Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Detecting Newcastle disease virus in combination of RT-PCR with red blood cell absorption.

ABSTRACT: Reverse transcription-polymerase chain reaction (RT-PCR) has limited sensitivity when treating complicated samples, such as feces, waste-water in farms, and nucleic acids, protein rich tissue samples, all the factors may interfere with the sensitivity of PCR test or generate false results. In this study, we developed a sensitive RT-PCR, combination of red blood cell adsorption, for detecting Newcastle disease virus (NDV). One pair of primers which was highly homologous to three NDV pathotypes was designed according to the consensus nucleocapsid protein (NP) gene sequence. To eliminate the interfere of microbes and toxic substances, we concentrated and purified NDV from varied samples utilizing the ability of NDV binding red blood cells (RBCs). The RT-PCR coupled with red blood cell adsorption was much more sensitive in comparison with regular RT-PCR. The approach could also be used to detect other viruses with the property of hemagglutination, such as influenza viruses.

SUBMITTER: Yi J

PROVIDER: S-EPMC3096100 | biostudies-literature | 2011

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Detecting Newcastle disease virus in combination of RT-PCR with red blood cell absorption.

Yi Jianzhong J Liu Chengqian C

Virology journal 20110502

Reverse transcription-polymerase chain reaction (RT-PCR) has limited sensitivity when treating complicated samples, such as feces, waste-water in farms, and nucleic acids, protein rich tissue samples, all the factors may interfere with the sensitivity of PCR test or generate false results. In this study, we developed a sensitive RT-PCR, combination of red blood cell adsorption, for detecting Newcastle disease virus (NDV). One pair of primers which was highly homologous to three NDV pathotypes wa ...[more]

PMID: 21535888

Similar Datasets

Development of strand-specific real-time RT-PCR to distinguish viral RNAs during Newcastle disease virus infection.

Project description:Newcastle disease virus (NDV) causes large losses in the global fowl industry. To better understand NDV replication and transcription cycle, quantitative detection methods for distinguishing NDV genomic RNA (gRNA), antigenomic RNA (cRNA), and messenger RNA (mRNA) in NDV-infected cells are indispensible. Three reverse transcription primers were designed to specifically target the nucleoprotein (NP) region of gRNA, cRNA, and NP mRNA, and a corresponding real-time RT-PCR assay was developed to simultaneously quantify the three types of RNAs in NDV-infected cells. This method showed very good specificity, sensitivity, and reproducibility. The detection range of the assay was between 5.5 × 10(2) and 1.1 × 10(9) copies/?L of the target gene. These methods were applied to investigate the dynamics of the gRNA, cRNA, and mRNA synthesis in NDV La Sota infected DF-1 cells. The results showed that the copy numbers of viral gRNA, cRNA, and NP mRNA all exponentially increased in the beginning. The viral RNA copy number then plateaued at 10'h postinfection and gradually decreased from 16 h postinfection. No synthesis priority was observed between replication (gRNA and cRNA amounts) and transcription (mRNA amounts) during NDV infection. However, the cRNA accumulated more rapidly than gRNA, as the cRNA copy number was three- to tenfold higher than gRNA starting from 2 h postinfection. Conclusion. A real-time RT-PCR for absolute quantitation of specific viral RNA fragments in NDV-infected cells was developed for the first time. The development of this assay will be helpful for further studies on the pathogenesis and control strategies of NDV.

| S-EPMC4212552 | biostudies-literature

Degenerate primers based RT-PCR for rapid detection and differentiation of airborne chicken Newcastle disease virus in chicken houses.

Project description:Airborne Newcastle disease (ND) viruses in the air of five chicken houses were detected and differentiated by reverse transcriptase polymerase chain reaction (RT-PCR) using degenerate primers. Fifteen air samples were collected with All Glass Impinger-30 (AGI-30) air samplers in each house. Airborne ND viruses were also isolated and virulence identified by in vivo tests. Avirulent viruses were detected both in air samples and swab samples in four houses by degenerate primers based RT-PCR. Virulent viruses were detected only in the air samples by degenerate primers based RT-PCR in two houses. Seven strains viruses were isolated from the RT-PCR positive air samples. Of the seven strains, three strains were virulent viruses and four strains were avirulent viruses identified by in vivo tests. The results showed that it was feasible to detect and differentiate NDV in the air samples using degenerate primers based RT-PCR. This technique could decrease the time it required identify NDV infected flocks while distinguishing between virulent and avirulent viruses. It will help effectively to control Newcastle disease.

| S-EPMC7112938 | biostudies-literature

A duplex real-time RT-PCR assay for detecting H5N1 avian influenza virus and pandemic H1N1 influenza virus.

Project description:A duplex real-time reverse transcriptase polymerase chain reaction (RT-PCR) assay was improved for simultaneous detection of highly pathogenic H5N1 avian influenza virus and pandemic H1N1 (2009) influenza virus, which is suitable for early diagnosis of influenza-like patients and for epidemiological surveillance. The sensitivity of this duplex real-time RT-PCR assay was 0.02 TCID50 (50% tissue culture infective dose) for H5N1 and 0.2 TCID50 for the pandemic H1N1, which was the same as that of each single-target RT-PCR for pandemic H1N1 and even more sensitive for H5N1 with the same primers and probes. No cross reactivity of detecting other subtype influenza viruses or respiratory tract viruses was observed. Two hundred and thirty-six clinical specimens were tested by comparing with single real-time RT-PCR and result from the duplex assay was 100% consistent with the results of single real-time RT-PCR and sequence analysis.

| S-EPMC2892456 | biostudies-literature

Newcastle Disease virus infection in chickens

Project description:Airway epithelial cells from 3 different breeds of chicken infected with Newcastle Disease virus were sequenced and compared to cells from uninfected control birds. The 3 breeds were an indigenous breed, commercial Rhode Island Reds and a hybrid breed (Kenbro).

