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Biomedical subjects

S U Emerson

Publications and source records attributed to S U Emerson.

At least 19 recordsLinked to original sources
PubMed2002

Four chimpanzee monoclonal antibodies isolated by phage display neutralize hepatitis a virus.

Chimpanzee immunoglobulins are virtually identical to human immunoglobulins and may have clinically useful applications. Four chimpanzee monoclonal antibodies (MAbs) to the hepatitis A virus (HAV) capsid were isolated from a combinatorial cDNA library of gamma1/kappa antibody genes using phage display. Competition assays indicated that three of the MAbs recognized the same or overlapping epitopes, whereas the fourth recognized a different, nonoverlapping epitope on the HAV capsid. All four MAbs neutralized the homologous HAV strain, HM-175, in a radioimmunofocus assay and two of the four MAbs neutralized a heterologous simian HAV strain, AGM-27. From these data, we conclude that the MAbs must recognize at least three epitopes on the HAV capsid. Furthermore, competition assays performed with neutralizing murine MAbs suggested that three of the chimpanzee MAbs recognized epitopes on the HAV capsid which have not been defined previously.

Amino Acid Sequence↗
PubMed2002

Prevalence of antibodies to hepatitis E virus in veterinarians working with swine and in normal blood donors in the United States and other countries.

Hepatitis E virus (HEV) is endemic in many developing and some industrialized countries. It has been hypothesized that animals may be the source of infection. The recent identification of swine HEV in U.S. pigs and the demonstration of its ability to infect across species have lent credence to this hypothesis. To assess the potential risk of zoonotic HEV infection, we tested a total of 468 veterinarians working with swine (including 389 U.S. swine veterinarians) and 400 normal U.S. blood donors for immunoglobulin G anti-HEV. Recombinant capsid antigens from a U.S. strain of swine HEV and from a human HEV strain (Sar-55) were each used in an enzyme-linked immunosorbent assay. The anti-HEV prevalence assayed with the swine HEV antigen showed 97% concordance with that obtained with the human HEV antigen (kappa = 92%). Among the 295 swine veterinarians tested from the eight U.S. states (Minnesota, Indiana, Nebraska, Iowa, Illinois, Missouri, North Carolina, and Alabama) from which normal blood donor samples were available, 26% were positive with Sar-55 antigen and 23% were positive with swine HEV antigen. In contrast, 18% of the blood donors from the same eight U.S. states were positive with Sar-55 antigen and 17% were positive with swine HEV antigen. Swine veterinarians in the eight states were 1.51 times more likely when tested with swine HEV antigen (95% confidence interval, 1.03 to 2.20) and 1.46 times more likely when tested with Sar-55 antigen (95% confidence interval, 0.99 to 2.17) to be anti-HEV positive than normal blood donors. We did not find a difference in anti-HEV prevalence between veterinarians who reported having had a needle stick or cut and those who had not or between those who spent more time (> or = 80% of the time) and those who spent less time (< or = 20% of the time) working with pigs. Similarly, we did not find a difference in anti-HEV prevalence according to four job categories (academic, practicing, student, and industry veterinarians). There was a difference in anti-HEV prevalence in both swine veterinarians and blood donors among the eight selected states, with subjects from Minnesota six times more likely to be anti-HEV positive than those from Alabama. Age was not a factor in the observed differences from state to state. Anti-HEV prevalence in swine veterinarians and normal blood donors was age specific and paralleled increasing age. The results suggest that swine veterinarians may be at somewhat higher risk of HEV infection than are normal blood donors.

Adult↗
PubMed2001

Immunogenicity and protective efficacy of a vaccine prepared from 53 kDa truncated hepatitis E virus capsid protein expressed in insect cells.

Hepatitis E virus (HEV) is an enterically transmitted virus that causes acute hepatitis. Expression of recombinant HEV capsid protein in insect cells results in two major proteolytically-processed products of 56 and 53kDa which consist of amino acids (aa) 112-607 and 112-578, respectively. The only neutralization epitope identified to date is located at least partially between amino acids 578 and 607 meaning it should be present only in the 56 and not in the 53kDa protein. Previously, it was shown that vaccination with the 56kDa protein greatly reduced virus shedding and protected Rhesus monkeys from hepatitis E when challenged with a high intravenous dose of homologous or heterologous HEV. To evaluate the immunogenicity and protective efficacy of the 53kDa protein, we vaccinated Rhesus monkeys with this protein and challenged them with a high or low dose of homologous virus. Vaccination with the 53kDa protein greatly reduced virus shedding but did not protect against hepatitis following the high dose challenge. Virus was not detected in the vaccinated animals following the low dose challenge, suggesting that sterilizing immunity may have been achieved.

