• Journal of Microbiology and Biotechnology
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• v.28 no.2
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• pp.227-235
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• 2018

Foodborne illness represents a major threat to public health and is frequently attributed to pathogenic microorganisms on fresh produce. Recurrent outbreaks often come from vegetables that are grown close to or within the ground. Therefore, the first step to understanding the public health risk of microorganisms on fresh vegetables is to identify and describe microbial communities. We investigated the phyllospheres on Chinese cabbage (Brassica rapa subsp. pekinensis, N = 54). 16S rRNA gene amplicon sequencing targeting the V5-V6 region of 16S rRNA genes was conducted by employing the Illumina MiSeq system. Sequence quality was assessed, and phylogenetic assessments were performed using the RDP classifier implemented in QIIME with a bootstrap cutoff of 80%. Principal coordinate analysis was performed using a weighted Fast UniFrac matrix. The average number of sequence reads generated per sample was 34,584. At the phylum level, bacterial communities were composed primarily of Proteobacteria and Bacteroidetes. The most abundant genera on Chinese cabbages were Chryseobacterium, Aurantimonadaceae_g, Sphingomonas, and Pseudomonas. Diverse potential pathogens, such as Pantoea, Erwinia, Klebsiella, Yersinia, Bacillus, Staphylococcus, Salmonella, and Clostridium were also detected from the samples. Although further epidemiological studies will be required to determine whether the detected potential pathogens are associated with foodborne illness, our results imply that a metagenomic approach can be used to detect pathogenic bacteria on fresh vegetables.

• Journal of Microbiology and Biotechnology
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• v.33 no.1
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• pp.51-60
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• 2023

The foodborne illness is the important public health concerns, and the livestock feces are known to be one of the major reservoirs of foodborne pathogens. Also, it was reported that 45.5% of foodborne illness outbreaks have been associated with the animal products contaminated with the livestock feces. In addition, it has been known that the persistence of a pathogens depends on many potential virulent factors including the various virulent genes. Therefore, the first step to understanding the public health risk of livestock feces is to identify and describe microbial communities and potential virulent genes that contribute to bacterial pathogenicity. We used the whole metagenome shotgun sequencing to evaluate the prevalence of foodborne pathogens and to characterize the virulence associated genes in pig and chicken feces. Our data showed that the relative abundance of potential foodborne pathogens, such as Bacillus cereus was higher in chickens than pigs at the species level while the relative abundance of foodborne pathogens including Campylobacter coli was only detected in pigs. Also, the microbial functional characteristics of livestock feces revealed that the gene families related to "Biofilm formation and quorum sensing" were highly enriched in pigs than chicken. Moreover, the variety of gene families associated with "Resistance to antibiotics and toxic compounds" were detected in both animals. These results will help us to prepare the scientific action plans to improve awareness and understanding of the public health risks of livestock feces.

• Journal of Microbiology and Biotechnology
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• v.27 no.10
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• pp.1753-1762
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• 2017

Sea cucumber (Apostichopus japonicus) is a popular seafood source in Asia, including South Korea, and its consumption has recently increased with recognition of its medicinal properties. However, because raw sea cucumber contains various microbes, its ingestion can cause foodborne illness. Therefore, analysis of the microbiota in the whole body of sea cucumber can extend our understanding of foodborne illness caused by microorganisms and help to better manage products. We collected 40 sea cucumbers from four different sites in August and November, which are known as the maximum production areas in Korea. The microbiota was analyzed by an Illumina MiSeq system, and bacterial amounts were quantified by real-time PCR. The diversity and bacterial amounts in sea cucumber were higher in August than in November. Alpha-, Beta-, and Gammaproteobacteria were common dominant classes in all samples. However, the microbiota composition differed according to sampling time and site. Staphylococcus warneri and Propionibacterium acnes were commonly detected potential pathogens in August and November samples, respectively. The effect of experimental Vibrio parahaemolyticus infection on the indigenous microbiota of sea cucumber was analyzed at different temperatures, revealing clear alterations of Psychrobacter and Moraxella; thus, these shifts can be used as indicators for monitoring infection of sea cucumber. Although further studies are needed to clarify and understand the virulence and mechanisms of the identified pathogens of sea cucumber, our study provides a valuable reference for determining the potential of foodborne illness caused by sea cucumber ingestion and to develop monitoring strategies of products using microbiota information.