2015-01-19 | E-MTAB-3068 | biostudies-arrayexpress

The use of RT-PCR in the diagnosis and differentiation of vaccine strains of chicken infectious bronchitis and Newcastle disease.

Project description:BackgroundInfectious diseases of young and adult birds with respiratory syndrome are a significant deterrent to the development of industrial poultry farming due to decreased productivity and significant mortality. The only effective method of combating viral diseases is timely and targeted vaccination, which largely depends on laboratory diagnostic results.AimThis article aims to study the real-time reverse transcription polymerase chain reaction, (RT-PCR) which has the prospect of more effective diagnosis of vaccine strains of chicken infectious bronchitis and Newcastle disease.MethodsThe fastest and most accurate method for the differential diagnosis of pathogens in an associative viral infection is RT-PCR. The method proposed in the article for selecting primers for amplification made it possible to use this method for the simultaneous interspecies differential diagnosis of two or more viral agents, significantly accelerating their diagnosis.ResultsThe correlation of the nucleotide sequence obtained from sequencing to a specific virus strain is complicated by the lack of a single nomenclature mechanism for separating genetic groups.ConclusionThe results of this study will allow easy and fast typing of sequences into known and databased virus strains and avoid further confusion in the nomenclature of genetic groups in the future.

| S-EPMC10399655 | biostudies-literature

FMD Virus RT-PCR Control Samples

Project description:This study involved utilization of a DNA Plasmid that contained a full-length FMDV genome. The DNA plasmid was grown in bacteria before ultra-deep sequencing on a Illumina GAII. The Plasmid was then PCR amplified and again sequenced on a GAII, to asses the impact of PCR on the amount of observed mutations in the sequence data. The Plasmid was then transcribed, then reverse transcribed and PCR amplified before sequenced on a GAII, to asses the impact of RT on the amount of observed mutations in the sequence data. These control samples essentially allow the impact of sample processing on sequencing errors to be assessed in terms of viral quasispecies analyses.

| PRJEB8601 | ENA

Transposon mutagenesis screening of Newcastle disease virus (NDV)

Project description:We report the genetic plasticity of Newcastle disease virus. We introduced insertional mutation in the virus genome and checked fitness by comparing distribution of mutants in passage 1 and passage 2.

2019-10-04 | GSE138366 | GEO

Combination of RT-PCR and proteomics for the identification of Crimean-Congo hemorrhagic fever virus in ticks.

Project description:Crimean-Congo hemorrhagic fever (CCHF) is an emerging tick-borne zoonotic disease caused by the CCHF virus (CCHFV). In this study, an experimental approach combining RT-PCR and proteomics was used for the identification and characterization of CCHFV in 106 ticks from 7 species that were collected from small ruminants in Greece. The methodological approach included an initial screening for CCHFV by RT-PCR followed by proteomics analysis of positive and control negative tick samples. This novel approach allowed the identification of CCHFV-positive ticks and provided additional information to corroborate the RT-PCR findings using a different approach. Two ticks, Dermacentor marginatus and Haemaphysalis parva collected from a goat and a sheep, respectively were positive for CCHFV. The sequences for CCHFV RNA segments S and L were characterized by RT-PCR and proteomics analysis of tick samples, respectively. These results showed the possibility of combining analyses at the RNA and protein levels using RT-PCR and proteomics for the characterization of CCHFV in ticks. The results supported that the CCHFV identified in ticks are genetic variants of the AP92 strain. Although the AP92-like strains probably do not represent a high risk of CCHF to the population, the circulation of genetically diverse CCHFV strains could potentially result in the appearance of novel viral genotypes with increased pathogenicity and fitness.

| S-EPMC5508474 | biostudies-literature

Development of a New RT-PCR with Multiple Primers for Detecting Southern African Territories Foot-and-mouth Disease Viruses.

Project description:Introduction:The extremely high genetic variation and the continuously emerging variants of foot-and-mouth disease virus (FMDV) of Southern African Territory (SAT) serotypes including SAT1, SAT2, and SAT3 make it necessary to develop a new RT-PCR for general use for monitoring viruses based on the updated genome information. Material and Methods:A FMDV SAT-D8 one-step RT-PCR was established based on the 1D2A2B genes of the SAT serotype viruses with a multiplex primer set. FMDV A, O, C, and Asia 1 serotypes, other vesicular disease viruses, inactivated SAT viruses, and 125 bovine, ovine, caprine and porcine tissue samples collected from the Chinese mainland were included for evaluating the assay. Results:The new RT-PCR was proven to be specific without cross-reactions with Eurasian FMDV, swine vesicular disease virus (SVDV), Seneca valley virus (SVV), or other common viral pathogens of cattle, sheep, goat, and pig. An around 257 bp-sized amplicon clearly appeared when the inactivated SAT viruses were detected. However, all 125 samples collected from FMDV-susceptible animals from the Chinese mainland which has not known SAT epidemics showed negative results. Conclusions:A FMDV SAT-D8 one-step RT-PCR is a promising method for primary screening for FMDV SAT serotypes.

| S-EPMC6364153 | biostudies-literature

Newcastle Disease virus infection in chickens

Project description:Newcastle Disease virus infection in chickens

| PRJEB7672 | ENA