Alanine Transaminase↗
PubMed2001

Recombinant hepatitis E virus genomes infectious for primates: importance of capping and discovery of a cis-reactive element.

Hepatitis E virus recombinant genomes transcribed in vitro from two cDNA clones differing by two nucleotides were infectious for chimpanzees. However, one cDNA clone encoded a virus that was attenuated for chimpanzees and unable to infect rhesus monkeys. The second cDNA clone encoded a virus that infected both chimpanzees and rhesus monkeys and caused acute hepatitis in both. One mutation differentiating the two clones identified a cis-reactive element that appeared to overlap the 3' end of the capsid gene and part of the 3' noncoding region. Capping of the RNA transcripts was essential for infectivity.

Animals↗
PubMed2001

A virus discovery method incorporating DNase treatment and its application to the identification of two bovine parvovirus species.

Identification of previously unrecognized viral agents in serum or plasma samples is of great medical interest but remains a major challenge, primarily because of abundant host DNA. The current methods, library screening or representational difference analysis (RDA), are very laborious and require selected sample sets. We have developed a simple and reproducible method for discovering viruses in single serum samples that is based on DNase treatment of the serum followed by restriction enzyme digestion and sequence-independent single primer amplification (SISPA) of the fragments, and have evaluated its performance on known viruses. Both DNA viruses and RNA viruses at a concentration of approximately 10(6) genome equivalents per ml were reproducibly identified in 50 microl of serum. While evaluating the method, two previously unknown parvoviruses were discovered in the bovine sera used as diluent. The near complete genome sequence of each virus was determined; their classification as two species (provisionally named bovine parvoviruses 2 and 3) was confirmed by phylogenetic analysis. Both viruses were found to be frequent contaminants of commercial bovine serum. DNase treatment of serum samples may prove to be a very useful tool for virus discovery. The DNase-SISPA method is suitable for screening of a large number of samples and also enables rapid sequence determination of high-titer viruses.

Animals↗
PubMed2001

Detection of antibodies to HAV 3C proteinase in experimentally infected chimpanzees and in naturally infected children.

Commercial assays for the diagnosis of hepatitis A detect antibody to hepatitis A virus (anti-HAV), but they cannot discriminate between antibody resulting from infection and antibody induced by inactivated vaccine. With the licensing and increasing use of inactivated hepatitis A vaccines, there is a need for a test to distinguish between infection and vaccination. Since antibodies to viral non-structural proteins are elicited by infection but not by vaccination with inactivated vaccine, we developed and evaluated a test for such antibodies. The antibody response to the non-structural 3C proteinase (anti-3C) of virus HAV was studied by ELISA in chimpanzees experimentally infected with virulent (wild type) or with attenuated HAV strains and in children who received inactivated HAV vaccine or placebo during a vaccination trial in Nicaragua. Anti-3C was detected in 89% of 18 chimpanzees infected with wild-type HAV strains and 27% of 26 chimpanzees infected with attenuated HAV strains. There was a direct correlation between severity of hepatitis and magnitude of the anti-3C response. In the vaccine trial, anti-3C was detected only in children who were infected with HAV during the study; IgG anti-3C persisted for at least 15 months after infection in one child. Vaccinated and uninfected children remained negative for anti-3C. The anti-3C response can be regarded as an indicator of viral replication. Its detection should be useful for distinguishing between antibody acquired in response to HAV infection and antibody induced by immunization with inactivated vaccine.

3C Viral Proteases↗
PubMed2001

Identification of the 5' terminal sequence of the SAR-55 and MEX-14 strains of hepatitis E virus and confirmation that the genome is capped.

