Disease condition and impact on patients HEV is an RNA virus and a leading cause of acute viral hepatitis worldwide (1). Hepatitis E disease presents as acute, viral hepatitis. During the first week of illness, many symptoms are nonspecific, including fever, malaise, nausea and vomiting. After the prodromal phase, patients experience a period of acute, icteric hepatitis, including jaundice, dark urine, pale stools and prolonged cholestasis with elevated liver enzymes. Symptoms can last 4–6 weeks (2). Most cases occur in older adolescents and adults. In general, these cases are mild and self-limiting, yet approximately 1–2% of cases die. However, some populations are more prone to severe disease. Women infected during pregnancy are at increased risk of fulminant hepatic failure and its associated complications, including hepatic encephalopathy, cerebral oedema and disseminated intravascular coagulation. HEV infection during pregnancy also has poor outcomes for the fetus, including low birth weight, small for gestational age, preterm birth and intrauterine death (3). A recent meta-analysis found the case fatality rate of hepatitis E during pregnancy to be 26% (IQR: 17–41%) for the mother, 33% (IQR: 19–37%) for the fetus and 8% (IQR 3–20%) for the neonate (4). HEV is a pathogen of global concern (5). However, the burden of disease is not distributed evenly; HEV is very common in low-income countries, where it causes substantial burden of disease, while relatively few cases are reported from high-income countries (5). There are four genotypes of HEV that infect humans. Genotypes 1 and 2 only infect humans and are thought to be transmitted primarily via contaminated drinking water. These genotypes are most common in South-East Asia and Africa (6). They are responsible for large outbreaks, with cases numbering in the tens of thousands. Outbreaks have been reported from East and South-East Asia, and protracted outbreaks have occurred in Africa, often affecting displaced populations (7). However, genotypes 1 and 2 also cause substantial disease outside of outbreaks (6). In India, between 25% and 50% of clinical hepatitis cases are caused by HEV, even in the absence of an outbreak (6). Genotypes 3 and 4 infect humans and a wide range of mammals, notably wild and domestic swine. These genotypes are usually transmitted zoonotically from eating infected meat and are largely reported from Europe, East Asia and the Americas. Here, HEV is responsible for sporadic cases and small foodborne outbreaks (8). Does the test meet a medical need? The availability of HEV diagnostic tests will improve differential diagnosis capability in the case of acute jaundice syndrome. Early diagnosis will alert authorities to the possibility of an outbreak, which can mobilize mitigation efforts. Due to the lack of diagnostic testing in areas where HEV is most common, the burden of HEV disease is underestimated and often poorly understood at the country level. This lack of awareness and understanding by country officials has prevented the use of a vaccine during or in endemic areas, to prevent outbreaks. How the test is used NATs are considered the gold standard for diagnosing HEV infection. However, due to the laboratory requirements, NATs are not always available. When a suspect HEV case presents at the community level, defined as any person presenting with an acute (recent, new or abrupt) onset of jaundice (yellowing of whites of eyes or skin) or dark urine and pale clay stools, an anti-HEV IgM RDT should be performed, if available, to determine the diagnosis (9, 10). If the rapid IgM test is positive, this can be considered case confirmation. If the RDT is positive early in the clinical course of infection, samples may still be sent to a reference laboratory for PCR, or IgM enzyme-linked immunosorbent assay (ELISA) confirmation. If the patient tested negative on the RDT and no other cause for the acute jaundice has been found, a NAT such as a PCR test to detect HEV RNA or an ELISA to detect anti-HEV IgM should be performed to confirm the negative diagnosis (see link to diagnostic algorithm below). In the case of immunocompromised individuals, antibody assays are less sensitive, and those with clinical suspicion should be confirmed via PCR (11). Patients who are immunocompromised should be tested for HEV RNA in three clinical settings: when the anti-HEV IgM is negative and alanine aminotransferase activity is elevated, when the HEV RNA in blood and stool persists for 3–6 months (to identify a chronic infection) and when a recent reduction in immunosuppression has been made or antiviral therapy has been started (to monitor chronic infection) (12). HEV RNA testing can also be useful in immunocompetent individuals to confirm ongoing infection in the absence of anti-HEV IgM when suspicion for infection is high or to perform viral genotyping. Note: the following algorithm was provided by the applicant as part of this application and is not a WHO algorithm. Diagnostic algorithm for HEV infection: https://docs.google.com/presentation/d/ 10Yd0I67LoepbYSHxIIZwrMEKUJttU9OA/edit#slide=id.p1 (accessed 1 August 2023).

Prevalence A commonly cited meta-analysis estimated that HEV causes 20.1 million infections, 3.4 million clinical cases, 70 000 deaths and 3000 stillbirths annually due to the epidemic-prone genotypes 1 and 2 (13). While there is increasing recognition of HEV in genotype 3 and 4 endemic areas, the burden of disease has not been well classified. However, the burden of disease of HEV is vastly underestimated due to the lack of diagnostic capabilities in areas where HEV is most common. Poor understanding of the global burden of disease has hindered efforts to implement control and prevention strategies. Socioeconomic impact Few studies have looked at the economic impact of hepatitis E. In Nepal, acute HEV infection led to an average of 10 bedridden days and 22 sick days; wage earners lost nearly 20% of their yearly income (14). An estimated 108 years of life lost (YLL) per 1000 individuals, 144 years lived with disability per 1000 individuals and 252 disability-adjusted life-years (DALYs) per 1000 individuals were attributed to an HEV outbreak in Uganda (15). However, this estimate used disability weights for untreated diarrhoeal disease and did not account for differential severity among pregnant women and neonates, therefore underestimating the true economic impact.

