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At least 631 records · Page 35Linked to original sources
PubMed1984

The presence of RNA complementary to HSV-2 (herpes simplex virus) DNA in cervical intraepithelial neoplasia after laser therapy.

In-situ hybridization of radiolabelled DNA probes to tissue sections detected RNA complementary to HSV-2 (herpes simplex virus) DNA in 65% (22 of 34) biopsies from cervical intraepithelial neoplasia (CIN) and in none of internally paired benign epithelia from individuals before treatment by laser vaporization cone. Laser therapy did not produce an increase in clinical or subclinical HSV infections of the cervix. Of 18 patients with RNA complementary to HSV-2 DNA in CIN biopsies before treatment, in whom treatment was successful, virus transcripts were detected at follow-up in only two of them.

Cervix Uteri↗
PubMed1994

Outcome of HCV infection after renal transplantation.

The fate and significance of hepatitis C virus (HCV) infection after renal transplantation (TP) remain debated. We therefore evaluated the incidence and outcome of HCV infection in 120 kidney graft recipients both at the time of TP and 54 +/- 28 (13 to 123) months later using ELISA-II and RIBA-II immunoblot. Furthermore, the presence of anti-HCV antibodies at follow-up was correlated with HCV viremia, as detected by the nested polymerase chain reaction (PCR), and with chronically abnormal ALT levels. At the time of TP, 17 patients (14.2%) had anti-HCV antibodies. Compared to anti-HCV (-) patients, anti-HCV (+) patients had a longer duration of pre-TP dialysis (P < 0.001) and had received more pre-TP blood transfusions (P < 0.01). After an average follow-up of 48.5 +/- 24.7 (21 to 97) months, all these patients remained anti-HCV (+) but only 10 were still RIBA "reactive" due to the loss of reactivity against the 5-1-1, C100 and C33 (but not the C22) antigens. Five initially anti-HCV (-) patients had become (+) at follow-up. Among the 22 patients anti-HCV (+) at follow-up, 20 had HCV-RNA detectable by PCR but only 10 had elevated ALT. Ten out of 13 HBsAg (-) patients with elevated ALT were anti-HCV (+). Our study indicates that disappearance of ELISA-II anti-HCV antibodies is rare in kidney recipients and that HCV infection may also occur after TP. Anti-HCV antibodies are likely to reflect a persisting infection as suggested by the frequent detection of HCV-RNA.(ABSTRACT TRUNCATED AT 250 WORDS)

Alanine Transaminase↗
PubMed1988

The detection of coxsackievirus RNA in cardiac tissue by in situ hybridization.

A cloned cDNA probe derived from coxsackie B4 virus-infected cell RNA was shown to hybridize to the RNA of a number of different enteroviruses including coxsackie A and B viruses, echoviruses and poliovirus. The probe was used to detect virus-specific RNA sequences in cardiac tissue obtained from patients diagnosed as having a coxsackievirus infection. Virus RNA was detected using the technique of in situ hybridization in 46% (6/13) cases, but none was found in normal, control, cardiac samples. Two distinct patterns of infection were observed. The significance of these differences and the possible uses of the technique are discussed.

Acute Disease↗
PubMed2001

Vaginal shedding of hepatitis C virus.

A group of 14 women infected with hepatitis C virus (HCV) was evaluated for vaginal shedding of the virus. The HCV-RNA detection was performed in plasma, peripheral blood mononuclear cells (PBMC) and in supernatant and pellet of vaginal washings. HCV-RNA positive results were obtained in all plasma samples, in 57% of the PBMC samples, in 36% pellets obtained following centrifugation of vaginal washings and in 36% of the supernatants. In 21% of the women a positive result was found, at the same time, in every analysed product. The HCV genotypes identified in the plasma samples (1a: 38%; 1b: 36%; 3a: 13% and 1b+3a: 14%) matched with those in PBMC and vaginal washings in every sample, with the exception of cases where mixed infection (1b+3a) was detected. In these, genotype 1b was only detected in plasma. HCV viral load was not associated with the presence of HCV-RNA in PBMC or in vaginal washings (either cells or supernatant).

