Extent of hepatitis E virus elimination is affected by stabilizers present in plasma products and pore size of nanofilters

Abstract

Background and ObjectiveTo investigate the physico-chemical properties of hepatitis E virus (HEV) with regard to inactivation/removal, we have studied four isolates with respect to sensitivity to heat during liquid/dry-heating as well as removal by nanofiltration. Materials and MethodsHepatitis E virus in an albumin solution or phosphatebuffered saline (PBS) was liquid-heated at 60 ° C for a preset time. HEV in a freezedried fibrinogen containing stabilizers was also dry-heated at 60 or 80 ° C for a preset time. In addition, to clarify the removal of HEV, the purified virus in PBS was filtered using several types of virus-removal filter (nanofilters) that have different pore sizes. HEV infectivity or genome equivalents before and after the treatments were assayed by a semiquantitative cell-based infectivity assay or quantitative polymerase chain reaction assay, respectively.Results Hepatitis E virus isolates in albumin solutions were inactivated slowly at 60 ° C for 5 h and the resultant log reduction factor (LRF) was from 1·0 to ≥ 2·2, whereas the virus in PBS was inactivated quickly to below the detection limit and the LRF was ≥ 2·4 to ≥ 3·7. The virus in a freeze dried fibrinogen containing trisodium citrate dihydrate and L -arginine hydrochloride as stabilizers was inactivated slowly and the LRF was 2·0 and 3·0, respectively, of the 72 h at 60 ° C, but inactivated to below the detection limit within 24 h at 80 ° C with an LRF of ≥ 4·0. The virus in PBS was also confirmed as to be approximately 35 nm in diameter by nanofiltration. These results are useful for evaluating viral safety against HEV contamination in blood products. ConclusionThe sensitivity of HEV to heat was shown to vary greatly depending on the heating conditions. On the other hand, the HEV particles were completely removed using 20-nm nanofilters. However, each inactivation/removal step should be carefully evaluated with respect to the HEV inactivation/removal capacity, which may be influenced by processing conditions such as the stabilizers used for blood products.

Hepatitis E in Developed Countries: Current Status and Future Perspectives

“…Little is known on the physico-chemical properties of HEV with regard to inactivation/removal during industrial processes. Current data indicate that the sensitivity of HEV to heat vary depending on the heating conditions and the composition of the sample [18]. HEV particles are completely removed using 20 nm nanofilters [18].…”

Thermal stability of hepatitis E virus assessed by a molecular biological approach

“…By monitoring of seroconversion of pigs experimentally inoculated with an HEV GT3-containing liver suspension it was shown that incubation at 56°C for one hour did not affect infectivity, whereas the suspension was no longer infective after heating at 191°C (internal temperature of 71°C) or above 100°C each for five minutes [22]. Yunoki et al showed that proteins or magnesium may act as stabilizers, which can increase heat stability of HEV [23]. …”

Hepatitis E Virus

“…However, neither kinetics of inactivation nor the influence of stabilisers such as those used in the production of clotting factors have been extensively investigated. Studies by a Japanese manufacturer of plasma products suggest a certain temperature resistance of HEV [223]. In these experiments the inactivation of HEV seemed to be limited to a factor of 2 log10 during pasteurisation of albumin (60 °C, 10 h).…”

“…Such filters are often used in the production of factor IX (FIX) and in part in the production of immunoglobulins or highly purified FVIII. Filtration of complex coagulation factors is only possible by using medium pore size filters (35-50 nm) that presumably will not remove HEV effectively [223] (PEI, unpublished data).…”