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PubMed1996

Serological and genomic characterization of porcine rotaviruses in Thailand: detection of a G10 porcine rotavirus.

A total of 557 fecal specimens collected from piglets with diarrhea in Thailand were examined for rotavirus RNA by polyacrylamide gel electrophoresis. Twenty-three, one, and two samples were positive for group A, group B, and group C rotaviruses, respectively. Two samples exhibited two segments found in picobirnavirus RNA. RNA electropherotyping of 23 group A rotaviruses showed that they were classified into five patterns. By serotyping by enzyme-linked immunosorbent assay and PCR, viruses in 3 and 14 specimens were found to be serotype G3 and serotype G10, respectively. For one specimen, containing a serotype G10 virus (strain P343), virus was isolated in MA-104 cells, and the nucleotide sequences of the VP7 and VP4 genes were determined. Comparative sequence analysis and cross-neutralization tests showed that strain P343 has B223-like G10 and UK-like P7 serotype (or VP4 genotype 5) specificities. Rotaviruses having such antigenic specificities have not been detected in piglets. Thus, the interspecies transmission of rotaviruses between cows and pigs was suggested.

Amino Acid Sequence↗

PubMed1996

Quantification of systemic and local immune responses to individual rotavirus proteins during rotavirus infection in mice.

The purpose of the present study was to develop a quantitative assay that could be used to measure the local and systemic immune responses to specific rotavirus proteins following rotavirus infection of adult mice. To measure these responses, we used an immunocytochemical staining assay of Spodoptera frugiperda (Sf-9) cells which were infected with recombinant baculovirus expressing selected rotavirus proteins. The specificity of the assay was documented by using a series of monoclonal antibodies to individual rotavirus proteins. We observed that the assay had high levels of sensitivity and specificity for a series of VP7- and VP4-specific neutralizing monoclonal antibodies which recognized conformation-dependent epitopes on their target proteins. We also studied immunoglobulin G (IgG) immune responses in serum and IgA immune responses in the stools of mice infected with wild-type murine rotavirus strain EHPw. In both sera and stools, the most immunogenic proteins were VP6 and VP4. VP2 was less immunogenic than VP6 or VP4, and the immune responses to VP7, NSP2, and NSP4 were very low in serum and undetectable in stools.

Animals↗

PubMed2006

Full genomic analysis of human rotavirus strain B4106 and lapine rotavirus strain 30/96 provides evidence for interspecies transmission.

The Belgian rotavirus strain B4106, isolated from a child with gastroenteritis, was previously found to have VP7 (G3), VP4 (P[14]), and NSP4 (A genotype) genes closely related to those of lapine rotaviruses, suggesting a possible lapine origin or natural reassortment of strain B4106. To investigate the origin of this unusual strain, the gene sequences encoding VP1, VP2, VP3, VP6, NSP1, NSP2, NSP3, and NSP5/6 were also determined. To allow comparison to a lapine strain, the 11 double-stranded RNA segments of a European G3P[14] rabbit rotavirus strain 30/96 were also determined. The complete genome similarity between strains B4106 and 30/96 was 93.4% at the nucleotide level and 96.9% at the amino acid level. All 11 genome segments of strain B4106 were closely related to those of lapine rotaviruses and clustered with the lapine strains in phylogenetic analyses. In addition, sequence analyses of the NSP5 gene of strain B4106 revealed that the altered electrophoretic mobility of NSP5, resulting in a super-short pattern, was due to a gene rearrangement (head-to-tail partial duplication, combined with two short insertions and a deletion). Altogether, these findings confirm that a rotavirus strain with an entirely lapine genome complement was able to infect and cause severe disease in a human child.

Amino Acid Sequence↗

PubMed2006

Physicians' knowledge and attitudes about rotavirus gastroenteritis and rotavirus vaccine.