Female↗
PubMed1997

Murine FGF-12 and FGF-13: expression in embryonic nervous system, connective tissue and heart.

The molecular cloning of cDNAs encoding murine fibroblast growth factor-13 (FGF-13/FHF-2) and three isoforms of murine FGF-12 (FHF-1) is described. Like their highly conserved human counterparts, murine FGF-12 and FGF-13 are part of a distinct subfamily of FGF-like proteins characterized by a greater degree of amino acid sequence cross-homology and by conserved N-terminal domains which do not include secretion signal sequences. In addition to their expression in several adult tissues, both of these FGF genes are prominently and regionally expressed in midgestation mouse embryos, as revealed by in situ hybridization. Fgf12 and fgf13. RNAs were detected in developing central nervous system in cells outside the proliferating ependymal layer, and fgf13 RNA was also found throughout the peripheral nervous system. Fgf12 is expressed in developing soft connective tissue of the limb skeleton and in presumptive connective tissue linking vertebrae and ribs. Both FGF genes are also expressed in the myocardium of the heart, with fgf12 RNA found only in the atrial chamber and fgf13 RNA detected in both atrium and ventricle. On the basis of their novel structure and patterns of expression, FGF-12 and FGF-13 are anticipated to perform embryonic functions distinct from other known FGF molecules.

Adult↗
PubMed1994

Ligation-mediated amplification of RNA from murine erythroid cells reveals a novel class of beta globin mRNA with an extended 5'-untranslated region.

Ligation-mediated RNA amplification was developed as a tool for analysis and determination of the termini of RNA molecules [Volloch et al. (1991) Proc. Natl. Acad. Sci. USA 88: 10671-10675]. In this approach, T4 RNA ligase is used to join cellular RNA with a defined ribo-oligonucleotide. Although several additional enzymatic steps are involved in this type of analysis, the reliability of the entire procedure is determined by the initial ligation step, which marks and preserves the termini of cellular RNA molecules. We applied this approach to the analysis of the 5' terminus of beta globin mRNA in various murine erythroid cells. As expected, we detected RNA molecules with 5' ends terminating at the regular cap site as well as globin RNA molecules truncated at the 5' end. Unexpectedly, we also detected a class of beta globin mRNA which is identical to regular beta globin mRNA in every respect but contains 17, 29, or 31 additional nucleotides 5' to the regular cap site. These extensions correspond precisely to the genomic segments just upstream of the regular cap site and are probably generated by initiation of transcription of the globin gene upstream from the regular cap site. It is likely that the extended globin RNA is transcribed not from the TATA promoter, which regulates the transcription of regular murine globin mRNA, but from the GATA regulatory element located 30 nucleotides upstream of the 31-nucleotide extension, in a position identical to that of the active GATA promoter of the TATA-less chicken beta globin gene. The evolutionary conservation of this relationship suggests the importance of the GATA promoter element of the mouse beta globin gene and its possible involvement in developmental regulation of expression of this gene.

Animals↗
PubMed1994

[Detection of plus and minus strand hepatitis C virus RNA in peripheral blood mononuclear cells and spermatid].

Plus and minus strands of HCV RNA in plasma, peripheral blood mononuclear cells (PBMC), sperm and spermatid of 8 patients with chronic hepatitis C were tested by the method of PCR to find out whether HCV exists and duplicates in extrahepatic cells or not. Plus HCV RNA was detected in all the plasma samples, but no minus HCV RNA was detected. Plus strand HCV RNA was detected in 5 of 8 PBMC samples and minus strand in 2 of the 5 samples. Three of 5 male chronic hepatitis C patients were willing to cooperate and offer their sperm for detection. In the patients, no minus but plus strand HCV RNA was detected in both supernatant of sperm and spermatid. HCV RNA was not detected in the last washing solution for spermatid. The results suggest that: (1). Probably, only plus HCV RNA exists in plasma; (2). HCV exists and probably duplicates in PBMC; (3). HCV exists in sperm. The possibility of transmission of hepatitis C by sexual intercourse is really existent. But, HCV probably don't duplicate in spermatid.

