Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UMLS:C0017160 (gastroenteritis)
 11,398 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

During a search for established cell lines to produce large quantities of porcine transmissible gastroenteritis virus (TGEV), we observed bright immunofluorescent staining 6- 12h after infection of pig kidney derived LLC-PK1 line. Infectious virus yield was, however, 2 log10 lower than that from secondary adult pig thyroid (APT/2) cell cultures, although small plaques were visible by three days in cultures maintained under agarose, suggesting limited replication. Attempts to adapt TGEV to the LLC-PK1 cell line by 10 serial 20h passes were unsuccessful. Procedures to purify virions from infected LLC-PK1 cells produced less than 1% of the particles isolated from parallel APT/2 cultures. Examination of intracellular viral RNA in actinomycin-D treated cells revealed similar amounts of genomic RNA and the 4 major subgenomic species in both cell types, suggesting that there was no defect in viral RNA replication. In vitro translation of polyadenylated RNA from infected APT/2 and LLC-PK1 cells, followed by immune precipitation of the products, showed similar profiles of precursors to structural polypeptides, confirming the functional integrity of the viral messengers in the restrictive cell. Comparison of the viral polypeptides synthesised following infection of the two cell types showed that similar species were synthesised in both, corresponding to a group of 28-30,000 mol. wt. envelope glycopolypeptides, a 47,000 mol. wt. nucleoprotein and peplomer glycopolypeptides of about 200,000 mol. wt. The rate of viral polypeptide synthesis in LLC-PK1 cells was reproducibly higher than in APT/2, resulting in the earlier detection of bands and greater incorporation of isotope. Tunicamycin at 1 microgram/ml had a similar effect in both cells, preventing glycosylation of the 26,000 mol. wt. precursor of the envelope glycopolypeptides and synthesis of the 200,000 peplomer glycoprotein. Degradation of the nucleoprotein from 47,000 to 42,000 mol. wt. although detectable in both cells was more marked in the LLC-PK1 cultures. Phosphorylation of these proteins was readily demonstrated in both cells, although phosphorylation of host proteins and, to some extent, viral envelope proteins was considerably greater in the LLC-PK1. The significance of this finding with respect to virus maturation is being investigated.
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PMID:Defective replication of porcine transmissible gastroenteritis virus in a continuous cell line. 633 Nov 29

Human noroviruses (HuNoVs) is the primary non-bacterial pathogen causing acute gastroenteritis worldwide. Molecular approaches have been mainly used for detection of HuNoVs. Aptamer-based assay has been also applied for detection of HuNoVs through affinity binding of viral capsid. In a conventional systematic evolution of ligands by exponential enrichment process, the target protein-bound sequences in the library were recovered by complicated process including affinity chromatography, extraction, membrane-filtration or antibody-conjugated magnetic beads. In this study, a novel approach was applied to select aptamers for HuNoVs. The new approach incorporated an in situ capture assay and next generation sequencing (NSG) for selecting the aptamers. P particles of HuNoV (GII.4) were purified and coated on the module to capture sequences that were specifically bound with the protein. The unbound sequences were easily removed by washing. The sequences with high affinity were amplified just in the wells and selected by repeated in situ selection process. From the total of 30,622,226 tested sequences, two aptamers, APTL-1 and APTL-6, were finally selected to incorporate with in situ capture RT-qPCR assay for detection of HuNoVs from clinical samples. The sensitivity of these two aptamers was compared with porcine gastric mucin (PGM) that contains well-known viral receptors, and the reported aptamer APT-M6-2. Both GI and GII HuNoVs could be detected from 5 clinical samples tested. The selected aptamer APTL-1 was comparable to PGM and slightly superior to the reported APTM6-2 aptamer for detection of HuNoVs from clinical samples. The results demonstrated that this in situ target-capture approach for aptamer selection is practicable.
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PMID:Development and evaluation of a novel in situ target-capture approach for aptamer selection of human noroviruses. 3036 91