The morbidity and mortality associated with rotavirus gastroenteritis are significant. Despite the past safety concerns with RotaShield, many physicians believed the need for rotavirus vaccination remains. Although most physicians were not aware of specific rotavirus gastroenteritis disease burden statistics, they perceive rotavirus gastroenteritis to pose a serious risk for children for which a vaccine would be beneficial. Proof of safety and efficacy, an ACIP/AAP/AAFP recommendation, and private insurance coverage/reimbursement were critical features that would allow physicians to overcome barriers to the adoption of a new rotavirus vaccine.

Attitude of Health Personnel↗

PubMed2005

Efficient intranasal immunization of newborn mice with recombinant adenovirus expressing rotavirus protein VP4 against oral rotavirus infection.

Efficacy of passive protection of newborn mice against rotavirus infection by the rotavirus VP4 protein expressed by an adenoviral vector in mice was studied. The VP4 gene was inserted into the E1 region of adenoviral vector pJM17. Recombinant adenovirus Ad5N/VP4 was grown in 293 cells. Intramuscular (i.m.), oral or intranasal (i.n.) immunization of newborn mice with Ad5/VP4 resulted in appearance of VP4-specific antibodies. Specific IgG antibodies were detected in the serum and intestine specimens of i.m. vaccinated mice. Oral immunization elicited serum IgG antibodies and intestinal IgG and IgA antibodies. Compared with i.m. and oral applications, i.n. immunization led to higher levels of serum IgG and intestinal IgG and IgA antibodies. Pups were challenged twice with simian rotavirus SA11 strain orally at the days 7 and 8 after birth. Pups born to i.n. immunized dams achieved 100% protection from rotavirus-induced diarrhea after both challenges. The protection of pups born to orally immunized dams was 80%, while only 30% of pups born to i.m. immunized dams were protected after both challenges. I.n. immunization was most efficient in inducing rotavirus VP4-specific serum, intestinal and milk IgG or IgA in mice that protected newborn mice completely.

Adenoviridae↗

PubMed1987

Candidate rotavirus vaccine (rhesus rotavirus strain) in children: an evaluation.

Fifty children, 3 months to 12 years of age, were given an experimental, orally administered, live attenuated rotavirus vaccine. Overall evidence of vaccine effectiveness as judged by vaccine virus shedding or a serologic response was seen in 82% of vaccinees. No clinical illness was seen in the rotavirus vaccinees when compared with 40 concurrently studied control children. No transmission to control children was observed even with close daily contact in a day-care setting. Young infants, generally less than 1 year of age, who had not previously experienced wild-type rotavirus infection shed significantly more vaccine virus. Limitation of virus shedding in those already exposed may be related to a prompt copro-IgA response which was significantly elevated by three days after vaccination. In summary, the development of this rotavirus vaccine, rhesus rotavirus-MMU-18006, is a promising step in the development of immunoprophylaxis against this major enteric pathogen.

Antibodies, Viral↗

PubMed1996

Assay of neutralizing antibodies to animal rotavirus strains and human rotavirus serotype G8 by a modified method in the residents of Pune, India.

In the western literature, four G serotypes (G1-G4) of human rotaviruses have been found to be of a major epidemiological importance. During the analysis of rotavirus serotypes from faecal samples in Pune, over 50% of specimens could not be serotyped with the available monoclonal antibodies against G1-G4 serotypes. The results prompted to look for the prevalence of neutralizing antibodies against serotypes other than the major human serotypes (G1-G4) in adults. Neutralizing antibodies against animal rotavirus serotype, viz. G3, G6 and G10, and human rotavirus serotype G8 were determined in adult sera, by a modified technique, which is ELISA-based and mechanized. The results showed that, of the 68 sera tested at 1:100 dilution, 65 (95.58%) were reactive for G3 (SA-11), 52 (76.47%) for G6 (Bovine Lincoln), 6 (8.82%) for G10 (B223), and 40 (58.82%) for G8 (M69) serotypes. It appears that the prevalence of rotaviruses in India may be quite different from that in the developed countries.

Adult↗

PubMed1994

Induction of rotavirus-specific cytotoxic T lymphocytes by vaccinia virus recombinants expressing individual rotavirus genes.

