• 제목/요약/키워드: PCR typing

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RAPD Analysis and Antimicrobial Susceptibility of Streptococcus equi subsp. zooepidemicus Isolated from Thoroughbred Horses (더러브렛 말에서 분리한 Streptococcus equi subsp. zooepidemicus의 RAPD 분석 및 약제 감수성)

  • Choi, Seong-Kyoon;Park, Yong-Soo;Cho, Kwang-Hyun;Cho, Gil-Jae
    • Journal of Life Science
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    • v.20 no.5
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    • pp.649-654
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    • 2010
  • A total of 68 samples were collected including vaginal mucosa (n=66) from Jangsu stud farm, an equine aborted fetus (n=1), and uterine contents (n=1) from Jeju island. Seventeen Streptococcus equi subsp. zooepidemicus (S. zooepidemicus) strains isolated from horses in Korea were identified as S. zooepidemicus by biochemical tests and sodA.seeI specific multiplex PCR. All isolated strains were divided into 4 clusters: group 1 (No. 2, 3, 5, 6, 7, 11, 12, 13, 14, 15), group 2 (No. 4, 9), group 3 (No. 10, 16, 17), and group 4 (No. 1, 8) by RAPD typing. In group 3, No. 10 isolate that was isolated from vaginal mucosa was indistinguishable from No. 16 and 17 isolates, which were isolated from the equine uterine contents and the equine aborted fetus, respectively. The results of this study suggest that a limited epidemiological relationship exists between the strains from Jangsu (No. 10) and Jeju (No. 16 and No. 17). All isolates showed a high susceptibility to ampicillin, cefoxitin, ceftiofur, cephalothin, florofenicol, gentamicin, nalidixic acid, oxacillin, penicillin, tiamulin, tylosin and vancomycin in antimicrobial susceptibility tests. These results may provide the basic information needed to establish strategies for the treatment and prevention of reproductive diseases in mares in Korea.

Colistin resistance and plasmid-mediated mcr genes in Escherichia coli and Salmonella isolated from pigs, pig carcass and pork in Thailand, Lao PDR and Cambodia border provinces

  • Pungpian, Chanika;Lee, Scarlett;Trongjit, Suthathip;Sinwat, Nuananong;Angkititrakul, Sunpetch;Prathan, Rangsiya;Srisanga, Songsak;Chuanchuen, Rungtip
    • Journal of Veterinary Science
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    • v.22 no.5
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    • pp.68.1-68.15
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    • 2021
  • Background: Colistin and carbapenem-resistant bacteria have emerged and become a serious public health concern, but their epidemiological data is still limited. Objectives: This study examined colistin and carbapenem resistance in Escherichia coli and Salmonella from pigs, pig carcasses, and pork in Thailand, Lao PDR, and Cambodia border provinces. Methods: The phenotypic and genotypic resistance to colistin and meropenem was determined in E. coli and Salmonella obtained from pigs, pig carcasses, and pork (n = 1,619). A conjugative experiment was performed in all isolates carrying the mcr gene (s) (n = 68). The plasmid replicon type was determined in the isolates carrying a conjugative plasmid with mcr by PCR-based replicon typing (n = 7). The genetic relatedness of mcr-positive Salmonella (n = 11) was investigated by multi-locus sequence typing. Results: Colistin resistance was more common in E. coli (8%) than Salmonella (1%). The highest resistance rate was found in E. coli (17.8%) and Salmonella (1.7%) from Cambodia. Colistin-resistance genes, mcr-1, mcr-3, and mcr-5, were identified, of which mcr-1 and mcr-3 were predominant in E. coli (5.8%) and Salmonella (1.7%), respectively. The mcr-5 gene was observed in E. coli from pork in Cambodia. Two colistin-susceptible pig isolates from Thailand carried both mcr-1 and mcr-3. Seven E. coli and Salmonella isolates contained mcr-1 or mcr-3 associated with the IncF and IncI plasmids. The mcr-positive Salmonella from Thailand and Cambodia were categorized into two clusters with 94%-97% similarity. None of these clusters was meropenem resistant. Conclusions: Colistin-resistant E. coli and Salmonella were distributed in pigs, pig carcasses, and pork in the border areas. Undivided-One Health collaboration is needed to address the issue.

