• Journal of Ginseng Research
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• v.45 no.1
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• pp.75-85
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• 2021

Background: Invasive infections due to foodborne pathogens, including Salmonella enterica serovar Typhimurium, are prevalent and life-threatening. This study aimed to evaluate the effects of ginsenoside Rg3 (Rg3) on the adhesion, invasion, and intracellular survival of S. Typhimurium. Methods: The impacts of Rg3 on bacterial growth and host cell viability were determined using the time kill and the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide assays, respectively. Gentamicin assay and confocal microscopic examination were undertaken to determine the effects of Rg3 on the adhesive and invasive abilities of S. Typhimurium to Caco-2 and RAW 264.7 cells. Quantitative reverse transcription polymerase chain reaction was performed to assess the expression of genes correlated with the adhesion, invasion, and virulence of S. Typhimurium. Results: Subinhibitory concentrations of Rg3 significantly reduced (p < 0.05) the adhesion, invasion, and intracellular survival of S. Typhimurium. Rg3 considerably reduced (p < 0.05) the bacterial motility as well as the release of nitrite from infected macrophages in a concentration-dependent manner. The expression of genes related to the adhesion, invasion, quorum sensing, and virulence of S. Typhimurium including cheY, hilA, OmpD, PrgK, rsgE, SdiA, and SipB was significantly reduced after Rg3 treatment. Besides, the compound downregulated rac-1 and Cdc-42 that are essential for actin remodeling and membrane ruffling, thereby facilitating Salmonella entry into host cells. This report is the first to describe the effects of Rg3 on "trigger" entry mechanism and intracellular survival S. Typhimurium. Conclusion: Rg3 could be considered as a supplement agent to prevent S. Typhimurium infection.

• Journal of Microbiology and Biotechnology
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• v.31 no.12
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• pp.1672-1683
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• 2021

Vibrio parahaemolyticus is recognized as one of the most important foodborne pathogens responsible for gastroenteritis in humans. The bla_CARB-17 gene is an intrinsic β-lactamase gene and a novel species-specific genetic marker of V. parahaemolyticus. In this study, a loop-mediated isothermal amplification (LAMP) assay combined with a lateral flow dipstick (LFD) was developed targeting this bla_CARB-17 gene. The specificity of LAMP-LFD was ascertained by detecting V. parahaemolyticus ATCC 17802 and seven other non-V. parahaemolyticus strains. Finally, the practicability of LAMP-LFD was confirmed by detection with V. parahaemolyticus-contaminated samples and natural food samples. The results showed that the optimized reaction parameters of LAMP are as follows: 2.4 mmol/l Mg²⁺, 0.96 mmol/l dNTPs, 4.8 U Bst DNA polymerase, and an 8:1 ratio of inner primer to outer primer, at 63℃ for 40 min. The optimized reaction time of the LFD assay is 60 min. Cross-reactivity analysis with the seven non-V. parahaemolyticus strains showed that LAMP-LFD was exclusively specific for V. parahaemolyticus. The detection limit of LAMP-LFD for V. parahaemolyticus genomic DNA was 2.1 × 10^-4 ng/μl, corresponding to 630 fg/reaction and displaying a sensitivity that is 100-fold higher than that of conventional PCR. LAMP-LFD in a spiking study revealed a detection limit of approximately 6 CFU/ml, which was similar with conventional PCR. The developed LAMP-LFD specifically identified the 10 V. parahaemolyticus isolates from 30 seafood samples, suggesting that this LAMP-LFD may be a suitable diagnostic method for detecting V. parahaemolyticus in aquatic foods.

• Parasites, Hosts and Diseases
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• v.59 no.6
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• pp.573-583
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• 2021

Toxoplasma gondii, an intracellular protozoan parasite that infects one-third of the world's population, has been reported to hijack host cell apoptotic machinery and promote either an anti- or proapoptotic program depending on the parasite virulence and load and the host cell type. However, little is known about the regulation of human FHs 74 small intestinal epithelial cell viability in response to T. gondii infection. Here we show that T. gondii RH strain tachyzoite infection or ESP treatment of FHs 74 Int cells induced apoptosis, mitochondrial dysfunction and ER stress in host cells. Pretreatment with 4-PBA inhibited the expression or activation of key molecules involved in ER stress. In addition, both T. gondii and ESP challenge-induced mitochondrial dysfunction and cell death were dramatically suppressed in 4-PBA pretreated cells. Our study indicates that T. gondii infection induced ER stress in FHs 74 Int cells, which induced mitochondrial dysfunction followed by apoptosis. This may constitute a potential molecular mechanism responsible for the foodborne parasitic disease caused by T. gondii.

