scholarly journals Detection of canine distemper virus (CDV) through one step RT-PCR combined with nested PCR

2001 ◽  
Vol 2 (1) ◽  
pp. 59 ◽  
Author(s):  
Yong Hwan Kim ◽  
Kyu Woan Cho ◽  
Hwa Young Youn ◽  
Han Sang Yoo ◽  
Hong Ryul Han
Keyword(s):  
Nested Pcr ◽  
Canine Distemper Virus ◽  
Canine Distemper ◽  
Rt Pcr ◽  
One Step

Comparison of one-step RT-PCR and a nested PCR for the detection of canine distemper virus in clinical samples

2004 ◽  
Vol 82 (1-2) ◽  
pp. 83-86 ◽  
Author(s):  
YJ SHIN ◽  
KO CHO ◽  
HS CHO ◽  
SK KANG ◽  
HJ KIM ◽  
...  
Keyword(s):  
Nested Pcr ◽  
Canine Distemper Virus ◽  
Clinical Samples ◽  
Canine Distemper ◽  
Rt Pcr ◽  
One Step

scholarly journals One-step triplex PCR/RT-PCR to detect canine distemper virus, canine parvovirus and canine kobuvirus

2019 ◽  
Vol 81 (7) ◽  
pp. 1040-1042 ◽  
Author(s):  
Dafei LIU ◽  
Fei LIU ◽  
Dongchun GUO ◽  
Xiaoliang HU ◽  
Zhijie LI ◽  
...  
Keyword(s):  
Canine Distemper Virus ◽  
Canine Parvovirus ◽  
Canine Distemper ◽  
Rt Pcr ◽  
One Step ◽  
Canine Kobuvirus

Serological and Nucleocapsid Gene Based Molecular Characterization of Canine Distemper Virus (CDV) Isolated from Dogs of Southern Gujarat, India

2020 ◽  
Author(s):  
Dhruv Desai ◽  
Irshadullakhan Kalyani ◽  
Jayesh Solanki ◽  
Dharmesh Patel ◽  
Pushpa Makwana ◽  
...  
Keyword(s):  
Cell Line ◽  
Mdck Cell ◽  
Canine Distemper Virus ◽  
Age Group ◽  
Canine Distemper ◽  
Rt Pcr ◽  
Serum Samples ◽  
Mdck Cell Line ◽  
One Step

Background: The present study was undertaken to diagnose and characterize canine distemper virus (CDV) isolated from dogs of southern Gujarat, India. CDV is lethal disease of canines and felines. Total of 40 different samples were collected from 18 suspected stray dogs having different clinical signs which were processed for diagnosis and characterization of CDV.Methods: All samples were processed by employing different methods like, Immunochromatography based lateral flow test (LFA), IgG based indirect enzyme linked immunosorbent assay (i-ELISA), one step RT-PCR, nested one step RT-PCR and virus isolation in MDCK cell line. Restriction endonuclease (RE) analysis was used to characterize CDV Nucleocapsid (N) gene. Conclusion: Only 04 samples (02 nasal and 02 ocular swabs) of 02 dogs found positive for LFA, while 14 serum samples out of 17 samples of 18 dogs found positive for IgG antibody. As all dogs were unvaccinated, serum samples found positive in IgG based ELISA considered for confirmative positive for CDV infection. Whereas 13 samples of 10 dogs found positive for one step RT-PCR and nested one step RT-PCR. In RE digestion, characteristic two bands were found. All representative CDV positive samples of 10 dogs showed characteristic cytopathic effect in MDCK cell line. On age group wise percent positivity was found 71.42 % (05/07) in 0 - ≤6 months, while 77.77% (07/09) in 6- ≤12 months of age group, whereas, both samples were found positive in 12 months and above group. Overall 77.77% (14/18) dogs found positive for CDV infection. To the best of our knowledge, this is the first report on study of CDV infection in dogs from Gujarat state, India.

scholarly journals A New Molecular Detection System for Canine Distemper Virus Based on a Double-Check Strategy

Viruses ◽  
2021 ◽  
Vol 13 (8) ◽  
pp. 1632
Author(s):  
Sabrina Halecker ◽  
Sabine Bock ◽  
Martin Beer ◽  
Bernd Hoffmann
Keyword(s):  
Canine Distemper Virus ◽  
Parallel Implementation ◽  
Detection System ◽  
Quantitative Polymerase Chain Reaction ◽  
Diagnostic Methods ◽  
Molecular Diagnostic ◽  
Wild Animals ◽  
Canine Distemper ◽  
One Step ◽  
Diagnostic Sensitivity And Specificity

Due to changing distemper issues worldwide and to inadequate results of an inter-laboratory study in Germany, it seems sensible to adapt and optimize the diagnostic methods for the detection of the canine distemper virus (CDV) to the new genetic diversity of virus strains. The goal of the project was the development, establishment and validation of two independent one-step reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) methods for the safe detection of CDV in domestic and wild animals. For this purpose, an existing CDV-RT-qPCR was decisively adapted and, in addition, a completely new system was developed. Both CDV-RT-qPCR systems are characterized by a very high, comparable analytical and diagnostic sensitivity and specificity and can be mutually combined with inhibition or extraction controls. The reduction in the master mix used allows for the parallel implementation of both CDV-RT-qPCR systems without significant cost increases. For validation of the new CDV-RT-qPCR duplex assays, a panel comprising 378 samples derived from Germany, several European countries and one African country were tested. A sensitivity of 98.9% and a specificity of 100% were computed for the new assays, thus being a reliable molecular diagnostic tool for the detection of CDV in domestic and wild animals.

