Screening of hepatitis C virus in dried plasma spots among people who inject drugs and attending syringe exchange programme in Delhi, India

Hepatitis C Virus (HCV) represents a major health burden worldwide.¹ In India, about 12–13 million HCV carriers have been identified. The prevalence of HCV infection in people who inject drugs (PWID) is more than in the general population. However, the majority of PWID are either unaware of the infection or have not received any treatment.^2,3 Some of the reasons for low testing involve barriers like lack of knowledge, fear of invasive tests, limited access to testing and treatment.⁴ Several strategies have been proposed to overcome these barriers. One of them is the use of dried plasma spots due to its scalability and ease of use. Dried plasma spots offer advantages like ease of collection, transportation, and storage. Thus, dried plasma spots hold a promising approach to HCV testing and treatment delivery among risk groups like PWID.⁵ It would be difficult and cumbersome to use traditional methods of blood collection, such as drawing of blood and storage of samples, in the community settings. In Melbourne, point-of-care testing of HCV, follow-up, and linkage to treatment was seen as a feasible and acceptable method to PWID, suggesting further evaluation of this approach.⁶ There is a lacuna of information on the use of dried plasma spots to identify the viral infection among PWID. This study aimed to explore the use of dried plasma spots as a reliable and feasible alternative for HCV screening among non-treatment seeking population of PWID attending the syringe exchange programme in Delhi, India.

The Institute’s Ethics Committee of All India Institute of Medical Sciences, New Delhi, approved the research protocol prior to the start of the study. A cross-sectional study was conducted from January to December 2025, by the National Drug Dependence Treatment Centre, AIIMS Delhi, India. Delhi State AIDS Control Society supports NGOs to run Targeted Intervention (TI) through which PWID are identified and registered in the community to provide HIV prevention services. The PWID registered with two such TIs, with a history of injection sharing, were recruited. After due consent, their sociodemographic and drug use history was recorded on a questionnaire. Blood samples (5 mL) were collected by venipuncture into ethylenediamine tetra acetic acid tubes. Separation of plasma samples was done immediately by centrifugation. DPS spots were prepared by spotting 40 uL plasma sample (5-10 circles) onto Whatman 903 filter paper. The remaining plasma samples were stored. Plasma spotted samples were air dried at room temperature (18-25°C) and stored in a plastic bag with a desiccator before being used. Both the plasma samples and dried plasma spots were transported to the centralised laboratory and stored.

On the day of analysis, the elution of HCV from dried plasma spots was efficiently done using normal saline with overnight incubation at 37°C. The elute was transferred into a fresh tube for HVC. For comparison, paired plasma samples were analysed simultaneously. A fully automated immunoassay analyser (Abbott-Architect, Illinois, USA) based on chemiluminescent microparticle immunoassay (CMIA) technology was used for anti-HCV screening. The interpretation of the anti-HCV result is >1.0 Signal to cutoff ratio (S/CO) as reactive and <1.0 S/CO as non-reactive. Plasma-based quality controls (negative and positive) were also run along with all the experiments (Supplementary Material).

In this study, total of 188 males with a history of injecting drug use were included, with a mean (SD) age of 27.44 (6.21) yr. The majority of the participants were unmarried (56.7%), fully employed (75.5%), and educated (68.9%) from primary to higher secondary. The drug use history revealed heroin as the most commonly used opioid, used daily by the injection route by the participants. The mean (SD) duration of injecting drug use was 29 (10.4) months. Prevalence of HCV among PWID was found to be 69.7% (131/188) in the plasma samples collected in the study. The DPS testing showed that 68.8% (129/188) were positive for HCV antibodies. The comparison of anti-HCV (S/CO) in DPS and paired plasma samples showed a linear correlation, correlation coefficient, r=0.90 (P<0.05) ( Figure). There were two positive plasma samples (S/CO=4.1 and 3.27) that tested negative in paired dried plasma spots samples (S/CO=0.85 and 0.63), respectively. None of the dried plasma spots samples tested false positive in the study. Further, the performance of the dried plasma spots assay was calculated as sensitivity 98.5%, specificity 100%, positive predictive value (PPV) 100%, negative predictive value (NPV) 96.6%, and accuracy 98.9%.

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Figure. The screening of HCV from DPS and paired plasma samples (n=188).

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This study was planned with an aim to explore the use of dried plasma spots as a reliable and feasible alternative for HCV screening among non-treatment seeking population of PWID. To the best of our knowledge, this is the first cross-sectional study performed in PWID from the community setting in India. The screening of HCV revealed high prevenance (69.7%) among PWID in this study. A recent study from a treatment setting in South India reported about 50% HCV seroprevalence among PWID.⁷ A high prevalence of HCV (69.7%) in this study further underscores the importance of screening in high-risk populations, especially from non-treatment-seeking PWID from the community.

The use of filter paper has been reported among PWID to screen for HCV. A recent study from China used DBS and DPS to screen HCV using ELISA assay.⁸ The present study uses a fully automated chemiluminescence-based assay for the efficient extraction and recovery from DPS. The plasma samples (n=2) reported negative in the paired DPS may be attributed to the weak anti-HCV immunoassay reactivity. Earlier studies also reported false negative from dried spots, which are rare incidents.^8,9 The anti-HCV screened between DPS and plasma showed good and strong correlation (r=0.90), which is similar to the previous reports.¹⁰ The sensitivity, specificity, and accuracy of the method being 98.5%, 100%, and 98.9%, respectively, which is in line with the previous reports on the use of DPS for anti-HCV screening.⁸

The strengths of this study include real time screening of PWID along with the use of an efficient extraction and screening method that can be scaled up in a resource-efficient, cheap, and patient-friendly manner. The limitations of the study are small sample size and the exploration of using dried blood spots samples for HCV screening. Additionally, this study is still ongoing, and the method to use dried blood spot is underway. This incremental work will pave the way to a finger-prick-based testing protocol from the community. The method developed has the potential to screen HCV from non- treatment seeking populations within the community. Further, this study adds to the limited information on the people who inject drug screening using dried plasma spots in a real-time setting.