• 대한수의학회지
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• 제48권2호
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• pp.191-195
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• 2008

This study was performed to detect Neospora caninum in blood of 61 Korean native cattle and 50 dairy cows in Chungnam province. All of them were healthy and did not show any clinical signs. DNA was isolated from blood samples and a 328 bp fragment was amplified by PCR using primer pair Np21 and Np6. The PCR positive rate was 14.8% in Korean native cattle and 0% in dairy cows. Cows with 15.6% were a little higher than bulls with 12.5% in gender. The detection rate of over 3-year-old Korean native cattle was 28.6% in age. The results demonstrate that N. caninum DNA can be detected in blood by PCR. PCR analysis in blood may be useful to annually screening test for N. caninum infection in clinically healthy cattle.

• 대한수의학회지
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• 제39권6호
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• pp.1151-1160
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• 1999

This study was conducted to detect the toxoplasma-specific DNA in peripheral blood collected from cats experimentally infected with Toxoplasma gondii (RH strain) and from domiciled cats by B1 gene-base polymerise chain reaction(PCR). The sensitivity of oligonucleotide primer, T-1 & T-2, designed from toxoplasma B1 gene amplification method was compared with parasite detection by mouse inoculation(MI). And also, latex agglutination test(LAT) and indirect fluorescent antibody test(IFAT) were conducted to detect the fluctuation of serum antibodies compared with the detection of toxoplasma by PCR and MI. Toxoplasma B1 gene PCR was shown consistently high sensitivity and the results obtained by PCR agreed completely with those from MI. All blood samples collected before infection with T gondii gave negative results by PCR and MI. Also, toxoplasma Bl gene PCR was not cross reaction with Neospora caninum DNA and normal cat leucocyte as controls. The toxoplasma-specific DNA was detected by PCR in blood of 5 cats experimentally infected with T gondii 6 days after infection and the detection of this specific-DNA was long lasted in blood for 64 days after infection. The detection of toxoplasma-specific DNA by PCR could be identified as few as 10 tachyzoites and the isolation of T gondii by MI could be isolated as few as 1 tachyzoite from tenfold serial dilution of T gondii with normal cat blood, respectively. In healthy domiciled cats, the toxoplasma-specific DNA and the parasite were detected and isolated in blood from 3 of 56(5.3%) cats by both PCR and MI, respectively. In the results of antibody test from the total 56 heads of healthy domiciled cats, the positive rates are 15(26.7%) by LAT and 19(33.9%) by IFAT. These results suggest that PCR detection of toxoplasma can be applied as a sensitive and specific diagnostic and research tool.

• Parasites, Hosts and Diseases
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• 제48권2호
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• pp.99-103
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• 2010

Malaria is endemic in the Cukurova region while it is sporadic in other regions of Turkey. Therefore, the laboratory and clinical diagnosis of malaria is important for the treatment of malaria. In this study, 92 blood samples that were taken from the suspected malaria patients for routine diagnosis in a period of 10 years between 1999 and 2009 were analyzed. All of these blood samples were examined by microscopic examinations using Giemsa-stained thick blood films, nested PCR, and real-time PCR. The sensitivity-specificity and positive-negative predictive values for these diagnostic tests were then calculated. It was found that the positive predictive values of microscopic examination of thick blood films, nested PCR, and real-time PCR were 47.8%, 56.5%, and 60.9% for malaria, respectively. The real-time PCR was found to have a specificity of 75% and sensitivity of 100%, while specificity and sensitivity of nested PCR was found 81.2% and 97.7% according to the microscopic examination of thick blood films, respectively.

