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DNA of human cytomegalovirus: size heterogeneity and defectiveness resulting from serial undiluted passage
Abstract:
The majority of DNA molecules associated with plaque-purified, low-multiplicity-passaged human cytomegalovirus (Towne strain) had a molecular weight of approximately 150 x 10(6) and a molecular complexity of approximately 140 x 10(6). Serial high-multiplicity passage resulted in the production of defective cytomegalovirions. An accumulation of smaller DNA molecules packaged into virions was directly correlated with a decrease in infectivity and an increase in the particle-to-PFU ratio. The majority of defective DNA molecules had a molecular weight of approximately 100 x 10(6). In addition, there were some viral DNA molecules of approximately 60 x 10(6). Restriction enzyme analysis of defective cytomegalovirus DNA detected unique DNA fragments, suggesting a specific alteration in the nucleotide sequence. Some reasons for the generation of defective cytomegalovirus DNA are discussed.
Insights
Human cytomegalovirus (Towne strain) passage generates defective virions with smaller DNA molecules. This DNA alteration correlates with reduced infectivity and increased particle-to-PFU ratios.
Area of Science:
- Virology
- Molecular Biology
Background:
- Human cytomegalovirus (HCMV) is a significant human pathogen.
- Understanding HCMV genome replication and virion assembly is crucial for antiviral strategies.
Purpose of the Study:
- To investigate the molecular characteristics of DNA within HCMV virions after serial high-multiplicity passage.
- To identify alterations in viral DNA associated with defective virion production and loss of infectivity.
Main Methods:
- Plaque purification and low-multiplicity passage of HCMV (Towne strain).
- Serial high-multiplicity passage to induce defective virion formation.
- Molecular weight and complexity analysis of viral DNA.
- Restriction enzyme analysis of viral DNA fragments.
Main Results:
- Low-multiplicity passage yielded HCMV DNA with ~150x10^6 Da molecular weight and ~140x10^6 Da complexity.
- High-multiplicity passage produced defective HCMV virions containing smaller DNA molecules (~100x10^6 Da and ~60x10^6 Da).
- Defective DNA accumulation correlated with decreased infectivity and increased particle-to-PFU ratios.
- Restriction enzyme analysis revealed unique DNA fragments in defective HCMV DNA, indicating sequence alterations.
Conclusions:
- Serial high-multiplicity passage of HCMV leads to the generation of defective virions.
- Alterations in viral DNA, including fragmentation and sequence changes, are associated with defective HCMV production.
- These findings provide insights into the mechanisms of defective viral genome generation in HCMV.