A reverse vaccinology approach identified novel recombinant tick proteins with protective efficacy against Rhipicephalus microplus infestation

Authors: DOMINGUES, LUISA - Boehringer Ingelheim; Bendele, Kylie; Bodine, Deanna; HALOS, LENAIG - Boehringer Ingelheim; CUTOLO, ANDRE - Boehringer Ingelheim; LIEBSTEIN, MARTIN - Boehringer Ingelheim; WIDENER, JUSTIN - Boehringer Ingelheim; FIGUEIREDO, MONICA - Boehringer Ingelheim; MORENO, YOVANY - Boehringer Ingelheim; EPE, CHRISTIAN - Boehringer Ingelheim; DA COSTA, ALVIMAR JOSE - Instituto Nacional De Pesquisas Da Amazonia (INPA); COSTA GOMES, LUCAS VINICIUS - Instituto Nacional De Pesquisas Da Amazonia (INPA); GUERRERO, FELIX - Retired ARS Employee

Submitted to: Ticks and Tick Borne Diseases
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/1/2024
Publication Date: 10/19/2024
Citation: Domingues, L.N., Bendele, K.G., Bodine, D.M., Halos, L., Cutolo, A.A., Liebstein, M., Widener, J., Figueiredo, M., Moreno, Y., Epe, C., Da Costa, A., Costa Gomes, L., Guerrero, F.D. 2024. A reverse vaccinology approach identified novel recombinant tick proteins with protective efficacy against Rhipicephalus microplus infestation. Ticks and Tick Borne Diseases. 15 (2024) 102403. https://doi.org/10.1016/j.ttbdis.2024.102403.
DOI: https://doi.org/10.1016/j.ttbdis.2024.102403

Interpretive Summary: The cattle tick, Rhipicephalus microplus, causes significant economic losses to the cattle industry. Tick control is predominately accomplished using pesticide applications, but alternative control methods such as vaccines are needed due to the tick's capacity to quickly develop pesticide resistance. A bioinformatic approach was used to screen, evaluate, and rank 121,768 sequences for their potential use as a vaccine candidate. Ten candidates were chosen to move forward with but only six were able to be successfully produced in the expression systems with three of the candidates confirmed using mass spectrometry. Four of the candidates were successfully produced in large volumes for trials in cattle. Three of the candidates produced significant antibody responses in animals in the trial. One candidate provided the same or better control of tick infestations than GAVAC, a commercially available vaccine, available for use in some countries.

Technical Abstract: The cattle tick, Rhipicephalus microplus, causes significant economic losses to the cattle industry. Tick control is predominately via pesticide applications. However, alternative control methods such as vaccines are needed due to the tick's capacity to quickly develop pesticide resistance. We used a reverse vaccinology approach composed of 3 vaccine prediction and 11 annotation tools to evaluate and rank 121,768 translated open reading frames (ORFs) from the tick's transcriptome for their potential use as anti-tick vaccine antigens. Following the in silico rankings, we manually annotated the 200 highest ranked antigens and used our scientific judgement to select 10 transcript ORFs as vaccine candidates for expression in Pichia pastoris or insect cells. Only 6 of the 10 vaccine candidate antigens could be successfully expressed and purified in vitro as recombinant proteins with > 1 mg quantity. The expression of these 6 transcripts and the proteins encoded by them were investigated in various tick tissues by reverse transcription-polymerase chain reaction (RT-PCR) and mass spectrometry (MS), respectively. RT-PCR confirmed the expression of all 6 transcripts in tick RNA. Three of the 6 transcripts' corresponding ORFs were confirmed by MS to be present in tick tissue protein extracts. Only 4 of the 6 vaccine candidate antigens, designated BI-TS002, BI-TS004, BI-TS008, and BI-TS009, were successfully expressed and purified in sufficient quantity (> 10 mg) for immunogenicity and efficacy trials in cattle. Cattle vaccinated with BI-TS004, BI-TS008 and BI-TS009 yielded a statistically significant difference in antibody response post-vaccination. BI-TS008 and BI-TS009, were formulated with adjuvant and cattle stall tests conducted over a 175 day period to evaluate vaccine efficacy against R. microplus infestations. Both initial (designated Phase 1 and calculated from tick collections of Days 60-94 days post-first vaccination) and long-term (designated Phase 2 and calculated from tick collections of Days 152-175 post-first vaccination) efficacies were determined. Both an adjuvant only negative control group and a positive control group that used the commercially available GAVAC anti-tick vaccine were used. The overall Phase 1 trial efficacies of BI-TS008, BI-TS009, and GAVAC were 44.0%, 23.9%, and 52.4%, respectively. The overall Phase 2 trial efficacies of BI-TS008, BI-TS009, and GAVAC were 46.1%, -16.4%, and 26.3%, respectively. Thus, the anti-tick vaccine antigen BI-TS008 provided equivalent or better control of R. microplus infestations than the GAVAC vaccine positive control.