HIV-1 drug resistance surveillance using dried whole blood spots.
BACKGROUND
Field-friendly methods for HIV drug resistance (HIVDR) surveillance in resource-limited regions are urgently needed. Despite evidence that dried blood spots (DBS) are suitable for HIV serology, viral load and CD4+ T-cell enumeration, no study has evaluated DBS for HIVDR genotyping. We assessed the feasibility of genotyping HIV-1 from field-collected DBS stored under challenging environmental conditions.


METHODS
We prospectively collected specimens from newly diagnosed, treatment-naive HIV-positive subjects in Mexico. Whole blood was spotted onto filter cards, air dried at ambient temperature and stored with desiccant at 37 degrees C and 85% humidity for 3 months. Genotypes obtained from DBS-extracted nucleic acids using an in-house nested reverse transcription-PCR method were compared to genotypes derived from matched plasma.


RESULTS
Genotypes from 103 phylogenetically matched plasma and DBS were compared. In total, 90.1% of all DBS specimens could be amplified in either the region of HIV protease or the region of reverse transcriptase. Failure to amplify from DBS did not correlate with low plasma viral loads. Between paired specimens, the median nucleotide similarity was 99.95%. In the nine specimens with drug resistance mutations, all differences between pairs were partial discordances. Mutations identified in plasma were found in the majority of replicate DBS amplifications.


CONCLUSIONS
The results suggest that genotypes obtained from DBS are equivalent to those from plasma. DBS are a promising public health tool for HIVDR surveillance of treatment-naive subjects, especially in regions where specimens might be exposed to severe environmental conditions and where logistical difficulties could prevent timely specimen processing. More studies are needed to validate DBS for patient monitoring.
Bertagnolio
Silvia
S
World Health Organization, HIV Department, Geneva, Switzerland. bertagnolios@who.int
Soto-Ramirez
Luis
L
Pilon
Richard
R
Rodriguez
Roberto
R
Viveros
Monica
M
Fuentes
Luis
L
Harrigan
P Richard
PR
Mo
Theresa
T
Sutherland
Don
D
Sandstrom
Paul
P
D000293:Adolescent; D000328:Adult; D000368:Aged; D001800:Blood Specimen Collection / Q000379:methods*; D024882:Drug Resistance, Viral / Q000235:genetics*; D005240:Feasibility Studies; D005260:Female; D005838:Genotype; D015658:HIV Infections / Q000097:blood / Q000188:drug therapy / Q000235:genetics / Q000821:virology*; D016333:HIV Protease / Q000235:genetics; D054303:HIV Reverse Transcriptase / Q000235:genetics; D015497:HIV-1 / Q000201:enzymology / Q000235:genetics*; D006801:Humans; D008297:Male; D008800:Mexico; D008875:Middle Aged; D009154:Mutation; D010802:Phylogeny; D011446:Prospective Studies; D012367:RNA, Viral / Q000097:blood*; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction
D012367:RNA, Viral; D054303:HIV Reverse Transcriptase; D016333:HIV Protease