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Journal of Clinical Microbiology, June 2003, p. 2465-2470, Vol. 41, No. 6
0095-1137/03/$08.00+0     DOI: 10.1128/JCM.41.6.2465-2470.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Quantitation of Human Immunodeficiency Virus Type 1 in Breast Milk

Department of Pediatrics,¹ Microbiology, University of Alabama at Birmingham, Birmingham, Alabama,⁵ Gertrude H. Sergievsky Center, College of Physicians and Surgeons, and Department of Epidemiology, Mailman School of Public Health, Columbia University, New York, New York,² Center for International Health, Boston University, Boston, Massachusetts,³ Center for International Health, Boston University, Boston, Massachusetts⁴

Received 19 November 2002/ Returned for modification 31 January 2003/ Accepted 24 March 2003

The distribution and stability of human immunodeficiency virus type 1 (HIV-1) in breast milk (BM) components remain largely unknown. Inhibitory effects, if any, of BM on HIV RNA and DNA PCR amplification are poorly understood. We have addressed these issues by using virus-spiked BM samples from HIV-negative women. BM samples from HIV-negative women were spiked with HIV-1 virions or cells containing a single integrated copy of HIV DNA (8E5/LAV). After incubation under different experimental conditions, viral RNA was detected by the Roche Amplicor UltraSensitive assay in whole-milk, skim milk, and lipid fractions. We found excellent correlation between HIV-1 input copy and recovery in whole milk (r = 0.965, P < 0.0001), skim milk (r = 0.972, P < 0.0001), and the lipid fraction (r = 0.905, P < 0.001). PCR inhibition was observed in less than 10% of the spiked samples. Similar levels of inhibition were noted in BM samples collected from HIV-infected women. HIV proviral DNA was detected in BM samples using real-time PCR (linear correlation between the threshold cycle versus log DNA copy number, >0.982). The effects of incubation duration and temperature and repeated freeze-thaw cycles on HIV RNA recovery were analyzed. HIV RNA levels were remarkably stable in whole milk after three freeze-thaw cycles and for up to 30 h at room temperature. Our findings improve the understanding of the dynamics of HIV detection in BM and the conditions for BM sample collection, storage, and processing.

Present address: University of Nebraska, Lincoln, Nebraska.

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