Genotype Frequency of Human Papillomavirus Determined by PCR and DNA Sequencing in Korean Women

  • Lee, Kyung-Ok;Seong, Hye-Soon;Chung, Soo-Jin;Jung, Na-Young;Lee, Hye-Jung;Kim, Kyung-Tae
    • Korean Journal of Clinical Laboratory Science
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    • v.38 no.2
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    • pp.99-104
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    • 2006
  • Human Papilloma viruses (HPVs) are etiological agents for cervical cancer and are classified into low- and high-risk categories. The aim of this study was to determine the frequency of the HPV genotype in the HPV screening test of Korean women using PCR-direct sequencing. Consensus primers of L1 legion were used for the amplification of HPV DNA and the PCR products (450 bps) obtained were analyzed by automatic sequencing. Sequences were compared with those in GenBank by using the BLAST program. Cervical swab samples of 3,978 women (20-73 years) were tested and the average age was 37.6 years. In this study, 1,174 samples were HPV positive out of 3,978 cervical swab samples screened (29.5%) and 136 samples (11.6%) showed a double infection. A total of 1,310 HPV genotypes were analyzed. The HPV positive rate was the lowest in the 20 years group (69.5%) and most of the samples of the > 60 years group were found HPV positive. Among thirty seven different HPV types identified by sequencing, 21 were HPV high risk types and 16 HPV low risk types were 69.8% (914/1,310) and 26.0% (340/1,310), respectively. In HPV high-risk types, 16 (13.21%), was the most frequently found. HPV 53 (9.62%) and 58 (9.24%) were also frequently found. This group was followed by HPV types 70 (5.50%), 33 (4.73%), 66 (4.20%), 18 (4.05%), 52 (4.05%), 31 (3.97%) and 56 (3.51%) in descending order of frequency. Among HPV low-risk types, 62 (4.20%), 6 (3.59%), 81 (3.59%), 84 (3.51%), and 11 (2.6%) were frequently found. In conclusion, PCR-direct sequencing could be used for quick and reliable typing of known and novel HPVs from clinical specimens. This data could be useful for epidemiological study of HPV and it also allows type-specific follow-up of women who have been treated for cervical intraepithelial neoplasia.

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Changes in Acidity and Distributions of the Vancomycin-Resistant Lactic Acid Bacteria in the Kimchi Fermented at Different Temperatures (발효 온도에 따른 김치의 산도 변화와 Vancomycin 내성 젖산균의 분포)

