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R F Bishop

Publications and source records attributed to R F Bishop.

At least 37 records · Page 2Linked to original sources
PubMed1997

Phase 1 trial of a candidate rotavirus vaccine (RV3) derived from a human neonate.

OBJECTIVE: To conduct a phase 1 safety and tolerability trial of an oral rotavirus vaccine candidate RV3 in healthy volunteers. METHODOLOGY: Double blind placebo controlled trial of a single 1 mL oral dose (6.5 x 10(5) fluorescing focus units [FFU]/mL) in 10 healthy young men, 10 3-4 year old children and 10 3 month old infants with a 4 week surveillance period. The study was undertaken at a children's hospital and nearby community in Melbourne, Australia. RESULTS: All subjects successfully completed the trial. There were no significant side-effects attributable to the vaccine preparation in any age group. No shedding of vaccine virus was detected by enzyme immunoassay. There was evidence of an immune response in serum and/or gut secretions in two of five vaccinees in each age group. CONCLUSION: RV3 rotavirus vaccine appears to be safe and well tolerated. Evidence of immunogenicity in some subjects after a single dose encourages further trials to determine immunogenicity after three doses, after reduction of viral dose, and without prior administration of buffer.

Administration, Oral↗
PubMed1997

Production of reassortant viruses containing human rotavirus VP4 and SA11 VP7 for measuring neutralizing antibody following natural infection.

The outer capsid proteins VP4 and VP7 of group A rotaviruses are both targets of neutralizing antibody produced following natural infection in humans. Of interest is the relative importance and immunodominance of each protein in the generation of a protective immune response. In order to measure neutralizing antibody responses to VP4 and VP7 separately, reassortants bearing VP4 of each of the major human rotavirus P types with VP7 of SA11 origin were successfully produced by neutralizing monoclonal antibody selection. The resulting reassortants, together with reassortants representing each of the major VP7 types, were antigenically characterized with serotype-specific neutralizing monoclonal antibodies and hyperimmune sera. The neutralization proteins of human rotavirus origin were found to be unaffected antigenically by reassortment. The abilities of these reassortants to discriminate between VP4 and VP7 immune responses were evaluated with postinfection sera collected from three patients infected with either a P1A[8],G1, a P1B[4],G2, or a P1A[8],G4 rotavirus strain. The reassortants were shown to be capable of separating the neutralizing antibody responses to VP4 and VP7, with each patient showing a different immune response with respect to VP4 or VP7 immunodominance. These reassortants can now be applied to analyses of individual immune responses to VP4 and VP7 proteins after primary rotavirus infections and reinfections in humans.

Antibodies, Monoclonal↗
PubMed1997

Comparison of enzyme immunoassay, PCR, and type-specific cDNA probe techniques for identification of group A rotavirus gene 4 types (P types).

This study was designed to evaluate three techniques most commonly used to identify the VP4 (P) types of human group A fecal rotaviruses. The techniques included PCR with nested primers and hybridization with PCR-generated probes (to determine the P genotypes). The results obtained by these genetic techniques were evaluated against those obtained by an enzyme immunoassay (EIA) incorporating neutralizing monoclonal antibodies (N-MAbs) reacting with three major human P serotypes (serotypes P1A, P1B, and P2A). The P types of the rotaviruses present in 102 fecal specimens were determined under code by each of the three assays. The specificity of each assay was evaluated against a "gold standard" putative P type (P serotype and genotype) deduced from knowledge of the VP7 (G) type and the origin of the fecal specimen. Overall comparison of the results showed respective sensitivities and specificities of 92 and 92% for reverse transcription-PCR, 80 and 99% for hybridization, and 73 and 91% for EIA with N-MAbs. The hybridization assay retained high sensitivity with specimens stored for > or = 10 years. Hybridization assays with nonradioactive probes are relatively inexpensive and are suited for use in developing countries. In summary, both genetic assays showed high sensitivities and specificities in assigning a P type to human fecal rotavirus strains. Further evaluation of the EIA with N-MAbs is required, together with incorporation of new N-MAbs for the detection of the additional P types detected in developing countries.

