Non-polio enteroviruses (NPEV) cause significant disease worldwide. Population-based sero-surveillance, by measuring antibodies against specific NPEV types, provides additional information on past circulation and the prediction for future upsurges. Virus neutralisation assays (VNA), the current method of choice for measuring NPEV type specific antibodies, are not entirely standardised. Via the European Non-Polio Enterovirus Network, we organised a VNA quality assessment in which twelve laboratories participated. We provided five echovirus (E) types (E1, E18, E30 G2, E30 G6 and E6) and intravenous immunoglobulins (IVIG) as a sample for the NPEV VNA quality assessment. Differences in VNA protocols and neutralising Ab (nAb) titres were found between the participating laboratories with geometric coefficients of variation ranging from 10.3-62.9 %. Mixed-effects regression analysis indicated a small but significant effect of type of cell line used. Harmonisation of cell line passage number, however, did not improve variation between laboratories. Calibration by making use of a reference sample, reduced variation between laboratories but differences in nAb titres remained higher than two log2 dilution steps. In conclusion, sero-surveillance data from different laboratories should be compared with caution and standardised protocols are needed.
- MeSH
- ECHO virové infekce virologie epidemiologie imunologie MeSH
- enterovirové infekce virologie imunologie MeSH
- enterovirus B lidský * imunologie MeSH
- lidé MeSH
- neutralizační testy * metody normy MeSH
- neutralizující protilátky * krev imunologie MeSH
- protilátky virové * krev imunologie MeSH
- séroepidemiologické studie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Evropa MeSH
Tick-borne encephalitis (TBE) is one of the leading and most dangerous human viral neuroinfections in Europe and north-eastern Asia. The clinical manifestations include asymptomatic infections, fevers and debilitating encephalitis that might progress into chronic disease or fatal infection. To understand TBE pathology further in host nervous systems, three human neural cell lines, neuroblastoma, medulloblastoma and glioblastoma, were infected with TBE virus (TBEV). The susceptibility and virus-mediated cytopathic effect, including ultrastructural and apoptotic changes of the cells, were examined. All the neural cell lines tested were susceptible to TBEV infection. Interestingly, the neural cells produced about 100- to 10,000-fold higher virus titres than the conventional cell lines of extraneural origin, indicating the highly susceptible nature of neural cells to TBEV infection. The infection of medulloblastoma and glioblastoma cells was associated with a number of major morphological changes, including proliferation of membranes of the rough endoplasmic reticulum and extensive rearrangement of cytoskeletal structures. The TBEV-infected cells exhibited either necrotic or apoptotic morphological features. We observed ultrastructural apoptotic signs (condensation, margination and fragmentation of chromatin) and other alterations, such as vacuolation of the cytoplasm, dilatation of the endoplasmic reticulum cisternae and shrinkage of cells, accompanied by a high density of the cytoplasm. On the other hand, infected neuroblastoma cells did not exhibit proliferation of membranous structures. The virions were present in both the endoplasmic reticulum and the cytoplasm. Cells were dying preferentially by necrotic mechanisms rather than apoptosis. The neuropathological significance of these observations is discussed.
- MeSH
- apoptóza MeSH
- buněčná membrána ultrastruktura MeSH
- buněčná smrt MeSH
- buněčné linie MeSH
- cytoplazma virologie MeSH
- cytoskelet metabolismus ultrastruktura MeSH
- endoplazmatické retikulum virologie MeSH
- lidé MeSH
- neuroglie ultrastruktura virologie MeSH
- neurony ultrastruktura virologie MeSH
- virion ultrastruktura MeSH
- viry klíšťové encefalitidy fyziologie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- práce podpořená grantem MeSH
Hepatitis C virus (HCV) is an important pathogen causing both acute and chronic infections in humans. The HCV polyprotein is synthesized by cap-independent translation initiation after ribosome binding to the highly structured internal ribosome entry site (IRES). The HCV IRES has been shown to have a low requirement for translation initiation factors and the ability to bind directly to the 40S ribosomal subunit. A novel yeast bicistronic reporter system, suitable for sensitive and accurate analysis of IRES activity, has been developed. It employs signal amplification based on the Gal4p transcription factor-mediated activation of a variety of secondary reporter genes. The system has a broad dynamic range and, depending on the nature of the particular secondary reporter, can be used both for precise measurements of IRES activity and for selection and screening for novel IRES variants and IRES trans-acting factors. By using this novel bicistronic system, it was shown that the HCV IRES is functional in yeast cells. Mutational analysis of the IRES loop IV and the adjacent region revealed that, in yeast, as in mammalian cells, translation initiates preferentially at the authentic (342)AUG codon and that disruption of the HCV IRES loop IV abrogates its function, whilst minor positional changes or substitutions of the initiation codon within loop IV are largely tolerated. These findings bring more general insights to translation initiation, but also open the door for utilization of yeast and its sophisticated genetics for searching for new antiviral drugs and HCV IRES trans-acting proteins.
- MeSH
- DNA virů genetika MeSH
- financování organizované MeSH
- genetické vektory MeSH
- geny hub MeSH
- Hepacivirus fyziologie genetika patogenita MeSH
- kodon iniciační MeSH
- lidé MeSH
- plazmidy genetika MeSH
- proteosyntéza MeSH
- rekombinantní proteiny biosyntéza genetika MeSH
- reportérové geny MeSH
- ribozomy metabolismus MeSH
- Saccharomyces cerevisiae genetika metabolismus MeSH
- sekvence nukleotidů MeSH
- techniky in vitro MeSH
- virové geny MeSH
- virové proteiny biosyntéza genetika MeSH
- Check Tag
- lidé MeSH