We determined the capacity of vaccinia virus recombinants expressing individual rotavirus genes to induce virus-specific cytotoxic T lymphocytes (CTLs) in mice. Mice were orally inoculated with vaccinia virus recombinants containing genes which encode rotavirus outer capsid proteins vp4 or vp7, single-shelled virus proteins vp1, vp2, or vp6, or rotavirus nonstructural proteins NS53, NS35, NS28, or NS26/NS12. We found that (i) the greatest frequencies of virus-specific CTLs were induced by vaccinia virus recombinants expressing vp7, (ii) transport of vp7 beyond the endoplasmic reticulum was not necessary for induction of CTLs, (iii) recombinants expressing vp7 induced CTLs which reacted with different rotavirus serotypes, and (iv) CTLs were induced among both intestinal and nonintestinal lymphocytes after oral inoculation. These findings may be relevant to vaccine strategies which utilize vectors expressing individual rotavirus genes.

Administration, Oral↗

PubMed1997

Maternal immunization of pregnant cattle with recombinant VP8* protein of bovine rotavirus elicits neutralizing antibodies to multiple serotypes. Colostral neutralizing antibody by rotavirus VP8*.

Neonatal calf diarrhoea caused by bovine rotavirus is one of the most common diseases in cattle. VP8 portion of the rotavirus VP4 protein is known to contain neutralizing epitopes and hemagglutination activity. We expressed the VP8* portion of bovine rotavirus strain C486 (G6P1 serotype) in E. coli, and examined potential of the recombinant VP8* protein for induction of neutralizing antibody responses in host animals. One hundred twenty pregnant beef cows were selected and immunized eight weeks prior to parturition with the recombinant VP8* protein. Colostral and milk samples were collected 12 hours and 10 days post-calving, respectively, and the virus neutralizing titers elicited by the recombinant subunit antigen were determined by plaque reduction assay. Colostral antibody titres of the vaccinated group were significantly higher than those of the unvaccinated control group, and these titers were equivalent to the titers elicited by a commercial vaccine. While titers of the control group rapidly decreased to 220 after 10 days of calving, neutralizing titers in the milk of the vaccinees remained 510. Rabbit and bovine antibodies induced by the recombinant VP8* protein were also able to neutralize bovine rotavirus P5 serotype (B641) at significant level and P11 serotype (B223) moderately. Our results suggest that the E. coli-produced recombinant VP8* protein can be an useful subunit vaccine candidate to prevent rotavirus infection in new-born calves.

Animals↗

PubMed2000

Characterization of human symptomatic rotavirus isolates MP409 and MP480 having 'long' RNA electropherotype and subgroup I specificity, highly related to the P6[1],G8 type bovine rotavirus A5, from Mysore, India.

In an epidemiological study of symptomatic human rotaviruses in Mysore, India during 1993 and 1994, isolates MP409 and MP480 were isolated from two children suffering from severe, acute dehydrating diarrhea. Both isolates exhibited 'long' RNA pattern and subgroup I specificity suggesting the likelihood of their animal origin. Both isolates did not react with monoclonal antibodies (MAbs) specific for serotypes G1 to G6 as well as G10. To determine the genetic origin of these isolates, complete nucleotide sequences of genes encoding the outer capsid proteins VP4 and VP7, nonstructural proteins NSP1 and NSP3 and viral enterotoxin protein NSP4 from MP409 and partial sequences of genes from MP480 were determined. Comparison of the 5' and 3' terminal sequences of 250 nucleotides revealed complete identity of the gene sequences in both strains suggesting that MP409 and MP480 are two different isolates of a single strain. Comparison of the nucleotide and deduced amino acid sequences of VP4, VP7, NSP1 and NSP3 of MP409 with published sequences of strains belonging to different serotypes revealed that both outer capsid proteins VP4 and VP7 and NSPI are highly related to the respective proteins from the P6[1], G8 type bovine rotavirus A5 isolated from a calf with diarrhoea in Thailand and that the NSP3 is highly homologous to that of bovine rotaviruses. The NSP4 protein showed greatest sequence identity with NSP4s belonging to the KUN genetic group to which NSP4s from human G2 type strains and bovine rotaviruses belong. MP409 and MP480 likely signify interspecies transmission of P6[1], G8 type strains from cattle to humans and represent the first P6[1] type rotaviruses isolated in humans. These and our previous studies on the asymptomatic neonatal strain 1321 are of evolutionary and epidemiological significance in the context of close association of majority of the Indian population with cattle.