Hepatitis E virus (HEV) is a nonenveloped virus with a genome of single-stranded, positive-sense RNA. The 5' terminal sequence of two HEV strains (SAR-55 and MEX-14) was determined by a 5' RNA ligase-mediated rapid amplification of cDNA ends (RACE) method designed to select capped RNAs. The 5' noncoding region of the SAR-55 and MEX-14 strains were amplified, confirming that the genomic RNA of HEV is capped. The 5' noncoding region of the SAR-55 strain had 25 nucleotides, which is two less than reported for the Burmese strain, and that of the MEX-14 strain had 24 nucleotides, which is 21 more than reported previously [Huang et al., 1992].

5' Untranslated Regions↗
PubMed2001

High-level expression of hepatitis C virus (HCV) structural proteins by a chimeric HCV/BVDV genome propagated as a BVDV pseudotype.

A chimeric cDNA genome was constructed in which the core, E1 and E2 genes of hepatitis C virus (HCV) replaced the core, E(rns), E1 and E2 genes of bovine viral diarrhea virus (BVDV). High levels of HCV structural proteins were expressed in a small number of human or bovine cells following transfection with chimeric RNA. However, in one cell line, bovine embryonic trachea cells [EBTr(A)], the number of cells expressing HCV proteins increased to greater than 70% following serial passage of culture medium. These cells were persistently infected with a non-cytopathogenic BVDV helper virus. In these cells, the chimeric genome was packaged into infectious particles that accumulated in the culture medium at a titer as high as 10(7)-10(9) genome equivalents per ml. The virus particles were pseudotypes, because they were neutralized by anti-BVDV but not by anti-HCV.

Animals↗
PubMed2001

Recombinant vaccines for hepatitis E.

Hepatitis E virus causes epidemics of acute hepatitis in many developing countries. It infrequently causes disease in developed countries, but avirulent strains might circulate. Some evidence suggests that hepatitis E might be a zoonosis. There is probably only a single serotype. A candidate vaccine consisting of baculovirus-expressed recombinant capsid protein protected macaques from hepatitis E--it passed phase I clinical trials and is currently scheduled for phase II/III clinical trials.

Animals↗
PubMed2001

Failure to infect rhesus monkeys with hepatitis C virus strains of genotypes 1a, 2a or 3a.

The chimpanzee is the only recognized animal model for the study of hepatitis C virus (HCV). However, recently it was reported that rhesus monkeys were susceptible to HCV and developed hepatitis during infection. In the present study, we inoculated two rhesus monkeys each with HCV strain H77 (genotype 1a), strain HC-J6 (genotype 2a) or strain S52 (genotype 3a). Weekly serum samples were tested for liver enzyme values, HCV antibodies and HCV RNA. We did not find evidence of HCV infection in any of the monkeys during 24 weeks of follow-up. Our study demonstrates that rhesus monkeys are not readily infected with HCV and apparently do not represent a useful animal model for the study of HCV.

Animals↗
PubMed2001

Animal models of hepatitis A and E.

Several useful animal models for both hepatitis A and E have been identified, characterized, and refined. At present, all of the best models utilize nonhuman primates: chimpanzees, tamarin species, and owl monkeys for hepatitis A; and macaque species, chimpanzees, and owl monkeys for hepatitis E. Pigs may prove useful for some studies of hepatitis E, and it is hoped that serological evidence of widespread infection of rats with an HEV-like agent may lead to the development of an animal model based on laboratory rats. As has been the case for each of the hepatitis viruses as they have been discovered, the development of useful and reproducible animal model systems has been critical for moving the field forward as expeditiously as possible.

Animals↗
PubMed2001

Comparative pathogenesis of infection of pigs with hepatitis E viruses recovered from a pig and a human.

