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Establishment of cell lines persistently infected with foot-and-mouth disease virus
Abstract:
Cell lines persistently infected with foot-and-mouth disease virus (FMDV) have been established by growth of BHK-21 (c-13) or IBRS-2 (c-26) that survived standard cytolytic infections with FMDV. They maintain cytoplasmic FMDV RNA sequences, as shown by dot blot hybridization tests, using cloned FMDV cDNA as probes. Cell line C1-BHK-Rc1 was derived by infection of cloned BHK-21 c1 cells and plaque-purified FMDV C-S8 c1. Indirect immunofluorescence assays indicated the presence of FMDV antigens. It was resistant to superinfection by FMDV C-S8 c1, O-S7, or A5, but not by encephalomyocarditis virus (EMCV), vesicular stomatitis virus (VSV), or Semliki forest virus (SFV). Infectious FMDV was detected in the culture medium only up to cell passage 65. The virus isolated from C1-BHK-Rc1 cells showed decreased plaque size and diminished yield in infections at 42 degrees. Multiple mutations in the intracellular FMDV RNA have been detected by T1 oligonucleotide fingerprinting of genomic RNA segments hybridized to FMDV cDNA fragments. At late cell passages, when no infectious FMDV is detected, cells continue to express viral antigens and FMDV RNAs with deletions of up to 3 kb have been identified by Northern blot analysis. We conclude that persistent infections of cell cultures with FMDV are readily established and that multiple genetic and phenotypic variations occur in the virus during persistence.
Insights
Persistent foot-and-mouth disease virus (FMDV) infections in cell cultures were established, showing viral RNA and antigens. The virus underwent genetic and phenotypic variations during prolonged cell culture, with infectious virus production ceasing over time.
Area of Science:
- Virology
- Cell Biology
- Molecular Biology
Background:
- Foot-and-mouth disease virus (FMDV) causes significant economic losses in livestock.
- Establishing persistent viral infections in cell culture is crucial for studying viral evolution and host-pathogen interactions.
Purpose of the Study:
- To establish and characterize persistent infections of cell lines with FMDV.
- To investigate the genetic and phenotypic changes of FMDV during prolonged cell culture.
Main Methods:
- Establishment of persistent FMDV infection in BHK-21 and IBRS-2 cell lines.
- Dot blot hybridization and indirect immunofluorescence assays to detect FMDV RNA and antigens.
- T1 oligonucleotide fingerprinting and Northern blot analysis to assess viral RNA mutations and deletions.
Main Results:
- Persistent FMDV infection was established in cell lines, maintaining viral RNA and antigens.
- Cells showed resistance to superinfection by homologous FMDV strains but not heterologous viruses.
- Infectious FMDV was intermittently detected, with isolated viruses exhibiting altered plaque morphology and reduced yield.
- Significant mutations and deletions (up to 3 kb) were observed in FMDV RNA during late passages.
Conclusions:
- Persistent FMDV infections are readily established in cell cultures.
- FMDV undergoes substantial genetic and phenotypic variation during persistent infection.
- Late-stage persistent infections are characterized by the presence of defective viral RNAs and antigens without infectious virus production.