Amino Acid Sequence↗

PubMed2004

Rotavirus VLP2/6: a new tool for tracking rotavirus in the marine environment.

The potential of rotavirus 2/6-virus-like-particles (VLP2/6) for use as tracers in the marine environment was investigated. The stability of bovine rotavirus (strain RF) and VLP2/6 in natural seawater at 25 degrees C for six days was studied. ELISA and western blot methods were used to quantify the particles. The rates of decline of rotavirus particles and VLP2/6 were similar (approximately 0.5log(10) per day). Western blot analysis showed that the integrity of capsid proteins VP2 and VP6 was conserved during the incubation time. These results demonstrate that VLP2/6 particles have the same stability in seawater as rotavirus particles. Thus, VLP2/6 can be used as a tracer, which should be of particular value for studying the fate of rotavirus particles in the marine environment.

Animals↗

PubMed1981

Rescue of noncultivatable human rotavirus by gene reassortment during mixed infection with ts mutants of a cultivatable bovine rotavirus.

Fastidious human rotaviruses that did not undergo productive infection in tissue culture were rescued by genetic reassortment during mixed infection with a temperature-sensitive (ts) mutant of a cultivatable bovine rotavirus. In this manner, the genes of the fastidious rotavirus that restricted growth in vitro were replaced by the corresponding genes from a tissue culture-adapted rotavirus. We recovered genetically reassorted viruses that grew to high titer and were neutralized specifically by hyperimmune guinea pig type 1 or type 2 human rotavirus antiserum. Preliminary RNA analysis of these clones disclosed that they were indeed viruses with reassorted genes.

Animals↗

PubMed2006

An outbreak of rotavirus infection among adults in an institution for rehabilitation: long-term residence in a closed community as a risk factor for rotavirus illness.

An outbreak of group A rotavirus infection resulted in gastroenteritis among disabled adults in an isolated rehabilitation institution in Kobe, Japan. Of the 95 residents, 16 were diagnosed with rotavirus illness. The causative agent was a single strain of typical human group A rotavirus belonging to VP7 serotype G2, VP4 genotype P[4], and NSP4 genotype A. Mean duration of stay was significantly longer for residents with rotavirus illness (22.1+/-11.8 years) than for residents without the disease (13.5+/-10.6 years; P=0.01). Age, sex, disability and location of resident rooms displayed no significant relationships with illness. These observations suggest that long-term residence in a closed community, which might be related to absence of immuno-stimulation, represents a risk factor for rotavirus illness.

Adult↗

PubMed1981

Isolation from chickens of a rotavirus lacking the rotavirus group antigen.

A virus, designated 132 virus, was isolated from the faeces of chickens in chick embryo liver cell cultures. The morphology and morphogenesis of 132 virus were indistinguishable from that of rotaviruses. The nucleic acid of 132 virus had the nuclease resistance of double-stranded RNA, and was separated by polyacrylamide gel electrophoresis into 11 segments with mol. wt. ranging from 2.07 x 10(6) to 0.20 x 10(6). SPF chickens were susceptible to oral infection with 132 virus, which replicated in the villous epithelial cells of the small intestine. 132 virus was therefore a rotavirus by morphological, biochemical and biological criteria. However, by immunofluorescence it was not possible to demonstrate an antigenic relationship between 132 virus and known avian and mammalian rotaviruses, indicating that 132 virus does not possess the group antigen shared by all previously characterized rotaviruses. This finding has implications for the diagnosis of rotavirus infections by serological tests.

Animals↗

PubMed1987

Immune response to rotavirus polypeptides after vaccination with heterologous rotavirus vaccines (RIT 4237, RRV-1).

