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Biomedical subjects

R L Ward

Publications and source records attributed to R L Ward.

At least 109 records · Page 6Linked to original sources
PubMed1992

Seroepidemiologic evaluation of antibodies to rotavirus as correlates of the risk of clinically significant rotavirus diarrhea in rural Bangladesh.

A case-control study was conducted among children and adult women in rural Bangladesh to evaluate whether serologic immunity to rotavirus was associated with a lower risk of rotavirus diarrhea of sufficient severity to cause patients to seek medical care. Acute-phase sera from 219 cases of rotavirus diarrhea, detected among patients treated in three diarrheal treatment centers, were compared with sera from 477 contemporaneously selected community controls. Overall, serum IgG antirotavirus antibody titers were nearly one-fourth as high in cases as in controls (107 vs. 417 units/ml; P less than .001). Among persons aged greater than or equal to 8 months, in whom titers of maternal antirotavirus antibodies should have been negligible, even the lowest range of detectable titers (100-200 units/ml) was associated with a substantial (75%, P less than .05) reduction of the risk of rotavirus diarrhea. We conclude that titers of serum IgG antirotavirus antibodies induced by earlier infection were inversely related to the risk of clinically significant rotavirus diarrhea.

Adult↗
PubMed1992

Evidence that protection against rotavirus diarrhea after natural infection is not dependent on serotype-specific neutralizing antibody.

This case-control study sought to determine whether protection against clinically significant rotavirus diarrhea in children aged 4-35 months correlated with titers of serum neutralizing antibody and, if so, whether this protection was serotype-specific. Titers of acute-phase sera from 156 cases of treated rotavirus diarrhea in rural Bangladesh were contrasted with titers from 312 contemporaneously selected, age-matched controls. Analyses of the culture-adapted rotaviruses from the cases revealed that 24%, 15%, 43%, and 17% belonged to serotypes 1-4, respectively. Titers of both homologous and heterologous neutralizing antibody in acute blood specimens of cases were significantly lower than those of matched controls. However, multivariate logistic regression models demonstrated that only antibody titers to heterotypic rotaviruses were independently associated with protection against rotavirus disease. These data, which indicate that the correlation of protection with neutralizing antibody titers is not serotype-specific, suggest that immunity to rotavirus disease may be mediated by other factors.

Algorithms↗
PubMed1992

Measurement of human phospholipase A2 in arthritis plasma using a newly developed sandwich ELISA.

A sandwich enzyme-linked immunosorbent assay (ELISA) for human non-pancreatic phospholipase A2 (PLA2) was developed using monoclonal antibodies raised against purified recombinant PLA2. This assay was shown to be specific for human non-pancreatic PLA2, showing no cross-reactivity with human pancreatic PLA2 or with snake venom PLA2 (Crotalus durissus). The immunoassay showed no cross-reactivity with plasma components, and was reproducible and quantitative between 39 pmol and 2.7 nmol PLA2/1. The levels of non-pancreatic PLA2 in the plasma of patients with arthritis was measured using this immunoassay. There were significantly higher levels of PLA2 in patients with rheumatoid arthritis than in those with osteoarthritis or healthy controls. Plasma PLA2 was highest in those patients with active rheumatoid arthritis.

Aged↗
PubMed1992

Inhibition of rotavirus in vitro transcription by optimal concentrations of monoclonal antibodies specific for rotavirus VP6.

Three monoclonal antibodies (MAbs) obtained from inoculation of mice with either a serotype 1 human rotavirus or rotavirus SA11 (serotype 3) inhibited the in vitro transcription of rotavirus SA11. Two of the MAbs exhibited a biphasic inhibitory response. Removal of antibody from MAb preparations by adsorption with Sepharose-Protein G reduced the inhibitory activity completely for all three MAb preparations. Analysis by radioimmunoprecipitation and Western blotting indicated that all three MAbs reacted with VP6. All MAbs also reacted with four group A rotavirus serotypes by ELISA, but did not cross-react with reovirus type 1, poliovirus type 2 or MA-104 cell lysates. Transcription of four rotavirus serotypes as well as epizootic diarrhoea of infant mice rotavirus was inhibited when tested with two of the MAbs. Transcription of both purified single-shelled virus and purified heat-activated double-shelled SA11 rotavirus was inhibited by purified MAb. Our results indicate that these MAbs can be used effectively to study the events associated with rotavirus transcription.

