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)

RNA sequences of five flaviviruses were detected by a modified polymerase chain reaction (PCR) that incorporated a reverse transcriptase and RNase inhibitor. Oligonucleotide primer pairs were synthesized to amplify sequences from St. Louis encephalitis (SLE), Japanese encephalitis (JBE), yellow fever (YF), dengue 2 (DEN-2), and dengue 4 (DEN-4) viruses. The amplified products were visualized as bands of appropriate size on ethidium bromide-stained agarose gels. The identity of these products was confirmed by restriction endonuclease cleavage to generate fragments of predicted lengths. The reverse-transcriptase PCR (RT-PCR) successfully amplified flavivirus sequences from cell cultures, frozen brain tissue, and formalin-fixed, paraffin-embedded brain tissue. The reactions were highly specific, and the method compared favorably to two conventional assays of viral infectivity. RT-PCR followed by PCR with nesting primers (N-PCR) was 1,000-fold more sensitive in detecting virus than classical infectivity titration by intracerebral inoculation of suckling mice and nearly 1,000-fold more sensitive than amplification of virus in cell culture followed by inoculation of mice.
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PMID:Detection of flaviviruses by reverse-transcriptase polymerase chain reaction. 171 65

A complementary DNA copy of DEN-1 RNA was synthesized using reverse transcriptase and a random primer. The double-stranded DNA copy was cloned at the Sma I site of the pUC13 vector and was used to transform Escherichia coli JM83. Among the transformants selected for characterization by nucleotide sequence analysis, we report here one that codes for a region of nonstructural hydrophilic protein, NS-3. Computer analysis of this sequence (967 bp) showed about 87% conservation at the amino acid level and 79% at the nucleotide level when compared with dengue serotypes 2, 3 and 4 and Japanese encephalitis virus. This suggests an important function which is common to all four serotypes. Comparison of the cloned region with sequences of the above strains also suggested conservation of hydrophobic regions.
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PMID:Partial sequence analysis of type 1 dengue virus coding for the nonstructural hydrophilic protein NS-3. 182 70

Japanese encephalitis (JE) virus genomic RNAs were purified from virions. Two hundred nucleotides at the 3' end of JE virus genomic RNA were directly sequenced by using reverse transcriptase. The nucleotide sequence at the 3' end of the viral RNA was conserved among four kinds of JE virus strains. The sequence has no AU-rich region that is present at the 3' termini of alphavirus RNAs. We also compared the nucleotide sequences at the 3' ends of RNAs from three different flaviviruses and found several common sequence elements. A secondary structure at the 3' end of JE virus genomic RNA was proposed that may be common among flavivirus genomic RNAs. Such structures and other common stretches of nucleotide sequences may be related to the biological properties of flaviviruses.
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PMID:Nucleotide sequence at the 3' end of Japanese encephalitis virus genomic RNA. 372 3

Detection and isolation of Japanese encephalitis (JE) virus were attempted from female mosquitoes collected in Kampong Pasir Panjang, Sabak Bernam, Selangor, from May to November 1992. A total of 7,400 mosquitoes consisting of 12 species in 148 pools were processed and inoculated into Aedes albopictus clone C6/36 cell cultures. Of these, 26 pools showed the presence of viral antigens in the infected C6/36 cells by specific immunoperoxidase staining using an anti-JE virus polyclonal antibody. Presence of JE virus genome was confirmed in the infected culture fluid for 16 pools by using reverse transcriptase-polymerase chain reaction and JE virus-specific primers. Of these, 3 pools were from Culex tritaeniorhynchus, 4 from Culex vishnui, 3 from Culex bitaeniorhynchus, 2 from Culex sitiens, one from Aedes species, and 3 from Culex species. Isolation of JE virus from Cx. sitiens, Cx. bitaeniorhynchus, and Aedes sp. (Aedes butleri and Ae. albopictus) is reported for the first time in Malaysia.
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PMID:Isolation of Japanese encephalitis virus from mosquitoes collected in Sabak Bernam, Selangor, Malaysia in 1992. 761 98

