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R H Purcell

Publications and source records attributed to R H Purcell.

At least 163 records · Page 9Linked to original sources
PubMed1991

A long-term study of hepatitis C virus replication in non-A, non-B hepatitis.

BACKGROUND: Although antibodies to the hepatitis C virus (HCV) are known to be associated with non-A, non-B hepatitis, little is known about the pattern of HCV replication, its relation to antibody levels, and the clinical course of non-A, non-B hepatitis. METHODS: We measured HCV RNA in serial serum samples from five patients with post-transfusion non-A, non-B hepatitis who were followed for 10 to 14 years after transfusion. We also studied four chimpanzees that were experimentally infected with serum from four of these patients. Serum HCV RNA was detected by a "nested" polymerase-chain-reaction (PCR) assay that used two sets of primers derived from the third (NS3) and fourth (NS4) non-structural gene regions of the HCV genome. RESULTS: HCV sequences were detected by PCR in only two of the five patients and two of the four chimpanzees with the set of primers corresponding to the NS3 region, but in all five patients (and in all four chimpanzees) with the primers from the NS4 region. Serum HCV RNA was first detected within three weeks of transfusion in all five patients and within one week in three patients. The viremia lasted less than 4 months in the patient (and two chimpanzees) with acute, self-limited hepatitis, whereas it persisted for 10 to 14 years in the four patients (and for 1 and 3 years in two chimpanzees) with chronic non-A, non-B hepatitis. Antibodies to HCV were first detected at week 12 to 14; they disappeared after nine years in the patient with self-limited disease and became borderline after five years in one of the patients with chronic disease. CONCLUSIONS: During the early phase of primary HCV infection, there is a period of several months of sero-negativity during which HCV RNA is the only diagnostic marker of infection. The disappearance of HCV RNA from serum appears to correlate with the resolution of non-A, non-B hepatitis, whereas viremia persists in patients whose disease progresses to chronic hepatitis. In contrast, antibody levels do not necessarily remain elevated in patients with chronic disease.

Acute Disease↗
PubMed1991

Nucleotide sequence and mutation rate of the H strain of hepatitis C virus.

Patient H is an American patient who was infected with hepatitis C virus (HCV) in 1977. The patient became chronically infected and has remained so for the past 13 years. In this study, we compared the nucleotide and predicted amino acid sequences of the HCV genome obtained from plasma collected in 1977 with that collected in 1990. We find that the two HCV isolates differ at 123 of the 4923 (2.50%) nucleotides sequenced. We estimate that the mutation rate of the H strain of HCV is approximately 1.92 x 10(-3) base substitutions per genome site per year. The nucleotide changes were exclusively base substitutions and were unevenly distributed throughout the genome. A relatively high rate of change was observed in the region of the HCV genome that corresponds to the non-structural protein 1 gene region of flaviviruses, where 44 of 960 (4.6%) nucleotides were different. Within this region there was a 39-nucleotide domain in which 28.2% of the nucleotides differed between the two isolates. In contrast, relatively few nucleotide substitutions were observed in the 5' noncoding region, where only 2 of 276 (0.7%) nucleotides were different. Our results suggest that the mutation rate of the HCV genome is similar to that of other RNA viruses and that genes appear to be evolving at different rates within the virus genome.

Amino Acid Sequence↗
PubMed1991

Attempts to transmit hepatitis B virus to chimpanzees by arthropods.

Bedbugs (Cimex lectularius L.) were fed on an infective blood-hepatitis B virus (HBV) mixture. Further bedbugs and tampan ticks (Ornithodoros moubata [Murray]) were fed on HBV-carrier chimpanzees. After a 10-13 day interval for oviposition, tests done on samples of individual arthropods showed that 53-85% of the bugs were HBsAg-positive and none HBeAg-positive, while 100% of the ticks were HBsAg-positive and 88% HBeAg-positive. The remaining arthropods were fed on 3 susceptible chimpanzees, which had failed to develop HBV infection after 11 months, indicating no transmission had occurred. Subsequently the presence of viable virus in the original infective meals was confirmed by inoculation of the relevant donor sera directly into the 3 still susceptible chimpanzees. HBV infections quickly followed in each animal. It is concluded that, while mechanical transmission of HBV is most unlikely after a 10-13-day interval between feedings in bedbugs and tampans, it is still possible that mechanical transmission between humans might occur during interrupted feeds.

