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R G Webster

Publications and source records attributed to R G Webster.

At least 37 records · Page 2Linked to original sources
PubMed2003

Differences between influenza virus receptors on target cells of duck and chicken and receptor specificity of the 1997 H5N1 chicken and human influenza viruses from Hong Kong.

To study whether influenza virus receptors in chickens differ from those in other species, we compared the binding of lectins and influenza viruses with known receptor specificity to cell membranes and gangliosides from epithelial tissues of ducks, chickens, and African green monkeys. We found that chicken cells contained Neu5Ac alpha(2-6)Gal-terminated receptors recognized by Sambucus nigra lectin and by human viruses. This finding explains how some recent H9N2 viruses replicate in chickens despite their human virus-like receptor specificity. Duck virus bound to gangliosides with short sugar chains that were abundant in duck intestine. Human and chicken viruses did not bind to these gangliosides and bound more strongly than duck virus to gangliosides with long sugar chains that were found in chicken intestinal and monkey lung tissues. Chicken and duck viruses also differed by their ability to recognize the structure of the third sugar moiety in Sia2-3Gal-terminated receptors. Chicken viruses preferentially bound to Neu5Ac alpha(2-3)Gal beta(1-4)GlcNAc-containing synthetic sialylglycopolymer, whereas duck viruses displayed a higher affinity for Neu5Ac alpha(2-3)Gal beta(1-3)GalNAc-containing polymer. Our data indicate that sialyloligosaccharide receptors in different avian species are not identical and provide a potential explanation for the differences between the hemagglutinin and neuraminidase proteins of duck and chicken viruses.

Animals↗
PubMed2003

The quest of influenza A viruses for new hosts.

There is increasing evidence that stable lineages of influenza viruses are being established in chickens. H9N2 viruses are established in chickens in Eurasia, and there are increasing reports of H3N2, H6N1, and H6N2 influenza viruses in chickens both in Asia and North America. Surveillance in a live poultry market in Nanchang, South Central China, reveals that influenza viruses were isolated form 1% of fecal samples taken from healthy poultry over the course of 16 months. The highest isolation rates were from chickens (1.3%) and ducks (1.2%), followed by quail (0.8%), then pigeon (0.5%). H3N6, H9N2, H2N9, and H4N6 viruses were isolated from multiple samples, while single isolates of H1N1, H3N2, and H3N3 viruses were made. Representatives of each virus subtype were experimentally inoculated into both quail and chickens. All the viruses replicated in the trachea of quail, but efficient replication in chickens was confined to 25% of the tested isolates. In quail, these viruses were shed primarily by the aerosol route, raising the possibility that quail may be the "route modulator" that changes the route of transmission of influenza viruses from fecal-oral to aerosol transmission. Thus, quail may play an important role in the natural history of influenza viruses. The pros and cons of the use of inactivated and recombinant fowl pox-influenza vaccines to control the spread of avian influenza are also evaluated.

Animals↗
PubMed2003

Multiple genotypes of nonpathogenic H6N2 influenza viruses isolated from chickens in California.

From February 2000 through September 2001, a limited number of H6N2 influenza viruses were isolated from chickens in California. This report describes the genetic characterization of nine of these H6N2 viruses. All of the viruses analyzed had phylogenetically similar hemagglutinin (HA) and neuraminidase molecules that suggested the viruses shared a recent common ancestor. The analysis of the HA sequence of these viruses with all available H6 viruses from different hosts and locations showed that these genes do not separate into well-defined North American and Eurasian lineages. The neuraminidase genes of the California viruses contain an 18 amino acid deletion, a possible adaptation to growth in chickens. Analysis of the remaining gene segments of the California viruses revealed that three distinct genotypes of H6N2 viruses were present.

Amino Acid Sequence↗
PubMed2003

Reassortants of H5N1 influenza viruses recently isolated from aquatic poultry in Hong Kong SAR.

The H5N1 virus (H5N1/97) that caused the bird flu incident in Hong Kong in 1997 has not been isolated since the poultry slaughter in late 1997. But the donor of its H5 hemagglutinin gene, Goose/Guangdong/1/96-like (Gs/Gd/96-like) virus, established a distinct lineage and continued to circulate in geese in the area. In 2000, a virus from the Goose/Guangdong/1/96 lineage was isolated for the first time from domestic ducks. Subsequently, it has undergone reassortment, and these novel reassortants now appear to have replaced Gs/Gd/96-like viruses from its reservoir in geese and from ducks. The internal gene constellation is also different from H5N1/97, but these variants have the potential for further reassortment events that may allow the interspecies transmission of the virus.