Antibodies, Monoclonal↗
PubMed1997

Sequence of the VP7 gene of an atypical human rotavirus: evidence for genetic and antigenic drift.

The nucleotide sequence of the gene encoding the outer capsid glycoprotein, VP7, isolated from a reassortant human rotavirus, M3014, was determined. The deduced amino acid sequence exhibited significant identity to the VP7 from a standard strain belonging to serotype G4, although the antigenic regions of the M3014 VP7 resembled sequences from both serotype G4 and G9 viruses. However, reactivity with G4 or G9 serotype-specific monoclonal antibodies was not observed. We suggest that the M3014 VP7 was derived from sequential mutation of a G4-like progenitor gene resulting in a protein with novel antigenic properties.

Amino Acid Sequence↗
PubMed1996

Natural history of human rotavirus infection.

Rotavirus infections occur repeatedly in humans from birth to old age. Most are asymptomatic or are associated with mild enteric symptoms. Infection in young children can be accompanied by severe life-threatening diarrhea, most commonly after primary infection. Annual childhood morbidity rates for severe diarrhea are similar worldwide. Mortality rates are low in developed countries but approach 1,000,000 annually in young children in developing countries. Rotaviruses can be classified into Groups A-E according to antigenic groups on VP6, the major capsid antigen. Only Group A,B and C rotaviruses have been shown to infect humans, and most human rotavirus disease is caused by Group A viruses. These are further classified into G and P types based on identification of antigens on the outer capsid proteins VP7 and VP4 respectively. Most severe infections in young children are caused by serotypes G1-4, and during the last two decades, G1 infections appear to have predominated worldwide. In general the more densely populated countries show the most complex patterns of occurrence of serotypes. Clinical rotavirus disease can be accompanied by shedding of > 10(12) rotavirus particles/gm feces. The virus is highly infectious and appears to retain infectivity over many months. In temperate climates, disease is most common during the colder months, when it is likely that rapid spread within families and communities occurs. Nosocomial infections are frequent, and rotaviruses can become endemic within obstetric hospital nurseries for the newborn. Few (if any) human rotavirus infections appear to be zoonoses, even though Group A rotaviruses are widespread in the young of all mammalian species. However infection of humans with reassortant rotavirus strains derived from human-animal sources can occur. The extent to which this contributes to new epidemic strains within particular countries (or worldwide) remains to be determined.

Age Factors↗
PubMed1996

G3P2 rotaviruses causing diarrhoeal disease in neonates differ in VP4, VP7 and NSP4 sequence from G3P2 strains causing asymptomatic neonatal infection.

During longitudinal epidemiological studies of rotavirus infections in children in Melbourne, Australia human G3P2 rotavirus strains causing asymptomatic or symptomatic infections have been identified. Eleven strains (AS strains) associated with asymptomatic infection of newborn babies from 1974-1984, and five strains (S strains) associated with symptomatic infection of newborn babies (4) or a 22 week old infant (1) during 1980-1986 were studied. The entire nucleotide sequences of genes coding for VP4, VP7, NSP4 and VP6 were derived for representative AS and S strains. The nucleotide sequences of neutralization epitope regions present on the outer capsid proteins VP4 and VP7 (regions C and F) showed extensive conservation of nucleotide and deduced amino acid sequence in all strains. Minor variations were observed over the 12 year period in VP7 epitope regions A and B in some strains. Specific conserved amino acids differences between the asymptomatic and symptomatic strains were observed in the genes encoding VP4 at aa133 and 303 (asparagine or threonine) and 380 (serine or isoleucine), VP7 at aa27 (threonine or isoleucine), aa29 (isoleucine or threonine), aa42 (valine or alanine) and aa238 (asparagine or aspartic acid/serine) and NSP4 at aa135 (isoleucine or valine). No amino acid changes were identified in gene 6. The observed amino acid differences occurred in proteins that have been implicated in virulence, and correlate with differences in clinical symptoms of infants infected with these strains. These results permit speculation about the genetic basis for virulence of human strains.

Amino Acid Sequence↗
PubMed1996

Human rotavirus VP4 contains strain-specific, serotype-specific and cross-reactive neutralization sites.