Adult↗
PubMed1988

Expression of human sequences related to those of mouse mammary tumor virus.

Sequences related to those of the mouse mammary tumor virus (MuMTV) genome have been cloned from human DNA by screening a library prepared from the DNA of a human breast cancer cell line with MuMTV gag-pol DNA. Nine distinct groups of (MuMTV-related) sequences were identified among 100 lambda recombinants by cross-hybridization experiments with subcloned fragments containing gag-pol-related DNA. The largest group, of 64 recombinants, contains the MuMTV-related sequences cloned by others. The other eight groups contain MuMTV-related sequences that have not been described previously. The gag-pol regions of one recombinant from each of the nine groups were hybridized to RNA prepared from five human breast cancer cell lines, from placenta, and from two cell lines derived from other malignancies. RNAs were detected by probes for seven of the groups. The RNAs ranged in size from 1.2 to 12 kilobases. Probes for six of the groups detected large RNAs that could represent transcripts of full-length proviral DNA. Two of the probes detected RNA in one breast cancer cell line only. Most of the RNAs were detected in more than one cell line.

Animals↗
PubMed1998

Hepatitis C virus RNA in dried serum spotted onto filter paper is stable at room temperature.

To overcome the instability of viral RNA, we carried out hepatitis C virus (HCV) RNA detection in dried serum spotted onto filter paper. The spotted serum samples were stored at room temperature and then processed for PCR assay at intervals of 1, 2, 3, and 4 weeks. The results showed that serum HCV RNA is stable in a dried condition, as it was detectable in spotted serum samples stored for 4 weeks at room temperature. Furthermore, although the HCV RNA titer showed an approximately 10-fold reduction in virus yield in dried serum stored at room temperature for 4 weeks, the PCR results of frozen serum samples and dried serum samples matched completely. This storage method facilitates transport and analysis by nucleic acid amplification techniques even when freezing conditions are not available.

Blood Specimen Collection↗
PubMed2003

Use and interpretation of hepatitis C virus diagnostic assays.

The HCV genotype, HCV RNA, HCV core antigen, and anti-HCV antibodies are the four biologic markers currently used in hepatitis C. Acute and chronic hepatitis C are diagnosed by anti-HCVantibody (enzyme immunoassay) and HCV RNA detection with sensitive molecular biology techniques. Other virologic tools include HCV genotype determination and HCV RNA quantification; these are used to guide the individual treatment choice, and also to monitor treatment efficacy. Overall, the management of HCV infection has been vastly improved by the use of virologic assays. These assays remain to be fully standardized and automated, however, and more clinically relevant cut-off values are required on which to base management recommendations. More sensitive and accurate HCV RNA assays will improve not only the assessment of the response to antiviral treatment, but also our understanding of antiviral resistance. These improvements, and the development of new antiviral drugs (see the article by Drs. DeFrancesco and Rice elsewhere in this issue), should help to optimize the treatment of HCV infection.

Hepacivirus↗
PubMed1991

Cloning and sequencing of the S RNA from a Bulgarian isolate of tomato spotted wilt virus.

Libraries of cloned cDNA were prepared from complete genomic RNA and isolated S RNA of the Bulgarian L3 isolate of tomato spotted wilt virus (TSWV-L3). Northern blotting of TSWV genomic RNA detected clones specific for the L, M and S RNAs in the library from complete RNA. S RNA-specific clones selected from both libraries covered approximately 2.8 kb (about 95%) of the S RNA. Sequencing of these clones showed TSWV-L3 S RNA to be ambisense. It contains two open reading frames (ORFs); one of 1401 nucleotides located on the viral RNA encodes an Mr 52,400 (52K) protein, and the other of 774 nucleotides on the complementary strand encodes an Mr 28,900 (29K) protein. Expression of the 29K ORF in bacteria and immunological analysis of the fusion protein synthesized confirmed that the 29K protein is the N protein of TSWV-L3. Comparison with the published sequence for the S RNA of a Brazilian TSWV isolate, CNPH1, revealed almost complete identity in the amino acid sequences for the 29K protein, but several clustered amino acid exchanges in the putative 52K protein. In addition, the separating non-translated intergenic region of the S RNA of the Bulgarian isolate is 81 nucleotides longer than that of CNPH1.