Animals↗
PubMed2003

[Antigenic structure of influenza A virus subtype H5 hemagglutinin: mechanism of acquiring stability to monoclonal antibodies in escape-mutants].

The analysis of escape mutants of the avian influenza virus of H5 subtype (strain A/Mallard/Pennsylvania/10218/84) revealed the location and structure of two antigenic sites in the hemagglutinin (HA) molecule. Several escape mutants exhibited unusual features in the reactions with monoclonal antibodies (Mabs), being completely resistant in the infectivity neutralization test to the Mabs used for their selection, and retaining the ability to bind the Mabs as revealed by enzyme-linked immunosorbent assay. An enhancement of the binding by an amino acid change in a different antigenic site was demonstrated, as well as a complete abolishment of the binding by a mutation selected by passage in the presence of an excess of the non-neutralizing Mab of high binding ability. The observed effects did not result from the changes in the affinity of the mutant HA toward sialic receptors. The data suggest that one amino acid change in HA may prevent the virus neutralization by different mechanisms for different Mabs: either the binding of the Mab to HA is prevented, or the bound Mab is unable to block the receptor-binding pocket of HA. Different mechanisms of the acquisition of resistance to Mabs in the course of the selection of escape mutants are discussed.

Antibodies, Monoclonal↗
PubMed2002

Emergence of multiple genotypes of H5N1 avian influenza viruses in Hong Kong SAR.

Although A/Hong Kong/156/97 (H5N1/97)-like viruses associated with the "bird flu" incident in Hong Kong SAR have not been detected since the slaughter of poultry in 1997, its putative precursors continue to persist in the region. One of these, Goose/Guangdong/1/96 (H5N1 Gs/Gd)-like viruses, reassorted with other avian viruses to generate multiple genotypes of H5N1 viruses that crossed to chickens and other terrestrial poultry from its reservoir in geese. Whereas none of these recent reassortants had acquired the gene constellation of H5N1/97, these events provide insight into how such a virus may have been generated. The recent H5N1 reassortants readily infect and kill chicken and quail after experimental infection, and some were associated with significant mortality of chickens within the poultry retail markets in Hong Kong. Some genotypes are lethal for mice after intra-nasal inoculation and spread to the brain. On this occasion, the early detection of H5N1 viruses in the retail, live poultry markets led to preemptive intervention before the occurrence of human disease, but these newly emerging, highly pathogenic H5N1 viruses provide cause for pandemic concern.

Animals↗
PubMed2002

Molecular evolution of H6 influenza viruses from poultry in Southeastern China: prevalence of H6N1 influenza viruses possessing seven A/Hong Kong/156/97 (H5N1)-like genes in poultry.

The A/teal/Hong Kong/W312/97 (H6N1) influenza virus and the human H5N1 and H9N2 influenza viruses possess similar genes encoding internal proteins, suggesting that H6N1 viruses could become novel human pathogens. The molecular epidemiology and evolution of H6 influenza viruses were characterized by antigenic and genetic analyses of 29 H6 influenza viruses isolated from 1975 to 1981 and 1997 to 2000. Two distinct groups were identified on the basis of their antigenic characteristics. Phylogenetic analysis revealed that all H6N1 viruses isolated from terrestrial poultry in 1999 and 2000 are closely related to A/teal/Hong Kong/W312/97 (H6N1), and the nucleotide sequences of these viruses and of A/Hong Kong/156/97 (H5N1) were more than 96% homologous. The hemagglutinin (HA) of the 1999 and 2000 terrestrial viruses does not have multiple basic amino acids at the site of cleavage of HA1 to HA2; however, a unique insertion of aspartic acid in HA1 between positions 144 and 145 (H3 numbering) was found. The neuraminidase of these terrestrial H6N1 viruses has a deletion of 19 amino acids characteristic of A/Hong Kong/156/97 (H5N1). Evolutionary analysis suggested that these H6N1 viruses coevolved with A/quail/Hong Kong/G1/97-like H9N2 viruses and became more adapted to terrestrial poultry. These terrestrial 1999 and 2000 A/teal/Hong Kong/W312/97 (H6N1)-like viruses, along with the H9N2 viruses, could have been involved in the genesis of the pathogenic H5N1 influenza viruses of 1997. The presence of H6N1 viruses in poultry markets in Hong Kong that possess seven of the eight genes of the A/Hong Kong/156/97 (H5N1) virus raises the following fundamental questions relevant to influenza pandemic preparedness: could the pathogenic H5N1 virus reemerge and could the H6N1 viruses directly cross the species barrier to mammals?