scholarly journals Canine distemper virus detection by different methods of One-Step RT-qPCR

2016 ◽  
Vol 46 (9) ◽  
pp. 1601-1606
Author(s):  
Claudia de Camargo Tozato ◽  
Vívian Ferreira Zadra ◽  
Caroline Rodrigues Basso ◽  
João Pessoa Araújo Junior
Keyword(s):  
Urine Sample ◽  
Canine Distemper Virus ◽  
Limit Of Detection ◽  
Analytical Sensitivity ◽  
Canine Distemper ◽  
System A ◽  
Virus Rna ◽  
One Step ◽  
Commercial Kits ◽  
The One

ABSTRACT: Three commercial kits of One-Step RT-qPCR were evaluated for the molecular diagnosis of Canine Distemper Virus. Using the kit that showed better performance, two systems of Real-time RT-PCR (RT-qPCR) assays were tested and compared for analytical sensitivity to Canine Distemper Virus RNA detection: a One-Step RT-qPCR (system A) and a One-Step RT-qPCR combined with NESTED-qPCR (system B). Limits of detection for both systems were determined using a serial dilution of Canine Distemper Virus synthetic RNA or a positive urine sample. In addition, the same urine sample was tested using samples with prior centrifugation or ultracentrifugation. Commercial kits of One-Step RT-qPCR assays detected canine distemper virus RNA in 10 (100%) urine samples from symptomatic animals tested. The One-Step RT-qPCR kit that showed better results was used to evaluate the analytical sensitivity of the A and B systems. Limit of detection using synthetic RNA for the system A was 11 RNA copies µL-1 and 110 RNA copies µl-1 for first round System B. The second round of the NESTED-qPCR for System B had a limit of detection of 11 copies µl-1. Relationship between Ct values and RNA concentration was linear. The RNA extracted from the urine dilutions was detected in dilutions of 10-3 and10-2 by System A and B respectively. Urine centrifugation increased the analytical sensitivity of the test and proved to be useful for routine diagnostics. The One-Step RT-qPCR is a fast, sensitive and specific method for canine distemper routine diagnosis and research projects that require sensitive and quantitative methodology.

scholarly journals Rapid and Sensitive Detection of Noroviruses by Using TaqMan-Based One-Step Reverse Transcription-PCR Assays and Application to Naturally Contaminated Shellfish Samples

2005 ◽  
Vol 71 (4) ◽  
pp. 1870-1875 ◽  
Author(s):  
Narayanan Jothikumar ◽  
James A. Lowther ◽  
Kathleen Henshilwood ◽  
David N. Lees ◽  
Vincent R. Hill ◽  
...  
Keyword(s):  
Reverse Transcription ◽  
Nested Pcr ◽  
Rt Pcr ◽  
Reverse Transcription Pcr ◽  
Specificity And Sensitivity ◽  
Routine Monitoring ◽  
The Family ◽  
One Step ◽  
Stool Specimens ◽  
Pcr Assays

ABSTRACT Noroviruses (NoV), which are members of the family Caliciviridae, are the most important cause of outbreaks of acute gastroenteritis worldwide and are commonly found in shellfish grown in polluted waters. In the present study, we developed broadly reactive one-step TaqMan reverse transcription (RT)-PCR assays for the detection of genogroup I (GI) and GII NoV in fecal samples, as well as shellfish samples. The specificity and sensitivity of all steps of the assays were systematically evaluated, and in the final format, the monoplex assays were validated by using RNA extracted from a panel of 84 stool specimens, which included NoV strains representing 19 different genotypes (7 GI, 11 GII, and 1 GIV strains). The assays were further validated with 38 shellfish cDNA extracts previously tested by nested PCR. Comparison with a recently described real-time assay showed that our assay had significantly higher sensitivity and was at least as sensitive as the nested PCR. For stool specimens, a one-step duplex TaqMan RT-PCR assay performed as well as individual genogroup-specific monoplex assays. All other enteric viruses examined were negative, and no cross-reaction between genogroups was observed. These TaqMan RT-PCR assays provide rapid (less than 90 min), sensitive, and reliable detection of NoV and should prove to be useful for routine monitoring of both clinical and shellfish samples.

scholarly journals Molecular analysis of the N gene of canine distemper virus in dogs in Brazil

2007 ◽  
Vol 59 (3) ◽  
pp. 654-659 ◽  
Author(s):  
J.G. Castilho ◽  
P.E. Brandão ◽  
P. Carnieli Jr ◽  
R.N. Oliveira ◽  
C.I. Macedo ◽  
...  
Keyword(s):  
Central Nervous System ◽  
Phylogenetic Analysis ◽  
Nervous System ◽  
Molecular Analysis ◽  
Canine Distemper Virus ◽  
Nucleotide Identity ◽  
N Gene ◽  
Canine Distemper ◽  
Gene Sequences ◽  
Rt Pcr