• Journal of Microbiology and Biotechnology
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• 제33권11호
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• pp.1457-1466
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• 2023

Enteric fever is caused by typhoidal Salmonella serovars (Typhi, Paratyphi A, Paratyphi B, and Paratyphi C). Owing to the importance of Salmonella serovars in clinics and public hygiene, reliable diagnostics for typhoidal serovars are crucial. This study aimed to develop a novel diagnostic tool for typhoidal Salmonella serovars and evaluate the use of human blood for clinically diagnosing enteric fever. Five genes were selected to produce specific PCR results against typhoidal Salmonella serovars based on the genes of Salmonella Typhi. Heptaplex PCR, including genetic markers of generic Salmonella, Salmonella enterica subsp. enterica, and typhoidal Salmonella serovars, was developed. Typhoidal Salmonella heptaplex PCR using genomic DNAs from 200 Salmonella strains (112 serovars) provided specifically amplified PCR products for each typhoidal Salmonella serovar. These results suggest that heptaplex PCR can sufficiently discriminate between typhoidal and non-typhoidal Salmonella serovars. Heptaplex PCR was applied to Salmonella-spiked blood cultures directly and provided diagnostic results after 12- or 13.5-h blood culture. Additionally, it demonstrated diagnostic performance with colonies recovered from a 6-h blood culture. This study provides a reliable DNA-based tool for diagnosing typhoidal Salmonella serovars that may be useful in clinical microbiology and epidemiology.

• 한국가축번식학회지
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• 제27권2호
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• pp.97-102
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• 2003

본 연구는 분만시 동복 신생자돈의 제대혈에서 추출한 genomic DNA를 PCR-RFLP 기법을 이용하여 농가수입증대를 위하여 육질이 불량한 PSS 돼지를 판별하는 방법을 개발하기 위한 기초실험으로써 그 결과를 요약하면 다음과 같다. 자돈 제대에서 혈액 genomic DNA를 추출하여 PCR에 의하여 증폭된 ryanodine receptor gene 영역의 산물은 자돈의 제대혈에서 1.8kb의 길이로 증폭되었음을 확인하였다. 제대혈에서 추출된 DNA 의 PCR 증폭 단편을 가지고 Hha I 제한효소로 digest 하여준 결과에서 PSS 돼지는 Yorkshire 종에서 출현하지 않았으나, Landrace 종과 Crossbred 종에서 각각 4.76%와 7.14%로 교잡종(LYD 또는 YLD)에서 더욱 많이 출현되었다. 이상의 결과로 볼 때 분만시 신생자돈의 제대혈을 채취하여 PSS 돼지를 조기에 선발하면 스트레스 감소는 물론 혈액채취의 간편성을 기대할 수 있을 것으로 판단된다.

• 한국수정란이식학회지
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• 제11권2호
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• pp.145-150
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• 1996

This study was conducted to detect the Y-specific DNA in the blood of the female calf in bovine heterosexual production. Genomic DNAs of the freemartin were isolated from the blood and amplified with Y-chromosome specific DNA primer(l4lbp). In order to estimate the lower limit for the detection of XY cells, blood from a hull was diluted in cow blood to 0.01%. DNA sequencing on the PCR products was shown the same sequences as Y chromosome DNA of the normal cows. The Y specific DNA hand by PCR was detected all blood of female calf suspected to have bovine freemar tin syndrom and the karyotyping with freemartin blood was identified as XX / XY chimerism. Therefore, the PGR methods used in this study was very useful technique for the detection of freemartin in Ranwoo and Holstein.

• 대한화학회지
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• 제47권2호
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• pp.127-132
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• 2003

소의 타일레리아병 진단을 위해 DNA 추출과정 없이 전혈에서 바로 중합효소연쇄반응(PCR)을 통하여 T. buffeli(buffeli/orientalis/sergenti)의 16S rRNA 유전자 단편을 증폭시킨 뒤, 증폭된 DNA를 전기영동법으로 분석하는 방법을 개발하였다. 특이유전자 단편의 증폭을 위해 formamide를 사용하여 혈액세포를 용해시켰으며, 단백질의 응고를 줄이기 위해 낮은 반응온도를 사용하는 FoLT(Formamide Low Temp.) PCR법을 이용하였다. 전혈 100-200 nL를 바로 PCR 증폭에 사용하였으며, PCR 산물(816-bp DNA)은 전기영동법으로 분석하였다. 본 결과는 T. buffeli에 감염된 소의 혈액으로부터 정제된 DNA를 사용하여 얻은 실험결과와 잘 일치하였다.

