• Title/Summary/Keyword: PCR-ELISA

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Detection of Cryptosporidium spp. in Diarrheic Immunocompetent Patients in Beni-Suef, Egypt: Insight into Epidemiology and Diagnosis

  • Gawad, Samah S. Abdel;Ismail, Mousa A.M.;Imam, Naglaa F.A.;Eassa, Ahmed H.A.;abu-Sarea, Enas Yahia
    • Parasites, Hosts and Diseases
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    • v.56 no.2
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    • pp.113-119
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    • 2018
  • Cryptosporidium species is an important cause of gastrointestinal infections globally. This study aimed to shed light on its role in diarrheic immunocompetent patients in Beni-Suef, Egypt and to compare three diagnostic methods. Two hundred diarrheic patients, $37{\pm}16.8year$ old, were enrolled. Stool samples were examined by light microscopy, using modified Ziehl-Neelsen stain (MZN) for Cryptosporidium spp. oocysts. Coproantigens were detected by sandwich ELISA. DNA molecular diagnosis was done by nested PCR. PCR yielded the highest detection rates (21.0%), compared to ELISA (12.5%) and MZN staining method (9.5%). The higher infection rates were in 20-40 year-old group, followed by 40-60 year-old. Association between epidemiologic factors was statistically not significant; positivity and gender, clinical manifestations, residence, source or water, or contact with animals. Cryptosporidiosis is an important enteric parasitic infection in Beni-Suef and PCR remains the gold standard for diagnosis.

An Effective Method of Diagnosis of Potato Leafroll Virus by RT-PCR (RT-PCR 방법을 이용한 효과적인 감자 잎말림 바이러스의 검정)

  • Jeon, Jae-Heung;Joung, Young-Hee;Choi, Kyung-Hwa;Kim, Hyun-Soon;Oh, Hyun-Woo;Park, Se-Won;Joung, Hyouk
    • Korean Journal Plant Pathology
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    • v.12 no.3
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    • pp.358-362
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    • 1996
  • 감자 잎말림 바이러스를 검정하기 위하여 ELISA 및 전자현미경에 의해 바이러스 감염이 확인된 기내 배양중인 감장의 줄기로부터 RT-PCR 분석을 수행하였다. 분리된 총 RNA들로부터 바이러스 cDNA를 합성하고 감자 잎말림 바이러스 외피단백질의 일부인 465bp를 특이하게 증폭하도록 고안한 두 primer를 사용하여 PCR 반응을 하였다. 증폭된 465pb의 DNA 절편의 염기서열을 분석한 결과 역시 감자 잎말림 바이러스임을 확인하였다. 바이러스 검정에 있어서 EL-ISA 방법과 RT-PCR 방법간의 민감도를 조사한 결과 RT-PCR 방법간의 민감도를 조사한 결과 RT-PCR 방법이 ELISA 방법보다 감자 잎말림 바이러스검정에 있어서보다 정확한 방법인 것으로 사료된다.

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Development of Single-tube Multiplex Immunocapture RT-PCR Assay for Simultaneous Detection of Two Pepper Tobamoviruses

  • Kim, Jae-Hyun;Choi, Gug-Seoun;Kim, Jeong-Soo;Lee, Sin-Ho;Choi, Jang-Kyung;Ryu, Ki-Ryun
    • The Plant Pathology Journal
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    • v.22 no.2
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    • pp.164-167
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    • 2006
  • An immunocapture reverse transcription-polymerase chain reaction (IC/RT-PCR) was developed for simultaneous detection of two pepper-infecting RNA viruses, Pepper mud mottle virus (PMMoV) and Tobacco mild green mosaic virus (TMGMV). The assay could be performed in a single tube for simultaneous and sensitive detection of these tobamoviruses. This detection system revealed thousand-fold increase in detection sensitivity compare to ELISA. This method could save time and reagent cost compare to common RT-PCR which needs several reactions and several procedures of viral RNA extractions for the same number of samples.

