• Title/Summary/Keyword: Multiplex PCR

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Epidemiologic Characteristics of Human Bocavirus-Associated Respiratory Infection in Children (소아 보카바이러스 호흡기 감염증의 역학적 특징)

  • Choi, Jae Hong;Paik, Ji Yeun;Choi, Eun Hwa;Lee, Hoan Jong
    • Pediatric Infection and Vaccine
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    • v.18 no.1
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    • pp.61-67
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    • 2011
  • Purpose : This study was performed to investigate the epidemiologic characteristics of human bocavirus (HBoV)-associated lower respiratory tract infections (LRTIs) in children. Methods : Nasopharyngeal aspirate samples were obtained from 658 children who had been hospitalized for LRTIs in Seoul National University (SNU) Children's Hospital and SNU Bundang Hospital from March 2000 to September 2005. Multiplex RT-PCR was performed to detect 11 respiratory viruses including respiratory syncytial virus, adenovirus, rhinovirus, parainfluenza viruses 1 and 3, influenza viruses A and B, human metapneumovirus, HBoV, human coronavirus (HCoV) OC43/ 229E, and HCoV-NL63. Clinical data were reviewed retrospectively. Results : Overall, respiratory viruses were detected in 325 (49.4%) among 658 patients. HBoV was detected in 62 cases (9.4%) and was responsible for 19.1% of virus-positive cases. HBoV was prevalent among infants and young children aged from 3 months to 5 years with the mean age of 25.3 months. Co-detection of HBoV and other respiratory viruses was observed in 37.1% which is significantly higher than average co-detection rate (12.3%) among overall virus-positive cases (P=0.000). HBoV was identified mainly in late spring and early summer from May to July. Conclusion : This study describes epidemiologic features of HBoV in Korean children compared with those associated with other respiratory viruses. HBoV was prevalent among LRTIs in childhood, especially in late spring and early summer season in Korea.

Two cases of Hemolytic Uremic Syndrome Associated with Esecherichia coli O114 (Escherichia coli O114에 의한 용혈성 요독 증후군 2례)

  • Ryu Su-Jeung;Hahn Hyewon;Yoo Soo Jin;Lee Byung-Sun;Lee Jae Kyoo;Kim Mi-Na;Kim Eui Chong;Park Young Seo
    • Childhood Kidney Diseases
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    • v.6 no.1
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    • pp.102-108
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    • 2002
  • We report two cases of hemolytic uremic syndrome (HUS) associated with Escherichia coli O114. Two cases were similar and showed the same clinical courses. After prodrome of diarrhea and vomiting lasting 1-2 days, azotemia persisted for about 10 days, and during that period, the patients were on peritoneal dialysis. They recovered without any sequelae after about 15 days. Direct multiplex PCR of stool culture revealed eae and stx2 gene and the result of ELISA done on the colony positive of one gene confirmed Escherichia coli O114. This is the first report of HUS associated with Escherichia coli O114. We recommend, Shiga toxin producing bacterial Infection must be considered and efforts should be made to scrutinize the organism in all diarrhea-prodrome HUS patients.(J Korean Soc Pediatr Nephrol 2002 ;6 : 102-8)

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Modulation of Pituitary Somatostatin Receptor Subtype (sst1-5) mRNA Levels by Growth Hormone (GH)-Releasing Hormone in Purified Somatotropes