Animals↗
PubMed2001

Emergence of influenza A viruses.

Pandemic influenza in humans is a zoonotic disease caused by the transfer of influenza A viruses or virus gene segments from animal reservoirs. Influenza A viruses have been isolated from avian and mammalian hosts, although the primary reservoirs are the aquatic bird populations of the world. In the aquatic birds, influenza is asymptomatic, and the viruses are in evolutionary stasis. The aquatic bird viruses do not replicate well in humans, and these viruses need to reassort or adapt in an intermediate host before they emerge in human populations. Pigs can serve as a host for avian and human viruses and are logical candidates for the role of intermediate host. The transmission of avian H5N1 and H9N2 viruses directly to humans during the late 1990s showed that land-based poultry also can serve between aquatic birds and humans as intermediate hosts of influenza viruses. That these transmission events took place in Hong Kong and China adds further support to the hypothesis that Asia is an epicentre for influenza and stresses the importance of surveillance of pigs and live-bird markets in this area.

Animals↗
PubMed2001

Structural features of HIV envelope defined by antibody escape mutant analysis.

Two neutralizing antibodies specific for the V3 sequence of HIV envelope were used to generate escape variants from the HTLV(IIIB) founder virus. The full gp120 sequence of each variant was then analyzed to identify mutations responsible for immune escape. As predicted, most escape variants harbored amino acid changes in the V3 crown sequence. However, one variant differed from its founder only within the conserved C2 region. These findings, when analyzed in conjunction with crystallographic data, suggest a new three-dimensional model for HIV envelope folding, in which the V3 loop extends across the CD4-binding face of gp120 to associate with discontinuous C2 residues. This envelope configuration may provide an effective immune defense mechanism for HIV, as the highly variable residues of the V3 loop may shield conserved amino acids pertinent to viral infection.

Amino Acid Sequence↗
PubMed2001

H9N2 influenza A viruses from poultry in Asia have human virus-like receptor specificity.

H9N2 influenza A viruses are currently widespread in chickens, quail, and other poultry in Asia and have caused a few cases of influenza in humans. In this study, we found that H9N2 viruses from Hong Kong live bird markets have receptor specificity similar to that of human H3N2 viruses. In addition, the neuraminidase of poultry H9N2 viruses has mutations in its hemadsorbing site, a characteristic resembling that of human H2N2 and H3N2 viruses but differing from that of other avian viruses. Peculiar features of surface glycoproteins of H9N2 viruses from Hong Kong suggest an enhanced propensity for introduction into humans and emphasize the importance of poultry in the zoonotic transmission of influenza viruses.

Amino Acid Substitution↗
PubMed2001

Universal primer set for the full-length amplification of all influenza A viruses.

To systematically identify and analyze the 15 HA and 9 NA subtypes of influenza A virus, we need reliable, simple methods that not only characterize partial sequences but analyze the entire influenza A genome. We designed primers based on the fact that the 15 and 21 terminal segment specific nucleotides of the genomic viral RNA are conserved between all influenza A viruses and unique for each segment. The primers designed for each segment contain influenza virus specific nucleotides at their 3'-end and non-influenza virus nucleotides at the 5'-end. With this set of primers, we were able to amplify all eight segments of N1, N2, N4, N5, and N8 subtypes. For N3, N6, N7, and N9 subtypes, the segment specific sequences of the neuraminidase genes are different. Therefore, we optimized the primer design to allow the amplification of those neuraminidase genes as well. The resultant primer set is suitable for all influenza A viruses to generate full-length cDNAs, to subtype viruses, to sequence their DNA, and to construct expression plasmids for reverse genetics systems.

Child↗
PubMed2001

Comparison of efficacies of RWJ-270201, zanamivir, and oseltamivir against H5N1, H9N2, and other avian influenza viruses.

The orally administered neuraminidase (NA) inhibitor RWJ-270201 was tested in parallel with zanamivir and oseltamivir against a panel of avian influenza viruses for inhibition of NA activity and replication in tissue culture. The agents were then tested for protection of mice against lethal H5N1 and H9N2 virus infection. In vitro, RWJ-270201 was highly effective against all nine NA subtypes. NA inhibition by RWJ-270201 (50% inhibitory concentration, 0.9 to 4.3 nM) was superior to that by zanamivir and oseltamivir carboxylate. RWJ-270201 inhibited the replication of avian influenza viruses of both Eurasian and American lineages in MDCK cells (50% effective concentration, 0.5 to 11.8 microM). Mice given 10 mg of RWJ-270201 per kg of body weight per day were completely protected against lethal challenge with influenza A/Hong Kong/156/97 (H5N1) and A/quail/Hong Kong/G1/97 (H9N2) viruses. Both RWJ-270201 and oseltamivir significantly reduced virus titers in mouse lungs at daily dosages of 1.0 and 10 mg/kg and prevented the spread of virus to the brain. When treatment began 48 h after exposure to H5N1 virus, 10 mg of RWJ-270201/kg/day protected 50% of mice from death. These results suggest that RWJ-270201 is at least as effective as either zanamivir or oseltamivir against avian influenza viruses and may be of potential clinical use for treatment of emerging influenza viruses that may be transmitted from birds to humans.

