Nucleotide sequences and mutations of the 5'-nontranslated region (5'NTR) of natural isolates of an epidemic echovirus 11' (prime).

An echovirus 11' (prime) virus caused an epidemic in Hungary in 1989. The leading clinical form of the diseases was myocarditis. Hemorrhagic hepatitis syndroms were also caused, however, with lethal outcome in 13 newborn babies. Altogether 386 children suffered from registered clinical disease. No accumulation of serous meningitis cases and intrauterine death were observed during the epidemic, and the monovalent oral poliovirus vaccination campaign has prevented the further circulation of the virus. The 5'-nontranslated region (5'-NTR) of 12 natural isolates were sequenced (nucleotides: 260-577). The 5'-NTR was found to be different from that of the prototype Gregory strain (X80059) of EV11 (less than 90% identity), but related to the swine vesicular disease virus (D16364) SVDV and EV9 (X92886) as indicated by the best fitting dendogram. The examination of the variable nucleotides in the internal ribosomal entry site (IRES) revealed, that the nucleotide sequence of a region of the epidemic 5'-NTR was identical to that of coxsackievirus B2. Five of the epidemic isolates were found to carry mutations. Seven EV11' IRES elements possessed identical sequences indicating, that the virus has evolved before its arrival to Hungary. The comparative examination of the suboptimal secondary structures revealed, that no one of the mutations affected the secondary structure of stem-loop structures IV and V in the IRES elements. Although it has been shown previously, that the echovirus group is genetically coherent and related to coxsackie B viruses the sequence differences in the epidemic isolates resulted in profound modification of the central stem (residues 477-529) of stem-loop structure No.V known to be affecting neurovirulence of polioviruses. Two alternate cloverleaf (stem-loop) structures were also recognised (nucleotides 376 to 460 and 540 to 565) which seem to mask both regions of the IRES element complementary to the 3'-end of the 18 S rRNA (460 to 466 and 561 to 570), thus probably diminishing initiation of translation. The possible biological importance of the alternative cloverleaf structures is supported by the fact that neither the 17 variable nucleotides nor the two mutations of epidemic isolates within the regions seem to modify the predicted alternative secondary structures in EV11, SVDV and CBV1-4.

Characterisation of an echovirus type 11' (prime) epidemic strain causing haemorrhagic syndrome in newborn babies in Hungary.

Echovirus 11' (prime) isolates from an epidemic of haemorrhagic syndrome in departments of obstetrics in Hungary have been characterised. The leading component of the clinical disease was carditis and its lethal outcome occurred in 13 newborn babies. Maternal immunity was found to be absent even in women of 41 years of age. The application of monovalent oral poliovirus type 1 vaccine prevented the progress of the epidemic within two weeks. Nevertheless, a serological survey among primovacinees of 3-15 months of age revealed that 20% of the babies seroconverted without clinical symptoms during the epidemic. Serological evidence showed that the echovirus 11' infection was unable to interfere with the efficacy of oral poliovirus vaccine (OPV), since seroconversion rates of primovaccinees did not differ significantly from those in the group seroconverted also to echovirus 11' during the vaccination campaign. A 440 nucleotide (nt) fragment of the 5'-non-translated region of 12 epidemic echovirus 11' isolates and 26 echovirus prototype strains was amplified by a nested reverse transcription-polymerase chain reaction (RT-PCR) and analysed using three different restriction endonucleases. The 5'-regions of the echovirus 11' isolates were found to be identical to each other but different from that of the prototype echovirus 11 (Gregory) strain. The results indicate that echovirus 11' isolates underwent genetic changes in the 5'-end and P1 region of the genome before the onset of the epidemic.

Detection of transfusion-associated hepatitis caused by non-A, non-B, non-C flavivirus.

Sera of patients suffering from acute hepatitis, and different forms of chronic hepatitis were found to be reactive to reagents prepared from the yellow fever virus (YF) vaccine strain. Serum samples of 1974 patients were tested, and 133 of them were positive. Hepatitis C virus specific antibodies were absent from the majority of them. The frequency of antibodies to other flaviviruses (tick-borne encephalitis, West Nile) and hepatitis B virus markers was similar to that measured among the population in Hungary positive for any of the surrogate markers of hepatitis infections. Results of both immunofluorescence tests, and Western blots suggest that there is a non-A, non-B, non-C hepatitis virus circulating among the Hungarian population, which possesses antigenic cross-reactivity with the yellow fever virus, but the identity to any of the known flaviviruses could not be verified yet. No history of yellow fever vaccination could be revealed in any of the patients included into this study. The anamnestic data on previous transfusions or surgical operations can be verified only in the case of the half of YFV-positive patients, nevertheless, the sexual transmission seems to be very infrequent. Attempts are continued in order to detect the viral RNA using polymerase chain reaction, and clone cDNA sequences for sequence analysis.