  • Park, Seung-Joon;Park, Hee-Soon;Lee, Mi-Na;Sohn, Sook-Jin;Kim, Eun-Hee;Jung, Jee-Chang;Frohman, Lawrence A.;Kineman, Rhonda D.
    • The Korean Journal of Physiology and Pharmacology
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    • v.7 no.2
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    • pp.79-84
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    • 2003
  • We have previously reported that expression of the somatostatin receptor subtypes, sst1-5, is differentially regulated by growth hormone (GH)-releasing hormone (GHRH) and forskolin (FSK), in vitro. GHRH binds to membrane receptors selectively located on pituitary somatotropes, activates adenylyl cyclase (AC) and increases sst1 and sst2 and decreases sst5 mRNA levels, without significantly altering the expression of sst3 and sst4. In contrast FSK directly activates AC in all pituitary cell types and increases sst1 and sst2 mRNA levels and decreases sst3, sst4 and sst5 expression. Two explanations could account for these differential effects: 1) GHRH inhibits sst3 and sst4 expression in somatotropes, but this inhibitory effect is masked by expression of these receptors in unresponsive pituitary cell types, and 2) FSK inhibits sst3 and sst4 expression levels in pituitary cell types other than somatotropes. To differentiate between these two possibilities, somatotropes were sequentially labeled with monkey anti-rat GH antiserum, biotinylated goat anti-human IgG, and streptavidin-PE and subsequently purified by fluorescent-activated cell sorting (FACS). The resultant cell population consisted of 95% somatotropes, as determined by GH immunohistochemistry using a primary GH antiserum different from that used for FACS sorting. Purified somatotropes were cultured for 3 days and treated for 4 h with vehicle, GHRH (10 nM) or FSK ($10{\mu}M$). Total RNA was isolated by column extraction and specific receptor mRNA levels were determined by semi-quantitative multiplex RT-PCR. Under basal conditions, the relative expression levels of the various somatostatin receptor subtypes were sst2>sst5>sst3=sst1> sst4. GHRH treatment increased sst1 and sst2 mRNA levels and decreased sst3, sst4 and sst5 mRNA levels in purified somatotropes, comparable to the effects of FSK on purified somatotropes and mixed pituitary cell cultures. Taken together, these results demonstrate that GHRH acutely modulates the expression of all somatostatin receptor subtypes within GH-producing cells and its actions are likely mediated by activation of AC.

Direct Deletion Analysis in Two Duchenne Muscular Dystrophy Symptomatic Females Using Polymorphic Dinucleotide (CA)n Loci within the Dystrophin Gene

  • Giliberto, Florencia;Ferreiro, Veronica;Dalamon, Viviana;Surace, Ezequiel;Cotignola, Javier;Esperante, Sebastian;Borelina, Daniel;Baranzini, Sergio;Szijan, Irene
    • BMB Reports
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    • v.36 no.2
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    • pp.179-184
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    • 2003
  • Duchenne muscular dystrophy (DMD) is the most common hereditary neuromuscular disease. It is inherited manifestations. In some rare cases, the disease can also be manifested in females. The aim of the present study was to determine the molecular alteration in two cases of nonrelated DMD symptomatic carriers with no previous history of DMD. Multiplex PCR is commonly used to search for deletion in the DMD gene of affected males. This method could not be used in females because the normal X chromosome masks the deletion of the mutated one. Therefor, we used a set of seven highly polymorphic dinucleotide $(CA)_n$ repeat markers that lie within the human dystrophin gene. The deletions were evidenced by hemizygosity of the loci under study. We localized a deletion in the locus 7A (intron 7) on the maternal X chromosome in one case, and a deletion in the region of introns 49 and 50 on the paternal X chromosome in the other. The use of microsatellite genotyping within the DMD gene enables the detection of the mutant allele in female carriers. It is also a useful method to provide DMD families with more accurate genetic counseling.

Epidemiological Characterization of Adenovirus and Human Bocavirus Detected Acute Respiratory Patients in Busan (부산지역 호흡기감염증 환자로부터 분리한 아데노바이러스와 보카바이러스의 유행양상 분석)

  • Hwang, Su-Jeong;Kim, Nam-Ho;Park, Dong-Ju;Ku, Pyung-Tae;Lee, Mi-Ok;Jin, Sung-Hyun
    • Journal of Life Science
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    • v.27 no.3
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    • pp.275-282
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    • 2017
  • Adenovirus (ADV) and human bocavirus (hBoV) cause acute respiratory tract infections, and are often associated with increased rates of hospitalization and death, particularly in infants and young children. The aim of this study was to analyze the clinical features and molecular phylogeny of ADV and hBoV isolated in Busan, from January 2011 to November 2013. In total, 3,230 specimens (throat swabs) were collected from patients with influenza-like illnesses and acute respiratory tract infections. Multiplex real-time RT-PCR was performed to detect eight respiratory viru [rhinovirus, adenovirus, respiratory syncytial virus, human coronavirus, human metapneumovirus, human bocavirus, parainfluenza virus and influenza virus] and detected 1,485(46.0%) cases. Among 1,485 positive specimens, 257(8.0%) cases were ADV and 68(2.1%) cases of hBoV. A significant clinical feature of ADV is fever and headache whereas hBoV is wheezing. Serotypic distributions of isolated ADV and hBoV were analyzed by sequencing of hexon and VP1/VP2 gene, respectively. ADV was identified seven different serotypes(1~6, 8), revealing a high similarity among the isolates (>97%). The predominant types of ADV were type 1 in 2011, type 3 and 4 in 2012, type 3 in 2013, respectively. ADV type 3 was major causative type during outbreaks in 2013. All of the hBoV was identified as hBoV type 1.