Acetamides↗
PubMed2001

Comparison of the activities of zanamivir, oseltamivir, and RWJ-270201 against clinical isolates of influenza virus and neuraminidase inhibitor-resistant variants.

RWJ-270201 is a novel cyclopentane inhibitor of influenza A and B virus neuraminidases (NAs). We compared the ability of RWJ-270201 to inhibit NA activity of clinical influenza isolates and viruses with defined resistance mutations with that of zanamivir and oseltamivir carboxylate. In NA inhibition assays with influenza A viruses, the median 50% inhibitory concentration (IC(50)) of RWJ-270201 (approximately 0.34 nM) was comparable to that of oseltamivir carboxylate (0.45 nM) but lower than that of zanamivir (0.95 nM). For influenza B virus isolates, the IC(50) of RWJ-270201 (1.36 nM) was comparable to that of zanamivir (2.7 nM) and less than that of oseltamivir carboxylate (8.5 nM). A zanamivir-resistant variant bearing a Glu119-to-Gly (Glu119-->Gly) or Glu119-->Ala substitution in an NA (N2) remained susceptible to RWJ-270201 and oseltamivir carboxylate. However, a zanamivir-selected variant with an Arg292-->Lys substitution in an NA (N2) showed a moderate level of resistance to RWJ-270201 (IC(50) = 30 nM) and zanamivir (IC(50) = 20 nM) and a high level of resistance to oseltamivir carboxylate (IC(50) > 3,000 nM). The zanamivir-resistant influenza B virus variant bearing an Arg152-->Lys substitution was resistant to each NA inhibitor (IC(50) = 100 to 750 nM). The oseltamivir-selected variant (N1) with the His274-->Tyr substitution exhibited resistance to oseltamivir carboxylate (IC(50) = 400 nM) and to RWJ-270201 (IC(50) = 40 nM) but retained full susceptibility to zanamivir (IC(50) = 1.5 nM). Thus, drug-resistant variants with substitutions in framework residues 119 or 274 can retain susceptibility to other NA inhibitors, whereas replacement of functional residue 152 or 292 leads to variable levels of cross-resistance. We conclude that RWJ-270201 is a potent inhibitor of NAs of wild-type and some zanamivir-resistant or oseltamivir-resistant influenza A and B virus variants.

Acetamides↗
PubMed2001

Efficacy of zanamivir against avian influenza A viruses that possess genes encoding H5N1 internal proteins and are pathogenic in mammals.

In 1997, an avian H5N1 influenza virus, A/Hong Kong/156/97 (A/HK/156/97), caused six deaths in Hong Kong, and in 1999, an avian H9N2 influenza virus infected two children in Hong Kong. These viruses and a third avian virus [A/Teal/HK/W312/97 (H6N1)] have six highly related genes encoding internal proteins. Additionally, A/Chicken/HK/G9/97 (H9N2) virus has PB1 and PB2 genes that are highly related to those of A/HK/156/97 (H5N1), A/Teal/HK/W312/97 (H6N1), and A/Quail/HK/G1/97 (H9N2) viruses. Because of their similarities with the H5N1 virus, these H6N1 and H9N2 viruses may have the potential for interspecies transmission. We demonstrate that these H6N1 and H9N2 viruses are pathogenic in mice but that their pathogenicities are less than that of A/HK/156/97 (H5N1). Unadapted virus replicated in lungs, but only A/HK/156/97 (H5N1) was found in the brain. After three passages (P3) in mouse lungs, the pathogenicity of the viruses increased, with both A/Teal/HK/W312/97 (H6N1) (P3) and A/Quail/HK/G1/97 (H9N2) (P3) viruses being found in the brain. The neuraminidase inhibitor zanamivir inhibited viral replication in Madin-Darby canine kidney cells in virus yield assays (50% effective concentration, 8.5 to 14.0 microM) and inhibited viral neuraminidase activity (50% inhibitory concentration, 5 to 10 nM). Twice daily intranasal administration of zanamivir (50 and 100 mg/kg of body weight) completely protected infected mice from death. At a dose of 10 mg/kg, zanamivir completely protected mice from infection with H9N2 viruses and increased the mean survival day and the number of survivors infected with H6N1 and H5N1 viruses. Zanamivir, at all doses tested, significantly reduced the virus titers in the lungs and completely blocked the spread of virus to the brain. Thus, zanamivir is efficacious in treating avian influenza viruses that can be transmitted to mammals.

