Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

To improve the detection and molecular identification of infectious bronchitis virus (avian coronavirus), two reverse transcriptase-polymerase chain reaction (PCR) assays were developed. As 'diagnostic#10; PCR', a set of consensus nested primers was selected from highly conserved stretches of the nucleocapsid (N) gene. As 'phylogeny' PCR, a fragment of the spike protein gene (S1) was amplified and the PCR products were directly sequenced. To study the phylogenetic relationships of the viruses from various outbreaks, studies of molecular epizootiology were performed in Sweden, a Nordic region, where the occurrence of natural cases of the disease is relatively low and the occasional use of live vaccine(s) is well recorded and monitored. The disease appeared in the region in 1994, associated with production problems among layers of various ages. During outbreaks in 1995 and 1997, both layers and broilers were affected. To reduce losses, a live attenuated vaccine has been applied since 1997. By examining 12 cases between 1994 and 1998, molecular epizootiology revealed that, before 1997, the viruses had gene sequences very similar to strains of the Massachusetts serotype. However, comparative sequence analysis of the S1 gene revealed that the identity was not 100% to any of the strains of this serotype that we analysed. A virus related to the Dutch-type strain, D274, was also identified on one farm. Surprisingly, from 1997, the year that vaccination commenced with a live Massachusetts serotype vaccine, the majority of viruses detected had S1 sequences identical to the live Massachusetts vaccine strain. This genetic relation to the vaccine virus was also confirmed by N gene sequence analysis. The studies of molecular epizootiology reveal a strong probability that the vaccination had lead to the spread of the vaccine virus, causing various disease manifestations and a confusing epizootiological situation in the poultry population.
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PMID:Molecular epizootiology of infectious bronchitis virus in Sweden indicating the involvement of a vaccine strain. 1239 45

A universal primer set was developed that amplifies a region covering hypervariable region (HVR) 1 and HVR 2 in the S1 gene of the infectious bronchitis virus (IBV). The universality of this primer set was confirmed by testing the reference strains of different serotypes or variants of the IBV present in the United States. An approximately 450-bp region containing HVR 1 and HVR 2 of 7 untyped field isolates obtained in 1999 and 2000 was amplified. Direct sequencing followed by phylogenetic analysis on that region allowed us to type those field isolates that were not typable by reverse transcriptase-polymerase chain reaction (RT-PCR) and restriction fragment length polymorphism (RFLP). Furthermore, it was found that typing by phylogenetic analysis of that region correlates with virus neutralization results. Together with RT-PCR and RFLP, this method will serve as a fast typing method for IBV diagnosis.
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PMID:Typing of field isolates of infectious bronchitis virus based on the sequence of the hypervariable region in the S1 gene. 1291 15

In this report, we describe a real-time reverse transcriptase-polymerase chain reaction (RRT-PCR) diagnostic test for infectious bronchitis virus (IBV) with the use of fluorescence resonance energy transfer (FRET) technology. Two primers that amplify a 383-base pair product between nucleotide positions 703 and 1086 relative to the start codon for the S1 gene of the Massachusetts 41 virus were designed and used to amplify the Beaudette, Massachusetts 41, Florida 18288, Connecticut, Iowa 97, Arkansas DPI, CA/NE95/99, DE/072/ 92, and GA/0470/98 strains of IBV. The primers were specific and did not amplify New Castle disease virus, Mycoplasma spp., or infectious laryngotracheitis virus. For RRT-PCR by FRET, an anchor probe conjugated to fluorescein and a detection probe conjugated to a red fluorophore were designed to anneal to a hypervariable region within the 383-base pair product. The level of sensitivity was 1 x 10(4) RNA molecules used as starting template. After amplification, a melting curve analysis was conducted to specifically identify IBV types. Because of sequence differences in the annealing position of the detection probe, the Arkansas, Connecticut, Beaudette, and Massachusetts 41 strains could be differentiated. No fluorescence was observed for the DE/072/ 92 and GA/0470/98 viruses with the anchor and detection probes. When the Beaudette strain was examined, two melting peaks were observed at 44 C and 51 C, indicating a quasispecies in that laboratory strain of IBV. Routine typing of vaccine strains of IBV was possible with this technology, but high standard deviations associated with the melting curve analysis of the FRET probes described herein made it difficult to use this test reliably for routine typing of IBV field isolates.
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PMID:Detection of infectious bronchitis virus by real-time reverse transcriptase-polymerase chain reaction and identification of a quasispecies in the Beaudette strain. 1456 2

