scholarly journals Evaluation of a novel TaqMan probe-based real-time polymerase chain reaction (PCR) assay for detection and quantitation of red sea bream iridovirus

2021 ◽  
Vol 24 (11) ◽  
pp. 351-359
Author(s):  
Guk Hyun Kim ◽  
Min Jae Kim ◽  
Hee Ju Choi ◽  
Min Ji Koo ◽  
Min Jeong Kim ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Red Sea ◽  
Sea Bream ◽  
Taqman Probe ◽  
Red Sea Bream ◽  
Chain Reaction ◽  
Pcr Assay ◽  
Polymerase Chain

scholarly journals Rapid diagnosis of red sea bream iridovirus infection using the polymerase chain reaction

1998 ◽  
Vol 32 ◽  
pp. 87-90 ◽  
Author(s):  
S Oshima ◽  
JI Hata ◽  
N Hirasawa ◽  
T Ohtaka ◽  
I Hirono ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Red Sea ◽  
Rapid Diagnosis ◽  
Sea Bream ◽  
Red Sea Bream ◽  
Chain Reaction ◽  
Polymerase Chain

scholarly journals Deteksi Spesies Megalocytivirus pada Budidaya Ikan Laut di Kepulauan Riau

2019 ◽  
Vol 13 (3) ◽  
pp. 265-274
Author(s):  
Achmad Bahtiar Rifai ◽  
Anak Agung Gede ◽  
Lucky Putra P Kusuma ◽  
Yani Maulani
Keyword(s):  
Polymerase Chain Reaction ◽  
Capsid Protein ◽  
Red Sea ◽  
Major Capsid Protein ◽  
Sea Bream ◽  
Red Sea Bream ◽  
Necrosis Virus ◽  
Chain Reaction ◽  
Polymerase Chain

Penyakit yang dapat menyebabkan kerugian ekonomi yang sangat tinggi adalah virus dan salah satunya adalah Megalocytivirus. Megalocytivirus mempunyai tiga spesies yaitu Infectious Spleen and Kidney Necrosis Virus (ISKNV), Red Sea Bream Iridovirus (RSIV), dan Turbot Redy Body Iridovirus (TRBIV). Gejala klinis pada ikan yang terinfeksi Megalocytivirus pada umumnya terlihat dari perubahan tingkah laku yaitu nafsu makan berkurang, berenang lemah, dan malas bergerak. Tubuh ikan yang terinfeksi terlihat lebih gelap baik pada permukaan tubuh sirip, dan bagian ekor. Metode uji yang digunakan pada penelitian ini adalah pengambilan sampel, pengujian histopatologi, deteksi spesies Megalocytivirus dengan polymerase chain reaction (PCR). Sampel ikan yang menunjukan gejala klinis diambil dari beberapa kabupaten/kota yang ada di kepulauan Riau antara lain Batam, Tanjungpinang, Bintan, Tanjung Balai Karimun dan Natuna. Sampel ikan adalah ikan kakap dan kerapu dari keramba jaring apung dan hatchery. Tujuan dari penelitian ini adalah mendeteksi spesies Megalocytivirus pada budidaya ikan laut yang ada di Kepulauan Riau. Pada uji histopatologi ditemukan lesio berupa badan inklusi, nekrosa, melano makrofag center pada hati, ginjal dan limpa. Badan inklusi dan nekrosa yang menyebar pada organ tersebut merupakan karakteristik Megalocytivirus. Hasil identifikasi PCR dengan menggunakan gen Major Capsid Protein pada panjang amplikon 777 bp berhasil mendeteksi Megalocytivirus. deteksi spesies Megalocytivirus dibeberapa Kabupaten/Kota di Kepulauan Riau menunjukan semua sampel ikan air laut terinfeksi ISKNV, sedangkan untuk spesies RSIV dan TRBIV tidak berhasil dideteksi pada penelitian ini. Infeksi Mengalocytivirus sudah menyebar pada budidaya ikan laut diseluruh kabupaten/Kota di Propinsi Kepulauan Riau. Spesies Megalocytivirus yang terdeteksi di Kepulauan Riau adalah ISKNV.