  • 정의숙;김기환;신원철;송광영;윤성식
    • Microbiology and Biotechnology Letters
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    • v.32 no.3
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    • pp.249-255
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    • 2004
  • Chinese cabbage ('Baechu') Kimchi was fermented at the three different temperatures right after it was prepared. Samples were taken everyday for measuring bacterial populations, pH, and titratable acidity through the whole periods of fermentation up to 50 days. pH values and developed acidity were significantly affected by the fermenting temperatures of 4, 10, and $20^{\circ}C$, suggesting that different bacterial flora has been established by the temperatures exposed. The modified MRS agar containing vancomycin (300 $\mu$g/mL) was used for isolating the vancomycin-resistant LAB strains and 127 isolates were finally obtained. Of the LAB isolates, 13 isolates were subjected to the identification experiments based on the biochemical characteristics and the molecular-typing approach, an ITS-PCR, whether they belong to the genus Leuconostoc or not. The data obtained from API 50 CHL kit resulted that six isolates were identified as the members of Leuconostoc and six as Lactobacillus brevis strains except for a single isolate YKI 30-0401, which was not able to be identified because its biochemical traits were not matched to the database of API 50 CHL kit. It was noted that some isolates were distinct in a couple of some biochemical characteristics compared with those of the reference Leuconostoc species. To overcome the limitations experienced in the commercial identification products above, an ITS-PCR experiment was also conducted for the isolates, resulting that eight isolates belong to Leu. mesenteroides ssp. mesenteroides or dextranicum with a single band of 564 bp, and four to L. brevis strains. The ITS-PCR profiles clearly differentiated the closely-related LAB isolates for which same results were obtained by the biochemical method. This molecular approach, however, failed to produce the amplicons for the YKI 20-1003, leaving the strain unidentified. Judging from the identification data obtained in the Kimchi fermented at $4^{\circ}C$ or $10^{\circ}C$, Leuconostoc spp. including Leu. mesenteroides/dextranicum were likely predominant species in the earlier stage and L. brevis occurred at the high level through the whole period. By contrast, L. brevis, as one of the major flora, possibly lead the fermentation from the beginning in the Kimchi fermented at $20^{\circ}C$.}C$.

Associated-Genes and Virulence Factors of Staphylococcus aureus Isolated from Nasal Cavity of Neonates (신생아 비강에서 분리된 황색포도구균의 병원성 인자와 관련 유전자)

  • Kim, Yung Bu;Moon, Ji Young;Park, Jae Hong
    • Clinical and Experimental Pediatrics
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    • v.46 no.1
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    • pp.24-32
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    • 2003
  • Purpose : Nosocomial infection with Staphylococcus aureus, especially methicillin resistant S. aureus, has become a serious concern in the neonatal intensive care unit. The aim of this study is to investigate the virulence factors, and the relationship between the antibiotic resistance and the associated genes of Staphylococcus aureus isolated from nasal cavity of neonates. Methods : Fifty one isolates of S. aureus were obtained from nasal swab taken in 28 neonates in the NICU and nursery of Pusan National University Hospital between February and May, 2001. They were tested in regard to antibiotic susceptibility, coagulase test and typing, plasmid DNA profile, as well as reactivity to enterotoxin A-E(sea, seb, sec, sed, see) genes and toxic shock syndrome toxin-1(tst) gene by polymerase chain reaction(PCR). Associated genes such as mecA, mecR1, mecI, and femA were also determined by PCR. The origin of MRSA strains was assessed using DNA fingerprinting by arbitrarily-primed polymerase chain reaction(AP-PCR). Results : Twenty three(45.1%) and six(11.8%) isolates were resistant to oxacillin and vancomycin respectively. Multidrug resistance to three or more of the antibiotics tested was observed in 51.0% of the isolates. Forty two isolates were coagulase positive and twenty two isolates had mecA gene. Sixteen isolates had both mecA and femA genes and had type I-III plasmids. 64.7% of isolates carried sec gene, and 80.4% carried tst gene. DNA fingerprinting by AP-PCR for 12 MRSA strains showed 10 distinct patterns, suggesting different origins. Conclusion : We confirmed that the prevalence of nasal carriage of S. aureus and the incidence of antimicrobial-resistant S. aureus, especially vancomycin resistance, is very high in neonates who were admitted in NICU and nursery. It is possible that these pathogens are responsible for serious nosocomial infections in neonates. The need for improved surveillance and continuous control of pathogens is emphasized.

