Sensitive and reproducible quantitation of mucosal HIV-1 RNA and DNA viral burden in patients with detectable and undetectable plasma viral HIV-1 RNA using endoscopic biopsies

Mucosal tissue is the main portal of entry for HIV-1 infection and, in macaques, has been demonstrated to be a significant compartment for viral replication and CD4 + T lymphocyte depletion. Quantitating tissue viral burden in addition to plasma viral load provides insights into HIV-1 pathogenesis a...

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Bibliographic Details
Published in:Journal of virological methods 2001-06, Vol.95 (1), p.65-79
Main Authors: Anton, Pete A, Poles, Michael A, Elliott, Julie, Mao, S.H, McGowan, Ian, Lenz, Heinz-Josef, Chen, Irvin S.Y
Format: Article
Language:English
Subjects:
Animal viral diseases
Biological and medical sciences
CD4 antigen
Colonoscopy
DNA, Viral - analysis
Female
Fundamental and applied biological sciences. Psychology
Gastrointestinal
Gastrointestinal associated lymphoid tissue
HIV Infections - pathology
HIV Infections - virology
HIV-1
HIV-1 - genetics
HIV-1 - physiology
Human immunodeficiency virus 1
Human viral diseases
Humans
Infectious diseases
Intestinal Mucosa - pathology
Intestinal Mucosa - virology
Male
Medical sciences
Microbiology
Mucosal
Polymerase Chain Reaction - methods
Quantitative
Reproducibility of Results
RNA, Viral - analysis
Sensitivity and Specificity
Tissue
viral burden
Viral diseases
Viral diseases of the lymphoid tissue and the blood. Aids
Viral Load
Virus Replication
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Summary:Mucosal tissue is the main portal of entry for HIV-1 infection and, in macaques, has been demonstrated to be a significant compartment for viral replication and CD4 + T lymphocyte depletion. Quantitating tissue viral burden in addition to plasma viral load provides insights into HIV-1 pathogenesis and an additional means to gauge antiretroviral response. The aim of this study was to develop reliable, reproducible, and sensitive assays to quantitate tissue viral burden of HIV-1 RNA and DNA using 1–3 endoscopically acquired, rectosigmoid biopsies. Total DNA and RNA were simultaneously extracted following homogenization from the same tissue samples. Quantitative polymerase chain reaction (PCR) assay in the HIV-1 LTR region was used to detect viral DNA and RT-PCR for viral RNA. It was determined that HIV-1 RNA and DNA can be reproducibly quantified from a single rectosigmoid biopsy with minimal intra-assay or intra-patient variability. These results reflect high recovery of extracted nucleic acids with calculated results accurately reflecting in vivo levels. The techniques outlined differ from currently available approaches by incorporating control standards to identify loss or degradation of RNA and DNA from acquisition through the in vitro assay and permit extraction with high yields of RNA and DNA from the same tissue sample.
ISSN:0166-0934
1879-0984
DOI:10.1016/S0166-0934(01)00295-6