scholarly journals Polymerase Chain Reaction (PCR) Amplification of DNA of Red Sea Bream Iridovirus (RSIV)

1998 ◽  
Vol 33 (1) ◽  
pp. 17-23 ◽  
Author(s):  
Jun Kurita ◽  
Kazuhiro Nakajima ◽  
Ikuo Hirono ◽  
Takashi Aoki
Keyword(s):  
Polymerase Chain Reaction ◽  
Red Sea ◽  
Pcr Amplification ◽  
Sea Bream ◽  
Red Sea Bream ◽  
Chain Reaction ◽  
Polymerase Chain

scholarly journals Development of Real-Time Polymerase Chain Reaction Assay with TaqMan Probe for the Quantitative Detection of Infectious Hypodermal and Hematopoietic Necrosis Virus from Shrimp

2006 ◽  
Vol 89 (1) ◽  
pp. 240-244 ◽  
Author(s):  
Zhi-Qin Yue ◽  
Hong Liu ◽  
Wei-Ji Wang ◽  
Zhi-Wen Lei ◽  
Cheng-Zhu Liang ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Real Time Pcr ◽  
Taqman Probe ◽  
Quantitative Detection ◽  
Necrosis Virus ◽  
Chain Reaction ◽  
Pcr Assay ◽  
The Real ◽  
Polymerase Chain

Abstract An assay was developed for the detection of infectious hypodermal and hematopoietic necrosis virus (IHHNV) based on real-time quantitative polymerase chain reaction (PCR). A pair of primers and a TaqMan probe were designed that are specific for the recognition of a conservative region in the IHHNV genome. The IHHNV real-time PCR assay had a detection limit of 9 DNA copies,with a dynamic range of detection between 9 106 and 9 DNA copies. The primer pairs and probe were specific to IHHNV and did not cross-reactwith shrimp genomic DNAor other shrimp viruses such as White Spot Syndrome Virus (WSSV), Monodon Baculovirus (MBV), and hepatopancreatic parvovirus (HPV). This assay has a broad application for basic and clinical investigations. For clinical samples, the real-time PCR assay detected all the positive samples screened by conventional PCR, which indicated the sensitivity of the real-time assay. The IHHNV real-time PCR assay with high sensitivity, specificity, wide range of detection ability, and simplicity is particularly useful for screening large numbers of specimens and measuring viral loads to monitor the broodstock.

scholarly journals A Quantitative Real-time Polymerase Chain Reaction Assay for Botrytis aclada in Onion Bulb Tissue

HortScience ◽  
2008 ◽  
Vol 43 (2) ◽  
pp. 408-413 ◽  
Author(s):  
Timothy W. Coolong ◽  
Ronald R. Walcott ◽  
William M. Randle
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Real Time Pcr ◽  
Taqman Probe ◽  
Chain Reaction ◽  
Pcr Assay ◽  
Polymerase Chain ◽  
And Storage ◽  
Assay Detection

A real-time polymerase chain reaction (PCR) assay has been developed for the detection and quantification of Botrytis aclada (Fresenius), a causal agent of neck rot in onion (Allium cepa L.) bulbs. The assay uses TaqMan probe-based chemistry to detect an amplicon from the L45-550 region of B. aclada while using a DNA sequence from the onion serine acetyl transferase gene (SAT1) as a control. The assay detection limits for B. aclada and onion were 10 pg·μL−1 of genomic DNA. The detection limit for lyophilized B. aclada mycelium was 1 μg. The presence of onion tissue in the samples did not affect the performance of the real-time PCR assay. The assay distinguished among different amounts of B. aclada mycelium growing on onion disks that were inoculated with 0, 102, or 104 B. aclada conidia. Visual observations during the incubation period corresponded with changes in real-time PCR results. This assay could be used to determine the amount of B. aclada mycelium in bulbs during growth, harvest, and storage, thus giving researchers an objective and efficient tool by which to quantify the growth rate and virulence of B. aclada strains in vivo.

scholarly journals Analysis and validation of a highly sensitive one-step nested quantitative real-time polymerase chain reaction assay for specific detection of severe acute respiratory syndrome coronavirus 2