Prevalence, Serotype Diversity, Genotype and Antibiotic Resistance of Listeria monocytogenes Isolated from Carcasses and Human in Korea

  • Oh, Hyemin;Kim, Sejeong;Lee, Soomin;Lee, Heeyoung;Ha, Jimyeong;Lee, Jeeyeon;Choi, Yukyung;Choi, Kyoung-Hee;Yoon, Yohan
    • Food Science of Animal Resources
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    • v.38 no.5
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    • pp.851-865
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    • 2018
  • This study investigated the prevalence of Listeria monocytogenes in slaughterhouses, and determined serovars and genotypes, and antibiotic resistance of the isolates obtained from slaughterhouses and humans in Korea. Two hundred ninety samples were collected from feces (n=136), carcasses [n=140 (cattle: n=61, swine: n=79)], and washing water (n=14) in nine slaughterhouses. Eleven human isolates were obtained from hospitals and the Korea Center for Disease Control and Prevention. Listeria monocytogenes was enriched and identified, using polymerase chain reaction (PCR) and 16S rRNA sequencing. Serovars and presence of virulence genes were determined, and genetic correlations among the isolates were evaluated by the restriction digest patterns of AscI. Antibiotic resistance of L. monocytogenes isolates were examined against 12 different antibiotics. Of 290 slaughterhouse samples, 15 (5.17%) carcass samples were L. monocytogenes positive. Most L. monocytogenes isolates possessed all the virulence genes, while polymorphisms in the actA gene were found between carcass and human isolates. Serovars 1/2a (33.3%) and 1/2b (46.7%) were the most frequent in carcass isolates. Genetic correlations among the isolates from carcass and clinical isolates were grouped within serotypes, but there were low geographical correlations. Most L. monocytogenes isolates were antibiotic resistant, and some strains showed resistance to more than four antibiotics. These results indicate that L. monocytogenes are isolated from carcass and human in Korea, and they showed high risk serotypes and antibiotic resistance. Therefore, intensive attentions are necessary to be aware for the risk of L. monocytogenes in Korea.

Characterization of antimicrobial resistance and application of RFLP for epidemiological monitoring of thermophilic Campylobacter spp. isolated from dogs and humans in Korea

  • Cho, Hyun-Ho;Kim, Sang-Hyun;Min, Wongi;Ku, Bok-Kyung;Kim, Jong-Hyun;Kim, Yong-Hwan
    • Korean Journal of Veterinary Research
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    • v.54 no.2
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    • pp.91-99
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    • 2014
  • An antimicrobial susceptibility test was conducted to compare the resistance rates among Campylobacter spp. isolates from dogs (n = 50) raised under diverse conditions and humans (n = 50). More than 60% of Campylobacter (C.) jejuni from dogs and humans showed resistance to nalidixic acid, enrofloxacin and ciprofloxacin. C. jejuni isolates from humans showed higher resistance to tetracycline (83.3%) and ampicillin (91.3%) than those from dogs. None of the C. jejuni or Campylobacter coli isolates from humans or dogs were resistant to erythromycin. Overall, 85% of Campylobacter spp. isolates showed a multidrug resistant phenotype. Nucleotide sequencing analysis of the gryA gene showed that 100% of $NA^R/CIP^R$ C. jejuni isolates from dogs and humans had the Thr-$86^{th}$-Ile mutation, which is associated with fluoroquinolone resistance. flaA PCR restriction fragment length polymorphism (RFLP) typing to differentiate the isolates below the species level revealed 12 different clusters out of 73 strains. The human isolates belonged to eight different RFLP clusters, while five clusters contained dog and human isolates.

Isolation and Identification of the Antilisterial Bifidobacterium Isolates from the Infants Fecal Samples (유아의 분변으로부터 항리스테리아 활성의 Bifidobacterium 속 균주의 분리 및 동정)

