Immunization with inactivated Middle East Respiratory Syndrome coronavirus vaccine leads to lung immunopathology on challenge with live virus

Abstract

To determine if a hypersensitive-type lung pathology might occur when mice were given an inactivated MERS-CoV vaccine and challenged with infectious virus as was seen with SARS-CoV vaccines, we prepared and vaccinated mice with an inactivated MERS-CoV vaccine. Neutralizing antibody was induced by vaccine with and without adjuvant and lung virus was reduced in vaccinated mice after challenge. Lung mononuclear infiltrates occurred in all groups after virus challenge but with increased infiltrates that contained eosinophils and increases in the eosinophil promoting IL-5 and IL-13 cytokines only in the vaccine groups. Inactivated MERS-CoV vaccine appears to carry a hypersensitive-type lung pathology risk from MERS-CoV infection that is similar to that found with inactivated SARS-CoV vaccines from SARS-CoV infection.

Keywords: coronavirus; Eosinophils; immunopathology; Middle East Respiratory Syndrome; vaccination.

Figure 1.

Mean serum-neutralizing antibody titers to MERS-CoV of vaccinated mice 3 weeks after the second immunization. Alum and MF59 are adjuvant only groups, WIV is whole inactivated vaccine (WIV) only, Alum/WIV is WIV formulated with Alum adjuvant, MF59/WIV is WIV formulated with MF59 adjuvant. The serum neutralizing antibody titers are expressed as Geometric Mean Titer (GMT) based on a 2-fold dilution sequence beginning at 1:2 (Log₂). * Significantly different (P < 0.01) after correcting for multiple comparisons.

Figure 2.

Mean viral titers of MERS-CoV on days 3 and 6 after intranasal challenge of vaccinated mice with 100 LD₅₀ of MERS-CoV. Lung homogenates and total RNAs extracted from tissues of vaccinated mice at days 3 and 6 post challenge with MERS-CoV were subjected to Vero E6 cell-based infectivity assay and one-step real-time RT-PCR analyses targeting the upE gene of MERS-CoV for assessing viral loads, as previously described (5,6). A serial 10-fold diluted MERS-CoV stock with a titer of 10⁷ TCID₅₀/ml was included in parallel during the quantitative PCR assays to calculate and express the levels of upE gene expression in individual specimens as log₁₀ TCID₅₀ equivalents per gram of tissue. Alum and MF59 are adjuvant-only groups, WIV is whole inactivated vaccine (WIV) only, Alum/WIV is WIV formulated with Alum, MF59/WIV is WIV formulated with MF59. A: Vero E6-based infectious viral titers at Day 3, B: Vero E6-based infectious viral titers at Day 6, C: RT-PCR-based viral load at Day 3, and D: RT-PCR-based viral load at Day 6. * Significantly different (P < 0.01) after correcting for multiple comparisons.

Figure 3.

Representative photomicrographs of lung tissue 3 days after challenge of previously vaccinated mice with MERS-CoV. Lung sections were stained with an antibody directed specifically against eosinophilic major basic protein as described (3); eosinophils are brown. The vaccine groups (alum only, MF59 only, WIV only, WIV plus Alum and WIV plus MF59) and the eosinophil infiltration severity score (E0 and E2) are noted on the micrograph; E0 is none, E2 is moderate.

Figure 4.

Mean lung cytokine levels on day 3 after challenge of vaccinated mice with MERS-CoV. Alum and MF59 are adjuvant only groups, WIV is whole inactivated vaccine (WIV) only, Alum/WIV is WIV formulated with Alum, MF59/WIV is WIV formulated with MF59. Test 1 and test 2 are separate day tests of the same lung tissue specimen. Results are mean fold increase over naïve transgenic mice based on ΔCt values of each group in reference to those of the internal mouse GAPDH gene. * Significantly greater than for the naïve mouse group (P < 0.01) after correcting for multiple comparisons; ** P = 0.026.