Eleven central-nervous-system samples collected from stray dogs between 2000 and 2004 were found positive by RT-PCR, which amplified a 480bp fragment of the N gene of canine distemper virus (CDV). Phylogenetic analysis based on partial N-gene sequences showed four major clusters. All dog strains segregated into cluster I, with a mean nucleotide identity of 95.8% and 95.6% with the Onderstepoort and Lederle vaccine strains, respectively. Cluster II contained all the raccoon-related strains, cluster III Orient strains and Cluster IV the Onderstepoort and Lederle vaccine strains, with a mean nucleotide identity of 99.7% between them. This is the first report of phylogenetic analysis of CDV strains in Brazil.

scholarly journals DETEKSI VIRUS DENGUE SEROTIPE-3 DAN PERAN SPERMATEKA DALAM PENULARAN SECARA TRANSVENEREAL PADA NYAMUK Aedes aegypti BETINA

2019 ◽  
Vol 12 (1) ◽  
pp. 130
Author(s):  
Devita Febriani Putri ◽  
Tusy Triwahyuni
Keyword(s):  
Aedes Aegypti ◽  
Nested Pcr ◽  
Rt Pcr ◽  
One Step

Penularan transvenereal berpotensi menyebarkan virus dengue melalui perilaku kawin. Pengendalian vektor Demam Berdarah Dengue (DBD) dengan strategi perilaku kawin nyamuk secara alami telah diterapkan untuk menurunkan perluasan daerah endemis DBD. Dengan dasar tersebut, pemahaman perilaku kawin nyamuk Ae. aegypti penting untuk diketahui. Penelitian ini bertujuan untuk mendeteksi virus dengue serotipe 3 (DENV-3) pada organ spermateka nyamuk Ae. aegypti betina yang telah terinfeksi DENV-3 secara transvenereal di laboratorium. Pembedahan organ spermateka pada nyamuk betina dilakukan setelah nyamuk Ae. aegypti betina kawin dengan nyamuk Ae. aegypti jantan yang positif DENV-3. Keberadaan DENV-3 pada organ spermateka nyamuk betina dilakukan dengan melakukan pengujian pooling sampel menggunakan metode One-Step RT-PCR untuk screening virus dengue (profil pita DNA spesifik 511 bp). Sampel yang hasil pengujiannya positif virus dengue, dilanjutkan dengan metode Semi-Nested PCR untuk serotyping DENV-3 (profil pita DNA spesifik 290 bp). Hasil penelitian dari 7 sampel pooling organ spermateka dari nyamuk betina positif DENV-3 hasil penularan transvenereal nyamuk jantan positif DENV-3 secara intratorakal menunjukkan tidak ada satupun sampel yang terdeteksi adanya DENV-3. Tidak ditemukan virus DENV-3 pada organ spermateka nyamuk Ae. aegypti betina yang telah terinfeksi DENV-3 secara transvenereal pada 7 sampel yang digunakan. Perlu pengujian lebih lanjut pada organ ovarium nyamuk betina untuk memastikan mekanisme terjadinya penularan transvenereal virus dengue pada Ae. aegypti dalam upaya mencari strategi baru dalam pengendalian vektor DBD

scholarly journals Rapid Detection and Molecular Typing of Dengue Virus by Using Multiplex-Nested-RT-PCR

2015 ◽  
Vol 11 (2) ◽  
Author(s):  
Nastiti Wijayanti ◽  
Hera Nirwati ◽  
Tri Wibawa ◽  
Aris Haryanto ◽  
S. Sutaryo
Keyword(s):  
Dengue Virus ◽  
Nested Pcr ◽  
Genotypic Variation ◽  
Clinical Samples ◽  
Rt Pcr ◽  
Virus Serotype ◽  
Pcr Multiplex ◽  
Pcr Products ◽  
One Step ◽  
Nested Pcr Assays

world. We have evaluated the combination of one-step RT-PCR and multiplex nested PCR assays for detectingdengue viruses from clinical samples. Twelve patients were screened for the dengue virus, using a pair of primersthat conserve for several Flavivirus. The results showed that in 12 suspect patients, 100% were positive for Flavivirusand there are some genotypic variation among them, that indicated by several RT-PCR products higher than 511 bp,the expected product for RT-PCR. Further assay was performed to clarify the presence and serotypes of dengue virususing multiplex nested PCR. Serotyping results indicated that 83,3% of samples can be confirmed for dengue virus.Among the dengue virus positive 16,7 % are dengue-2, 16.7 % are dengue-3, and the most common 50% are dengue-4,whereas dengue-1 were not found among the patients. The combination of RT-PCR and multiplex nested PCR assaycan be used for rapid analysis dengue samples in early phase which is potentially useful for clinical, epidemiologyand also evolutionary studies.Key words: Flavivirus, dengue virus, serotype, RT-PCR, multiplex nested PCR

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