Prevalence of bovine tuberculosis, brucellosis and Q fever in Korean black goats (국내 흑염소에서의 소결핵, 브루셀라, Q fever 유병률 연구)

  • Kim, Hyobi;Kim, Seongjoon;Kim, Kina;Kim, Byeol;Chang, Byungjoon;Choe, Nong-Hoon
    • Korean Journal of Veterinary Research
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    • v.56 no.4
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    • pp.249-254
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    • 2016
  • As the meat of black goats has become popular as a healthy food, domestic goat meat-related industries are steadily growing. However, previous studies are scarce of informations about the zoonotic disease originated from the black goat in Korea. In this study, we investigated Korean black goat's infectious diseases representing bovine tuberculosis, brucellosis, and Q fever. One hundred and eighty samples were collected from a local slaughter house located in Jeollanam-do. Three typical zoonotic diseases were separately examined by carrying out enzyme linked immunosorbent assay (ELISA) and polymerase chain reaction (PCR). Histopathological test was additionally performed in tuberculosis. In case of tuberculosis, results of the PCR and histopathological test were negative but the ELISA results were positive in eight samples. In case of brucellosis, one out of the total samples was shown to be positive in the ELISA and none in the PCR. In case of Q fever, there were forty one positive in the ELISA and twenty positive in the real-time PCR. Those results indicate that the Korean black goat could be a natural reservoir in the possible chain-infections among human, cows and goats. Thus, further study needs in order to improve productivity as well as to prevent the zoonosis spreading and circulation of other livestock with the black goat in this country.

Investigation of infection rate and genetic sequence analysis of chicken infectious anemia virus (닭 전염성빈혈 감염률 및 유전자 분석)

  • Chu, Keum-Suk;Kang, Mi-Seon;Song, Hee-Jong;Lee, Jeong-Won
    • Korean Journal of Veterinary Service
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    • v.33 no.1
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    • pp.13-21
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    • 2010
  • Chicken anemia virus (CAV) has been recognized as an immunosuppressive agent and plays role as an etiological agent of multifactorial diseases in chicken. In this study, we investigated distribution of CAV antibody by ELISA and the virus gene by PCR in poultry farms in Jeongeup, Jeonbuk province. In the test using ELISA kit, 41 (95.3%) of 43 flocks and 88.6% of the individual chickens were positive, respectively. By PCR, 90.9% of the broiler breeders and 75.0% of White-semi breeders were found positive, respectively. All hatchery was negative by PCR. Of the clinical cases from 49 poultry flocks, 87.5% of flocks and 54.7% for each samples were found positive by ELISA, respectively. By PCR test, 21 (42.9%) of 49 flocks were positive. Major clinical signs of the infected flocks were growth retardation, femoral subcutaneous bleeding, depression, limping, and continuing selection. The genetic analysis of separate N genes of CAV showed highly homologous each other. The nucleotide sequence of field isolates had homology ranged from 99.9% to 97.5% with Chinese strains, and 99.9% to 99.6% with Japanese strain. Phylogenetic analysis based on the N gene of CAV isolates showed the closely relation with Chinese strains. The results of this survey could be used as basic data for development of vaccine.

Prevalence of paratuberculosis of dairy cattle in Kangwon area (강원지역 젖소의 요네병 감염실태)

  • Kim, Doo;Jeon, Kwan-joon;Kim, Jong-taek;Shin, Kwang-soon;Shin, Myung-kyun;Chang, Guk-hyun;Kim, Jeung-ki;Kim, Och-sung;Jung, Jae-young
    • Korean Journal of Veterinary Research
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    • v.42 no.1
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    • pp.81-88
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    • 2002
  • The purpose of this study was to conduct diagnosis of bovine paratuberculosis in Kangwon area. Blood samples were collected from 2,261 dairy cows of 162 herds, and the ELISA and immunoblotting using recombinant 34KDa protein of M. paratuberculosis were conducted. The feces collected from dairy cows were cultured on HEY medium with mycobactin-J and PCR was conducted with washing solution of medium 4 weeks after culture. The ELISA had sensitivity of 83.3% and specificity of 96.7%. And the immunoblotting had sensitivity of 83.3% and specificity of 100%. Of the 2,261 dairy cows, 371 cows(16.4%) were positive in ELISA and 75 cows(3.3%) were positive in immunoblotting. And of the 162 herds, 109 herds(67.3%) had an apparent paratuberculosis prevalence by ELISA and 40 herds(24.7%) by immunoblotting. The geographic distribution of herds with paratuberculosis was not uniform. Of the 241 feces samples including 110 feces from ELISA positive cow, 9 feces were positive in culture and PCR. PCR was able to detect the growth of M. paratuberculosis as early as 4 weeks of culture.