This study reports on the development of a reverse transcriptase-polymerase chain reaction (RT-PCR) for the specific detection of turkey coronavirus (TCoV). Of the several sets of primers tested, 1 set of primers derived from the P gene and 2 sets derived from the N gene of TCoV could amplify the TCoV genome in the infected samples. The RT-PCR was sensitive and specific for TCoV and did not amplify other avian RNA and DNA viruses tested except the infectious bronchitis virus (IBV). To overcome the problem of IBV amplification, a set of separate primers was designed from the spike protein gene of IBV. The RT-PCR under the same conditions as above could effectively differentiate between TCoV and IBV. The closely related bovine coronavirus and transmissible gastroenteritis virus of pigs were differentiated from TCoV using the same RT-PCR with slight modifications. The results of RT-PCR correlated well with the results of the immunofluorescent test for the same samples tested at the Purdue University Animal Disease Laboratory, West Lafayette, Indiana. The nucleotide sequence and projected amino acid sequence comparison of the P gene of different isolates of TCoV from 5 different states in the United States revealed a close association among the different isolates of TCoV.
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PMID:A reverse transcriptase-polymerase chain reaction assay for the diagnosis of turkey coronavirus infection. 1466 27

The possibility of genomic recombination among different strains of infectious bronchitis virus (IBV) was examined in eve by coinfecting specific pathogen free embryonating chicken eggs with commonly used, embryo-adapted vaccine strains of IBV (Arkansas, Massachusetts, and Connecticut), and a Delaware-072-like field virus isolated from a layer farm in Minnesota. Recombination was observed between th e Massachusetts and the Delaware-072-like strains of the virus. The recombination event was assessed by reverse transcriptase-polymerase chain reaction (RT-PCR) using a combination of specific primers designed to flank a known recombination hot spot of the viral genomic sequence that codes for the S1 subunit of the spike envelope protein. The use of these primers allowed the detection of viruses that have undergone recombination around this hot spot. Cloning and sequencing of the RT-PCR product obtained was performed to confirm these results.
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PMID:A recombination event, induced in ovo, between a low passage infectious bronchitis virus field isolate and a highly embryo adaptedvaccine strain. 1470 73

An infectious bronchitis virus (IBV) was isolated from commercial broilers from the state of California exhibiting respiratory distress, inflamed tracheas, airsaculitis, and edematous lungs. After reverse transcriptase-polymerase chain reaction (RT-PCR), the California isolate exhibited an identical restriction fragment length polymorphism (RFLP) pattern to some isolates obtained from California, known as California 99 isolates. Commercial Mass-Conn and Mass-Ark vaccines were used to vaccinate commercial broiler chickens via eye drop once at 1 or 10 days of age or twice at 1 and 10 days of age. At 27 days of age the birds were challenged via eye drop with the isolated IBV California 99 strain. Protection was measured by failure to reisolate the challenge virus from tracheas 5 days postchallenge and complemented withthe tracheal and epithelium thickness scores. When the Mass-Ark vaccine was included in the vaccination programs, there was protection against challenge with the IBV California 99 isolate. The Mass-Conn vaccine conferred protection when used once at 1 day of age and twice at 1 and 10 days of age. However, no total protection was achieved when used as the only vaccine at 10 days of age, since one of the replicates was positive for virus isolation. Significant differences (P < 0.05) in the epithelium thickness and tracheal scores were observed between the unvaccinated-unchallenged group and the groups vaccinated once or twice with the Mass-Conn vaccine. Based on these results, all chickens were protected against the California 99 isolate when the IBV Arkansas type was used as a vaccine.
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PMID:Evaluation of the protection conferred by commercial vaccines against the California 99 isolate of infectious bronchitis virus. 1470 75