Isolation of Japanese encephalitis virus (JEV) from mosquitoes in Sabak Bernam, Selangor, Malaysia, was attempted. An aliquot of homogenate from each pool of mosquitoes, 50 per tube, was inoculated into Aedes albopictus clone C6/36 cells for virus isolation. Each cell culture was tested for the presence of viral antigen by immunoperoxidase staining using an anti-JEV polyclonal antibody. Out of 4 Culex sitiens mosquito pools, 2 pools were positive for JEV by cell culture. Presence of JEV genome in the cell cultures for Cx. sitiens was confirmed by using reverse transcriptase-polymerase chain reaction and JEV-specific primers. This is the first report on the isolation of JEV from Cx. sitiens.
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PMID:Isolation of Japanese encephalitis virus from Culex sitiens mosquitoes in Selangor, Malaysia. 896 71

Mosquitoes were collected using CO2 and octenol-baited light traps during an outbreak of Japanese encephalitis (JE) on Badu Island in the Torres Strait, Australia in April 1995. A total of 13,300 mosquitoes comprising 12 species were processed for virus isolation. Eight isolates of JE virus were obtained from Culex annulirostris, with a carriage rate of 2.97:1,000; this mosquito also yielded one Sindbis virus isolate. A reverse transcriptase-polymerase chain reaction was used to sequence the JE viruses, which were compared with published sequences. The eight isolates were 90% homologous with known JE strains but only 68% homologous with other flaviviruses. Among the isolates, 99% homology was obtained, suggesting a common point of origin.
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PMID:Isolation of Japanese encephalitis virus from Culex annulirostris in Australia. 906 67

A reverse transcriptase-polymerase chain reaction (RT-PCR) method was developed as a rapid diagnostic test of dengue viremia. To detect dengue viruses in serum or plasma specimens, a pair of universal primers was designed for use in the RT-PCR. Using these primers, the 3'-noncoding region of dengue virus types 1, 2, 3, and 4 could be amplified, but not those of other flaviviruses, such as West Nile virus, Japanese encephalitis virus, and yellow fever virus, or the alphavirus Sindbis virus. The sensitivity of the RT-PCR assay was similar to that of a quantitative fluorescent focus assay of dengue viruses in cell culture. Combining a silica method for RNA isolation and RT-PCR dengue virus could be detected in a 6-hr assay. In a preliminary study using this method, we detected dengue virus in 38 of 39 plasma specimens from which dengue virus had been isolated by mosquito inoculation. We then applied this method for detecting dengue viremia to 117 plasma samples from 62 children with acute febrile illnesses in a dengue-endemic area. We detected dengue viremia in 19 of 20 samples obtained on the day of presentation, which had been confirmed as acute dengue infection by mosquito inoculation and antibody responses. The overall sensitivity of this method was 91.4% (32 of 35; 95% confidence interval [CI] = 82.2-100%). The results from testing plasma samples from febrile nondengue patients showed a specificity of 95.4% (42 of 44; 95% CI = 89.3-100%).
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PMID:Rapid diagnosis of dengue viremia by reverse transcriptase-polymerase chain reaction using 3'-noncoding region universal primers. 915 52