• Korean Journal of Food Science and Technology
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• v.37 no.4
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• pp.611-617
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• 2005

Dispersed repetitive DNA elements in genomes of microorganisms differ among and within species. Because distances between repetitive sequences vary depending on bacterial strains, genomic fingerprinting with interspersed repetitive sequence-based probes can be used to distinguish unrelated organisms. Among well-known bacterial repetitive sequences, Repetitive Extragenic Palindromic (REP) sequence has been used to identify environmental bacterial species and strains. We applied REP-PCR to detect and differentiate four major Gram-negative food-borne bacterial pathogens, E. coli, Salmonella, Shigella, and Vibrio. Target DNA fragments of these pathogens were amplified by REP-PCR method. PCR-generated DNA fragments were separated on 1.5% agarose gel. Dendrograms for PCR products of each strain were constructed using photo-documentation system. REP-PCR reactions with primer pairs REP1R-I and REP2-I revealed distinct REP-PCR-derived genomic fingerprinting patterns from E. coli, Salmonella, Shigella, and Vibrio. REP-PCR method provided clear distinctions among different bacterial species containing REP-repetitive elements and can be widely used for typing food-borne Gram-negative strains. Results showed established REP-PCR reaction conditions and generated dendrograms could be used with other supplementary genotyping or phenotyping methods to identify isolates from outbreak and to estimate relative degrees of genetic similarities among isolates from different outbreaks to determine whether they are clonally related.

• Journal of Microbiology and Biotechnology
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• v.28 no.8
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• pp.1318-1331
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• 2018

Lettuce (Lactuca sativa L.) is a major ingredient used in many food recipes in South Korea. Lettuce samples were collected during their maximum production period between April and July in order to investigate the microbiota of lettuce during different seasons. 16S rRNA gene-based sequencing was conducted using Illumina MiSeq, and real-time PCR was performed for quantification. The number of total bacterial was greater in lettuce collected in July than in that collected in April, albeit with reduced diversity. The bacterial compositions varied according to the site and season of sample collection. Potential pathogenic species such as Bacillus spp., Enterococcus casseliflavus, Klebsiella pneumoniae, and Pseudomonas aeruginosa showed season-specific differences. Results of the network co-occurrence analysis with core genera correlations showed characteristics of bacterial species in lettuce, and provided clues regarding the role of different microbes, including potential pathogens, in this microbiota. Although further studies are needed to determine the specific effects of regional and seasonal characteristics on the lettuce microbiota, our results imply that the 16S rRNA gene-based sequencing approach can be used to detect pathogenic bacteria in lettuce.

• Food Science of Animal Resources
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• v.25 no.4
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• pp.513-520
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• 2005

The purpose of this study is to measure the total bacterial count, the number of foodborne pathogens and the change of PH by the addition of grapefruit seed extract (GFSE) in kanjang Paste and kochujang paste, respectively. The change of inoculated foodborne pathogens such as Salmonella enteritidis IFO 3313, Staphlococcus aureus IFO 12732, Listeria monocytogenes ATCC 19115, Escherichia coli O157:H7 ATCC 43894 in kanjang Paste and kochujang paste were measured for 14 days at the storage temperatures of $4^{\circ}\;and\;20^{\circ}C$. In kanjang paste, the changes of pH were not showed between the control and the addition of GFSE at $4^{\circ}C$. However, pH were decreased about 2 pome in the control and the addition of GFSE 250 ppm at $20^{\circ}C$. In the addition of GFSE 500 ppm, pH decreased about 1.2 points at $20^{\circ}C$. In the growth-inhibitory effect on foodborne pathogens, when comparing with the con01 and the addition of GFSE 250 ppm and 500 ppm the addition of GFSE was decreased more than the control in kanjang paste at storage temperatures of $4^{\circ}C\;and\;20^{\circ}C$. Otherwise, there were no differences of the number of foodborne Pathogens in kochujang paste as additions. But in kochujang paste stored at storage temperatiues of $4^{\circ}C\;and\;20^{\circ}C$ there were differences of the number of foodborne pathogens. When kochujang paste stored $20^{\circ}C$ at least for 10 days, tested all foodborne Pathogens were not detected.

