Convalescent human plasma candidate reference materials protect against Crimean-Congo haemorrhagic fever virus (CCHFV) challenge in an A129 mouse model

Abstract

Crimean-Congo Haemorrhagic Fever Virus (CCHFV) is spread by infected ticks or direct contact with blood, tissues and fluids from infected patients or livestock. Infection with CCHFV causes severe haemorrhagic fever in humans which is fatal in up to 83 % of cases. CCHFV is listed as a priority pathogen by the World Health Organization (WHO) and there are currently no widely-approved vaccines. Defining a serological correlate of protection against CCHFV infection would support the development of vaccines by providing a 'target threshold' for pre-clinical and clinical immunogenicity studies to achieve in subjects and potentially obviate the need for in vivo protection studies. We therefore sought to establish titratable protection against CCHFV using pooled human convalescent plasma, in a mouse model. Convalescent plasma collected from seven individuals with a known previous CCHFV virus infection were characterised using binding antibody and neutralisation assays. All plasma recognised nucleoprotein and the Gc glycoprotein, but some had a lower Gn glycoprotein response by ELISA. Pooled plasma and two individual donations from convalescent donors were administered intraperitoneally to A129 mice 24 h prior to intradermal challenge with CCHFV (strain IbAr10200). A partial protective effect was observed with all three convalescent plasmas characterised by longer survival post-challenge and reduced clinical score. These protective responses were titratable. Further characterisation of the serological reactivities within these samples will establish their value as reference materials to support assay harmonisation and accelerate vaccine development for CCHFV.

Keywords: CCHF; CCHFV; Convalescent; Immunity; Infection; Mouse model; Vaccine.

Fig. 1

Characterisation of donations by ELISA using in-house developed ELISAs for a) Gn, b) Gc or c) Np; or commercially available assays (d and e). f) Parallel line analysis of two donation serial dilutions in commercial assays.

Fig. 2

Neutralisation of HAZV by individual donations. a) Mean integrated intensity for each donation including the negative control serum and b) mean integrated intensity expressed relative to negative control serum. Bars represent means +/- SEM. c) CCHFV neutralisation curves, dotted line represents 50 % neutralisation.

Fig. 3

Passive protection of anti-CCHFV plasma in the A129 mouse model conferred by a pooled material (Pool) or individual donations 004(2015) or 005(2017). Parameters evaluated in the model are shown including survival, weight change (as a% change from time of challenge), temperature and clinical score. The level of anti-CCHFV antibody detectable for each individual by Np ELISA is also shown.

Fig. 4

Viral RNA levels in blood, liver and spleen by rt-qPCR. Open circles represent individuals terminated at the end of the experiment (14 days) and closed circles represent individuals that met humane end points during the study. Bars represent the mean of the group and dotted lines represent the limit of detection for the assay. Asterisk denotes significantly lower level of detectable virus in mice receiving 005 (2017) neat plasma compared to the control group (p = 0.030).

Fig. 5

Histology images of liver and spleen from mice receiving neat plasma. Scale bars represent 100 µM. Upper panel shows H&E staining of liver and RNAScope of liver sections. Lower panel shows spleen H&E staining with RNAScope images below. Representative images from mice that met humane end points on comparable days (day of termination and animal ID noted below treatment group).

Fig. 6

Histology scores and levels of RNAScope staining. Open circles represent individuals terminated at the end of the experiment (14 days) and closed circles represent individuals that met humane end points during the study. Bars represent the mean of the group (+/- SE). Asterisk denotes significantly lower level of detectable pathology in mice receiving 005 (2017) neat plasma compared to the control group by ANOVA with post-hoc Bonferroni (liver histopathology p = 0.036 and spleen viral RNA p = 0.019).

Fig. 7

Ranking of samples by CCHFV antibody reactivity in different assays.