Hepatitis C virus (HCV) infection impacts tens of millions of people worldwide[1] with one million incident infections annually.[2, 3] As the first step in the care cascade, screening and diagnosis of HCV are key to elimination; however, there is considerable loss to follow-up at the screening and diagnosis steps.[4] A major barrier to streamlined diagnosis and treatment in the United States (U.S.) is the lack of a point-of-care (POC) test to establish the presence of HCV RNA. POC RNA tests, where results are available within an hour, are approved for the diagnosis of acute and chronic HCV infections in other countries across the globe[5] and offer several advantages. For example, POC testing can be utilized in CLIA-waived settings, thereby greatly expanding the locations in which screening can effectively occur and the number of individuals treated. Additionally, POC testing is associated with improved time to treatment initiation.[6, 7]

The development of the Xpert^® HCV VL Fingerstick paved the way for the Xpert^® HCV test.[5, 8], and the Xpert^® HCV VL Fingerstick was one of the first molecular point-of-care (POC) HCV RNA tests to receive CE marking,[9] with extensive studies confirming its clinical performance.[10, 11] Other POC HCV RNA tests have also been evaluated, but uptake has been limited largely due to a more complicated workflow.[11, 12] There are several notable differences between the Xpert^® HCV VL Fingerstick and Xpert^® HCV tests. Xpert^® HCV is designed to be simpler to perform and is a qualitative test that can be run by untrained users in CLIA-waived settings. The Xpert^® HCV test also has an early termination feature, whereby results are available in approximately 40 minutes for HCV RNA-detectable samples and 60 minutes for HCV RNA-undetectable samples. Another notable difference is the sample collection; 250-500 μl are collected into a BD Microtainer^® collection tube, 100μl of which is pipetted into the Xpert^® HCV cartridge. This difference in fingerstick collection procedures was required since the Minivette^® POCT used in sample collection for the Xpert^® HCV VL Fingerstick is not FDA-cleared.

The purpose of this study was to evaluate the clinical performance of the Xpert^® HCV test in K2-EDTA fingerstick blood collected from symptomatic and / or individuals at risk of HCV infection when performed by untrained users on the GeneXpert^® Xpress System in a CLIA-waived setting.

This multicenter, prospective clinical study of 985 participants at 15 CLIA-waived sites across the U.S. demonstrated good performance of the Xpert^® HCV test relative to PIS (PPA of 93.4%; NPA of 99.8%). The performance of the Xpert^® HCV test was consistent with studies evaluating the molecular performance of the Xpert^® HCV VL Fingerstick.[5] A systematic review of the diagnostic accuracy of the Xpert^® HCV VL Fingerstick test across seven studies had a sensitivity and specificity for HCV RNA detection of 99% (95% CI: 97%–99%) and 99% (95% CI: 94%–100%), respectively.[10] Additionally, a systematic review of the diagnostic accuracy of POC HCV RNA assays for diagnosis of HCV demonstrated a pooled sensitivity of 99% (95% CI: 98−99%) and specificity of 99% (95% CI: 99−100%).[11] The NPA of 99.8% in our study is similar to these studies but the PPA of 93.4% in our study is lower. This could be due to slight differences in the LoD of the qualitative Xpert^® HCV test and the quantitative Xpert^® HCV VL Fingerstick test, differences in the participant population in the study, differences in the collection device, and/or differences in the comparator methods.

The FDA reviewed the results of this study under its De Novo premarket review pathway and found that the Xpert^® HCV test provides reasonable assurance of safety and effectiveness for the intended use. The FDA authorized the Xpert^® HCV in June of 2024.[18] This is the first HCV RNA test approved from fingerstick blood, without specimen processing required. In October 2024, the CDC published guidelines[19] for consideration of implementation for POC HCV RNA testing, highlighting that setting characteristics influence the HCV testing approach with the following settings favoring POC RNA testing: high HCV prevalence, brief encounter settings, and settings with no phlebotomy or laboratory access. The CDC and AASLD/IDSA also emphasized the benefit of POC testing is maximized when paired with highly accessible HCV treatment.[17, 20, 21]