Amino Acid Sequence↗
PubMed1991

[Detection of hepatitis C virus RNA in serum during treatment of chronic hepatitis C with interferon alpha].

We estimated HCV RNA in serum of 20 patients with chronic hepatitis C during interferon therapy using a combination assay of reverse transcription and polymerase chain reaction (RT-PCR). RNA was extracted from 200 microliters of serum. The primers were chosen from the NS3 region of prototype HCV RNA sequence. After 40 cycles of PCR, the products were analyzed by Southern hybridization. HCV RNA was detected in 18 of 20 (90%) patients before therapy. In cases with improvement of serum GPT level, HCV RNA became undetectable at 4 weeks of the therapy. However, HCV RNA reappeared within 4 weeks after the therapy in cases with relapse. In the no response group, HCV RNA did not disappear during the therapy. Interferon therapy is beneficial in improvement of viremia of HCV, and the detection of HCV RNA in serum is useful to evaluate the antiviral effect of interferon.

Adult↗
PubMed2000

Evaluation of automated nucleic acid extraction devices for application in HCV NAT.

BACKGROUND: To further improve the safety of the blood supply, various national blood transfusion organizations presently use or are in the process of implementing routine HCV NAT in minipools. According to the Committee for Proprietary Medicinal Products (CPMP) of the European Union, the HCV NAT detection limit of the assay should be 100 IU per mL (270 geq/mL) for testing initial plasma pools. Paul Ehrlich Institute (PEI) regulations stipulate that 5000 IU per mL (13,500 geq/mL) must be detected to calculate the amount contributed by individual donations composing the minipool. The sensitivity for HCV RNA extraction achieved by three commercially available laboratory kits was compared. STUDY DESIGN AND METHODS: Nucleic acids from 1-in-3 serial dilutions of an HCV RNA run control (Pelispy, CLB) were extracted with three kits (Cobas Amplicor, Roche Diagnostic Systems; BioRobot 9604, Qiagen; and NucliSens Extractor, Organon Teknika). HCV PCR of all extracts was performed using a second-generation Cobas Amplicor HCV test and the Cobas Amplicor analyzer. RESULTS: The manual Cobas Amplicor, the BioRobot 9604, and the NucliSens Extractor setups allow a 95-percent HCV RNA detection limit of 129, 82, and 12 geq per mL, respectively. The maximal pool size for the manual Cobas Amplicor, the BioRobot 9604, and the NucliSens Extractor kits that would still meet the PEI criteria for HCV NAT in minipools was calculated at 104, 164, and 1125 donations, respectively. CONCLUSION: All three HCV NAT kits evaluated meet the criteria set by CPMP and PEI. The highest sensitivity for HCV NAT screening can be achieved with the high-volume NucliSens Extractor method in combination with the Cobas Amplicor HCV v2.0 test on the Cobas Amplicor analyzer.

Blood Donors↗
PubMed2003

Evaluation of a transcription-mediated amplification-based HCV and HIV-1 RNA duplex assay for screening individual blood donations: a comparison with a minipool testing system.