The use of PCR to diagnose acute hepatitis E in suspected cases is supported by WHO guidance on recognition, investigation and control of waterborne outbreaks of hepatitis E (9, 10). NATs are an alternative test that can be used in serum, plasma or stool.

There are currently no systematic reviews that detail the clinical accuracy of HEV PCR tests. The applicant performed a systematic review to identify primary studies on the clinical accuracy of HEV PCR tests. A systematic search was conducted in PubMed using the search terms: ((HEV) OR (Hepatitis E Virus)) AND ((PCR) OR (polymerase chain reaction test) OR (Polymerase Chain Reaction) [Mesh]) AND ((clinical accuracy) OR (clinical precision) OR (Sensitivity and Specificity) [Mesh]) and ended up with a result of 330 studies. Results were limited to the English language. Thirty articles were found eligible for full-text review after the screening. Five primary studies were found relevant to report the clinical accuracy of PCR tests. This review identified several primary studies that examined the clinical accuracy of different commercially available NATs. All studies used PCR products, including multiplex reverse transcriptase (RT)-PCR for simultaneous amplification of HBV, HCV and HEV (16), one-step real-time RT-PCR for rapid and sensitive detection (17), analytical sensitivity and performance of three HEV RT-PCR assays (18), and comparison between real-time RT-PCR and conventional RT-PCR (19) and RT-PCR-ELISA (20). Given the costs associated with establishing conventional single-target PCR, a study was designed to develop a reliable and cost-effective multiplex PCR assay (Bangalore Genei, India). The multiplex PCR was compared to in-house single-virus PCR assays in serum samples from acute hepatitis patients from India. The clinical accuracy of the multiplex compared to single-virus detection was reported as follows: sensitivity for HEV was 74%, with 100% specificity. The PPV was found to be 100%, whereas NPV was 83.8 ± 12% (16). Another study used the QuantiTect Probe RT-PCR Master Mix (Qiagen, Valencia, CA, United States). This assay was found to correctly detect 13 of 13 HEV isolates of four different HEV genotypes with 100% sensitivity. This in vitro study added HEV RNA to samples to evaluate the PCR assay (17). A separate study examined the analytical sensitivity and performance of three HEV RT-PCR assays in asymptomatic blood donors from Europe: RealStar HEV RT-PCR, hepatitisE@ceeramTools HEV RT-PCR and ampliCube HEV RT-PCR. For individual sample screening, the RealStar HEV RT-PCR assay consistently had the highest sensitivity of 37.8 IU/mL (95% CI: 22.2– 671.2), followed by the hepatitisE@ceeramTools kit with 86.8 IU/mL (95% CI: 68.9–124.7) and the ampliCube HEV RT-PCR kit with 180.4 IU/mL (95% CI: 128.5–355.2). Intra-assay and inter-assay variations were in acceptable ranges, with variation coefficients < 5% (18). Ahn et al. compared real-time RT-PCR and nested RT-PCR in Asian serum samples. They found a significant difference in the limit of detection from 1.68 × 101 copies of RNA in real-time RT-PCR to 1.68 × 104 copies of RNA in nested RT-PCR (19). In a study that used the MJ Mini thermal cycler (Bio-Rad, CA, United States), RT-PCR-ELISA was found to be 10–100 times more sensitive than nested RT-PCR with a 0.01 ng/μL limit of detection and did not falsely detect HEV in terms of specificity. This study used swine stool samples to detect HEV genotype 3, a genotype that infects both humans and swine (20).

A systematic search across several databases at the time of submission of this application found no systematic reviews that examined the direct clinical utility of HEV PCR tests on patient care and management. There is, however, one recent systematic review which examined data on the epidemiology of HEV in LMICs of Africa and Asia based on seroprevalence, outbreaks and risk factors for infection (21). The lack of routine testing is a major limiting factor to understanding the burden of HEV disease in LMICs. Ninety-one studies from 29 countries examined seroprevalence, which generally increased by age. Forty- nine completed or ongoing HEV outbreaks from 18 countries were reported from 1988 to 2017. However, most outbreaks are not reported in the published literature, and alternative sources were not used for this systematic review. Risk factors for HEV infection included increased exposure to contaminated water sources and poor hygiene (although not all studies found this association). Risk factor data suggested an increased likelihood of current or recent HEV infection and disease associated with faecal-oral transmission of HEV, as well as exposures to blood and animals. However, the authors emphasized that most data on HEV prevalence and risk factors come from specialized studies or outbreak reports. There are no primary studies specific to the clinical utility of HEV PCR tests on patient management and care. However, there are several preventative strategies that could reduce the number of future HEV cases and deaths, especially if these preventative measures are introduced early in an outbreak. The applicant identified 12 studies that point to the clinical utility of an HEV diagnosis and greater understanding of the epidemiology and burden of HEV disease in the local context. A better understanding of the local epidemiology and disease burden caused by HEV will allow country officials to make evidence-based decisions on intervention implementation both prior to and during outbreaks.

No data available.

HEV PCR tests are needed to diagnose immunocompromised patients, who may not form sufficient antibodies. PCR tests require advanced laboratory infrastructure and trained personnel and therefore may be inaccessible to some populations. Increased availability of diagnostic testing would allow country officials to better understand the local burden of HEV, and therefore to make evidence-based decisions regarding use of vaccines both routinely and during outbreaks. No ethical issues were identified.