Variant Of Echovirus Research Articles

A single amino acid substitution in viral VP1 protein alters the lytic potential of clone‐derived variants of echovirus 9 DM strain in human pancreatic islets

In vitro studies with primary human pancreatic islets suggest that several enterovirus serotypes are able to infect and replicate in beta cells. Some enterovirus strains are highly cytolytic in vitro whereas others show virus replication with no apparent islet destruction. The capability to induce islet destruction is determined only partially by the virus serotype, since strain specific differences have been detected within some serotypes including echovirus 9 (E-9). In this study, the viral genetic factors determining the outcome of islet infection (i.e., destructive vs. benign) were investigated by constructing parallel infectious clones of lytic E-9-DM strain that was isolated from a small child at the clinical onset of type 1 diabetes. The capabilities of these clone-derived viruses to induce islet destruction were monitored and the lytic potential of clones was modified by site-directed mutagenesis. The lytic capabilities of these clone-derived viruses in human pancreatic islets were modified by a single amino acid substitution (T81A) in the capsid protein VP1. The data presented outline the importance of amino acid point mutations in the pathogenetic process leading to islet necrosis. However, although the amino acid substitution (T81A) modifies the lytic capabilities of E-9-DM strain-derived microvariant strains, it is likely that additional viral genetic determinants of pancreatic islet pathogenicity exist in other E-9 strains.

A new variant of echovirus 4 associated with a large outbreak of aseptic meningitis.

A large outbreak of aseptic meningitis which began in April 1997 involved hundreds of cases in all geographical regions of Israel and the Palestinian Authority, peaked between June and September, and lasted until December. We have investigated the virus associated with the outbreak to determine its serotype and molecular type and to establish epidemiological links. Virus strains isolated from 210 clinical samples were serotyped by neutralization using LBM and WHO antiserum pools and two echovirus 4 (EV4)-specific antisera, and by immunofluorescence using a monoclonal antibody. RNA was extracted and a 435 base long fragment derived from the 5'UTR of the genome was amplified by RT-PCR using common primers, and sequenced. Sequences were compared to echoviruses 4, 6 and 7 prototypes from ATCC, and to other echoviruses sequences from the EMBL/Genbank data base. The outbreak isolates were identified by the EV4 type-specific antisera and the monoclonal antibody but not with the WHO pools. Very few isolates could be typed by the LBM pools. The EV4 isolates accounted for 68% of all enterovirus isolates in our laboratory in 1997. The age distribution of the patients was: 0-11 month, 11.2%; 1-4 years, 16.1%; 5-9 years, 31.8%; 10-14 years, 9.9%; 15-20 years, 9.5%; 21-44 years, 21.5%; and > 45 years, 0%. Males between 1 and 14 years of age were affected more frequently than females of the same age. The sequences of 25 of 28 EV4 isolates analyzed were closely related to each other (> 95% homology) and the remaining three isolates had < 95% homology to the others and to each other. Interestingly, the outbreak strains were less closely related to the EV4 prototype, than to several other echoviruses. Three closely related subgroups were identified which correlated with geographical distribution but the temporal distribution did not reveal links leading to the source of the outbreak. The outbreak was caused by a variant of EV4 which apparently did not circulate in the area before and thus was capable of causing a widespread infection.

Echovirus 11-Epidemie bei frühgeborenen auf einer neonatal-intensiv-station

During a period of 3 weeks, 17 premature neonates 4 of them weighing less than 1000 g at birth became infected with echovirus 11 in a neonatal intensive-care unit. Beside 3 inapparent infections, severe 'septicaemic' illness was observed among 5 premature neonates combined with meningitis and apnoeic attacks. 4 neonates presented meningitis as the predominant clinical feature and 3 showed gastrointestinal symptoms without neurological involvement. 2 children experienced a febrile illness with apnoea and bradycardia. All babies recovered. Echovirus 11 could be isolated in 51 of 74 (68.9%) specimens, mainly from stool samples. The source of this epidemic outbreak has remained unclear. Virus isolations and serological examinations of antibodies of the IgM class showed that one mother and her eldest child and 3 staff members of the unit had been infected during this epidemic outbreak. The agent was identified as an antigenic variant of echovirus 11 resembling other echoviruses of this type which were isolated in other children hospitals in Bremen during this outbreak.

Isolation and characterization of an echovirus, intratypic variant of echovirus 33.

An echovirus, isolated from children, residents of Cairo, Egypt, U.A.R., was characterized and was shown to be an intratypic variant strain of echovirus type 33.

Isolation of a specific enhancer for echovirus 6 from uninfected permissive host cells.

A specific enhancer for the m+ variant of echovirus 6 was isolated from uninfected, permissive host cells. The enhancer transferred the susceptibility to virus infection from permissive cells to less susceptible cells. Enhancer activity in cultured, human cells (WISH) was released by these cells into extracellular fluids at a linear rate. Maximal enhancer activity was recovered from cell monolayers that were extracted with buffered salt solutions (pH 6.6) for 5 h at 37 C. Crude enhancer preparations contained substances that reduced virus titers in permissive host cells and suppressed some of the enhancer activity. Viral inhibitory activity was removed from cellular extracts by either differential centrifugation or ion exchange chromatography. The enhancer, in contrast to inhibitory substances, remained in supernatant fractions after centrifugation at 110,000 x g for 4 h and adsorbed to diethylaminoethyl-Sephadex columns. Enhancer preparations that were devoid of inhibitory activity increased the virus titers in less susceptible simian cell cultures (approximately 100-fold) to the titers attained by fully permissive human cell cultures. A direct relationship between protein concentration and enhancer activity in cellular extracts was demonstrated. Partial purification of the enhancer by differential centrifugation and ammonium sulfate precipitation increased the specific activity of enhancer (units/mg of protein) by 13-fold.

In vitro interactions between virus and enhancer.

Cell-free supernatant fluids of permissive host cells contained an enhancer for the m+ variant of echovirus 6. The enhancer activity remained in supernatant fractions after centrifugation at 110,000 x g for 4 hr. Under in vitro conditions, a complex between enhancer and virus formed rapidly. The complex was not dissociated by dilution and was sedimented by centrifugation. The effect of enhancer on the distribution of virus in CsCl density gradients was determined. In the absence of enhancer, virus was recovered in one peak at 1.33 g/ml. Incubation of enhancer with virus before centrifugation resulted in the formation of two peaks of infectivity at 1.33 g/ml and 1.24 g/ml. The heavier virus peak represented 43% of the recovered infectivity and could be enhanced. The light virus peak contained 26% of the infectivity and could not be enhanced. The density of enhancer was 1.20 g/ml and approximated the density of the light virus population.