• Journal of Microbiology and Biotechnology
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• v.32 no.10
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• pp.1307-1314
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• 2022

In this study, we sought to investigate the various characteristics of Salmonella spp. isolated from raw chicken meats available in Korean markets. The data collected, such as food source of isolation, sampling information, serotype, virulence, and genetic profile including sequence type, were registered in the database for further comparative analysis of the strains isolated from the traceback investigation samples. To characterize serotype, virulence and gene sequences, we examined 113 domestically distributed chicken meat samples for contamination with Salmonella spp. Phylogenetic analysis was conducted on 24 strains (21.2%) of Salmonella isolated from 113 commercially available chicken meats and by-products, using pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). Serotyping of the isolated Salmonella spp. revealed S. Enteritidis in 11 strains (45.8%), S. Virchow in 6 strains (25%), S. Montevideo in 2 strains (8.3%), S. Bsilla in 2 strains (8.3%), S. Bareilly in 1 strain (4.2%), S. Dessau in 1 strain (4.2%), and S. Albany in 1 strain (4.2%). The genetic correlation indicated that 24 isolated strains were classified into 18 clusters with a genetic similarity of 64.4-100% between them. Eleven isolated S. Enteritidis strains were classified into 9 genotypes with a sequence identity of 74.4%, whereas the most distantly related S. Virchow was divided into five genotypes with 85.9% identity. Here, the MLST analysis indicated that the major Sequence Type (ST) of the Salmonella spp. isolated from domestic chicken sold in Chungcheong Province belongs to the ST 11 and 16, which differs from the genotype of Salmonella isolated from imported chicken. The differential sequence characteristics can be a genetic marker for identifying causative bacteria for epidemiological investigations of food poisoning.

• Journal of Veterinary Science
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• v.21 no.6
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• pp.88.1-88.13
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• 2020

Background: Listeria monocytogenes is a gram-positive bacterium that causes listeriosis mainly in immunocompromised hosts. It can also cause foodborne outbreaks and has the ability to adapt to various environments. Peptide uptake in gram-positive bacteria is enabled by oligopeptide permeases (Opp) in a process that depends on ATP hydrolysis by OppD and F. Previously a putative protein Lmo2193 was predicted to be OppD, but little is known about the role of OppD in major processes of L. monocytogenes, such as growth, virulence, and biofilm formation. Objectives: To determine whether the virulence traits of L. monocytogenes are related to OppD. Methods: In this study, Lmo2193 gene deletion and complementation strains of L. monocytogenes were generated and compared with a wild-type strain for the following: adhesiveness, invasion ability, intracellular survival, proliferation, 50% lethal dose (LD₅₀) to mice, and the amount bacteria in the mouse liver, spleen, and brain. Results: The results showed that virulence of the deletion strain was 1.34 and 0.5 orders of magnitude higher than that of the wild-type and complementation strains, respectively. The function of Lmo2193 was predicted and verified as OppD from the ATPase superfamily. Deletion of lmo2193 affected the normal growth of L. monocytogenes, reduced its virulence in cells and mice, and affected its ability to form biofilms. Conclusions: Deletion of the oligopeptide transporter Lmo2193 decreases the virulence of L. monocytogenes. These effects may be related to OppD's function, which provides a new perspective on the regulation of oligopeptide transporters in L. monocytogenes.

• Journal of Environmental Health Sciences
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• v.48 no.3
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• pp.176-182
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• 2022

Background: The risk of imported infectious diseases has been increasing with the annual rise in the number of international travelers. Objectives: This study aims to analyze the distribution and characteristics of intestinal bacteria isolated in 2019 from residents of Chungcheongnam-do Province with experience of travelling overseas. Methods: Twenty-three former overseas travelers with diarrhea were analyzed to detect viruses and bacteria according to the Manual for Detection of Foodborne Pathogens at Outbreaks. Additionally, antibiotic susceptibility tests and 16s rRNA sequencing were performed. Results: Twenty-five strains of ten pathogens were isolated from 18 samples. Pathogenic E. coli was the most common at 57.7%, followed by Clostridium perfringens (15.4%), Campylobacter spp. (7.7%), and Salmonella spp. (7.7%). The serotype of Salmonella was confirmed as Salmonella Braenderup, II 9,46:g,[m],[s],t:[e,n,x]. Conclusions: It was confirmed that the major enteric bacterial pathogens isolated from overseas travelers in Chungcheongnam-do Province were pathogenic E. coli, as found in other studies. The study on Plesiomonas shigelloides is meaningful in that it is reported as a rare case of infection in Korea. Antibiotic resistance and 16s rRNA analysis were performed, which is expected to provide important basic data for the prevention of traveler's diarrhea.