Development of a Microsatellite Marker Set for the Individual Identification and Parentage Verification of Korean Native Black Goats (재래흑염소 개체식별과 친자확인을 위한 Microsatellite Marker Set 개발)

  • Lee, Sang-Hoon;Kang, Ho-Chan;Lee, Sung-Soo;Lee, Jinwook;Kim, Eun-Ho;Myung, Hyun-Cheol;Kim, Kwan-Woo;Lim, Hyun-Tae
    • Journal of Life Science
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    • v.30 no.10
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    • pp.912-918
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    • 2020
  • The Korean native black goat (Capra hircus coreanae) is the goat species to be officially registered in Korea under the Food and Agriculture Organization. The object of this study is to establish a set of microsatellite (MS) markers for the individual identification and parentage verification of goats. In this study, we analyzed alleles of MS markers in crosses between Korean native black goats and crossbred goats (n=304 animals), and, based on the diversity of alleles for each marker, we selected 11 MS markers for individual identification and parentage verification. Using these 11 MS markers, the probabilities of different individuals with the same genotype being found within random and half-sib mating populations were 5.58×10-10 and 1.15×10-7, respectively. The parentage verification accuracy was 0.999996 when information about the parents was available and 0.999833 with no information. Thus, even given the total rearing population of 576,150 animals in South Korea, we concluded that these markers could be used for the individual identification and parentage verification of goats. Moreover, by analyzing the genetic relationships between the four lines of Korean native black goats and the crossbred goats, we verified the genetic characteristics of Korean native black goats, confirming their conservation value as a unique genetic resource.

The Prevalence and Distribution of the P and G Genotypes of a Group A Rotavirus Detected in Acute Gastroenteritis Patients from Incheon (인천지역 급성 설사환자의 group A rotavirus 감염 실태 및 P와 G 유전자형 분포)

  • Choi, Hye-Jin;Oh, Bo-Young;Lee, Mi-Yeon;Koh, Yeon-Ja;Gong, Young-Woo;Hur, Myung-Je;Lee, Jea-Mann;Kim, Young-Hee;Jeong, Hye-Sook;Cheon, Doo-Sung
    • Journal of Life Science
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    • v.22 no.5
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    • pp.600-604
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    • 2012
  • Rotavirus is the main cause of severe diarrhea in infants and young children of the world. However, the frequency of genetic alterations makes it hard to control the prophylaxis. Therefore, continuous monitoring of the rotavirus's genetic change is inevitable to prevent disease prevalence and is useful in inventing an efficient vaccine. From January 2005 to December 2010, we investigated 11,607 stool samples of acute gastroenteritis patients in the Incheon metropolitan area. About 13.18% (1,530 stool samples) of all samples had a positive reaction against rotavirus using an antigen capture enzyme-linked immunosorbent assay (ELISA). Then, the 160 stool samples were searched for subtypes of group A rotavirus by using a reverse transcription polymerase chain reaction (RT-PCR) and a nested multiplex RCR. In P sub-typing, P8 (56.3%) was an extremely prevalent genotype, followed by P6 (21.3%), and P1A (10.0%). G1 (39.4%) was most widespread in the G subtype, followed by G4 (25.0%) and G3 (18.8%). G1P8 (35.5%) was the most common G and P subtype combination, followed by G4P6 (19.3%) and G3P8 (13.1%). These results might be useful data for understanding the epidemiological status of group A-rotavirus dispersion in the Incheon metropolitan area.