• 한국동물위생학회지
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• 제34권1호
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• pp.13-18
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• 2011

The purpose of this study was to apply a nested-polymerase chain reaction (nPCR) assay to detect and differentiate PCV 2a and PCV 2b. The compared with nPCR and one-step PCR and nPCR showed more sensitive in the detection of PCV-2 from tissue and blood samples. The total of 52 tissue samples was collected from postweanning pigs from 2006 to 2010. All tissue samples showed positive for PCV-2 in one-step PCR and nPCR, followed by the nPCR in order to identify the genotypes of PCV-2. 2 samples (3.8%) showed positive for PCV 2a, and 35 samples were positive for PCV 2b (67.3%), 15 samples (28.9%) were positive the dual genotypes. In addition, 42 blood samples which were collected from the 5 different swine farms were compared figure out the detection rates of nPCR and one-step PCR. The PCV 2 was positive by one-step PCR in 21 samples (50.0%) and nPCR was positive in 37 samples (88.1%). The PCV 2 genotypes in blood samples and 32 samples (76.2%) were positive for PCV 2b and none were positive for PCV 2a, 5 samples (11.9%) were positive for dual genotypes. These results suggest that the nPCR is very efficient for genotyping blood samples and differentiating the genotypes of PCV-2 from field samples.

• 한국동물위생학회지
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• 제32권4호
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• pp.335-341
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• 2009

Canine brucellosis produce abortions and infertility in dogs and is currently diagnosed by serological methods such as rapid slide agglutination test with 2-mercaptoethanol (2-ME RSAT) and immunochromatographic assay (ICA). Bacterial isolation is considered gold standard for Brucella diagnosis and the polymerase chain reaction (PCR) is an alternative method to bacterial isolation. A total of 36 whole blood samples were collected from dogs reared in area of Chuncheon and were subjected to serology (2-ME RSAT and ICA for B. canis, Rose Bengal test and C-ELISA for B. abortus), blood culture and 3 types of PCRs (BSCP31, 16s rRNA, and OMP-2). All blood samples were negative by serology and blood cultures. The BCSP31 and the OMP-2 PCR detected 5 samples were positive whereas the 16S rRNA PCR detected all samples were negative as serological methods and blood culture did. From the results observed in the present study, we conclude that 16S rRNA PCR could be used for direct PCR for canine blood samples.

• 대한수의학회지
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• 제41권1호
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• pp.89-98
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• 2001

This study was conducted to detect the toxoplasma specific-DNA in circulating blood and organs collected from slaughtered pigs at slaughtering house and experimentally infected pigs with Toxoplasma gondii tachyzoites by polymerase chain reaction(PCR), and also PCR was applied to diagnose for acute phase of swine toxoplasmosis as a newly developed diagnostic test. The sensitivity of oligonucleotide primer, T-1 & T-2, designed from toxoplasma B1 gene amplification method was compared with Tp parasite detection by mouse inoculation(MI). On the other hand, latex agglutination test(LAT) was conducted to detect the serum antibodies comparing with the detection of toxoplasma by PCR and MI. The results obtained were summarized as follows. PCR was able to determine at the lowest level of $10^0/ml$ T. gondii in blood samples which were blended with a serial diluted T gondii in vitro. On the other hand, $10^2/5g$ of T gondii could detect from a variety of tissues including lung, diaphragm, liver, heart, spleen and brain in vitro. The primer was proved to specifically determine T gondii in blood and tissues in vitro but it did not detect Neospora caninum used as a negative control. DNA of T. gondii was effectively extracted by freezing, thawing and grinding twice both tissues mixed with T gondii in vitro and in experimentally infected pig's tissues. PCR detected specific DNA in the blood of experimentally infected pigs at 108 hrs and 120 hrs post-infection, it was the same time that the pigs showed fever and parasitaemia. In case of tissue, specific DNA was, however, detected only lung from experimentally infected pigs. Even though the duration of acute phase was from 3 to 7 days post-infection, but the latex agglutination test (LAT) results appeared from 8 days post-infection. A comparison of sensitivity in determining T gondii in blood samples between PCR and MI, PCR positive rate ranged from 25 to 33.3%, but that of MI covered from 75 to 100%.