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WHO Guidelines on Hepatitis B and C Testing. Geneva: World Health Organization; 2017 Feb.

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WHO Guidelines on Hepatitis B and C Testing.

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11DETECTION OF VIRAEMIC HCV INFECTION – to guide who to treat

11.1. Recommendations

Detection of viraemic infection
  • Directly following a reactive HCV antibody serological test result, the use of quantitative or qualitative NAT for detection of HCV RNA is recommended as the preferred strategy to diagnose viraemic infection.
    Strong recommendation, moderate/low quality of evidence
  • An assay to detect HCV core (p22) antigen, which has comparable clinical sensitivity to NAT, is an alternative to NAT to diagnose viraemic infection1.
    Conditional recommendation, moderate quality of evidence
1

A lower level of analytical sensitivity can be considered if an assay is able to improve access (i.e. an assay that can be used at the point of care or suitable for dried blood spot [DBS] specimens) and/or affordability. An assay with a limit of detection of 3000 IU/mL or lower would be acceptable and would identify 95% of those with viraemic infection, based on available data.

11.2. Background

Detection of antibodies to HCV is used to determine current or past HCV infection (i.e. exposure to HCV infection), and therefore to triage those who require further evaluation to determine if active viral replication is present. Approximately 15–45% of persons who are infected with HCV will spontaneously clear the infection (267). These persons remain HCV antibody positive but are no longer infected with HCV. Diagnosis of viraemic HCV infection in those who are HCV antibody positive will distinguish persons with viraemic HCV infection and in need of treatment from those who have cleared the infection. This is generally done using NAT technologies to detect HCV RNA, but an alternative and potentially less costly option to NAT is to conduct testing to detect HCV core (p22) antigen.

Nucleic acid testing (NAT)

Both quantitative and qualitative methods are available for the detection of viraemic HCV infection. Quantitative NAT has been widely used for measuring viral load and identifying those in need of treatment, as well as in assessing treatment response (5, 15). Qualitative NAT allows for rapid and sensitive detection of the virus as well as evidence of a decline in viral RNA level below a defined threshold. There are currently five quantitative HCV RNA (viral load) assays that are commercially available with another two in the pipeline (268). At present, there has been limited comparison of the two methods. Although NAT technologies are very sensitive and specific for the detection of viraemia, they require sophisticated laboratory equipment and skilled staff. Assays to detect HCV RNA that may be used at or near the point of care have recently become commercially available. A comprehensive review of the HCV diagnostics landscapes by UNITAID is available (268).

HCV core (p22) antigen testing

In addition to NAT, it is possible to assess for viraemic infection by testing for HCVcAg – an HCV nucleocapsid peptide 22 (p22), which is released into the plasma during viral assembly and can be detected both early on and throughout the course of HCV infection (269). Serological methods that test for detection of HCVcAg have the potential to be less costly and centralized than NAT, but evaluation has been limited in low-resource settings. There are now several assays commercially available for stand-alone detection of HCVcAg (270). Detection of HCVcAg has also been used as an additional marker in a fourth-generation HCV Ag/Ab serological assay, because HCVcAg is detectable earlier than antibodies to HCV. However, the addition of core antigen was intended to increase sensitivity of the assay in early infection and not to differentiate seropositivity from active viraemic HCV infection.

11.3. Summary of the evidence

Two systematic reviews were undertaken, which evaluated the diagnostic accuracy for detection of viraemic HCV infection of (i) qualitative versus quantitative NAT (see Web annex 5.7); and (ii) HCVcAg testing versus NAT (see Web annex 5.8) (271).

Diagnostic accuracy and limit of detection of HCV NAT assays

The systematic review identified four eligible studies (272–275) that compared the performance of three quantitative HCV RNA NAT assays to a reference qualitative NAT (two assays used). Although early-generation qualitative NAT assays were able to detect the presence of HCV in plasma at concentrations a full log lower (i.e. about 10-fold less) than quantitative NAT assays, the lower limit of quantification of new versions of quantitative assays is now comparable to most commercial qualitative assays, i.e. 15 IU/mL.