The antibody response to individual rotavirus polypeptides after vaccination with live attenuated heterologous rotavirus vaccines was studied with a radioimmunoprecipitation assay (RIPA). Following vaccination with bovine rotavirus strain RIT 4237 and rhesus monkey rotavirus strain RRV-1, an antibody response was demonstrated mainly against the inner capsid polypeptides VP2 and VP6 and to a lesser extent against VP3/VP4. There was no qualitative difference between the vaccines in the antibody response; both vaccines induced antibodies against the same polypeptides. Quantitatively the antibody response to RRV-1 was somewhat stronger. All prevaccination sera contained by RIPA low levels of rotavirus antibodies. The effect of these antibodies on the 'take' of the vaccines is discussed.

Antibodies, Viral↗

PubMed1999

Impact of rotavirus disease in Spain: an estimate of hospital admissions due to rotavirus.

The epidemiology of rotavirus infection in Spain was investigated using information from existing surveillance systems. Reports from laboratories and data on hospital admissions were used to estimate the number of hospital admissions due to rotavirus infection. Between January 1989 and December 1995 there were 8265 reports of rotavirus identification in the Sistema de Información Microbiológica, a voluntary national laboratory surveillance system. Where age was reported, 89.2% were in children under 5 y old. This represents 21.8% of all pathogens identified in faecal specimens in children of this age group. Between January and December 1994 there were 5639 admissions coded as "infectious intestinal disease" and 8225 as "non-infective gastroenteritis" in children under 5 y of age in the Conjunto Mínimo Básico de Datos, an obligatory hospital admissions surveillance system which covers 83% of public hospitals. Using regression analysis of admission data and laboratory reports it was estimated that rotavirus infection accounted for 3519 (25.3%) of the hospital admissions. We estimate that 4239 admissions attributable to rotavirus occurred in children under 5 y of age in Spain in 1994, a rate of 2.5/1000 population in this age group. These estimates are lower than those published for other countries, where the burden of disease is considered high enough to warrant a national vaccination programme. A cost-effectiveness study with Spanish information is essential before embarking on a national vaccination programme.

Child, Preschool↗

PubMed1982

RNA of rotavirus: comparison of RNAs of human and animal rotaviruses.

Single-stranded RNA (ssRNA) was transcribed in vitro from inner-shell particles of human rotavirus strain Wa (HRV-Wa) and a bovine rotavirus (neonatal calf diarrhea virus [NCDV]) by virion-associated RNA polymerase activity. The ssRNA product consisted of 11 RNA segments which were separated by polyacrylamide gel electrophoresis. In vitro-transcribed 32P-labeled ssRNA was used to study the genetic relatedness between rotaviruses by annealing with genomic double-stranded RNA (dsRNA) of homologous or heterologous rotavirus. All segments of HRV-Wa ssRNA were hybridized with dsRNA of HRV TK80, collected from the feces of a gastroenteritis patient, at the level of 88 to 100% of the homologous reaction. On the other hand, no segments of ssRNA from HRV-Wa hybridized with dsRNA of NCDV or simian rotavirus (simian agent 11). Similarly, ssRNA from NCDV did not hybridize with dsRNA of HRV-Wa, but hybridized with dsRNA of simian agent 11 at the level of 30% of the homologous value.

Nucleic Acid Hybridization↗

PubMed1979

Use of antisera against bovine (NCDV) and simian (SA11) rotaviruses in ELISA to detect different types of human rotavirus.

Two ELISA systems for the detection of human rotaviruses were developed. In the first system antibodies to Nebraska calf diarrhea virus (NCDV) were used for coating the solid matrix and for the preparation of the enzyme conjugate. In the second system antibodies to human rotavirus and antibodies to simian rotavirus (SA11) were used for coating the solid matrix and for the preparation of the enzyme conjugate respectively. The second ELISA system proved to have a broader spectrum for the detection of human rotaviruses. By using the two ELISA systems, the different types of human rotavirus could be distinguished. The ELISA tests developed were 8 to 64 times as sensitive as electron microscopy (EM) and (or) counterimmunoelectrophoresis (CIEP). The antigen detected by ELISA was shown to be different from that detected by the hemagglutination test.

Animals↗