The neutralization epitopes of human rotavirus VP4 were studied by using a panel of neutralization monoclonal antibodies previously shown to be strain-specific (RV-3:3), serotype-specific (RV-5:2, ST-3:3) or cross-reactive (F45:4). Antigenic variants of human rotaviruses RV-3, ST-3, RV-5 and F45 resistant to neutralization by the appropriate of VP4 specific monoclonal antibodies (RV-3:3, ST-3:3, RV-5:2 and F45:4 respectively) were selected. By nucleotide sequence analysis and single strand conformational polymorphism analysis of these variants, three sites of neutralization on VP5* and one site on VP8* were identified. At or near to the putative fusion region on VP5*, a strain-specific site (aa383), a serotype P1A-P2 cross-reactive site (aa392) and a serotype P2-specific site (aa397) were found. On VP8*, a serotype P1B-specific site at aa148 was detected. These results confirmed the importance of the putative fusion region in neutralization and have identified a new neutralization site in the hypervariable region of VP8* which is specific for serotype P1B human rotaviruses.

Amino Acid Sequence↗
PubMed1996

Serum, fecal, and breast milk rotavirus antibodies as indices of infection in mother-infant pairs.

Sixty-eight mother-infant pairs were followed for 12-17 months after birth. Rotavirus infections in children were detected by EIA of weekly fecal antigen and anti-rotavirus IgA levels, by EIA of anti-rotavirus IgG in sera at birth, 6, or 12-17 months of age, and by anti-rotavirus EIA IgA and neutralizing antibody (NA) in monthly samples of maternal breast milk. Primary rotavirus infection was detected in 26 children (in 15 [58%] by fecal excretion, 12 [46%] by IgG seroconversion, and 22 [85%] by elevations of IgA anti-rotavirus antibodies [IgA coproconversion] in consecutive fecal specimens). Rotavirus "challenge" was detected by rises in levels of NA in breast milk in 9 (47%) of 19 mothers, including 5 (26%) from pairs in which there was no other evidence of rotavirus infection. Reinfections were detected in 2 children by rotavirus excretion and in 4 by coproconversion. IgA coproconversion is the most sensitive technique for detection of symptomatic and asymptomatic rotavirus infection in young children.

Age Factors↗
PubMed1996

Amino acids involved in distinguishing between monotypes of rotavirus G serotypes 2 and 4.

Neutralizing monoclonal antibodies (N-MAbs) to serotype G2 and G4 rotaviruses were used to study intraserotypic variation by selection and characterization of N-MAb-resistant antigenic variants and reaction of N-MAbs with prototype rotavirus strains. Two G2-specific N-MAbs reacted with G2 rotaviruses S2, DS-1, RV-5 and RV-6 but not with 1076. Sequence analysis of the gene encoding VP7 of 1076 virus showed that the differences in amino acid sequence between 1076 virus and the other G2 strains at position 147, 213 and 217 correlated with the loss of N-MAb reactivity. Rotavirus variant mutation mapping data suggested that the amino acid difference at position 213 was likely to be of greatest importance. Rotavirus 1076 was defined as monotype b within G2 strains, whereas S2, DS-1, RV-5 and RV-6 belong to monotype a. The molecular basis for G4 subtypes/monotypes was also studied. The monotype G4b N-MAb 3A3 selected an antigenic variant with an amino acid mutation at position 96, whereas variants of the G4a-reactive N-MAb ST-3:1 showed a mutation at position 94, which produced a new, utilized glycosylation site. Neutralization by N-MAb ST-3:1 was also affected by amino acid changes at position 96. Reactions with these N-MAbs show that serotype G2 viruses can be divided into monotypes and confirm the observation that serotype G4 rotaviruses can be subdivided into subtypes/monotypes a and b. The G2 monotypes relate to differences at particular amino acids within antigenic region C and possibly region B, whereas antigenic region A is most important for G4 monotype differentiation.

Amino Acid Sequence↗
PubMed1996

Multiple-gene rotavirus reassortants responsible for an outbreak of gastroenteritis in central and northern Australia.