Specific-pathogen-free pigs were inoculated with one of two hepatitis E viruses (HEV) (one recovered from a pig and the other from a human) to study the relative pathogenesis of the two viruses in swine. Fifty-four pigs were randomly assigned to three groups. Seventeen pigs in group 1 served as uninoculated controls, 18 pigs in group 2 were intravenously inoculated with the swine HEV recovered from a pig in the United States, and 19 pigs in group 3 were intravenously inoculated with the US-2 strain of human HEV recovered from a hepatitis patient in the United States. Two to four pigs from each group were necropsied at 3, 7, 14, 20, 27, or 55 days postinoculation (DPI). Evidence of clinical disease or elevation of liver enzymes or bilirubin was not found in pigs from any of the three groups. Enlarged hepatic and mesenteric lymph nodes were observed in both HEV-inoculated groups. Multifocal lymphoplasmacytic hepatitis was observed in 9 of 17, 15 of 18, and 16 of 19 pigs in groups 1 to 3, respectively. Focal hepatocellular necrosis was observed in 5 of 17, 10 of 18, and 13 of 19 pigs in groups 1 to 3, respectively. Hepatitis lesions were very mild in group 1 pigs, mild to moderate in group 2 pigs, and moderate to severe in group 3 pigs. Hepatic inflammation and hepatocellular necrosis peaked in severity at 20 DPI and were still moderately severe at 55 DPI in the group inoculated with human HEV. Hepatitis lesions were absent or nearly resolved by 55 DPI in the swine-HEV-inoculated pigs. All HEV-inoculated pigs seroconverted to anti-HEV immunoglobulin G. HEV RNA was detected by reverse transcriptase PCR in feces, liver tissue, and bile of pigs in both HEV-inoculated groups from 3 to 27 DPI. Based on evaluation of microscopic lesions, the US-2 strain of human HEV induced more severe and persistent hepatic lesions in pigs than did swine HEV. Pig livers or cells from the livers of HEV-infected pigs may represent a risk for transmission of HEV from pigs to human xenograft recipients. Since HEV was shed in the feces of infected pigs, exposure to feces from infected pigs represents a risk for transmission of HEV, and pigs should be considered a reservoir for HEV.

Animals↗
PubMed2001

Studies of hepatitis C virus in chimpanzees and their importance for vaccine development.

Persistent infection with hepatitis C virus (HCV) is an important cause of chronic liver disease worldwide. Therefore, the development of vaccines to prevent HCV infection, or at least to prevent progression to chronicity, is a major goal. Potential HCV vaccine candidates include recombinant proteins, recombinant viruses, DNA constructs, synthetic peptides and virus-like particles. Various vaccine candidates have been shown to generate humoral and cellular immune responses in animals, primarily in mice. However, the efficacy of most vaccine candidates in protecting against HCV has not been tested because the chimpanzee, the only animal other than humans that is susceptible to HCV, is not readily available, requires special facilities, and is very expensive. The course of infection in chimpanzees is similar in its diversity to that in humans and detailed studies in this model are beginning to define the immune responses that can terminate HCV infection. Of relevance for vaccine evaluation was the titration in chimpanzees of different HCV variants to provide well-characterized challenge pools. In addition, monoclonal virus pools generated from chimpanzees infected with cloned viruses make it possible now to examine immunity to HCV without the confounding factor of antigenic diversity of the challenge virus (quasispecies). The vaccine trials performed in chimpanzees to date all have tested the efficacy of immunizations with various forms of the envelope proteins of HCV.

Animals↗
PubMed2000

Hepatitis C virus envelope protein E2 binds to CD81 of tamarins.

Since recombinant envelope glycoprotein E2 of hepatitis C virus (HCV) binds to CD81 on human and chimpanzee cells, it has been suggested that CD81 may be a receptor for HCV. Humans and chimpanzees are the only species known to be susceptible to HCV infection. E2 has been reported not to bind to CD81 of the African green monkey, mouse, or rat, suggesting that binding of HCV to CD81 is species specific and may determine susceptibility to infection with HCV. We investigated the interaction between E2 of HCV and CD81 of tamarins, a group of small New World monkeys frequently used for the study of human viruses. Tamarins are not susceptible to HCV infection. Nonetheless, we found that three different forms of HCV E2 (intracellular, secreted, and cell surface-displayed) bound more efficiently to recombinant tamarin CD81 than to human CD81, as determined by ELISA and immunofluorescence. The affinity of the interaction was approximately 10-fold higher for tamarin than for human CD81. Binding of E2 to CD81 on cultured or primary tamarin cells was demonstrated by flow cytometry. In contrast to previous reports, there was also a low-affinity interaction between E2 and African green monkey CD81. Thus, the HCV E2 interaction with CD81 is not limited to humans and chimpanzees and does not predict susceptibility to HCV infection.

