Abstract
Hepatitis E virus (HEV) is a major concern in public health worldwide. Infections with HEV genotypes 3, 4, or 7 can lead to chronic hepatitis while genotype 1 infections can trigger severe hepatitis in pregnant women. Infections with all genotypes can worsen chronic liver diseases. As virions are lipid-associated in blood and naked in feces, efficient methods of propagating HEV clinical strains in vitro and evaluating the infectivity of both HEV forms are needed. We evaluated the spread of clinical strains of HEV genotypes 1 (HEV1) and 3 (HEV3) by quantifying viral RNA in culture supernatants and cell lysates. Infectivity was determined by endpoint dilution and calculation of the tissue culture infectious dose 50 (TCID50). An enhanced HEV production could be obtained varying the composition of the medium, including fetal bovine serum (FBS) and dimethylsulfoxide (DMSO) content. This increased TCID50 from 10 to 100-fold and allowed us to quantify HEV1 infectivity. These optimized methods for propagating and measuring HEV infectivity could be applied to health safety processes and will be useful for testing new antiviral drugs.
Highlights
The hepatitis E virus (HEV) is one of the most common causes of hepatitis worldwide
We have developed an endpoint-dilution method for calculating tissue culture infectious dose 50 (TCID50) that is based on Hepatitis E virus (HEV) culture and quantifying HEV RNA
Dulbecco’s modified Eagle’s medium (DMEM), Medium 199 (M199), William’s E medium (32551), fetal bovine serum (FBS), 0.05% trypsin-EDTA containing phenol red, phosphate-buffered saline without calcium and magnesium (PBS), were all purchased from Thermo Fisher Scientific, Life
Summary
The hepatitis E virus (HEV) is one of the most common causes of hepatitis worldwide. HEV is classified in the Orthohepevirus A species that includes at least eight genotypes, five of which (HEV1 to HEV4 and HEV7) infect humans [1,2]. HEV1 and 2 are restricted to humans; their transmission is linked to poor sanitation and contaminated water. HEV3 and 4 are prevalent in industrialized countries where they are zoonotic; their reservoir includes pigs, wild boar, deer, and rabbits [3,4,5,6]. HEV7 was identified in camel and was found in an immunocompromised patient [7]. HEV can be transmitted by blood transfusion [8]
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