Two rotavirus strains, E210 and E212, implicated in epidemics of gastroenteritis in children in central and northern Australia during 1993-1994, exhibited the unusual combination of a 'short' RNA electrophoretic pattern and subgroup II specificity. The outer capsid protein VP7 was found by PCR typing and sequence analysis to be related to that of serotype G2 viruses. Both strains displayed a novel pattern of reactivity to G2-specific monoclonal antibodies that correlated with sequence variation in the antigenic regions of VP7. The VP4 serotype of E210 and E212 was determined as P1B in an enzyme immunoassay, consistent with other G2 viruses. Analysis of the VP6 gene indicated significant identity (98-99%) with other human subgroup II viruses. Northern hybridization analysis of E210 RNA using total genome probes derived from the prototype strains RV4 and RV5 indicated that E210 was derived from multiple gene reassortment between rotaviruses belonging to different genetic types.

Amino Acid Sequence↗
PubMed1996

Annual incidence, serotype distribution, and genetic diversity of human astrovirus isolates from hospitalized children in Melbourne, Australia.

The incidence of astrovirus infection in children under 5 years of age hospitalized for acute gastroenteritis in Melbourne, Australia, during 1995 was determined. Astrovirus was detected in 16 fecal specimens by Northern (RNA) dot blot analysis of RNA isolated from feces with an astrovirus-specific cDNA probe. The incidence of astrovirus infection was determined as 4.2% (16 of 378 total samples) compared with rates of 63.2, 3.7, and 4.2% for rotavirus, adenovirus, and all bacterial pathogens, respectively. Astrovirus was detected during the winter season and mainly in infants between 6 and 12 months of age. Serotyping of samples was carried out by reverse transcriptase PCR and direct sequencing of a 348-bp region of the capsid protein gene. Type 1 strains predominated (11 of 13 typeable samples), although type 4 isolates were also detected. Astrovirus was retrospectively identified in 13 fecal samples collected from hospitalized infants between 1980 and 1985 and shown to contain small viruses by electron microscopy. Type 1 isolates were again the most common, although a type 5 strain was also found. Comparative sequence analysis indicated that type 1 astroviruses exhibited up to 7% sequence divergence over a 15-year period; however, all mutations were silent. The incidence of astrovirus reported here indicates that the virus is a significant cause of severe diarrhea in young children. The genetic analysis also provides important molecular epidemiological information relevant to the development of preventative therapies.

Astroviridae Infections↗
PubMed1996

Oscillatory fluctuations in the incidence of rotavirus infections by serotypes 1, 2, 3, and 4.

The statistical evidence for regularity in the epidemic cycles of rotavirus infection for serotypes 1, 2, 3, and 4 was examined. Hospitalization longitudinal data of the monthly incidence of rotavirus infections from the city of Melbourne, Australia during 1977-1993 were used. Periodograms were used for exploring seasonal and longer-term cycles (interepidemic periods) of rotavirus infection. There was a satisfactory agreement between the interepidemic period estimated by means of periodograms with the one predicted by theoretical epidemiological studies. Thus, there is a clear evidence of a biennial peak in the epidemiology of rotavirus. Results of the study show an evidence of the likely existence of an interepidemic cycle of 4.6-5.2 years of duration. The finding of this interepidemic cycle was unexpected, and does not arise from the alternating incidence of the 4 serotypes since this peak appears in the periodogram of each serotype.

Australia↗
PubMed1995

Genetic and antigenic characterization of a serotype G6 human rotavirus isolated in Melbourne, Australia.

An unusual rotavirus strain, MG6, was isolated from a 16-month-old child admitted to hospital with acute gastroenteritis. The virus could not be serotyped (G-typed) by enzyme immunoassay using standard reagents specific for common serotypes of human Group A rotaviruses. Nucleotide sequencing of cDNA derived from the gene encoding the outer capsid protein, VP7, and deduction of the VP7 amino acid sequence indicated that this strain belonged to serotype G6, a serotype normally associated with viruses causing disease in cattle. This was confirmed by polymerase chain reaction typing and enzyme immunoassay using a G6-specific monoclonal antibody. The VP4 genotype of MG6 was determined by hybridization of its VP4 cDNA to genomic RNA isolated from standard strains of defined P-types. This analysis, confirmed by deduced amino acid sequence analysis, classified MG6 into the novel genotype P13. MG6, therefore, is related to the previously described G6P13 human strain PA169, isolated in Italy. The emergence of strain MG6, the first human G6 rotavirus identified in Australia, provides further evidence of reassortment between human and animal rotaviruses.

