Periodic Reporting for period 2 - REPRODIVAC (Next-generation vaccines and diagnostics to prevent livestock reproductive diseases of worldwide impact)
Reporting period: 2024-03-01 to 2025-08-31
Endemic and zoonotic infectious reproductive diseases of livestock cause major economic losses globally and threaten food security and public health. REPRODIVAC is developing new and improved vaccines and diagnostic to better control four priority abortifacient diseases: porcine reproductive and respiratory syndrome (caused by PRRSV), Q fever (caused by Coxiella burnetii), ovine enzootic abortion (OEA) (caused by Chlamydia abortus), and porcine brucellosis (caused by Brucella suis). The consortium spans academia and industry with expertise in structural and molecular biology, microbiology, immunology, plant and veterinary sciences, diagnostic test and vaccine development. This interdisciplinary approach enables to address the target reproductive disorders by: 1) applying reverse and structural vaccinology to select and design vaccine and diagnostic candidate antigens; 2) exploiting relevant protein expression systems for the production of these antigens; 3) producing rationally attenuated and viral vectored vaccines; 4) utilising established animal models for the rational evaluation of candidate vaccines; and 5) developing a suite of molecular and immunological diagnostic tests, including point-of-care tests (PoC) to discriminate vaccinated from infected animals (DIVA). We will develop a broadly protective PRRSV vaccine and a serological ELISA that will reliably correlate with PRRSV neutralising antibody titres. We will identify protective subunit vaccine candidates for Q fever and validate a companion serological DIVA test. We will refine an OEA vaccine for commercialisation and pursue a complementary approach towards a DIVA-compatible next-generation subunit vaccine and serological and molecular DIVA/PoC tests. We will develop live attenuated B. suis vaccines and brucellosis diagnostic tests with improved specificity.
In RP2, activities have focused on the: i) production of selected antigens; ii) isolation and production of neutralising antibodies for the refinement of antigenic targets; iii) experiments to assess the antigenicity and immunogenicity of selected antigens and vaccine candidates; and iv) further development and evaluation of diagnostic assays.
• Single B cells have been sorted from pigs, whose sera broadly neutralise PRRSV and recombinant monoclonal antibodies generated and expressed to be characterised for specificity and neutralising capacity. Experiments are ongoing to provide a high-resolution model of the virion by cryo-electron tomography, and a new associated partner involved to support structure-guided vaccine design. Recombinant pseudorabies virus vectoring PRRSV glycoprotein or epitopes have been tested in pigs for immunogenicity, as a pilot proof-of-principle study.
• Live attenuated B. suis vaccine candidates were further characterised in THP-1 macrophages and evaluated in a mouse model for cross-protection against the three biovars causing porcine brucellosis. Selected antigens for delayed-type hypersensitivity (DTH) skin test and serological diagnosis were produced and characterised. Preliminary in vivo DTH and serological tests were performed.
• Rationally selected C. burnetii antigens have been produced and used to immunise sheep and polyclonal antibodies generated against these vaccine candidates tested for their ability to neutralize the bacteria through a novel assay developed within REPRODIVAC. In addition, candidate diagnostic antigens have been evaluated for serodiagnostic potential using an indirect ELISA platform and a panel of serum samples from ruminant livestock with known C. burnetii infection and vaccination status
• Recombinant C. abortus antigens have been produced and their immunoreactivity evaluated using convalescent serum and peripheral blood cells from immune sheep. Selected antigens have been screened in the surrogate C. abortus mouse models. The protocol to produce COMC has been provided to one of the industry partners to support the commercialisation of this new vaccine. A molecular point-of-care loop-mediated isothermal assay (LAMP) has been developed and optimised for the detection of C. abortus DNA compatible with a direct extraction method and is currently undergoing validation using samples from experimental challenge studies.
• Single B cells have been sorted from pigs, whose sera broadly neutralise PRRSV and recombinant monoclonal antibodies generated and expressed to be characterised for specificity and neutralising capacity. Experiments are ongoing to provide a high-resolution model of the virion by cryo-electron tomography, and a new associated partner involved to support structure-guided vaccine design. Recombinant pseudorabies virus vectoring PRRSV glycoprotein or epitopes have been tested in pigs for immunogenicity, as a pilot proof-of-principle study.
• Live attenuated B. suis vaccine candidates were further characterised in THP-1 macrophages and evaluated in a mouse model for cross-protection against the three biovars causing porcine brucellosis. Selected antigens for delayed-type hypersensitivity (DTH) skin test and serological diagnosis were produced and characterised. Preliminary in vivo DTH and serological tests were performed.
• Rationally selected C. burnetii antigens have been produced and used to immunise sheep and polyclonal antibodies generated against these vaccine candidates tested for their ability to neutralize the bacteria through a novel assay developed within REPRODIVAC. In addition, candidate diagnostic antigens have been evaluated for serodiagnostic potential using an indirect ELISA platform and a panel of serum samples from ruminant livestock with known C. burnetii infection and vaccination status
• Recombinant C. abortus antigens have been produced and their immunoreactivity evaluated using convalescent serum and peripheral blood cells from immune sheep. Selected antigens have been screened in the surrogate C. abortus mouse models. The protocol to produce COMC has been provided to one of the industry partners to support the commercialisation of this new vaccine. A molecular point-of-care loop-mediated isothermal assay (LAMP) has been developed and optimised for the detection of C. abortus DNA compatible with a direct extraction method and is currently undergoing validation using samples from experimental challenge studies.
The REPRODIVAC project envisaged that vaccines and diagnostics have the potential to:
• Increase the availability of veterinary medicinal products for animal species (e.g. sheep and goats) traditionally overlooked by pharmaceutical companies due to the small market size.
• Progress the implementation of preventive veterinary medicine programs by improving external and internal biosecurity in farms by: (i) correctly identifying infected animals to avoid transmission to naïve ones; (ii) protecting animals with efficacious vaccines to decrease the probability of becoming sick/dying and transmitting the pathogen; (iii) correctly treating infected animals with suitable medicinal products.
• Improve animal welfare by decreasing the prevalence of the selected diseases at a population level (better control of the disease) and decrease the clinical signs by the protection conferred by the vaccines.
• Positively impact human health by reducing and preventing the circulation of the zoonotic pathogens C. abortus, C. burnetii and B. suis, all of which can affect humans and their foetuses to different levels of severity.
• Increase the availability of veterinary medicinal products for animal species (e.g. sheep and goats) traditionally overlooked by pharmaceutical companies due to the small market size.
• Progress the implementation of preventive veterinary medicine programs by improving external and internal biosecurity in farms by: (i) correctly identifying infected animals to avoid transmission to naïve ones; (ii) protecting animals with efficacious vaccines to decrease the probability of becoming sick/dying and transmitting the pathogen; (iii) correctly treating infected animals with suitable medicinal products.
• Improve animal welfare by decreasing the prevalence of the selected diseases at a population level (better control of the disease) and decrease the clinical signs by the protection conferred by the vaccines.
• Positively impact human health by reducing and preventing the circulation of the zoonotic pathogens C. abortus, C. burnetii and B. suis, all of which can affect humans and their foetuses to different levels of severity.