Animals↗
PubMed1991

The nature of genetic variation among viruses.

Genetic variation among viruses may seem unimportant and academic--related only to pedagogical classification of things--but accurate determination of genetic relationships can have important implications, from characterizing the molecular basis of attenuation of viral vaccines to furthering knowledge about origins of viruses and even of life itself. It can even help to establish priority in the discovery of viruses when properly applied. The purpose of this brief review is to demonstrate how viruses change and what implications these changes can have on the delicate balance between the viral parasite and its host. Examples will be drawn from the hepatitis viruses when possible.

Animals↗
PubMed1991

Simian hepatitis A virus (HAV) strain AGM-27: comparison of genome structure and growth in cell culture with other HAV strains.

Fragments of cDNA representing greater than 99% of the entire genome of wild-type hepatitis A virus (HAV) strain AGM-27, isolated from an African green monkey, were obtained by the polymerase chain reaction and sequenced. Comparison with other HAV isolates revealed differences in the predicted amino acid sequence in functionally critical parts of the genome. Comparison of the biological properties of AGM-27 with those of human wild-type and cell culture-adapted HM-175 strains revealed that AGM-27 grew in cell culture significantly better than did wild-type HM-175, but not as well as cell culture-adapted HM-175. AGM-27 and cell culture-adapted HM-175 were distinguishable by their differential growth in CV-1, FRhK-4 and primary AGMK cells.

Amino Acid Sequence↗
PubMed1991

Mutations responsible for adaptation of hepatitis A virus to efficient growth in cell culture.

Chimeric genomes of hepatitis A virus strain HM-175 were constructed from cDNA clones of the wild-type virus and its cell culture-adapted variant. RNA transcribed in vitro from each construct was assayed for infectivity by transfection of cultured cells. RNA transcribed from the wild-type cDNA clone was minimally infectious and produced virus that grew inefficiently in vitro, whereas that transcribed from certain chimeric genomes consistently produced virus that grew efficiently in cultured cells. Mutations in the P2 region were found to be necessary for efficient virus growth in vitro, while mutations in the 5' noncoding region imparted a conditional enhancement of growth in vitro.

Animals↗
PubMed1990

Interrelationship of blood transfusion, non-A, non-B hepatitis and hepatocellular carcinoma: analysis by detection of antibody to hepatitis C virus.

To clarify the relationship between hepatitis C virus infection and the development of hepatocellular carcinoma as sequelae of non-A, non-B posttransfusion hepatitis, 231 patients with chronic non-A, non-B hepatitis (96 with chronic hepatitis, 81 with cirrhosis and 54 with hepatocellular carcinoma) were analyzed for antibody to hepatitis C virus and were compared with 125 patients with chronic hepatitis B (50 with chronic hepatitis, 46 with cirrhosis and 29 with hepatocellular carcinoma). Antibody to hepatitis C virus was detected in 89.6%, 86.4% and 94.4% of patients with non-A, non-B hepatitis-related chronic hepatitis, cirrhosis and hepatocellular carcinoma, respectively, compared with 6%, 17.4% and 34.5% with similar diseases related to hepatitis B. A history of transfusion was documented in 52%, 33% and 42% of anti-hepatitis C virus-positive cases of chronic hepatitis, cirrhosis and hepatocellular carcinoma. The mean intervals between the date of transfusion and the date of diagnosis of anti-hepatitis C virus-positive chronic hepatitis, cirrhosis and hepatocellular carcinoma were 10, 21.2 and 29 yr, respectively. In 21 patients with transfusion-associated hepatocellular carcinoma, anti-hepatitis C virus was present in each serial sample available for testing, including samples obtained up to 14 yr before the diagnosis of hepatocellular carcinoma. These data suggest the slow, sequential progression from acute hepatitis C virus-related non-A, non-B hepatitis through chronic hepatitis and cirrhosis to hepatocellular carcinoma and support a causal association between hepatitis C virus and hepatocellular carcinoma.