Diagnosis of Viral Infection of Pensaeide (보리새우류의 바이러스 감염증의 진단)

  • 허문수;정초록;장경립
    • Journal of Life Science
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    • v.9 no.4
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    • pp.453-459
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    • 1999
  • Baculovirus(WSBV) was isolated from infected Penaeide was collected from shrimp farm at southern sea of Korea from 1993 to 1995. The Infectious virus was purified and used for diagnosis of infected shrimp. Anti-viral serum were used for immunological detection as enzyme linked immunoabsorbent assay(ELISA) and indirect fluorescent antibody technique(IFAT). In IFAT, stomach, lymphoid organ and antenae gland of infected shrimp showed fluorescent reaction. In ELISA, tissues of spontaneously infected shrimp appeared higher O.D. values than in artificial infected shrimp. Primer set was constructed from sequence of 420bp of cloned Baculovirus(WSBV) genome. Specific band for infected shrimp was detected in Polymerase chain reaction(PCR)

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Detection of Toxigenicity of Cyanobacteria by Molecular Method (분자생물학적 방법에 의한 남조류의 독성 생성능의 확인)

  • Lee, Kyung-Lak;Jheong, Weon-Hwa;Kim, Jong-Min;Kim, Han-Soon
    • Korean Journal of Ecology and Environment
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    • v.40 no.1
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    • pp.149-154
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    • 2007
  • In the present study, we performed the PCR assay using TOX2P/TOX2M primer targeting a specific region within mcyB gene to identify potential microcystin-producing cyanobacteria. TOX2P/TOX2M primer set was effective in amplifying mcy gene in the field samples containing Microcystis spp. of 1,000 cells per mL. Moreover, the results from the PCR assay agreed with those of the ELISA analysis. Consequently, this study demonstrated that TOX2P/TOX2M primer set can be used as a genetic probe for the early detection of cyanobacterial toxigenicity in Korean water bodies.

Effects of bacterial LPS and DNA on the induction of IL-1β, IL-10 and IL-12 by mouse peritoneal macrophages in vitro

  • Samad, D. Abdel;Abdelnoor, AM
    • Advances in Traditional Medicine
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    • v.6 no.2
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    • pp.134-143
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    • 2006
  • The capacities of bacterial DNA, extracted from Salmonella typhimurium, and lipopolysaccharide (LPS), extracted from Salmonella minnesota, to activate mouse peritoneal macrophages in vitro were compared. Activation was assessed by estimating e levels of 3 cytokines, IL-10, IL-12, and $IL-1{\beta}$, at time intervals of 3, 6, 9, and 24 h after addition of LPS and/or DNA to macrophage cultures. Cytokine levels in culture supernatants were determined by enzyme-linked immunosorbent assay (ELISA) and cytokine mRNA levels were estimated based on band intensity in cultured cells by reverse transcriptase-polymerase chain reaction (RT-PCR). Results obtained demonstrated the ability of DNA and LPS to elicit increased production of all 3 cytokines as compared to controls. In the amount tested, LPS appeared to be a more potent inducer of IL-12, and $IL-1{\beta}$, whereas DNA induced higher levels of IL-10. DNA and LPS, used in combination, exhibited neither an additive nor a synergistic effect. Rather, an antagonist effect appeared to occur. RT-PCR results correlated well with ELISA.