JAC Advance Access originally published online on April 29, 2009
Journal of Antimicrobial Chemotherapy 2009 64(1):33-36; doi:10.1093/jac/dkp150
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Original research |
Rapid decline in the efficiency of HIV drug resistance genotyping from dried blood spots (DBS) and dried plasma spots (DPS) stored at 37°C and high humidity
1 Laboratory Branch, Division of HIV/AIDS Prevention, National Center for HIV/AIDS, Viral Hepatitis, STD and TB Prevention, Centers for Disease Control and Prevention, Atlanta, GA, USA 2 Department of Immunology/Microbiology, Rush Medical College, Chicago, IL, USA
Received 9 January 2009; returned 8 March 2009; revised 11 March 2009; accepted 5 April 2009
* Corresponding author. Laboratory Branch, Division of HIV/AIDS Prevention, Centers for Disease Control and Prevention, 1600 Clifton Rd, Atlanta, GA 30333, USA. Tel: +1-404-639-4987; Fax: +1-404-639-1174; E-mail: GGarcia-Lerma{at}cdc.gov
Objectives: Dried blood spots (DBS) and dried plasma spots (DPS) are considered convenient alternatives to serum and plasma for HIV drug resistance testing in resource-limited settings. We sought to investigate how extreme conditions could affect the short-term ability to amplify and genotype HIV from DBS.
Methods: A panel of six matched DPS/DBS was generated using blood collected from HIV-infected donors. Replicate cards were prepared in 903 filter paper using 50 µL of blood and stored at either –20°C or at 37°C/100% humidity. Nucleic acids were extracted at baseline and after 1, 2, 8 and 16 weeks of storage and were amplified and sequenced using an in-house RT-nested PCR method or the ViroSeq assay.
Results: HIV-1 pol was successfully amplified in all DBS/DPS at baseline and in those stored for up to 16 weeks at –20°C by the in-house assay. In contrast, amplification was rapidly lost during storage at 37°C/100% humidity with only 6/6 and 4/6 DBS specimens amplifiable by the in-house assay at weeks 1 and 2, respectively. Similarly, only two DPS stored at 37°C/100% humidity were amplified by the in-house assay at week 1.
Conclusions: We show that resistance testing from DBS and DPS is severely compromised after 2 and 1 weeks of storage at 37°C/100% humidity with desiccant, respectively. These findings underscore the importance of temperature and humidity for the efficient genotyping of HIV-1 from DBS and DPS, and reiterate the need to rapidly transport specimens from collection sites to locations that have appropriate storage conditions such as –20°C.
Keywords: drug resistance surveillance , 903 filter paper , RNA stability , DBS storage
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