[Transfusion-associated non-A, non-B, non-C hepatitis caused by flaviviruses]. / A non-A, non-B, non-C flavivírus által okozott inokulációs hepatitis.

Hepatitis C virus was shown to be a member of the flavivirus family. Tick-borne encephalitis virus and West Nile virus, members of the same family occur in Hungary, too. Serum samples from patients suffering from transfusion associated hepatitis were tested with yellow fever virus antigens for specific IgG, and IgM using immunofluorescence test. Eight hundred serum samples were tested. Yellow fever virus related IgG antibodies were found in 232 sera. In the case of 72 patients specific IgM antibodies could also be detected. The majority of the IgM positive patients underwent surgical operation and/or blood transfusion 1 to 2 months before the onset of the disease. Fifty-four sera positive for yellow fever virus-related antibodies were tested with HCV reagents, but only 13 were found to be positive, or cross-reacting. The 20 patients with yellow fever related antibodies were controlled with tick-borne encephalitis antigens, too. Nevertheless, no measurable cross-reaction could be detected. No measurable cross-reaction could be detected with the West Nile virus. The hepatitis B markers also were tested in 44 sera positive for yellow fever antibodies. There was only one, which contained HBsAg, and 10 of them proved to be positive for anti-HBcAg. The results indicate, that a non-A, non-B, non-C flavivirus is also present in the Hungarian population, which can be detected on the basis of the antigenic cross-reactivity with the attenuated yellow fever virus. This virus seems to be responsible for every 11th transfusion associated hepatitis examined.(ABSTRACT TRUNCATED AT 250 WORDS)

Prevalence of antibody to hepatitis C virus in blood donors, high-risk groups and patients with liver diseases in Hungary. A multicentre study using ABBOTT EIA test and a comparison with an ORTHO ELISA test system.

Serum samples from 1185 individuals (blood donors, health-care workers, patients on haemodialysis, those from other high-risk groups and those suffering from non-A, non-B hepatitis or other liver diseases) were examined for antibody to a recombinant HCV antigen. An ABBOTT HCV EIA system was used throughout and in addition a parallel study with ORTHO HCV ELISA was done in 380 of the samples to compare the two anti-HCV tests. A confirmatory neutralizing ABBOTT ELISA probe was also performed in 45 cases. The anti-HCV test was positive in 1.60% of the healthy blood donors and in 9% of subjects excluded from donation for elevated aminotransferase. In patients on haemodialysis 47%, in other high-risk-group subjects 33% anti-HCV prevalence was found. Patients with acute and chronic post-transfusion NANB hepatitis showed 40% and 70% prevalence, respectively. The two ELISA tests revealed 95% agreement in the parallel determinations. Serial end-point-dilution studies of anti-HCV-positive sera suggest that the ABBOTT test was of superior sensitivity. The results of the confirmatory test suggest that reactive (positive) samples of low optical density near to the cut-off value, required a confirmation with the naturalization test. HCV infection seems to be a common aetiological factor in PT-NANB hepatitis in Hungary, therefore, screening of blood donors for anti-HCV may be justified.

The use of ion exchange chromatography for demonstration of rubella-specific IgM antibodies.

IgM and IgG immunoglobulins of human sera were separated by stepwise column chromatography in QAE-Sephadex A-25 ion exchanger gel bed. The procedure resulted within 30 min in a fraction suitable for direct titration of rubella-specific IgM antibodies by haemagglutination inhibition test. The method proved to be a useful diagnostic tool for primary rubella. Serum samples of 13 individuals with previously acquired immunity, 152 patients with a recent rubella-like illness, and 194 pregnant women exposed to rubella infection were tested for the presence of rubella-specific IgM antibodies. Sera of individuals with previous immunity proved to be negative for specific IgM antibodies. Specific IgM titre was demonstrated in the blood of all the 25 patients with significant titre-rise tested because of rubella-like illness, and also in the sera of additional 8 patients whose serum samples were taken too late for demonstration of a rise in titre. Significant titre-rises were found in 5 women exposed to rubella infection, but only two of them exhibited rubella-specific IgM antibodies. The absence of specific IgM antibodies refers presumably to subclinical reinfection in the other three cases.