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EVALUATION OF DBS HCV RNA QUANTIFICATION AND GENOTYPING IN RESOURCE LIMITED SETTINGS
Jayaseelan Boobalan1, Thongadi R. Dinesha1, Sathasivam Sivamalar1, Selvamurthi Gomathi1, Rajendran Revathi1, Aylur K. Srikrishnan1, Amrose Pradeep1, Shruti H. Mehta2, Sunil S. Solomon2, Pachamuthu Balakrishnan1, Shanmugam Saravanan1
1YR Gaitonde Center for AIDS Research and Education, Chennai, India,2Johns Hopkins University, Baltimore, MD, USA
High burden of HCV infection in people who inject drugs (PWID) in resource limited settings (RLS) like India coupled with low access to HCV services necessitates an urgent need for evaluating less invasive DBS based HCV virological assays for appropriately tailored therapy and successful treatment scale up programs in RLS.
Paired 36 plasma/DBS samples were collected from PWID. Two blood spots were excised into 1.7 mL of Abbott lysis buffer. RNA extracted from plasma and DBS according to the standard 0.5 mL HCV RNA Abbott extraction protocol (Abbott Molecular Inc, IL, USA). In addition, patient with >3 log10 IU/mL HCV RNA were further subjected to in-house one-step core/E1 RT-PCR followed by Sanger's sequencing. All the values were log transformed and analysed on GraphPad Prism 5.0, A p <0.005 was considered significant.
There was a good correlation between standard Abbott plasma and DBS HCV assay (r = 0.97, p<0.001).Median PVL of standard plasma and DBS HCV RNA by Abbott Real-time PCR was 4.45(IQR 2.54-5.43) and 4.18 (IQR 2.24-5.74) log IU respectively. The mean difference between plasma and DBS HCV RNA assays were 0.250 and the upper and lower 95% Limit of Agreement -0.7321 to 1.23374. Out of 36 samples, 21 (58%) [Median = 5.74, (IQR=4.96-5.91)] of them were able to do HCV genotyping and majority of them were genotype 3b (42.8%) followed by 6xa (33%).
This study supports the use of DBS as an alternate to plasma for the reliable quantification of HCV RNA and genotyping. Therefore, the DBS specimens could be effectively used in resource-limited settings for therapeutic and research purposes.