BACKGROUND: NAT was introduced for HCV RNA in 1999 to screen blood donations and improve the safety of the blood supply. STUDY DESIGN AND METHODS: The performance of a NAT multiplex for HCV and HIV-1 RNA based on transcription-mediated amplification (TMA) was assessed with various sensitivity panels and by screening 50,000 serologically unscreened, first-time donor plasma samples. Results were compared with a routine NAT screening for HCV RNA by RT-PCR in pools of 96 plasma samples. RESULTS: The TMA multiplex 95 percent sensitivity ranged between 22 and 54 IU per mL for HIV-1 and 15 and 20 IU per mL for HCV RNA. The rate of test failure was 8.6 percent but decreased to 4.7 percent when results of two critical periods of equipment malfunction were excluded. Test failure was related to human error, minute control contamination, and insufficient mixing of reagents at the extraction stage. All 31 repeatedly reactive samples (0.06%) were seropositive for HCV (29) or HIV-1 (2) and contained RNA detectable by discriminatory TMA and confirmatory RT-PCR, indicating 100 percent specificity. A direct comparison of TMA in individual samples and RT-PCR in plasma pools was possible on 27 HCV RNA-containing samples. Twenty-six samples were detected in plasma pools; the lack of detection of 1 sample was due to an identification error at the pooling stage. CONCLUSION: The HCV and HIV-1 multiplex NAT had high specificity and sensitivity.

Base Sequence↗
PubMed1994

Hepatitis C virus infection and liver disease: peculiar epidemiological and clinicopathological features.

Hepatitis C virus (HCV) infection is associated with a wide spectrum of liver disease ranging from asymptomatic carriage to severe forms of chronic hepatitis. HCV is not invariably pathogenic and genetic heterogeneity of HCV could be a major cause of such a variability. In clinical practice this means that presence and replication of the virus do not invariably imply a virus-induced liver damage. IgM antibodies that are the best diagnostic tools for the other forms of viral hepatitis are not sensitive and specific enough for hepatitis C, therefore we have to look for alternatives. Detection of anti-HCV does not help to distinguish past from present infections and only anti-HCV seroconversion in previously negative patients can indicate a recent HCV infection. However, the significant association between serum anti-C100-3 and HCV-RNA suggests that anti-HCV can be considered an indirect marker of HCV infectivity. In anti-HCV-negative infections and early acute hepatitis cases HCV-RNA detection will represent a valid diagnostic alternative. In patients undergoing antiviral therapy monitoring anti-HCV by immunoblotting assays and HCV-RNA by quantitative assays represent a valid tool to predict response that invariably has occurred in patients who had undetectable serum HCV-RNA and/or decreasing anti-HCV titres. Assays that detect multiple anti-HCV antibodies all together appear unsuitable for monitoring because they miss the disappearance of single antibodies. Anti-C22 appears the most frequent and earliest to be detected and usually it has the highest titre. Anti-C100 titres decrease earlier than anti-C33 and anti-C22 in patients with chronic HCV hepatitis who respond to antiviral therapy. The natural course of HCV infection appears to be characterized by three consecutive phases: disease, asymptomatic carrier and recovery. If transition from the first to the last occurs very slowly or the disease phase persists for years it may warrant in susceptible hosts severe forms of liver disease.

Hepacivirus↗
PubMed2025

First surveillance study of avian orthoavulavirus type 1 in wild birds in Morocco: Insights and implications for future monitoring.