• Journal of Microbiology and Biotechnology
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• v.32 no.11
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• pp.1462-1470
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• 2022

Natural antimicrobial substances are needed as alternatives to synthetic antimicrobials to protect against foodborne pathogens. In this study, a bacteriocin-producing bacterium, Bacillus subtilis HD15, was isolated from doenjang, a traditional Korean fermented soybean paste. We sequenced the complete genome of B. subtilis HD15. This genome size was 4,173,431 bp with a G + C content of of 43.58%, 4,305 genes, and 4,222 protein-coding genes with predicted functions, including a subtilosin A gene cluster. The bacteriocin was purified by ammonium sulfate precipitation, Diethylaminoethanol-Sepharose chromatography, and Sephacryl gel filtration, with 12.4-fold purification and 26.2% yield, respectively. The purified protein had a molecular weight of 3.6 kDa. The N-terminal amino acid sequence showed the highest similarity to Bacillus subtilis 168 subtilosin A (78%) but only 68% similarity to B. tequilensis subtilosin proteins, indicating that the antimicrobial substance isolated from B. subtilis HD15 is a novel bacteriocin related to subtilosin A. The purified protein from B. subtilis HD15 exhibited high antimicrobial activity against Listeria monocytogenes and Bacillus cereus. It showed stable activity in the range 0-70℃ and pH 2-10 and was completely inhibited by protease, proteinase K, and pronase E treatment, suggesting that it is a proteinaceous substance. These findings support the potential industrial applications of the novel bacteriocin purified from B. subtilis HD15.

• Journal of Microbiology and Biotechnology
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• v.33 no.3
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• pp.329-338
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• 2023

Enterohemorrhagic Escherichia coli (EHEC) is a foodborne pathogen that produces attaching and effacing lesions on the large intestine and causes hemorrhagic colitis. It is primarily transmitted through the consumption of contaminated meat or fresh produce. Similar to other bacterial pathogens, antibiotic resistance is of concern for EHEC. Furthermore, since the production of Shiga toxin by this pathogen is enhanced after antibiotic treatment, alternative agents that control EHEC are necessary. This study aimed to discover alternative treatments that target virulence factors and reduce EHEC toxicity. The locus of enterocyte effacement (LEE) is essential for EHEC attachment to host cells and virulence, and most of the LEE genes are positively regulated by the transcriptional regulator, Ler. GrlA protein, a transcriptional activator of ler, is thus a potential target for virulence inhibitors of EHEC. To identify the GrlA inhibitors, an in vivo high-throughput screening (HTS) system consisting of a GrlA-expressing plasmid and a reporter plasmid was constructed. Since the reporter luminescence gene was fused to the ler promoter, the bioluminescence would decrease if inhibitors affected the GrlA. By screening 8,201 compounds from the Korea Chemical Bank, we identified a novel GrlA inhibitor named Grlactin [3-[(2,4-dichlorophenoxy)methyl]-4-(3-methylbut-2-en-1-yl)-4,5-dihydro-1,2,4-oxadiazol-5-one], which suppresses the expression of LEE genes. Grlactin significantly diminished the adhesion of EHEC strain EDL933 to human epithelial cells without inhibiting bacterial growth. These findings suggest that the developed screening system was effective at identifying GrlA inhibitors, and Grlactin has potential for use as a novel anti-adhesion agent for EHEC while reducing the incidence of resistance.

• Advances in nano research
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• v.14 no.4
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• pp.391-397
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• 2023

This manuscript describes the one-step eco-friendly green fabrication of silver nanoparticles (AgNPs) through the in-situ bio-reduction of an aqueous solution of silver nitrate using Syzygium aromaticum leaf extract. UV-vis spectroscopy shows a characteristic SPR peak around 442 nm. FTIR spectroscopy showed that the AgNPs were capped with bioactive phyto-molecules. TEM images revealed oval and spherical particles with a mean diameter of ~12.6 nm. XRD analysis revealed crystalline and face-cantered cubic AgNPs. The phytosynthesized AgNPs showed broad-spectrum anti-microbial activity against two foodborne pathogenic bacteria, Listeria monocytogenes and Staphylococcus aureus. The AgNPs showed a prominent ability to inhibit biofilms formed by L. monocytogenes and S. aureus in laboratory conditions through a crystal violet assay. The results suggest that the AgNPs could be a novel nanotool to develop effective antimicrobial and anti-biofilm agents in food preservation.