Diagnostic accuracy and limits of detection of HCVcAg assays

There were 50 studies that evaluated seven commercial HCVcAg assays. There was significant variation in performance between the different assay brands (Table 11.1) (271). The pooled sensitivity and specificity with 95% CI were: ARCHITECT 93.4% (95% CI: 88.7–96.2) and 98.7% (95% CI: 96.9–99.4); Ortho ELISA 93.2% (95% CI: 81.6–97.7) and 99.2% (95% CI: 87.9–100); and Hunan Jynda 59.5% (95% CI: 46–71.7) and 82.9% (95% CI: 58.6–94.3). The sensitivity for the Lumipulse was 95% (95% CI: 90.2– 99.8) in one study, but specificities could not be calculated. The estimates for the ARCHITECT assay were more homogeneous and precise as this assay has been the most extensively studied.

TABLE 11.1. Summary of diagnostic accuracy of HCV core antigen assays compared to NAT.

TABLE 11.1

Summary of diagnostic accuracy of HCV core antigen assays compared to NAT.

A pooled quantitative analysis of data available from three studies demonstrated a close correlation between HCVcAg and HCV RNA at viral loads above 3000 IU/mL. The LoD for the most sensitive assay is 3 fmol/L HCVcAg or 0.06 pg/mL, which equates to an LoD of about ∼1000–3000 IU/mL by NAT, and is consistent with the analytical sensitivity (LoD) reported by the manufacturer.

NAT assays are considered the reference standard for the detection of viraemia, but the quality of studies comparing quantitative versus qualitative assays for detection of viraemia was rated as low because of small numbers of studies and heterogeneity in populations.

The overall quality of the evidence for the recommendation to use HCVcAg was rated as low to moderate because of inconsistency and imprecision.

11.4. Rationale for the recommendations

Balance of benefits and harms

Use of quantitative or qualitative NAT assays for detection of HCV RNA

The Guidelines Development Group made a strong recommendation for the use of a NAT assay (either qualitative or quantitative) as the preferred strategy for diagnosis of viraemic HCV infection based on moderate-/low-quality evidence for several reasons:

  1. The new generation of quantitative and qualitative assays have the same LoD, which is around 15 IU/mL. However, quantitative assays are a reproducible method to detect and quantify HCV RNA in plasma or serum.
  2. A supplementary review of the literature showed that that 95% of those with chronic infection have a viral load >10 000 IU/mL except, temporarily, a minority with partial viral control between 5 and 12 months post infection. Therefore, the range of clinically observed HCV viral loads is rarely below the lower range of the limit of quantification (LoQ) of quantitative assays, and most NAT assays (quantitative or qualitative) will capture the majority of viraemic infections.
  3. Although quantitative RNA assays are considered the gold standard assays for the diagnosis and monitoring of HCV, the high cost of these assays and laboratory requirements means that they are not readily available in resource-limited settings. However, new NATs for use at or near the point of care for quantitation of HCV RNA are already available. These devices are easier to use than the laboratory-based NAT assays and can potentially improve access to diagnosis of viraemic HCV infection.

Use of HCV core antigen for detection of HCV RNA

The Guidelines Development Group recognized that there is limited access to NAT assays in resource-limited settings and that this represents an important barrier to antiviral treatment. The Group made a conditional recommendation to consider use of HCVcAg assays as an alternative to NAT to diagnose viraemic HCV infection, based on moderate-quality evidence, for several reasons:

  1. HCVcAg assays can utilize existing serological testing platforms and are potentially lower-cost options than NAT. They could serve as a more affordable replacement to NAT for HCV detection in the future.
  2. Although HCVcAg testing is currently limited to only a few platforms and even those with the highest performance do not reach the sensitivity of NAT, some well-performing HCVcAg assays have high sensitivity (up to 93.4% for certain commercial assays and high specificity (>98%), and good correlation with HCV RNA to an LoD of roughly 3000 IU/mL, which will detect over 95% of chronic HCV infections. However, it was noted that there was wide variation in sensitivity/specificity between assays and also within the same brand of assay for all but the Abbott ARCHITECT.
  3. HCVcAg tests also offer the potential in the future to be applied as a one-step screening test as HCVcAg appears earlier than HCV antibodies (1–2 days after HCV RNA appears), has a high specificity, and so does not require any further confirmatory testing. However, such a strategy would be cost–effective only in very high-prevalence settings.