• Food Science of Animal Resources
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• v.36 no.4
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• pp.463-468
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• 2016

Contamination by foodborne pathogens and mycotoxins was examined in 475 eggs and 20 feed samples collected from three egg layer farms, three egg-processing units, and five retail markets in Korea. Microbial contamination with Salmonella species, Escherichia coli, and Arcobacter species was examined by bacterial culture and multiplex polymerase chain reaction (PCR). The contamination levels of aflatoxins, ochratoxins, and zearalenone in eggs and chicken feeds were simultaneously analyzed with high-performance liquid chromatography coupled with fluorescence detection after the post-derivatization. While E. coli was isolated from 9.1% of eggs, Salmonella species were not isolated. Arcobacter species were detected in 0.8% of eggs collected from egg layers by PCR only. While aflatoxins, ochratoxins, and zearalenone were found in 100%, 100%, and 85% of chicken feeds, their contamination levels were below the maximum acceptable levels (1.86, 2.24, and 147.53 μg/kg, respectively). However, no eggs were contaminated with aflatoxins, ochratoxins, or zearalenone. Therefore, the risk of contamination by mycotoxins and microbes in eggs and chicken feeds is considered negligible and unlikely to pose a threat to human health.

• Food Engineering Progress
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• v.21 no.3
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• pp.191-200
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• 2017

There have been great efforts to develop a rapid and sensitive detection method to monitor the presence of pathogenic bacteria in food. While a number of methods have been reported for bacterial detection with a detection limit to a single digit, most of them are suitable only for the bacteria in pure culture or buffered solution. On the other hand, foods are composed of highly complicated matrices containing carbohydrate, fat, protein, fibers, and many other components whose composition varies from one food to the other. Furthermore, many components in food interfere with the downstream detection process, which significantly affect the sensitivity and selectivity of the detection. Therefore, isolating and concentrating the target pathogenic bacteria from food matrices are of importance to enhance the detection power of the system. The present review provides an introduction to the representative sample preparation strategies to isolate target pathogenic bacteria from food sample. We further describe the nucleic acid-based detection methods, such as PCR, real-time PCR, NASBA, RCA, LCR, and LAMP. Nucleic acid-based methods are by far the most sensitive and effective for the detection of a low number of target pathogens whose performance is greatly improved by combining with the sample preparation methods.

• Journal of Microbiology and Biotechnology
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• v.32 no.8
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• pp.1003-1010
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• 2022

The purpose of this study was to examine the phytochemical compounds and antibacterial activity of Coffea robusta leaf extract (RLE). The results indicated that chlorogenic acid (CGA) is a major component of RLE. The minimum inhibitory concentrations (MICs) of RLE against Staphylococcus aureus, Bacillus subtilis, Escherichia coli, and Salmonella Typhimurium were 6.25, 12.5, 12.5, and 12.5 mg/ml, respectively. RLE effectively damages the bacterial cell membrane integrity, as indicated by the high amounts of proteins and nucleic acids released from the bacteria, and disrupts bacterial cell membrane potential and permeability, as revealed via fluorescence analysis. Cytotoxicity testing showed that RLE is slightly toxic toward HepG2 cells at high concentration but exhibited no toxicity toward Caco2 cells. The results from the present study suggest that RLE has excellent potential applicability as an antimicrobial in the food industry.

• Journal of Microbiology and Biotechnology
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• v.33 no.1
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• pp.83-95
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• 2023

These days, bacterial detection methods have some limitations in sensitivity, specificity, and multiple detection. To overcome these, novel detection and identification method is necessary to be developed. Recently, NGS panel method has been suggested to screen, detect, and even identify specific foodborne pathogens in one reaction. In this study, new NGS panel primer sets were developed to target 13 specific virulence factor genes from five types of pathogenic Escherichia coli, Listeria monocytogenes, and Salmonella enterica serovar Typhimurium, respectively. Evaluation of the primer sets using singleplex PCR, crosscheck PCR and multiplex PCR revealed high specificity and selectivity without interference of primers or genomic DNAs. Subsequent NGS panel analysis with six artificially contaminated food samples using those primer sets showed that all target genes were multi-detected in one reaction at 10⁸-10⁵ CFU of target strains. However, a few false-positive results were shown at 10⁶-10⁵ CFU. To validate this NGS panel analysis, three sets of qPCR analyses were independently performed with the same contaminated food samples, showing the similar specificity and selectivity for detection and identification. While this NGS panel still has some issues for detection and identification of specific foodborne pathogens, it has much more advantages, especially multiple detection and identification in one reaction, and it could be improved by further optimized NGS panel primer sets and even by application of a new real-time NGS sequencing technology. Therefore, this study suggests the efficiency and usability of NGS panel for rapid determination of origin strain in various foodborne outbreaks in one reaction.