Field study of dried blood spot specimens for HIV-1 drug resistance genotyping.

Dried blood spots (DBS) are an alternative specimen type for HIV drug resistance genotyping in resource-limited settings. Data relating to the impact of DBS storage and shipment conditions on genotyping efficiency under field conditions are limited. We compared the genotyping efficiencies and resistance profiles of DBS stored and shipped at different temperatures to those of plasma specimens collected in parallel from patients receiving antiretroviral therapy in Uganda. Plasma and four DBS cards from anti-coagulated venous blood and a fifth card from finger-prick blood were prepared from 103 HIV patients with a median viral load (VL) of 57,062 copies/ml (range, 1,081 to 2,964,191). DBS were stored at ambient temperature for 2 or 4 weeks or frozen at -80 °C and shipped from Uganda to the United States at ambient temperature or frozen on dry ice for genotyping using a broadly sensitive in-house method. Plasma (97.1%) and DBS (98.1%) stored and shipped frozen had similar genotyping efficiencies. DBS stored frozen (97.1%) or at ambient temperature for 2 weeks (93.2%) and shipped at ambient temperature also had similar genotyping efficiencies. Genotyping efficiency was reduced for DBS stored at ambient temperature for 4 weeks (89.3%, P = 0.03) or prepared from finger-prick blood and stored at ambient temperature for 2 weeks (77.7%, P < 0.001) compared to DBS prepared from venous blood and handled similarly. Resistance profiles were similar between plasma and DBS specimens. This report delineates the optimal DBS collection, storage, and shipping conditions and opens a new avenue for cost-saving ambient-temperature DBS specimen shipments for HIV drug resistance (HIVDR) surveillances in resource-limited settings.

Evaluation of non-centrifuged dried plasma spots versus centrifuged and non-centrifuged plasma for determination of HIV-1 viral load.

Accurate viral load measurement in plasma specimens is subject to the transport conditions applied since the stability of HIV-1 RNA can be at risk. Also, except during the primary infection, HIV is unlikely to be free in circulation because most patients produce specific antibodies in the weeks following primary infection. This study evaluated non centrifuged dried plasma spots versus centrifuged and non centrifuged plasma in the determination of HIV-1 viral load. A total of 40 patients infected with HIV were bled and three groups of samples were prepared from each patient. The first group was centrifuged at 1500×g for 20min, the second was not centrifuged but left to sediment by gravity for up to 3h, and the third was for dried plasma spots prepared from the same non centrifuged plasma. HIV RNA quantitation in plasma and dried plasma spots was evaluated by the Pearson correlation and a T-test. The three groups yielded average viral loads of 58,249; 83,355 and 116,963 copies/ml for centrifuged, non centrifuged and dried plasma spot samples respectively. The correlation for centrifuged versus non centrifuged was R(2)=0.78, that of centrifuged and dried plasma spots was R(2)=0.72 and finally R(2)=0.81 between non centrifuged and dried plasma spot samples. A significant difference in viral load results of centrifuged and DPS samples prepared from non centrifuged plasma was observed.