  • Kim, Song-Yi;Kim, Ki-Hwan;Youn, Soon-Yong;Yoon, Sung-Sik
    • Journal of Dairy Science and Biotechnology
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    • v.24 no.1
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    • pp.19-28
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    • 2006
  • This study was conducted to isolate antilisterial strains of the Bifidobacterium isolates from the infant feces. The bifidobacteria were isolated anaerobically on BL agar and screened for their inhibitory activity on the MRS-cysteine medium against three foodborne pathogens: Listeria monocytogenes, Bacillus cereus, and Staphylococcus aureus. Among the 52 bifidobacterial isolates, 5 strains(A24, Bl, B6, B10, and Bl2) were finally selected based on their stronger antilisterial activity against Listeria monocytogenes than other isolates tested. Morphologically, all the isolates were typically shown Y-and V-shaped under electron microscopic examination. Each isolate was primarily subjected to identification by a polymerase chain reaction(PCR) using a genus-specific primer designed for targeting the 16S rRNA gene sequence, and confirmed the primary identification data using an API-kit(Biomeriuex, France), commercially available product for identification based on biochemical and physiological traits. Of the isolates with antilisterial activity, strain A24 was finally confirmed as the Bifidobacterium longum A24.

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Genetic characteristics of the Korean isolate KI -1 of Toxoplasma gondii

  • LIN Aifen;SHIN Eun-Hee;KIM Tae-Yun;PARK Jae-Hwan;GUK Sang-Mee;CHAI Jong-Yil
    • Parasites, Hosts and Diseases
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    • v.43 no.1 s.133
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    • pp.27-32
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    • 2005
  • Toxoplasma gondii tachyzoites were isolated from an ocular patient in the Republic of Korea and maintained in the laboratory (designated KI-1). In the present study, its genotype was determined by analyzing dense granule antigen 6 (GRA6) gene and surface antigen 2 (SAG2) gene as typing markers. Digestion of the amplification products of GRA6 and of the 5' and 3' ends of SAG2, respectively, with Mse I, Sau3A I, and Hha I, revealed that KI-1 is included in the genotype I, which includes the worldwide virulent RH strain. In addition, when the whole sequences of the coding regions of SAG1, rhoptry antigen 1 (ROP1), and GRA8 genes of KI-1 were compared with those of RH, minor nucleotide polymorphisms and amino acid substitutions were identified. These results show that KI-1 is a new geographical strain of T. gondii that can be included in the genotype I.

Coexistence of Malaria and Thalassemia in Malaria Endemic Areas of Thailand

  • Kuesap, Jiraporn;Chaijaroenkul, W.;Rungsihirunrat, K.;Pongjantharasatien, K.;Na-Bangchang, Kesara
    • Parasites, Hosts and Diseases
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    • v.53 no.3
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    • pp.265-270
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    • 2015
  • Hemoglobinopathy and malaria are commonly found worldwide particularly in malaria endemic areas. Thalassemia, the alteration of globin chain synthesis, has been reported to confer resistance against malaria. The prevalence of thalassemia was investigated in 101 malaria patients with Plasmodium falciparum and Plasmodium vivax along the Thai-Myanmar border to examine protective effect of thalassemia against severe malaria. Hemoglobin typing was performed using low pressure liquid chromatography (LPLC) and ${\alpha}$-thalassemia was confirmed by multiplex PCR. Five types of thalassemia were observed in malaria patients. The 2 major types of thalassemia were Hb E (18.8%) and ${\alpha}$-thalassemia-2 (11.9%). There was no association between thalassemia hemoglobinopathy and malaria parasitemia, an indicator of malaria disease severity. Thalassemia had no significant association with P. vivax infection, but the parasitemia in patients with coexistence of P. vivax and thalassemia was about 2-3 times lower than those with coexistence of P. falciparum and thalassemia and malaria without thalassemia. Furthermore, the parasitemia of P. vivax in patients with coexistence of Hb E showed lower value than coexistence with other types of thalassemia and malaria without coexistence. Parasitemia, hemoglobin, and hematocrit values in patients with coexistence of thalassemia other than Hb E were significantly lower than those without coexistence of thalassemia. Furthermore, parasitemia with coexistence of Hb E were 2 times lower than those with coexistence of thalassemia other than Hb E. In conclusion, the results may, at least in part, support the protective effect of thalassemia on the development of hyperparasitemia and severe anemia in malaria patients.