heart disorders. Most probiotic products administered to lower cholesterol are dairy products which are not suitable for lactose-intolerant individuals. In this study, we assessed the cholesterol-lowering efficacy of LAB isolated from traditionally fermented drinks in diet-induced rats and determine their efficacy in the production of non-dairy, probiotic formulations using papaya juice. LAB were isolated from palm wine and corn beer on MRS agar using a pour-plate technique. Identification was carried out using 16S rRNA gene sequencing. A hypercholesterolemia model in which diet-induced Wistar albino rats were assigned into four groups was established. Oral gavage was carried out for 30 days. On the 31^st day, the rats were dissected and the serum lipid profile was analyzed using biochemical kits. A 10⁶ cfu/ml of a 24-h-old culture of selected lactobacilli was used to inoculate papaya juice and incubated at 37℃. Microbial and chemical changes were assessed during papaya fermentation and after four weeks of cold storage. Two selected isolates (Pw1 and Cb4) had in vitro cholesterol reduction of > 80%. These two isolates lowered lipid profile (triglyceride, total cholesterol, LDL-c) significantly, and increased HDL-c levels (p < 0.5) in the rat sera. Phylogenetic analysis showed that Pw1 was 98.86% similar to Limosilactobacillus fermentum, while Cb4 was 99.54% similar to Enteroccocus faecium. Both strains fermented papaya juice with cell viability reaching 8.92 × 10⁸ cfu/ml and 25.3 × 10⁸ cfu/ml respectively, and were still viable after 4 weeks of cold storage.