Mycoplasma contamination in cell culture of Crohn's disease material.
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Biomedical subjects
Publications and source records attributed to R G Wyatt.
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The possibility of immunizing human infants against rotaviruses, which cause severe dehydrating diarrheal disease, may depend on the use of a related rotavirus, derived from another animal species, as a source of antigen. To test the feasibility of this approach, calves were infected in utero with a bovine rotavirus and challenged with bovine or human type 2 rotavirus shortly after birth. Infection in utero with bovine rotavirus induced resistance to diarrheal disease caused by the human virus as well as the homologous bovine virus. These data suggest that the bovine virus is sufficiently related antigenically to the human type 2 virus to warrant further evaluation of the former as a source of vaccine.
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Twenty-five separate outbreaks of nonbacterial gastrointestinal illnesses were studied serologically for evidence of infection with the Norwalk virus and the rotaviruses that affect humans. Eight of 25 outbreaks appeared to be related to the Norwalk virus. In one of the 25 outbreaks, there was evidence of rotavirus infection. These observations suggest that the Norwalk virus or serologically related agents play an important role in epidemic nonbacterial gastroenteritis in adults and older children.
Human rotavirus (HRV) type 1 or 2, adenovirus, or non-cultivatable 27 nm virus-like particles were demonstrated by electron microscopy and/or rotavirus ELISA in fecal samples from 45.5% of 604 gastroenteritis inpatients, 25.0% of 200 gastroenteritis outpatients and 6.0% of 812 control subjects, all sampled at Children's Hospital National Medical Center. Washington, DC. Rotaviruses were the most common pathogens detected as 39% and 22% of gastroenteritis inpatients and outpatients, respectively, shed HRV. About three-fourths of the rotaviruses were type 2, which was prevalent during five successive epidemic years from January, 1974, through June, 1978. HRV type 1 was detected in the last four successive epidemic years and represented nearly half of the HRV infections observed among gastroenteritis inpatients during the year 1977--1978. Both rotavirus serotypes were detected most often in the month of January, when 71% of 123 gastroenteritis inpatients and 62% of 34 gastroenteritis outpatients shed one of these viruses. Uncultivatable adenoviruses were detected significantly more frequently in stools from patients with gastroenteritis (3.9%) than from control subjects (0.6%), suggesting that these viruses played a role in acute enteric disease. The frequency of detection of 27 nm particles was not significantly different in gastroenteritis and control patients. Numerically, HRV infection was detected most often in gastroenteritis inpatients who were 10 through 12 months of age. The group of gastroenteritis inpatients with the highest percentage of HRV infection was 13 through 15 months of age. The excess of type 2 HRV infection relative to type 1 infection was especially large in those aged 7 through 24 months. Lower socioeconomic status or greater crowding appeared to be associated with the occurrence of rotavirus infection earlier in life and earlier in the epidemic year.
Serum samples from children and adults from several countries were tested by radioimmunoassay for antibody to the Norwalk virus. Antibody was commonly found in adults from all the countries tested. Antibody appears to be acquired more rapidly in children from underdeveloped countries than in children from the United States.
Infection of cell cultures with human rotavirus preparations was attempted and the effects of trypsin and low-speed centrifugation on antigen incorporation, as demonstrated by immunofluorescence and radioimmunoassay, were determined. In addition, the effect of viral aggregation on antigen incorporation was investigated by filtering viral preparations. Four strains of human rotavirus were employed, and the results were compared to those obtained with two tissue culture-adapted animal rotaviruses. Centrifugation and trypsin appeared to have little or no effect on infectivity of the tissue culture-adapted (simian rotavirus) or -adaptable (Nebraska calf diarrhea virus) strains, whereas centrifugation and viral aggregation appeared to be essential for the human viruses. In addition, trypsin enhanced antigen incorporation of the human strains to some extent. Infectivity for cell cultures and in vitro human rotavirus protein formation was demonstrated by [35S]methionine incorporation, and the specificity of this human viral protein was established by radio-immunoprecipitation.
A population of 24 infants and young children followed prospectively during the first 3 years of life was studied for the occurrence of rotavirus infection by using enzyme-linked immunosorbent assay to detect virus in stools. Infection with rotavirus was associated with 26 (14.2%) of 183 selected diarrheal episodes. Twenty of the 24 infants and young children had diarrhea associated with rotavirus on at least one occasion and six had two such episodes. Rotavirus infection was documented in over 50% of the dehydrating episodes studied, thus further indicating the importance of rotavirus in this population.