2020 ◽  
Vol 17 (1) ◽  
Author(s):  
Yang Zhang ◽  
Chunyang Dai ◽  
Huiyan Wang ◽  
Yong Gao ◽  
Tuantuan Li ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Quantitative Polymerase Chain Reaction ◽  
Clinical Samples ◽  
Chain Reaction ◽  
Pcr Assay ◽  
Qrt Pcr ◽  
Highly Sensitive ◽  
One Step ◽  
Polymerase Chain

Abstract Background Coronavirus disease 2019 (COVID-19), caused by SARS-CoV-2, is posing a serious threat to global public health. Reverse transcriptase real-time quantitative polymerase chain reaction (qRT-PCR) is widely used as the gold standard for clinical detection of SARS-CoV-2. Due to technical limitations, the reported positive rates of qRT-PCR assay of throat swab samples vary from 30 to 60%. Therefore, the evaluation of alternative strategies to overcome the limitations of qRT-PCR is required. A previous study reported that one-step nested (OSN)-qRT-PCR revealed better suitability for detecting SARS-CoV-2. However, information on the analytical performance of OSN-qRT-PCR is insufficient. Method In this study, we aimed to analyze OSN-qRT-PCR by comparing it with droplet digital PCR (ddPCR) and qRT-PCR by using a dilution series of SARS-CoV-2 pseudoviral RNA and a quality assessment panel. The clinical performance of OSN-qRT-PCR was also validated and compared with ddPCR and qRT-PCR using specimens from COVID-19 patients. Result The limit of detection (copies/ml) of qRT-PCR, ddPCR, and OSN-qRT-PCR were 520.1 (95% CI: 363.23–1145.69) for ORF1ab and 528.1 (95% CI: 347.7–1248.7) for N, 401.8 (95% CI: 284.8–938.3) for ORF1ab and 336.8 (95% CI: 244.6–792.5) for N, and 194.74 (95% CI: 139.7–430.9) for ORF1ab and 189.1 (95% CI: 130.9–433.9) for N, respectively. Of the 34 clinical samples from COVID-19 patients, the positive rates of OSN-qRT-PCR, ddPCR, and qRT-PCR were 82.35% (28/34), 67.65% (23/34), and 58.82% (20/34), respectively. Conclusion In conclusion, the highly sensitive and specific OSN-qRT-PCR assay is superior to ddPCR and qRT-PCR assays, showing great potential as a technique for detection of SARS-CoV-2 in patients with low viral loads.

A Duplex SYBR Green I-based Real-time Polymerase Chain Reaction Assay for Rapid Detection of Canine Kobuvirus and Canine Circovirus

2021 ◽  
Author(s):  
Yang Pan ◽  
Jing Chen ◽  
Junhuang Wu ◽  
Yongxia Wang ◽  
Junwei Zou ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Rapid Detection ◽  
Simultaneous Detection ◽  
Sybr Green ◽  
Sybr Green I ◽  
Chain Reaction ◽  
Pcr Assay ◽  
Polymerase Chain ◽  
Canine Kobuvirus

Abstract Background: Canine Kobuvirus (CaKoV) and Canine Circovirus (CaCV) are viruses that infect dogs causing diarrheal symptoms that are very similar. However, there is no clinical method to detect a co-infection of these two viruses.Results: In this study, a duplex SYBR Green I-based quantitative real-time polymerase chain reaction (PCR) assay for the rapid and simultaneous detection of CaKoV and CaCV was established. CaKoV and CaCV were distinguished by their different melting temperature which was 86℃ for CaKoV and 78℃ for CaCV. The assay was highly specific, with no cross-reactivity with other common canine viruses and demonstrated high sensitivity. The detection limits of CaKoV and CaCV were 8.924 × 101 copies/μL and 3.841 × 101 copies/μL, respectively. The highest intra- and inter-assay Ct value variation coefficients (CV) of CaKoV were 0.40% and 0.96%, respectively. For CaCV, the highest intra- and inter-assay Ct value variation coefficients were 0.26% and 0.70%, respectively. In 57 clinical samples, positive detection rates of CaKoV and CaCV were 8.77% (7/57) and 15.79% (9/57), respectively. The co-infection rate was 7.02% (4/57). Conclusions: The duplex SYBR Green I-based real-time PCR assay established in this study is a fast, efficient, and sensitive method for the simultaneous detection of the two viruses and provides a powerful tool for the rapid detection of CaKoV and CaCV in clinical practice.