BACKGROUND: Wild birds, particularly migratory species, can act as natural reservoirs and vectors of avian orthoavulavirus type 1 (AOAV-1) or Newcastle disease virus (NDV), contributing to its spread across regions and potentially threatening domestic poultry populations. AOAV-1, also known as NDV, is a major pathogen affecting avian species and poses a global threat to poultry production. It belongs to the Paramyxoviridae family and is an RNA virus encoding six key proteins, including the fusion (F) protein, which determines pathogenicity. AOAV-1 is classified into three pathotypes based on virulence: velogenic (highly pathogenic), mesogenic (moderately pathogenic), and lentogenic (mild or asymptomatic). In Morocco, AOAV-1 is endemic in poultry production systems, as evidenced by recent studies reporting a 52.1% seroprevalence and active viral RNA detection in backyard chickens in the Khemisset and Skhirat-Temara provinces; however, effective vaccination strategies have contributed to controlling the clinical signs and widespread dissemination of the virus. AIM: The main objective of this study was to investigate the presence of AOAV-1 in wild bird populations across Morocco, providing insights into possible transmission of infection affecting domestic poultry. METHODS: From November 2016 to April 2022, a total of 1984 samples were collected from 840 individual birds, encompassing 79 species, 25 families, and 12 orders. The majority of the samples belonged to Charadriiformes, Anseriformes, Pelecaniformes, and Passeriformes. Sampling was conducted at 17 wetlands and six additional locations throughout Morocco. Viral detection was performed using real-time reverse transcriptase PCR (RT-qPCR) targeting Matrix (M) and RNA polymerase (L) genes to confirm the presence of AOAV-1. RESULTS: Although the study spanned 6 years and included a large number of samples from bird orders considered primary AOAV-1 reservoirs, all samples tested negative for NDV RNA using both M and L gene targets. CONCLUSION: This study represents the first effort in Morocco to monitor wild birds for AOAV-1. The samples analyzed were initially collected for avian influenza surveillance, which shares epidemiological similarities with Newcastle's disease. However, to improve future surveillance efforts, sample collection should be optimized to target scenarios with the highest probability of virus detection.

Animals↗
PubMed2005

Stability of HCV, HIV-1 and HBV nucleic acids in plasma samples under long-term storage.

The implementation of nucleic acid amplification technology (NAT) for detection of HCV, HIV-1 and HBV has undoubtedly contributed to the viral safety of blood, reducing the window period. One important matter related to the stability of RNA/DNA is the effect of the storage conditions on samples. In a previous work, we studied the stability of HCV RNA in plasma samples after storage at different temperatures. This work is an update on the follow-up of a sample containing 100 IU/ml HCV RNA for 5 years at -20 degrees C, showing no decrease in the initial titre. The nucleic acid stability of other viruses, such as HIV-1 and HBV, has also been studied. At -20 degrees C, samples containing HIV-1 were followed up for approximately 3 years and the results obtained show no decay in HIV-1 RNA detectability. Regardless of the HIV-1 RNA concentration, samples stored at 5 degrees C maintain their titre for at least 14 days. At 25 degrees C, the HIV-1 RNA half-life was determined at nearly 7 days. The HBV DNA, at 5 degrees C and 25 degrees C, is stable for at least 28 days, regardless of the initial titre.

Cold Temperature↗
PubMed1986

Retrovirus-mediated transfer of human adenosine deaminase gene sequences into cells in culture and into murine hematopoietic cells in vivo.

Deficiency of the enzyme adenosine deaminase (adenosine aminohydrolase, EC 3.5.4.4; ADA) leads to severe combined immunodeficiency, a disorder that potentially could be corrected by gene transfer into hematopoietic cells. We have constructed retroviruses containing human ADA cDNA and a dominant selectable marker, a mutated dihydrofolate reductase gene (DHFR*) encoding methotrexate resistance. Human ADA cDNA was inserted alone (DHFR*-ADA) or with a simian virus 40 (SV40) promoter (DHFR*-SVADA). Although NIH 3T3 cells infected with either construct produced human ADA activity, substantially greater levels were attained with DHFR*-SVADA. Infection of murine lymphoid cells in culture with DHFR*-SVADA led to expression of human enzyme at a level well above the mouse endogenous level. ADA activity was also increased after infection of a human ADA-deficient B-cell line. Lethally irradiated mice that were reconstituted with syngeneic marrow infected with the DHFR*-SVADA virus contained unrearranged, integrated proviral DNA in total spleen DNA or in spleen hematopoietic stem cell (CFU-S)-derived colonies. Nevertheless, no human ADA was detectable. RNA analysis showed relatively low and variable expression from the retroviral long terminal repeat, and no detectable expression from the internal SV40 promoter. These data suggest that intrinsic biologic differences exist between cultured cells and CFU-S in vivo.

Adenosine Deaminase↗