Aged↗
PubMed1990

Riboprobe assay for HDV RNA: a sensitive method for the detection of the HDV genome in clinical serum samples.

We have used a new hybridization assay for the detection of the genome of hepatitis delta virus (HDV) in serum using a strand-specific RNA probe obtained by transcription of a recombinant riboprobe. This assay was tested on a panel of 30 sera from HBsAg carriers with hepatitis delta antigen (HDAg) in the liver. The riboprobe assay detected HDV RNA in the serum of 83% of the patients, while 63% were positive using the DNA hybridization assay. HDAg was detected in 73% of the same sera by immunoblotting. The riboprobe assay was also compared to other assays on serial samples from an experimentally infected chimpanzee. These results demonstrate that the Northern blot assay using the RNA probe is more sensitive than the homologous DNA probe for the detection of HDV in serum and is also more sensitive than the immunoblot assay for HDAg. The riboprobe assay is the most sensitive of currently available methods to measure HD viremia.

Adult↗
PubMed1990

Hepatitis B virus DNA detection and comparison with hepatitis B surface antigen.

Hepatitis B surface antigen (HBsAg) has been used for the detection of hepatitis B virus (HBV). Recently, HBV detection using the polymerase chain reaction (PCR) has been shown to be a direct measure of complete virions and to be potentially a very sensitive method. Therefore, we attempted to analyze the relationship between HBsAg detection and HBV DNA assay by PCR. We tested HBV DNA by a modification of the PCR technique in serial sera from five chimpanzees experimentally infected with HBV and 29 human sera. Our new method of PCR analysis is as sensitive as former methods but is more simple and rapid than existing procedures. In chimpanzee studies, HBV DNA was detected 2-3 weeks before the appearance of HBsAg, and continued to be detectable 2 weeks after the production of antibody to HBsAg. Also, 3 of 11 chronic hepatitis B patients who lost HBsAg were positive for HBV DNA in serum, while all of patients who recovered from acute viral infection were negative for HBV DNA in serum. These results indicate that HBV DNA detection by PCR analysis is the most sensitive method currently available to detect the presence of complete virions in serum.

Animals↗
PubMed1990

Detection of hepatitis B virus DNA in serum by polymerase chain reaction. Application for clinical diagnosis.

Standard methods of virus DNA detection using the polymerase chain reaction can be time consuming and can involve multiple steps in which contamination with exogenous DNA can occur. Therefore, we developed a simplified method for detecting hepatitis B virus DNA in serum. The main advantages of this method are that it can be performed rapidly, consists of only several steps, and has a false positive rate of less than 0.1% in our laboratory. In testing serial samples from five chimpanzees experimentally infected with hepatitis B virus, we found that hepatitis B virus DNA was detected 2-3 weeks before the appearance of hepatitis B surface antigen, and it continued to be detectable 1-3 weeks after the production of antibody to hepatitis B surface antigen. In testing serum from 84 human patients, we found hepatitis B virus DNA in all patients who had hepatitis B surface antigen and hepatitis B e antigen in serum and in 64% of the patients with hepatitis B surface antigen with antibody to hepatitis B e antigen in serum. Also, 3 out of 11 patients who were chronic hepatitis B carriers and who subsequently lost hepatitis B surface antigen were found to test positive for hepatitis B virus in serum. In contrast, all patients who lost hepatitis B surface antigen after acute hepatitis B virus infection or those classified as having non-A, non-B hepatitis tested negative for hepatitis B virus DNA. Thus, the modified PCR technique is a sensitive and rapid method for detecting hepatitis B virus DNA-containing virions in serum.

Animals↗
PubMed1990

The significance of infections with two types of viral hepatitis demonstrated by histologic features in chimpanzees.