Administration, Intranasal↗
PubMed2001

Characterization of a porcine lung epithelial cell line suitable for influenza virus studies.

We established a porcine lung epithelial cell line designated St. Jude porcine lung cells (SJPL) and demonstrated that all tested influenza A and B viruses replicated in this cell line. The infectivity titers of most viruses in SJPL cells were comparable to or better than those in MDCK cells. The propagation of influenza viruses from clinical samples in SJPL cells did not lead to antigenic changes in the hemagglutinin molecule. The numbers of both Sia2-3Gal and Sia2-6Gal receptors on SJPL cells were greater than those on MDCK cells. Influenza virus infection of SJPL cells did not lead to apoptosis, as did infection of MDCK cells. No porcine endogenous retrovirus was detected in SJPL cells, and in contrast to MDCK cells, SJPL cells did not cause tumors in nude mice.

Animals↗
PubMed2001

Cocirculation of avian H9N2 and contemporary "human" H3N2 influenza A viruses in pigs in southeastern China: potential for genetic reassortment?

Pigs are permissive to both human and avian influenza viruses and have been proposed to be an intermediate host for the genesis of pandemic influenza viruses through reassortment or adaptation of avian viruses. Prospective virological surveillance carried out between March 1998 and June 2000 in Hong Kong, Special Administrative Region, People's Republic of China, on pigs imported from southeastern China, provides the first evidence of interspecies transmission of avian H9N2 viruses to pigs and documents their cocirculation with contemporary human H3N2 (A/Sydney/5/97-like, Sydney97-like) viruses. All gene segments of the porcine H9N2 viruses were closely related to viruses similar to chicken/Beijing/1/94 (H9N2), duck/Hong Kong/Y280/97 (H9N2), and the descendants of the latter virus lineage. Phylogenetic analysis suggested that repeated interspecies transmission events had occurred from the avian host to pigs. The Sydney97-like (H3N2) viruses isolated from pigs were related closely to contemporary human H3N2 viruses in all gene segments and had not undergone genetic reassortment. Cocirculation of avian H9N2 and human H3N2 viruses in pigs provides an opportunity for genetic reassortment leading to the emergence of viruses with pandemic potential.

Animals↗
PubMed2001

Cross-reactive, cell-mediated immunity and protection of chickens from lethal H5N1 influenza virus infection in Hong Kong poultry markets.

In 1997, avian H5N1 influenza virus transmitted from chickens to humans resulted in 18 confirmed infections. Despite harboring lethal H5N1 influenza viruses, most chickens in the Hong Kong poultry markets showed no disease signs. At this time, H9N2 influenza viruses were cocirculating in the markets. We investigated the role of H9N2 influenza viruses in protecting chickens from lethal H5N1 influenza virus infections. Sera from chickens infected with an H9N2 influenza virus did not cross-react with an H5N1 influenza virus in neutralization or hemagglutination inhibition assays. Most chickens primed with an H9N2 influenza virus 3 to 70 days earlier survived the lethal challenge of an H5N1 influenza virus, but infected birds shed H5N1 influenza virus in their feces. Adoptive transfer of T lymphocytes or CD8(+) T cells from inbred chickens (B(2)/B(2)) infected with an H9N2 influenza virus to naive inbred chickens (B(2)/B(2)) protected them from lethal H5N1 influenza virus. In vitro cytotoxicity assays showed that T lymphocytes or CD8(+) T cells from chickens infected with an H9N2 influenza virus recognized target cells infected with either an H5N1 or H9N2 influenza virus in a dose-dependent manner. Our findings indicate that cross-reactive cellular immunity induced by H9N2 influenza viruses protected chickens from lethal infection with H5N1 influenza viruses in the Hong Kong markets in 1997 but permitted virus shedding in the feces. Our findings are the first to suggest that cross-reactive cellular immunity can change the outcome of avian influenza virus infection in birds in live markets and create a situation for the perpetuation of H5N1 influenza viruses.

Adoptive Transfer↗