The risk in the use of HCVcAg is of potentially missing cases due to reduced clinical sensitivity. A further consideration is that it is preferable to select an assay that can be used for both diagnosis of viraemia and for test of cure to simplify the diagnostic pathway. On the basis of the limited current evidence, the HCVcAg assay cannot be recommended as a monitoring test.

Acceptability, values and preferences

The values and preferences survey identified preferences for future HCV testing strategies among respondents. Key preferences were for a single-step HCV diagnostic strategy with a low-cost point-of-care test for confirming viraemic infection (48% of respondents). Of these, 52% opted for an HCV RNA test because of its high sensitivity, and 35% for an HCVcAg assay because of its lower cost and ease of use. More than half the respondents were prepared to compromise on sensitivity down to 95% in order to gain a reduction in the price of the test. Forty-seven per cent of respondents also indicated a preference for a test that uses capillary blood and therefore could be more easily performed in point-of-care settings, even at the expense of test sensitivity. A short turnaround time (at least same day) was identified as another key consideration to reduce loss to follow up, cost of transportation, and enable providers to see more patients within a day.

Feasibility

The survey of hepatitis testing experience in 19 LMICs found that NAT for HCV RNA is available at a third of the sites but 40% of respondent countries do not have access to NAT for HCV diagnosis in their countries. The HCVcAg assay was not available at any site.

Resource considerations

The resources required for quantitative NAT were considered to be substantial, with the cost per test ranging from US$ 30 to US$ 200. Furthermore, the laboratory equipment is expensive and requires technicians with specialized training. The cost of testing for HCVcAg is currently US$ 25–50 (MSF data), which is comparable to qualitative NAT (US$ 43–51), but this is still a major barrier to its use.

11.5. Implementation considerations

  1. Near patient or point-of-care (POC) technologies. The development of reliable and affordable POC NAT and HCVcAg tests able to diagnose viraemic infection in field settings will be crucial for expanding hepatitis testing services (see Chapter 17.3.5 – diagnostic innovations). These devices offer the possibility of a same-day diagnosis of viraemic infection either alone or when combined with an HCV antibody RDT and for test of cure. Since they are also more affordable than the laboratory-based assays, they can potentially improve access to early diagnosis, monitoring and linkage to care and treatment services, as well as reduce loss to follow up.
  2. Immediate NAT directly after a positive serological result. The value of prompt testing for viraemia after a positive antibody result was highlighted, as patients with resolved HCV infection following spontaneous clearance can be reassured and those with viraemic infection could be promptly referred for care and treatment.
  3. Genotyping. In most countries, there is a mix of HCV genotypes among persons with chronic HCV infection. The 2016 Hepatitis C treatment guidelines (5) provide recommendations on the preferred and alternative DAA regimens by HCV genotype. Therefore, knowing a patient's genotype is still important for determining the most appropriate treatment regimen. Genotype determination, however, is expensive and not available in all settings. Where genotype information is unavailable, pragmatic decision-making may be required, taking into account the common genotypes circulating in the affected population. However, as pangenotypic regimens become available over the next year, this will no longer be required.

Research gaps

  • Establish the proportion of patients with chronic HCV infection with low viral loads that may be missed by HCV RNA or cAg assays that have a higher limit of detection (i.e. 3000 IU/mL).
  • Evaluate the diagnostic accuracy, cost, cost–effectiveness and impact of HCVcAg or HCV RNA assays as a one-step diagnostic strategy.
  • Assess the impact of HIV or HBV coinfection or genotype (particularly genotypes 4, 5 and 6, on which there are limited data) on detection of viraemia.
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