scholarly journals Novel genotype ofTritrichomonas foetusfrom cattle in Southern Africa

Parasitology ◽  
2016 ◽  
Vol 143 (14) ◽  
pp. 1954-1959 ◽  
Author(s):  
ANDREA CASTERIANO ◽  
UMBERTO MOLINI ◽  
KORNELIA KANDJUMBWA ◽  
SIEGFRIED KHAISEB ◽  
CAROLINE F. FREY ◽  
...  
Keyword(s):  
Real Time ◽  
Cysteine Proteases ◽  
Taqman Probe ◽  
Tritrichomonas Foetus ◽  
Chain Reaction ◽  
Pcr Assay ◽  
New Genotype ◽  
Reproductive Disease ◽  
Polymerase Chain ◽  
Novel Genotype

SUMMARYBovine trichomonosis caused byTritrichomonas foetusis a significant reproductive disease of cattle. Preputial samples were collected using sheath washing technique in bulls in Namibia. Thirty-six trichomonad cultures were characterized using the TaqMan-probe commercial real-time polymerase chain reaction (PCR) diagnostic assay (VetMAX™-Gold Trich Detection Kit) and CYBR real-time PCR assay based on TFR3/4 primers. Diagnostic real-time PCRs and DNA sequencing of the internal transcribed region confirmed presence ofT. foetusin 35 out of 36 samples. Multilocus genotyping using cysteine proteases (CP1, CP2, CP4, CP5, CP6, CP7, CP8, CP9) and malate dehydrogenase (MDH1) gene sequences demonstrate that theT. foetusin Namibia are genetically distinct from those characterized elsewhere. We report the discovery of a novel genotype ofT. foetusin Namibian cattle, distinct from otherT. foetusgenotypes in Europe, South and North America and Australia. We suggest recognition of a ‘Southern African’ genotype ofT. foetus. Identification of the new genotype ofT. foetusdemonstrates the need for wider global sampling to fully understand the diversity and origin ofT. foetuscausing disease in cattle or cats.

scholarly journals Collaborative Trial for Validation of a Real-Time Reverse Transcriptase-Polymerase Chain Reaction Assay for Detection of Central Nervous System Tissues as Bovine Spongiform Encephalopathy RiskMaterial: Part 1

2006 ◽  
Vol 89 (5) ◽  
pp. 1335-1340
Author(s):  
Amir Abdulmawjood ◽  
Holger Schnenbrcher ◽  
Michael BÜlte
Keyword(s):  
Central Nervous System ◽  
Polymerase Chain Reaction ◽  
Real Time ◽  
Meat Products ◽  
Spongiform Encephalopathy ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Pcr Assay ◽  
Heat Treated ◽  
Polymerase Chain

Abstract A collaborative trial was conducted to evaluate a real-time reverse transcriptase-polymerase chain reaction (RT-PCR) assay for detection of central nervous system (CNS) tissues in meat products (e.g., sausages). The method is based on the detection of ruminant glial fibrillary acidic protein (GFAP) mRNA by applying real-time RT-PCR. The assay was evaluated through a multicenter trial involving 12 participating laboratories that received coded cDNA obtained from 3 different types of sausages. The participants used 5 different real-time detection systems. The results obtained in this validation revealed that this real-time RT-PCR assay performed well in the different laboratories with a detection limit of at least 0.1% CNS in those test materials that contained strongly heat-treated samples (sausages cooked at 120C) and the medium heat-treated samples (sausages cooked at 80C). The detection limit of liver sausages was determined to be 0.2% of CNS. Neither the samples with no CNS additive nor the bovine DNA and the negative control containing 100% swine brain gave any positive signals. The presented results indicate that the real-time RT-PCR assay was just as reproducible between laboratories, as repeatable within a laboratory, could reliably be used for detection of bovine spongiform encephalopathy risk material in meat and meat products, and signify that it may be used with confidence in any laboratory.

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