Source: KANSAS STATE UNIV submitted to NRP
EFFICACY OF PROTOTYPE LIVE-VECTORED POLYVALENT AFRICAN SWINE FEVER VIRUS VACCINES
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
COMPLETE
Funding Source
AFRI COMPETITIVE GRANT
Reporting Frequency
Annual
Accession No.
1012784
Grant No.
2016-67015-26873
Cumulative Award Amt.
$449,044.19
Proposal No.
2017-03271
Multistate No.
(N/A)
Project Start Date
Feb 1, 2017
Project End Date
Jan 31, 2022
Grant Year
2021
Program Code
[A1221]- Animal Health and Production and Animal Products: Animal Health and Disease
Recipient Organization
KANSAS STATE UNIV
(N/A)
MANHATTAN,KS 66506
Performing Department
Diagnostic Medicine/Pathobiolo
Non Technical Summary
The African Swine Fever Virus is a high-consequence pathogen that causes hemorrhagic fever in pigs and some isolates are capable of killing nearly all infected pigs. The U.S.A swine industry is the leading pork exporter, accounting for >28% of global exports, and is thus a major source of food, revenue, and employment. The pathogen poses a high risk to the U.S.A swine industry as it continues to spread globally and therefore, it is important that effective countermeasures are developed to safeguard the industry. There is no vaccine or treatment available for controlling this disease and therefore, development of an effective and safe vaccine is necessary for use in case there is an outbreak in North America. We have developed two rationally designed vaccine candidates and tested their ability to safely induce immune responses in commercial pigs. The goal of the current project is to test whether these vaccine candidates can confer protection of pigs against the virus.There is scientific evidence to show that protection against the African swine fever virus can be stimulated with a vaccine since pigs that recover from infection with isolates that do not cause disease are protected against isolates that cause disease. Due to safety concerns, development of a vaccine based on carefully selected virus proteins, as opposed to an attenuated live virus vaccine, is more attractive for use in U.S.A since the virus is not present in North America. However, the current challenges are identification of suitable virus vaccine candidate proteins and an effective way for immunizing pigs. Candidate vaccines formulated using one or two African swine fever virus proteins have so far failed to induce acceptable protection. It is envisaged that development of an effective vaccine will require identification and validation of multiple suitable candidate proteins that will induce significant protection in majority of the vaccinated pigs.The vaccine candidates that we have developed contain multiple promising African swine fever virus proteins and immunization of commercial pigs with the vaccines induced promising immune responses that increased over a ten week period after inoculation of a single dose. Overall, the vaccines were well tolerated and no serious negative effects were observed. Notably, all the pigs responded well to each virus component included in the vaccine. In addition, the induced responses increased dramatically after inoculation of a booster dose. More importantly, the induced immune responses strongly recognized the actual African swine fever virus. Taken together, the outcomes from this study showed that the vaccine is capable of safely inducing strong immune responses in commercial pigs. The next logical step is to test whether the vaccines can confer protection to pigs against the virus. The study will be conducted by immunizing pigs with the candidate vaccines. Naïve pigs will serve as negative controls. The pigs will be challenged with a lethal dose of the virus to evaluate protection. Successful development of an effective vaccine is critical for the containment of an African swine virus outbreak and safeguarding of the U.S.A pig industry. In addition, such a vaccine will play a critical role in controlling global spread of the virus and reduce outbreaks in endemic regions.
Animal Health Component
(N/A)
Research Effort Categories
Basic
100%
Applied
(N/A)
Developmental
(N/A)
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
31135991090100%
Knowledge Area
311 - Animal Diseases;

Subject Of Investigation
3599 - Swine, general/other;