Antibodies, Monoclonal↗
PubMed1991

Development of an improved method for measuring neutralizing antibody to rotavirus.

An improved assay was developed for measuring neutralizing antibody titers against rotaviruses. This procedure used the same initial steps as performed to determine antibody titers by the focus reduction neutralization (FRN) assay. However, instead of counting infected cells after staining with fluorescein, reductions in virus infectivity by neutralizing antibody were determined by quantitation of viral antigen production using an ELISA. A linear relationship was found between ELISA absorbance values and focus forming units for each of four prototype rotaviruses, representative of serotypes 1-4. Thus, the serum dilution that resulted in neutralization of 60% of infectious virus (i.e. the neutralizing antibody titer) could be readily determined from absorbance values. Titers found by this method were similar to and as reproducible as those found by the FRN assay. Because this method is less laborious and the results are obtained by objective rather than subjective methods, it represents an improvement over the FRN assay.

Adult↗
PubMed1991

Protection from rotavirus reinfection: 2-year prospective study.

To measure protection induced by natural rotavirus infection, 163 infants enrolled in a rotavirus vaccine trial were prospectively followed for 2 years. Serotype 1 rotaviruses were the predominant circulating strains during the study. Over the 2 years of observation, significantly fewer infants infected before enrollment developed a symptomatic reinfection (0 of 21) or any reinfection (4 of 21) compared with previously uninfected infants (P = .0003). Of the 60 infants who developed a primary rotavirus infection in the first year (40 symptomatic, 20 asymptomatic) only 4 were reinfected in the second year compared with 29 of 82 subjects not previously infected (P = .00003). Asymptomatic primary infection appeared to be as protective as symptomatic primary infection. The only symptomatic reinfections occurred in 2 subjects who did not develop rotavirus antibody after the initial detection of rotavirus. An age-related reduction in the ratio of symptomatic to asymptomatic primary rotavirus infection was also detected. In this study, protection against homotypic serotype 1 reinfection appeared to last greater than or equal to 2 years.

Age Factors↗
PubMed1991

Reactivities of serotyping monoclonal antibodies with culture-adapted human rotaviruses.

Rotaviruses collected in Bangladesh during 1985 to 1986 were culture adapted and used in a comparative serotyping study with three groups of monoclonal antibodies, all of which reacted with the major neutralization protein (VP7) of serotype 1, 2, 3, or 4. The goals were to determine which monoclonal antibodies most accurately predicted the serotype and why large variations in serotyping efficiencies have occurred with these monoclonal antibodies in previous studies. The 143 rotavirus isolates used in this study belonged to 69 different electropherotypes; and 44, 23, 21, and 55 isolates were identified as serotype 1 through 4, respectively, by neutralization with serotype-specific hyperimmune antisera. Serotyping specificity by enzyme-linked immunosorbent assay with monoclonal antibodies was 100% consistent with results found by neutralization with polyclonal antisera, but large differences were observed in the sensitivities of the different monoclonal antibodies. Monoclonal antibodies 5E8 (serotype 1), 1C10 (serotype 2), 159 (serotype 3), RV3:1 (serotype 3), ST-3:1 (serotype 4), and ST-2G7 (serotype 4) reacted with all the isolates of the corresponding serotype for which there were sufficient infectious particles. Monoclonal antibody 2F1 (serotype 2) was much less sensitive and reacted with only five serotype 2 isolates, but these were among those with the highest titers. Monoclonal antibodies RV4:2 (serotype 1), KU6BG (serotype 1), RV5:3 (serotype 2), and S2-2G10 (serotype 2), on the other hand, failed to react with between one and three isolates of the corresponding serotypes which had high titers, apparently because of epitope changes in these isolates. Effects of epitope variation were, however, most apparent with monoclonal antibodies 2C9 (serotype 1) and YO-1E2 (serotype 3), which reacted with one and no isolates of the corresponding serotypes, respectively. Cross-neutralization of escape mutants indicated that the serotype 1 monoclonal antibodies 5E8, 2C9, and RV4:2 reacted with different but probably overlapping epitopes, as did serotype 2 monoclonal antibodies 2F1, 1C10, and RV5:3, finding that were consistent with the enzyme-linked immunosorbent assay data. Because of epitope variations between rotavirus strains, serotyping with several monoclonal antibodies directed at different epitopes may increase the sensitivity of the method.