Acute Disease↗
PubMed1995

Patients with enteric adenovirus gastroenteritis admitted to an Australian pediatric teaching hospital from 1981 to 1992.

During the period 1981 to 1992, 4,473 fecal specimens collected from children hospitalized with acute gastroenteritis at the Royal Children's Hospital, Melbourne, Australia, were examined by electron microscopy. A monoclonal antibody enzyme immunoassay for enteric adenovirus (EAd) types 40 (Ad40) and 41 (Ad41) was used when adenoviruses were visualized. Fecal samples were positive for adenovirus by both electron microscopy and enzyme immunoassay in 138 patients (3.1%). Ad40 was identified in 19 children (14%), and Ad41 was identified in 119 children (86%). These EAd were identified during each of the 12 years surveyed. EAd were present year-round, but the annual number of hospitalizations was not constant. Yearly prevalence varied from 0.7% (1981) to 6.5% (1985). This was associated with monthly fluctuations in Ad41 activity, with overall peak monthly prevalence in May (late autumn). By contrast, Ad40 numbers remained low and constant year-round. The frequency of Ad41 relative to Ad40 increased from 25% in 1981 to exceed 75% after 1983. Children admitted with EAd infection were more likely to have diarrhea for more than 5 days (P < 0.001) but less likely to be febrile or dehydrated (P < 0.05) than children with rotavirus infection. EAd are responsible for enteric symptoms of only a fraction of hospitalized children with infectious diarrhea but result in a more-protracted illness than rotavirus. Their relationship to persistent diarrhea requires further investigation.

Adenoviridae↗
PubMed1994

Genetic analysis of NSP1 genes of human rotaviruses isolated from neonates with asymptomatic infection.

The nucleotide sequences of the genomic RNA segments 5 (gene 5) encoding the non-structural protein NSP1 of rotavirus strains M37 and ST3, isolated from neonates with asymptomatic infection, were determined. The sequences were similar overall (95% identity) as were the deduced amino acid sequences of NSP1 (93%). However, the M37 and ST3 NSP1 proteins shared only 82% and 81% identity, respectively, with the corresponding protein from another strain isolated from a neonate with asymptomatic infection (I321). Differences (of between 15% and 31%) were found in comparison with NSP1 sequences of rotaviruses isolated from older children with symptomatic infection (Wa, DS1 and IGV-80-3). Using an M37 gene 5-derived probe, Northern hybridization analysis of total genomic RNA extracted from viruses isolated from older children (Wa, RV4, RV5 and P) and neonates (M37, ST3, RV3 and 1076), representatives of the most common human G and P types, further indicated that while the gene 5 alleles of strains M37, ST3 and RV3 had a high degree of identity, no significant identity between 1076 and M37 was observed. In addition, cross-hybridization between the M37 probe and RNA of strains from older children (Wa, RV4 and P) was evident. Thus, neonatal human rotavirus strains do not carry a common NSP1 gene.

Amino Acid Sequence↗
PubMed1994

Sequences of VP6 genes of human rotavirus strain RV3 and its vaccine derivative.

The nucleotide sequence of the genomic segment encoding the major antigen, VP6, of human rotavirus strain RV3 was determined for the virus isolated from faeces. This was compared with the sequence of the cognate gene of tissue culture-adapted candidate vaccine virus that was derived from RV3 and had been passaged 30 times. A single silent nucleotide difference was detected between the two genes and the deduced amino acid sequence for both showed the highest degree of identity (> or = 97.5%) with the VP6 proteins of rotavirus strains belonging to the same subgroup. The amino acid residues that might affect VP6 subgroup epitope type from RV3 and other viruses of the same or different subgroups were compared. The commonality of particular amino acids among strains of different subgroups suggests that the presence of all subgroup-specific amino acids may be necessary for subgroup determination.

Amino Acid Sequence↗