We investigated genetic and serological characteristics of a human rotavirus isolate from Indonesia which had a "super short" RNA electrophoretic pattern (A.
SUMMARYPolyacrylamide gel electrophoresis of bovine rotavirus or neonatal calf diarrhoea virus (NCDV) grown in cell culture resolved eight species of polypeptide. The inner shell particles contained five polypeptides and the outer shell three potypeptides. A major polypeptide of the outer shell was glycosylated. The infectivity of NCDV was enhanced by treatment with trypsin in vitro. All eight polypeptides were affected by trypsin treatment as judged by diminished intensity of polypeptide bands by radiography and several new bands appeared. The intracellular synthesis of NCDV polypeptides was studied by pulse and pulsechase experiments. Infected cells contained all eight virus capsid proteins and, in addition, three presumably virus-specific polypeptides which were non-capsid polypeptides (NCVP). There was no evidence that any of these polypeptides was processed after synthesis. It is suggested, therefore, that all these polypeptides are primary gene products.
Direct rotavirus serotyping (VP7, G type) in stool specimens was carried out by reverse transcription and polymerase chain reaction amplification (RT-PCR) and compared to serotyping by enzyme immunoassay with monoclonal antibodies (EIA-MAb). The methods used for double-stranded (ds) RNA extraction, RT-PCR amplification, and the primers used were modified from previous reports [Gouvea et al.: Journal of Clinical Microbiology 29:519-523, 1990; Gentsch et al.: Journal of Clinical Microbiology, 1992]. For samples that were positive by both methods, the serotypes obtained were identical, however RT-PCR typing was found to be considerably more sensitive (70.4% samples serotyped) than EIA-MAb (35.6% of samples serotyped). The overall sensitivities for detection of rotavirus in stool samples by latex agglutination, enzyme immunoassay, electron microscopy, polyacrylamide gel electrophoresis, and RT-PCR were essentially the same. The results confirm that RT-PCR typing (genotyping) is extremely valuable for G typing of samples which cannot be typed by EIA-MAb. We also developed a PCR confirmation technique for serotypes 1, 2, and 4.
We succeeded in isolating human rotaviruses from the feces of gastroenteritis patients by using roller cultures of primary cynomolgus monkey kidney cells with trypsin in the maintenance medium but without concentration and trypsin treatment of the inocula at each passage level. These cells were found to be more sensitive than MA-104 cells (derived from fetal rhesus monkey kidney) for the propagation of human rotaviruses. Polyacrylamide gel electrophoresis of the genome RNA revealed that there were small differences in the migration pattern of the segments among all the strains isolated from 1976 to 1981. The cultivation of human rotaviruses in primary cell cultures might aid in developing a liver rotavirus vaccine.
Four human group C rotaviruses were detected in Tokyo in 1987 and 1988 during a survey over 7 years. Among the four rotaviruses, two electrophoretic patterns were indicated by polyacrylamide gel electrophoretic (PAGE) analyses. Clinical symptoms, signs, family history, and patients' ages varied. Group C rotaviruses were found also in other parts of Japan in 1988. It was suspected that group C rotaviruses would continue to spread throughout Japan within the near future.
Serotyping of human rotavirus in the Tokyo area was conducted from 1990 to 1993 by enzyme immunoassay with monoclonal antibodies (EIA-MAbs) against VP7 and by reverse transcription and polymerase chain reaction (RT-PCR) amplification of the VP4 and VP7 genes. The results by EIA-MAbs were very similar to those obtained by RT-PCR. Evidence of intraserotypic variations was suggested because strains of undetermined serotypes were detected by either EIA-MAb or RT-PCR. This kind of study is required for vaccine development.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.