Antibodies, Monoclonal↗
PubMed1991

Culture adaptation and characterization of group A rotaviruses causing diarrheal illnesses in Bangladesh from 1985 to 1986.

Group A rotaviruses collected between 1985 and 1986 during comprehensive surveillance of treated diarrheal episodes occurring in a rural Bangladesh population were culture adapted and characterized by electropherotype, serotype, and subgroup. Of 454 episodes of rotavirus-associated diarrhea, rotaviruses were culture adapted from 381 (84%), and 335 contained 11 electrophoretically identical segments in unpassaged and cultured preparations. These 335 comprised 69 different electropherotypes with between 1 (32 isolates) and 79 representatives. The persistence of specific rotavirus strains within the study population, as defined by the detection of viruses with particular electropherotypes, was generally limited to a period of only a few months. All 335 isolates were serotyped by neutralization with hyperimmune antisera to prototype rotavirus strains representative of serotypes 1 to 4, i.e., Wa, DS-1, P, and ST-3. It was found that 80, 48, 119, and 88 isolates belonged to serotypes 1 to 4, respectively. The concentrations of hyperimmune antisera required to neutralize these isolates, however, were at least threefold greater than those needed to neutralize the homologous strains. Therefore, the isolates appeared to have altered neutralization epitopes from their prototype strains. Furthermore, the serotype 4 isolates were consistently shown to be much more closely related to the serotype 4B VA70 strain than the serotype 4A ST-3 strain. All but two isolates identified as serotypes 1, 3, or 4 had long electropherotypes and were subgroup II, and all but one serotype 2 isolate were subgroup I and had short electropherotypes. The three disparate strains appeared to be genetic reassortants. Evidence is presented that dual infections required for reassortant formation were not uncommon. Thus, formation of multiple reassortants may have been a cause for the observed rapid shift in viral strains within the study population.

Age Factors↗
PubMed1991

Prevention of surface-to-human transmission of rotaviruses by treatment with disinfectant spray.

A model was developed to examine the effects of disinfectants on the transmission of infectious rotavirus from a dried surface to humans. The initial experiments were designed to find a method of preserving rotavirus infectivity during drying. Culture-adapted human rotavirus (CJN strain) was dried at room temperature in different organic suspensions, including fecal matter, several laboratory media, and nonfat dry milk (NDM). Recoveries of infectious virus were then compared. Fecal matter provided little protection in this study relative to distilled water, but the other suspensions were quite protective, especially NDM, which consistently allowed recoveries of greater than 50%. When 10(3) focus-forming units of unpassaged CJN virus were dried in NDM and administered to subjects who licked the dried material, 100% (8 of 8) became infected. The effect of Lysol brand disinfectant spray (LDS) was next examined. Although NDM provided some protection against inactivation by LDS, spraying under conditions recommended by the manufacturer consistently caused the CJN virus titer to decrease greater than 5 log10. Consumption of CJN virus (10(3) focus-forming units) sprayed with LDS caused no infection in 14 subjects, whereas 13 of 14 subjects who consumed the unsprayed virus became infected (P less than 0.00001). The methods developed in this study could be used to test the effects of other disinfectants on the spread of infectious rotavirus from inanimate surfaces to humans.

Adolescent↗
PubMed1991

Formation and selection of intergenogroup reassortants during cell culture adaptation of rotaviruses from dually infected subjects.

A previous study showed that intergenogroup reassortants of human rotaviruses can persist in nature (R.L. Ward, O. Nakagomi, D.R. Knowlton, M.M. McNeal, T. Nakagomi, J.D. Clemens, D.A. Sack, and G.M. Schiff, J. Virol. 64:3219-3225, 1990), but the mechanisms involved in their formation and selection had not been determined. In this study it was shown that, during cell culture adaptation of rotaviruses belonging to different genogroups from stools of dually infected subjects, intergenogroup reassortants were formed and selected, presumably mimicking the processes that occur in nature.

Animals↗
PubMed1990

Protection of adults rechallenged with a human rotavirus.

