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A Restriction Enzyme Based Cloning Method to Assess the In vitro Replication Capacity of HIV-1 Subtype C Gag-MJ4 Chimeric Viruses
Published on: August 31, 2014
Generation of a monkey-tropic human immunodeficiency virus type 1 carrying env from a CCR5-tropic subtype C clinical
Hiroyuki Otsuki1, Mai Yoneda1, Tatsuhiko Igarashi1
1Laboratory of Primate Model, Experimental Research Center for Infectious Diseases, Institute for Virus Research, Kyoto University, 53 Shogoin Kawara-cho, Sakyo-ku, Kyoto 606-8507, Japan.
Abstract:
Several derivatives of human immunodeficiency virus type 1 (HIV-1) that evade macaque restriction factors and establish infection in pig-tailed macaques (PtMs) have been described. These monkey-tropic HIV-1s utilize CXCR4 as a co-receptor that differs from CCR5 used by most currently circulating HIV-1 strains. We generated a new monkey-tropic HIV-1 carrying env from a CCR5-tropic subtype C HIV-1 clinical isolate. Using intracellular homologous recombination, we generated an uncloned chimeric virus consisting of at least seven types of recombination breakpoints in the region between vpr and env. The virus increased its replication capacity while maintaining CCR5 tropism after in vitro passage in PtM primary lymphocytes. PtM infection with the adapted virus exhibited high peak viremia levels in plasma while the virus was undetectable at 12-16 weeks. This virus serves as starting point for generating a pathogenic monkey-tropic HIV-1 with CCR5-tropic subtype C env, perhaps through serial passage in macaques.
Insights
Researchers created a novel CCR5-tropic, monkey-tropic HIV-1. This adapted virus showed high replication in pig-tailed macaques, offering a new model for HIV-1 pathogenesis research.
Area of Science:
- Virology
- Primate Models
Background:
- Human immunodeficiency virus type 1 (HIV-1) strains that infect macaques often use CXCR4, unlike most circulating strains using CCR5.
- Developing macaque-infecting HIV-1 models is crucial for studying viral pathogenesis and vaccine development.
Purpose of the Study:
- To generate a novel monkey-tropic HIV-1 strain that utilizes the CCR5 co-receptor.
- To investigate the adaptation and replication of this CCR5-tropic virus in pig-tailed macaques (PtMs).
Main Methods:
- Construction of a chimeric HIV-1 by incorporating the env gene from a CCR5-tropic subtype C isolate.
- Generation of an uncloned virus with multiple recombination breakpoints using intracellular homologous recombination.
- In vitro passage in PtM primary lymphocytes and subsequent infection of PtMs.
Main Results:
- The chimeric virus adapted and increased replication capacity while retaining CCR5 tropism after in vitro passage.
- Infection of PtMs with the adapted virus resulted in high peak plasma viremia.
- Viral detection decreased to undetectable levels by 12-16 weeks post-infection.
Conclusions:
- The generated CCR5-tropic, monkey-tropic HIV-1 serves as a valuable starting point for creating pathogenic macaque models.
- Further serial passage in macaques may lead to the development of a more virulent, CCR5-tropic subtype C HIV-1 env strain.