In view of the recognized importance of necroinflammatory episodes in chronic hepatitis B virus (HBV) infection, chimpanzees, either HBV surface antigen (HBsAg) carriers or noninfected (naive), were infected with other primary hepatotropic viruses to evaluate histologic alterations and changes in virologic and biochemical markers of infection. The advantages of studies on chimpanzees are the availability of serial biopsy specimens and the viral type-specific histologic lesions, not as well recognized in humans. Infection with hepatitis A and non-A, non-B (NANB) agents produced more severe lesions in chronic HBsAg carrier chimpanzees than in naive animals. During this superinfection, the specific expression of the second agent was predominant, indicating that the exacerbation is caused by the second agent, but that carriers are prone to more severe disease than the naive chimpanzees. Hepatitis delta virus (HDV) infections were always coexistant with HBV and superinfection of carriers produced histologic changes more severe than those seen in any other type of viral hepatitis. Such HDV infections revealed less evidence of lymphocytotoxicity but rather of cytotoxicity, and sometimes resembled in appearance the histopathology of NANB. Coinfection of HDV and HBV and superinfection of HBV-carriers with NANB resulted in hepatitis that was far less severe than superinfection of HDV in HBV carriers, greatly in keeping with human experiences. HBV replication was suppressed transiently in both NANB and HDV superinfection. This implies that in exacerbations during chronic HBV infections of humans, suppression of HBV replication markers indicates superinfection, for instance, by NANB for which markers are so far not widely available; by contrast, elevated markers of HBV replication suggest reactivation of the original HBV infection.

Animals↗
PubMed1990

Evaluation in chimpanzees of vaccinia virus recombinants that express the surface glycoproteins of human respiratory syncytial virus.

The immunogenicity and protective efficacy of recombinant vaccinia viruses that express the two major protective antigens of human respiratory syncytial virus (RSV), the F and G glycoproteins, were evaluated in chimpanzees. In previous studies in rodents and monkeys the F and G proteins expressed by the same recombinants were highly immunogenic and induced high levels of resistance to RSV replication following subsequent challenge. In contrast, in chimpanzees, a single intradermal immunization induced only moderate levels of F and G-specific serum antibodies as measured by an enzyme-linked immunosorbent assay, and these antibodies did not efficiently neutralize RSV infectivity in vitro. This poor antibody response in chimpanzees to the F and G glycoproteins occurred despite efficient replication of the vaccinia virus vector as evidenced by lesion size and serum antibody response to vaccinia virus. Upon intranasal RSV challenge, it was observed that prior immunization with the F and G recombinants effected only a marginal reduction in the magnitude and duration of RSV shedding from the nose and trachea and did not reduce illness. However, the RSV challenge induced a strong secondary antibody response, resulting in very high titres (greater than 8000 reciprocal mean titre) of serum neutralizing antibodies. The poor protective efficacy observed here is discussed with regard to the permissiveness of the chimpanzee to RSV replication, the general requirements for effective immunization against RSV, and the limitations of experimental animals for evaluating candidate RSV vaccines.

Animals↗
PubMed1990

Early events in hepatitis C virus infection of chimpanzees.

The cytoplasmic antigen and ultrastructural changes we described previously for chimpanzees (Pan troglodytes) infected with hepatitis C virus (HCV) or with hepatitis D virus have recently been shown to be indirect measures of viral replication and appear to represent a host response to the expression or action of interferon. The time of appearance of these changes in hepatocytes during HCV infection, when compared with similar changes in hepatitis D virus infection, suggests a very early replicative phase for HCV. To investigate the early events in HCV infection, we infected two chimpanzees with HCV and obtained blood and liver biopsy samples from them daily during the first 10 days of infection. The early stage of infection with regard to HCV replication, antigen expression, and ultrastructural changes was similar in both chimpanzees. When tested by cDNA/polymerase chain reaction, HCV sequences became detectable in the serum as early as 3 days after inoculation and remained positive through the peak of aminotransferase elevations. In one chimpanzee the peak of virus production appeared to be 7 weeks after inoculation, which was coincident with rising enzyme values. The cytoplasmic antigen, detected by immunofluorescence, and ultrastructural changes, detected by electron microscopy, became positive in hepatocytes 3 and 6 days, respectively, after HCV sequences were first detected in serum. Circulating anti-HCV appeared 13 weeks and 32 weeks after inoculation, respectively, in the chimpanzees. These data indicate a very early replicative phase for HCV and a potentially long period of infectivity before the appearance of anti-HCV.

Animals↗