Previous studies of adults challenged with human rotavirus (CJN strain) showed that 74% became infected and 55% of those infected experienced illness. Protection against infection correlated with rotavirus antibody, most significantly (P = .005) serum rotavirus IgG. In this study, 20 previously challenged subjects were reinoculated with the same virus 9-12 months after their initial challenge. Only 1 of 8 subjects not infected after the initial challenge and 2 of 12 infected after the first inoculation became infected after reinoculation; none became ill. Titers of rotavirus antibodies (serum, jejunal, and stool) at the time of reinoculation were about as high as or higher than they were before the initial inoculation. This correlated with greater protection, but the extent of protection was significantly greater (P less than .0001) than predicted based on a previous model relating protection and preinoculation titers of serum rotavirus IgG. Thus, inoculation with human rotavirus provided homotypic protection for at least 9-12 months, and protection remained correlated with higher concentrations of rotavirus antibody. However, the specific relationship between protection and rotavirus antibody was altered after the initial inoculation.

Adult↗
PubMed1990

Evaluation of WC3 rotavirus vaccine and correlates of protection in healthy infants.

The safety, immunogenicity, and efficacy of WC3 rotavirus vaccine was evaluated in a double-blind, placebo-controlled trial of healthy infants 2-12 months of age; 103 received one dose of vaccine and 103 received placebo. Vaccination appeared to be safe and induced an antibody response (WC3 neutralizing antibody) in 97% of vaccinees. Only 9 (9%) of these, however, produced antibody to human rotavirus serotypes; at least 7 of the 9 were naturally infected before vaccination. Neither the number of symptomatic episodes of rotavirus diarrhea (21 vs 25) nor the number of moderate to severe rotavirus illnesses (9 vs. 15) was significantly different in vaccine or placebo recipients, respectively, during a predominantly serotype 1 rotavirus season. A slight but significant decrease in mean symptom score was detected in vaccine recipients. Despite an overall lack of efficacy, protection could be correlated to previous rotavirus infection, high levels of WC3 neutralizing antibody, and preexisting (maternal) serotype 1 neutralizing antibody with a titer greater than or equal to 30.

Antibodies, Viral↗
PubMed1990

Effect of vaccination on serotype-specific antibody responses in infants administered WC3 bovine rotavirus before or after a natural rotavirus infection.

In an evaluation of WC3 bovine rotavirus (serotype 6) vaccine in infants, some subjects experienced a natural serotype 1 rotavirus infection before vaccination and others after. Therefore, the effects of both WC3 and natural rotavirus strains as either primary or boosting immunogens on serotype-specific neutralizing antibody responses could be determined. After primary natural infection (symptomatic or asymptomatic), neutralizing antibody titers were highest to serotype 1 but were consistently high to serotype 3, and low titers (greater than or equal to 20) to serotypes 2 and 4 were often detected. Previous vaccination with WC3 had little effect on the magnitude of these responses. In contrast, subjects infected with serotype 1 strains before vaccination experienced large (average, 12-fold) rises in neutralizing antibody to human serotypes 1-4 when vaccinated with WC3. Thus, although WC3 and the natural strains are distinct serotypes, their epitopes were sufficiently similar that reinfection with WC3 could boost neutralizing antibody titers to human serotypes in subjects primed by a previous natural infection.

Antibodies, Viral↗
PubMed1990

Development of an adult mouse model for studies on protection against rotavirus.

Although mice have been used as an animal model for studies on rotavirus disease, these studies have been limited by the short time period after birth during which mice are susceptible to rotavirus illness (i.e., approximately 15 days). To overcome this limitation, an adult mouse model was developed in which the endpoint was infection rather than illness. The model developed utilized a strain of mouse rotavirus (EDIM) adapted to grow in culture by multiple passages in MA104 cells. The second cell culture passage of EDIM caused severe diarrhea in neonatal BALB/c mice, and little or no amelioration of disease was observed after nine cell culture passages, even when this preparation was plaque purified. Oral administration of 2 x 10(3) PFU of passage 9 also consistently caused infection of mice 4, 10, 15, 30, 60, 120, and 180 days of age as determined by viral shedding and seroconversion. Reinoculation of these mice with the same virus preparation at 2, 3, or 4 months after the first inoculation produced no evidence of reinfection. In contrast, infection of neonatal mice with the heterotypic WC3 bovine rotavirus did not prevent reinfection with culture-adapted EDIM. Thus, this strain of EDIM caused consistent infection of previously uninoculated neonatal and adult BALB/c mice and produced homotypic but not heterotypic protection against reinfection.