To determine the relative importance of two known serotypes of human rotavirus, we developed an enzyme-linked immunosorbent assay to differentiate serotype-specific rotavirus antigen and antibody. Using this technic, we studied the epidemiology of the two serotypes in acute gastroenteritis. Seventy-seven per cent of 414 rotavirus isolates were Type 2, and the remainder were Type 1. The serotype distribution was similar in specimens from children in Washington, D.C., and other parts of the world. Sero-epidemiologic studies revealed that most children living in the Washington, D.C., area acquired antibody to both types by the age of two years. An analysis of children who were reinfected indicated that sequential infections usually involved different serotypes and that illness caused by one serotype did not provide resistance to illness caused by the other serotype. These results suggest that, to be completely effective, a vaccine must provide resistance to both serotypes.
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Rotaviruses cause gastroenteritis in man and a wide variety of animal species. They cross-react in many immunologic tests and have a similar appearance by electron microscopy, making differentiation among them difficult. Rotaviruses derived from different host species were distinguished by postinfection serum blocking virus activity in an enzyme-linked immunosorbent assay (ELISA). Thirty-three rotavirus isolates from children living in three different parts of the world could not be differentiated by this technique, but they were distinct from four strains recovered from calves, and a series of strains isolated from piglets, foals, monkeys, and infant mice. The four bovine strains were similar, but they could be differentiated from the other animal strains, each of which exhibited a distinct pattern when tested by the ELISA blocking technique.
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A fecal filtrate of human origin containing the Norwalk agent of epidemic viral gastroenteritis was administered by stomach tube to chimpanzees in an attempt to induce diarrheal disease. Significant postchallenge serum antibody rises against Norwalk viral antigens were demonstrated in all animals using the techniques of immune electron microscopy and radioimmunoassay. In addition, viral antigens were detected in feces from five of nine animals using radioimmunoassay. Clinical illness characterized by diarrhea and/or vomiting did not occur. Infection was transmitted subsequently by feeding four additional chimpanzees a fecal filtrate prepared from one of the previously infected animals. Development of an antibody response in four animals and detection of viral antigen in two animals that received this passage filtrate indicated that viral replication had occurred in the absence of clinical illness. The availability of the chimpanzee as an experimental animal host susceptible to infection with the Norwalk agent should facilitate the study of epidemic viral gastroenteritis.
The development of microtiter solid-phase radioimmunoassays for the detection of Norwalk antigen and its antibody is described. The tests are simple to perform and are sensitive and specific. The test for antigen can be used on crude stool filtrates and suspensions. Both tests are at least as sensitive as immune electron microscopy and more sensitive than immune adherence assay.
An immune adherence hemagglutination assay (IAHA) for the detection of antibody to the Norwalk agent of acute epidemic nonbacterial gastroenteritis was developed using as antigen virus purified from stool from an experimentally infected volunteer. The assay was sensitive and specific and was efficient for detecting Norwalk antibody seroresponses. The prevalence of Norwalk antibody in various groups in the United States was studied. Antibody to the Norwalk agent was acquired gradually, beginning slowly in childhood and accelerating in the adult period so that by the fifth decade 50% possessed antibody. This pattern of antibody acquisition contrasted sharply with that for the human rotavirus of infantile gastroenteritis. Rotavirus antibody was acquired during early childhood by almost all individuals in the pediatric groups studied. Antibody to the Norwalk agent was also found in rural Bangladesh; in a small prevalence survey of 39 children and adults 21% possessed Norwalk IAHA antibody, whereas 95% possessed antibody to the human rotavirus.
Human milk contains antibodies to a variety of enteropathic agents. We utilized the method of enzyme-linked immunosorbent assay to investigate anti-rotavirus secretory IgA in 113 human milk and colostral specimens from a rural area in Guatemala, 32 colostral specimens from an urban area of Costa Rica, and 12 from an urban area of the United States. Anti-rotavirus SCIgA was found in all colostral samples and in 94% of the milk specimans. Both the absolute concentration of anti-rotavirus SCIgA and concentration relative to total SCIgA were highest in colostrum, falling to lower but detectable levels from one week to two years after birth. No significant differences were noted in the results from the specimens from the three different geographic areas. The possible role of this antibody in immunity to rotavirus infections is discussed.