Fifteen isolates of Infectious bronchitis virus (IBV) were obtained from the kidney, trachea, and cecal tonsil of IB suspected chickens between 2001 and 2002 years in Korea. The S1 glycoprotein gene of IBV isolates were amplified by reverse transcriptase - polymerase chain reaction (RT-PCR) and analyzed by restriction fragment length polymorphism (RFLP) analysis. Fifteen Korean IBV isolates were classified into 4 groups by their RFLP patterns using restriction enzymes, HaeIII, BstYI, and XcmI. The RFLP patterns for 3, 1, and 1 of 15 isolates corresponded to the patterns of IBV Arkansas, Connecticut, and Massachusetts strains, respectively. Ten of 15 isolates generated unique KM91 RFLP pattern that was observed in the IBV KM 91 strain previously isolated in Korea. To confirm genetic diversity in the S1 genes of IBV isolates, viral RNAs of representative 9 of 15 IBV isolates were amplified, cloned, sequenced and compared with published sequences for non-Korean IBV strains. Korean IBV isolates showed amino acid sequence similarity between 61.8% (K446-01 and K161-02) and 96.1% (K281-01 and K210-02) with each other and they showed amino acid sequence similarity between 42.9% (K161-02 and GA980470) and 96.5% (K203-02 and KB8523) compared to non-Korean IBV strains. By phylogenetic tree analysis, Korean IBV field isolates were branched into five clusters in which 3 clusters were differentiated from non-Korean IBV strains. Especially, Korean IBV isolates K069-01, K507-01, K774-01 and K142-02 formed a separate cluster. It seems that IBVs continue to evolve and IBVs showing various genetic differences may cocirculate in Korea.
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PMID:S1 glycoprotein gene analysis of infectious bronchitis viruses isolated in Korea. 1499 38

A method is described for enabling safe transit of denatured virus samples for polymerase chain reaction (PCR) identification without the risk of unwanted viable viruses. Cotton swabs dipped in avian infectious bronchitis virus (IBV) or avian pneumovirus (APV) were allowed to dry. Newcastle disease virus and avian influenza viruses were used as controls. Autoclaving and microwave treatment for as little as 20 sec destroyed the infectivity of all four viruses. However, both IBV and APV could be detected by reverse transcriptase (RT)-PCR after autoclaving and as long as 5 min microwave treatment (Newcastle disease virus and avian influenza viruses were not tested). Double microwave treatment of IBV and APV with an interval of 2 to 7 days between was tested. After the second treatment, RT-PCR products were readily detected in all samples. Swabs from the tracheas and cloacas of chicks infected with IBV shown to contain infectious virus were microwaved. Swabs from both sources were positive by RT-PCR. Microwave treatment appears to be a satisfactory method of inactivating virus while preserving nucleic acid for PCR identification.
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PMID:Microwave or autoclave treatments destroy the infectivity of infectious bronchitis virus and avian pneumovirus but allow detection by reverse transcriptase-polymerase chain reaction. 1522 57

Five strains of infectious bronchitis virus (IBV) were isolated from five layer flocks that had nephropathogenic infection in four provinces in China. Among them, three of the five flocks had been vaccinated against infectious bronchitis. Virulence studies indicated that the five Chinese IBV isolates caused 10 to 30% mortality in 15-day-old specific pathogen free chickens and gross lesions were mainly confined to the kidneys in all of the dead chickens. Two oligonucleotide pairs, S1Uni2 and S1Oligo3' or S1Oligo5' and S1Oligo3', were used after propagation of the isolates in embryonated eggs to amplify the S1 protein genes of the spike protein. The cDNA derived by reverse transcriptase-polymerase chain reaction was cloned and sequenced. The nucleotide and amino acid sequence of S1 protein gene had a similar degree of identity (> or =92%) among the five Chinese IBV isolates. The nucleotide and amino acid identity of the S1 protein gene between the five Chinese IBV isolates and 16 strains of other IBVs varied from 60 to 81%. This clearly showed that the five Chinese IBV isolates comprised a separate genotype. These results demonstrated, for the first time, that there is a new genotype of nephropathogenic IBV circulating in vaccinated and non-vaccinated flocks in China.
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PMID:A new genotype of nephropathogenic infectious bronchitis virus circulating in vaccinated and non-vaccinated flocks in China. 1522 61

A novel infectious bronchitis variant, designated as IS/885/00, associated with nephritis, was isolated from outbreaks in 23 broiler farms in Israel. The virus was first identified by reverse transcriptase-polymerase chain reaction and showed a distinct restriction fragment length polymorphism pattern from previously described Israeli isolates. Sequence analysis of the S1 gene and the deduced amino acid sequence revealed 97.2% protein similarity to genotype IS/ 720/99 and 71.6% similarity to the vaccine strain H120, the only strain permitted for use in this country. A database search in GenBank revealed a closely related isolate from Egypt, Egypt/Beni-Seug/01, with 96.6% similarity. Other published nephropathogenic infectious bronchitis virus strains/isolates shared less than 77% similarity with IS/885/00. A vaccine protection test in specific-pathogen-free chicks indicated 91% protection to the trachea and only 25% protection to the kidneys in vaccinated birds challenged with IS/885/00.
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PMID:Identification of a novel nephropathogenic infectious bronchitis virus in Israel. 1552 87


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