Several characteristics of the Xpert^® HCV test make it useful in these settings. Venous blood access can be challenging among many individuals, and many settings serving high risk individuals do not have access to phlebotomy. The Xpert^® HCV test uses a fingerstick sample and approximately 10% of participants were excluded due to insufficient fingerstick sample. The proportion of insufficient sample collection in this study of Xpert^® HCV is higher than studies of the Xpert^® HCV VL Fingerstick[5, 8] which is likely due to differences in sample collection procedures with the microtainer stipulating minimum collection of 250μl (compared to 100μl). For this study, inadequate sample collection was more frequent at two (2) of the 15 sites accounting for approximately 40% of this sample set. Within the two sites, a single untrained self-trained user was responsible for many (approximately 39.1%) of the samples with insufficient volume. . It should be noted that sufficient sample collection often improves over time with sufficient training and experience.[22]. Subsequent to study completion, fingerstick collection training materials in addition to the manufacturer’s instructions have been updated to aid in sample collection.

A prospective study done on the Xpert^® HCV test prior to the clinical trial established the sample stability for fingerstick blood of 4 hours which enables re-testing from the same sample collected if initial testing produces in a non-determinant result.[23] Of note, the final proportion of non-determinant results of 1.2% was low, consistent with prior studies,[8] increasing the utility of the test in these settings.

There are limitations to the Xpert^® HCV test. There is a potential for false positive results due to contamination during sample collection, however, due to the closed cartridge design of the Xpert^® HCV test, the possibility of amplicon contamination is very low. Additionally, there is the potential for false negative results which may occur when very low levels of HCV RNA are not detected.[24] In this study, investigation into discrepant results did not provide sufficient evidence to ascertain a root cause; however, differences in the LoD between the qualitative Xpert^® HCV test (35 to 135 IU/mL)[23] in fingerstick blood and the quantitative cobas^® HCV test (13.7 IU/mL) in serum likely explains some false negative results. Additionally, the four specimens with the cobas detectable, not quantifiable results, is interpreted analytically within the manufacturer’s instructions as “HCV RNA detected but not quantified”. This may demonstrate low-level viraemia which could be indicative of previous infection with limited current clinical relevance.

The GeneXpert^® Xpress System has limited capacity for high volume testing, as the device runs up to four samples simultaneously. Additionally, depending on the setting, the price of the instrument and ongoing service may make testing with the Xpert^® HCV more expensive than other diagnostic strategies. There is data, however, supporting cost-effectiveness of a one-step HCV RNA POC strategy in high-risk populations. A study utilizing a health economic model to evaluate the cost and outcomes of a one-step HCV RNA POC diagnostic strategy compared to SoC among PWID in the Medicaid population, SoC suggested that a POC testing strategy could yield improved outcomes at lower cost in a high-risk population.[25] Additional studies also suggest that, compared to SoC, one-step HCV RNA POC diagnosis may be cost effective[26–28] However, studies that have evaluated cost effectiveness of strategies for diagnosing HCV infection in other countries, found that a two-step POC-based strategy (POC HCV antibody followed by either POC or dried blood spot-based HCV RNA testing) may be most effective.[26–28] The limitations of the HCV Xpert^® test were taken into account in the 2024 CDC guidance on implementation of HCV RNA POC testing which describes settings that favor POC HCV RNA and describes approaches to POC HCV RNA instrument and cartridge procurement.[19]

In conclusion, this study demonstrated that the Xpert^® HCV test has the ability to detect HCV RNA in individuals with active HCV infection, regardless of HCV antibody status. Data for this study led to the FDA authorization for clinical use. This is a critically important step in HCV elimination in the U.S. given the need for screening efforts to be coupled with building infrastructure to support testing, linkage to care, and treatment of infected individuals. The availability of POC HCV RNA testing has significant potential to improve treatment access and move the United States closer to elimination of HCV.