This paper presents the isolation and identification of subgenus B adenovirus during a fatal outbreak of enterovirus-71-associated hand, foot, and mouth disease in Sarawak, Malaysia. Two groups of patients were included in this study: children who had an unexplained sudden pediatric death after a febrile illness; children with acute flaccid paralysis (AFP) during the outbreak who did not die. Both groups were admitted to Sibu Hospital from April 14 to the end of September 1997. Serum and cerebrospinal fluid samples were tested for IgM antibodies to Japanese encephalitis and dengue viruses. Isolated viruses were identified by immunofluorescence, reverse transcriptase PCR, or PCR and DNA sequencing. The enterovirus was isolated in 3 (19%) of the 16 children who died and in 1 of the 8 surviving children with AFP. Moreover, another agent that was initially difficult to identify was found in 10 (63%) children who died and 5 (63%) surviving children who had AFP. The agents isolated from 10 (66.7%) of these 15 children were eventually identified as adenoviruses and were isolated primarily from clinical important sterile sites or tissues. All the enterovirus-positive children who died had this second agent.
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PMID:Isolation of subgenus B adenovirus during a fatal outbreak of enterovirus 71-associated hand, foot, and mouth disease in Sibu, Sarawak. 1067 93

As part of an evaluation of potential vectors of arboviruses during a Rift Valley fever (RVF) outbreak in the Nile Valley of Egypt in August 1993, we collected mosquitoes in villages with known RVF viral activity. Mosquitoes were sorted to species, pooled, and processed for virus isolation both by intracerebral inoculation into suckling mice and by inoculation into cell culture. A total of 33 virus isolates was made from 36,024 mosquitoes. Viruses were initially identified by indirect fluorescent antibody testing and consisted of 30 flaviviruses (all members of the Japanese encephalitis complex, most probably West Nile [WN] virus) and three alphaviruses (all members of western equine encephalitis complex, most probably Sindbis). The identity of selected viruses was confirmed by reverse transcriptase-polymerase chain reaction and sequencing. Culex antennatus (Becker) and Culex perexiguus Theobald accounted for five (17%) and 23 (77%) of the WN virus isolations, respectively. Despite isolation of viruses from 32 pools of mosquitoes (both WN and Sindbis viruses were isolated from a single pool), RVF virus was not isolated from these mosquitoes, even though most of them are known competent vectors collected during an ongoing RVF outbreak. Thus, it should be remembered, that even during a known arbovirus outbreak, other arboviruses may still be circulating and causing disease.
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PMID:Isolation of west nile and sindbis viruses from mosquitoes collected in the Nile Valley of Egypt during an outbreak of Rift Valley fever. 1193 Dec 67

As part of an evaluation of the ecology of arthropod-borne diseases in the Republic of Korea (ROK), we examined 8,765 mosquitoes captured in Paju County, Gyonggi Province, ROK, for the presence of viruses. Mosquitoes were captured in propane lantern/human-baited Shannon traps, Mosquito Magnet traps, or American Biophysics Corporation (East Greenwich, RI) miniature light traps with or without supplemental octenol bait and/or dry ice. Mosquitoes were identified to species, placed in pools of up to 40 mosquitoes each, and tested on Vero cells for the presence of virus. A total of 15 virus isolations were made from 293 pools of mosquitoes. Viruses were identified by reverse transcriptase-polymerase chain reaction and sequencing and consisted of 14 isolations of Japanese encephalitis (JE) virus and one isolation of Getah (GET) virus. All JE isolates were from Culex tritaeniorhynchus Giles, and the isolate of GET was from Aedes vexans (Meigen). The minimum field infection rate for JE in Cx. tritaeniorhynchus was 3.3 per 1,000, whereas the GET virus infection rate for Ae. vexans was 0.2 per 1,000. Isolation of JE and GET indicated that both viruses were actively circulating in northern Gyonggi Province, ROK. The lack of human cases of JE among the Korean population probably is because of an effective government-mandated vaccination program. The reason for no cases among >10,000 United States military and others that reside or train nearby is unknown, but may be related to personnel protection measures (permethrin-impregnated uniforms and use of deet repellent), adult mosquito control, mosquito selection of nonhuman hosts (unpublished data), and the low symptomatic to asymptomatic ratio of disease in adults.
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PMID:Isolation of Japanese encephalitis and Getah viruses from mosquitoes (Diptera: Culicidae) collected near Camp Greaves, Gyonggi Province, Republic of Korea, 2000. 1468 Jan 30


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