Animals↗
PubMed1990

Serum-neutralizing antibody to VP4 and VP7 proteins in infants following vaccination with WC3 bovine rotavirus.

Serum specimens from infants 2 to 12 months old vaccinated with the WC3 bovine rotavirus were analyzed to determine the relative concentrations of neutralizing antibody to the VP4 and VP7 proteins of the vaccine virus. To do this, reassortant rotaviruses that contained the WC3 genome segment for only one of these two neutralization proteins were made. The segment for the other neutralization protein in these reassortants was from heterotypic rotaviruses that were serotypically distinct from WC3. Sera were examined from 31 infants who had no evidence of a previous rotavirus infection and the highest postvaccination WC3-neutralizing antibody titers (i.e., 160 to 600) of the 103 subjects administered the vaccine. A reassortant (3/17) that contained both neutralization proteins from the heterotypic rotaviruses, i.e., EDIM (EW strain of mouse rotavirus) VP7 and rhesus rotavirus VP4, was not neutralized by these sera (geometric mean titer [GMT], less than 20). A reassortant (E19) that contained EDIM VP7 and WC3 VP4 was also very poorly neutralized by these antisera (GMT = 20). In contrast, antibody titers to a reassortant (R20) that contained WC3 VP7 and rhesus rotavirus VP4 were higher than those against WC3 (GMTs of 458 and 313, respectively). Thus, VP7 appeared to be the dominant immunogen for production of neutralizing antibody after intestinal infection of previously uninfected infants vaccinated with WC3 bovine rotavirus.

Animals↗
PubMed1990

Evidence for natural reassortants of human rotaviruses belonging to different genogroups.

Of 335 rotavirus isolates associated with diarrheal disease in Bangladesh that were culture adapted and subsequently characterized for electropherotype, subgroup, and serotype, 9 had properties that suggested they may be natural reassortants between human rotaviruses belonging to different "genogroups." Two of these were examined in greater detail by RNA-RNA hybridization with prototype strains representative of each of the three proposed human rotavirus genogroups. One subgroup II isolate, 248, with a "long" electrophoretic pattern was neutralized by hyperimmune antisera to both serotype 2 and 4 strains. Consistent with these results, seven RNA segments of this isolate formed hybrids with human strains belonging to the Wa genogroup and four segments hybridized with strains belonging to the DS-1 genogroup. The second isolate examined, 456, belonged to subgroup II and had a long electrophoretic pattern but was found to be a serotype 2 strain. This isolate also appeared to be an intergenogroup reassortant because three of its segments formed hybrids with strains belonging to the Wa genogroup and eight hybridized with viruses of the DS-1 genogroup. On the basis of the relative migration rates of these RNA-RNA hybrids during gel electrophoresis, a suggested origin for each gene segment was proposed which was consistent with the results expected from electrophoretic, subgroup, and serotypic analyses.

Blotting, Northern↗
PubMed1990

Neutralization epitopes on rotavirus SA11 4fM outer capsid proteins.

The VP7 and VP4 genes of seven antigenic mutants of simian rotavirus SA11 4fM (serotype 3) selected after 39 passages in the presence of SA11 4fM hyperimmune antiserum, were sequenced. Nucleotide sequence analysis indicated the following. (i) Twice as many amino acid substitutions occurred in the VP7 protein than in VP4, which has a molecular weight twice that of VP7. (ii) Most amino acid changes that occurred clustered in six variable regions of VP7 and in two variable regions of VP4; these variable regions may represent immunodominant epitopes. (iii) Most amino acid substitutions that occurred in VP7 and VP4 of these mutants were also observed in antigenic mutants selected with neutralizing monoclonal antibodies (NMAbs); however, some amino acid substitutions occurred that were not selected for NMAbs. (iv) On VP7, some of the neutralization epitopes appeared to be interrelated because amino acid substitution in one site affected binding of specific NMAbs to other sites, while other neutralization epitopes on VP7 appeared to be independent, in that amino acid substitution in one site did not affect the binding of NMAbs to another distant site.

Animals↗