Amino Acid Sequence↗
PubMed2000

Hepatitis C virus lacking the hypervariable region 1 of the second envelope protein is infectious and causes acute resolving or persistent infection in chimpanzees.

Persistent infection with hepatitis C virus (HCV) is among the leading causes of chronic liver disease. Previous studies suggested that genetic variation in hypervariable region 1 (HVR1) of the second envelope protein, possibly in response to host immune pressure, influences the outcome of HCV infection. In the present study, a chimpanzee transfected intrahepatically with RNA transcripts of an infectious HCV clone (pCV-H77C) from which HVR1 was deleted became infected; the DeltaHVR1 virus was subsequently transmitted to a second chimpanzee. Infection with DeltaHVR1 virus resulted in persistent infection in the former chimpanzee and in acute resolving infection in the latter chimpanzee. Both chimpanzees developed hepatitis. The DeltaHVR1 virus initially replicated to low titers, but virus titer increased significantly after mutations appeared in the viral genome. Thus, wild-type HCV without HVR1 was apparently attenuated, suggesting a functional role of HVR1. However, our data indicate that HVR1 is not essential for the viability of HCV, the resolution of infection, or the progression to chronicity.

Animals↗
PubMed2000

Vaccination of chimpanzees with plasmid DNA encoding the hepatitis C virus (HCV) envelope E2 protein modified the infection after challenge with homologous monoclonal HCV.

Hepatitis C virus (HCV) is an important cause of chronic liver disease worldwide. Development of vaccines to prevent HCV infection, or at least prevent progression to chronicity, is a major goal. In mice and rhesus macaques, a DNA vaccine encoding cell-surface HCV-envelope 2 (E2) glycoprotein stimulated stronger immune responses than a vaccine encoding intracellular E2. Therefore, we used DNA encoding surface-expressed E2 to immunize chimpanzees 2768 and 3001. Chimpanzee 3001 developed anti-E2 after the second immunization and antibodies to hypervariable region 1 (HVR1) after the third immunization. Although chimpanzee 2768 had only low levels of anti-E2 after the third immunization, an anamnestic response occurred after HCV challenge. CTL responses to E2 were not detected before challenge, but a strong response was detected after HCV challenge in chimpanzee 2768. An E2-specific CD4+ response was detected in chimpanzee 2768 before challenge and in both chimpanzees postchallenge. Three weeks after the last immunization, animals were challenged with 100 50% chimpanzee-infectious doses (CID(50)) of homologous monoclonal HCV. As a control, a naive chimpanzee was inoculated with 3 CID(50) of the challenge virus. The vaccine did not generate sterilizing immunity because both vaccinated chimpanzees were infected. However, both vaccinated chimpanzees resolved the infection early whereas the control animal became chronically infected. Compared with the control animal, hepatitis appeared earlier in the course of the infection in both vaccinated chimpanzees. Therefore, DNA vaccine encoding cell surface-expressed E2 did not elicit sterilizing immunity in chimpanzees against challenge with a monoclonal homologous virus, but did appear to modify the infection and might have prevented progression to chronicity.

Animals↗
PubMed2000

High prevalence of TT virus (TTV) in naive chimpanzees and in hepatitis C virus-infected humans: frequent mixed infections and identification of new TTV genotypes in chimpanzees.

A recently discovered DNA virus, TT virus (TTV), is prevalent in humans. In the present study, the genetic heterogeneity of TTV was evaluated in hepatitis C virus (HCV)-infected patients and in chimpanzees. TTV DNA was detected by PCR in serum samples from all ten HCV-infected patients studied; at least five major TTV genotypes, all previously identified in humans, were recovered. Eight patients were infected with multiple variants of TTV. TTV DNA was detected by PCR in serum samples from 11 (65%) of 17 naive chimpanzees bred in captivity; a persistent infection was present in three of six animals. At least five chimpanzees were infected with more than one TTV variant. Detection of TTV DNA in chimpanzee faecal samples suggests the possibility of faecal-oral transmission. Phylogenetic analysis of ORF1 sequences amplified from chimpanzees identified three major genotypes which had not previously been recognized in humans.

Amino Acid Sequence↗