Analysis of Small Fragment Deletions of the APC gene in Chinese Patients with Familial Adenomatous Polyposis, a Precancerous Condition

  • Chen, Qing-Wei;Zhang, Xiao-Mei;Zhou, Jian-Nong;Zhou, Xin;Ma, Guo-Jian;Zhu, Ming;Zhang, Yuan-Ying;Yu, Jun;Feng, Ji-Feng;Chen, Sen-Qing
    • Asian Pacific Journal of Cancer Prevention
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    • v.16 no.12
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    • pp.4915-4920
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    • 2015
  • Background: : Familial adenomatous polyposis (FAP) is an autosomal dominant inherited disease mainly caused by mutations of the adenomatous polyposis coli (APC) gene with almost complete penetrance. These colorectal polyps are precancerous lesions that will inevitable develop into colorectal cancer at the median age of 40-year old if total proctocolectomy is not performed. So identification of APC germline mutations has great implications for genetic counseling and management of FAP patients. In this study, we screened APC germline mutations in Chinese FAP patients, in order to find novel mutations and the APC gene germline mutation characteristics of Chinese FAP patients. Materials and Methods: The FAP patients were diagnosed by clinical manifestations, family histories, endoscope and biopsy. Then patients peripheral blood samples were collected, afterwards, genomic DNA was extracted. The mutation analysis of the APC gene was conducted by direct polymerase chain reaction (PCR) sequencing for micromutations and multiplex ligation-dependent probe amplification (MLPA) for large duplications and/or deletions. Results: We found 6 micromutations out of 14 FAP pedigrees, while there were no large duplications and/or deletions found. These germline mutations are c.5432C>T(p. Ser1811Leu), two c.3926_3930delAAAAG (p.Glu1309AspfsX4), c.3921_3924delAAAA (p.Ile1307MetfsX13), c3184_3187delCAAA(p.Gln1061AspfsX59) and c4127_4126delAT (p.Tyr1376LysfsX9), respectively, and all deletion mutations resulted in a premature stop codon. At the same time, we found c.3921_3924delAAAA and two c.3926_3930delAAAAG are located in AAAAG short tandem repeats, c3184_3187delCAAA is located in the CAAA interrupted direct repeats, and c4127_4128 del AT is located in the 5'-CCTGAACA-3', 3'-ACAAGTCC-5 palindromes (inverted repeats) of the APC gene. Furthermore, deletion mutations are mostly located at condon 1309. Conclusions: Though there were no novel mutations found as the pathogenic gene of FAP in this study, we found nucleotide sequence containing short tandem repeats and palindromes (inverted repeats), especially the 5 bp base deletion at codon 1309, are mutations in high incidence area in APC gene,.

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.

Prevalence and Antimicrobial Resistance of Enterococus faecalis and Enterococcus faecium Isolated from Beef, Pork, Chicken and Sashimi (시판 축산물 및 수산물에서 Enterococcus faecalis와 Enterococcus faecium 분포 및 항생제 감수성에 관한 연구)

  • Sung, Chang-Hyun;Chon, Jung-Whan;Kwak, Hyo-Sun;Kim, Hyunsook;Seo, Kun-Ho
    • Food Science of Animal Resources
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    • v.33 no.1
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    • pp.133-138
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    • 2013
  • In this study, a total of 256 samples of retail raw meats (beef, pork and chicken) and sashimi were investigated for the presence of Enterococcus faecalis and Enterococcus faecium. We isolated a total of 117 E. faecalis and E. faecium from the samples, with contamination rates ranging from 18.8% for sashimi samples to 68.8% of chicken samples. E. faecalis was the predominant species recovered from all of the retail raw meats beef (42.2%), pork (42.2%), chicken (65.6%) and sashimi (12.5%). Among 117 isolates, 61 isolates (52.1%) were resistant to tetracycline, 32 isolates (27.4%) were resistant to erythromycin, 23 isolates (19.7%) were resistant to chloramphenicol, 16 isolates (13.7%) were resistant to ripampin, 10 isolates (8.5%) were resistant to gentamycin, 9 isolates (7.7%) were resistant to ciprofloxacin and 1 isolate (0.9%) was resistant to ampicillin and penicillin G. No resistance to amoxicillin + clavulanic acid and vancomycin was observed. Although no strain was resistant to vancomycin, the vanB gene was observed in 9 of 117 of Enterococcus (7.7%) demonstrating potential risk of vancomycin-resistant Enterococcus (VRE). Our results indicate that E. faecalis and E. faecium were highly prevalent in retail raw meats, but most strains were sensitive to tested antibiotics.