Field Of Science
1090 - Immunology;
Goals / Objectives
Since African Swine Fever Virus is a high-consequence Transboundary Animal Disease pathogen, it poses a threat to the U.S.A swine industry, the leading pork exporter, a major source of food and employment. The pathogen is spreading globally and given that development of effective vaccines takes time, it is imperative that knowledge is generated to inform vaccine development since there is no vaccine or treatment available for controlling this disease. We have identified several promising vaccine candidates and shown that they can safely induce immune responses in commercial pigs. The major goal of this project is to test whether two experimental vaccines formulated using these candidates can confer protection of pigs against the virus.The specific Project Objectives are;I) Evaluate protective efficacy of adenovirus-encoded ASFV multivalent cocktail-I. We will determine whether or not, immunization of pigs with a cocktail of adenovirus expressing ten lead candidate antigens will confer protection in a majority of vaccinees upon lethal challenge; II) Evaluate protective efficacy of adenovirus-encoded ASFV multivalent cocktail-II. We will determine whether or not, immunization of pigs with a second cocktail of adenovirus expressing ten select candidate antigens will confer protection in a majority of vaccinees upon lethal challenge.
Project Methods
Study I:Experimental Design and Procedures: Stocks of recombinant adenoviruses evaluated in a previous immunogenicity and safety study were used to generate bulk vaccine virus needed for the efficacy study I. The scaled up virus was titrated and underwent quality control tests to confirm protein expression and authenticity. We have also validated that progeny virus is still replication-incompetent in non-complementing cells. In addition, affinity purified recombinant proteins have been generated using recombinant baculoviruses that were generated and used to produce proteins used in the immunogenicity studies mentioned above. The recombinant proteins will be used for immune readouts in the proposed efficacy studies.Immunization of piglets: Two groups, A and B, of age-matched ASFV sero-negative and virus-free piglets (n=15) will be used in this study. Each piglet in group A will be inoculated intramuscular (IM) with the adenovirus-vectored ASFV multivalent cocktail-I (1 x 1011 ifu of each recombinant adenovirus) formulated using adjuvant as per the protocol we used in the immunogenicity studies. Negative control piglets (n=15) will similarly be inoculated, but with an equivalent dose of Adenoluciferase virus. The piglets will be boosted 28 days post-priming using cognate immunizing dose and then challenged one month post-boost by intramuscular inoculation of 104 HAD50 ifu of a well-characterized ASFV isolate, which we have previously demonstrated to induce lethal infections in naive pigs.Readouts post-immunization and post-challenge: Antibody and T cell responses, post-prime, post-boost, and post-challenge (in survivors) will be evaluated weekly by ELISA and by IFN-g EliSpot. Following challenge, clinical signs will be monitored daily by a Veterinarian and recorded, and time to death will be the terminal readout. In addition, viremia in blood and nasal swabs will be quantified by PCR. Splenocytes and skin fibroblasts from each pig will be used to evaluate ASFV antigen-specific CTL responses to identify antigen-specific responses that correlate with protection or significant delay in time to death post-challenge. In addition, antigen-specific antibody profiles in terminal sera will be used to determine whether there will be a correlation with protection or significant delay in time to death post-challenge. The significance of the differences in immune readouts, clinical scores, and viremia between the treatment (group A) and the control (group B) will be analyzed using Analysis of Variance followed by determining Fishers Least Significant Difference if distributional assumptions for ANOVA are met or a Kruskal-Wallis test (non-parametric equivalent of ANOVA) if ANOVA is not appropriate. A significance level of P<0.05 will be used for all analyses.Study II:Experimental Design and Procedures: The ability of the chaperoned ASFV multivalent cocktail to confer protection will be evaluated in piglets (n=15) as above, whereas the negative controls (n=15) will receive adeno-luciferase sham treatment as above. Viremia, immune and clinical readouts will be evaluated and analyzed by ANOVA as above. We expect that this cocktail will confer protection in the majority of the vaccinees upon challenge. However, if no significant protection or significant delay to death is demonstrated in the treatment group compared to the negative control, we will conclude that the antigens in this cocktail alone are not sufficient to confer significant protection in pigs.

Progress 02/01/17 to 01/13/22

Outputs
Target Audience:1. Fellow NIFA National Project Directors and National Program Leaders, and CRWAD attendees during the 2021 virtual meeting. One abstract was accepted and one poster was presented virtually. 2. The broader audience was the scientific community which was reached through publication of our research findings. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Several people received hands-on training and acquired new research skills. Specifically, one research scientist [Tae Kim]; three PhD students [Michelle Zajac, Rakshith Kumar, Huldah Sang]; one PhD/DVM student [Jaydent McCall]; and one MSC student [Leeanna Burton] received hands-on training in this project [generation/QC/titration of adenovirus-vectored ASFV antigen expression viruses, handling/immunization/bleeding of pigs, processing sera and PBMCs, and isolation of splenocytes and kidney fibroblasts. Five undergraduate students [Kylynn Mallen, Rachel Brown, Brandon Green, Debra Holliman, Tristan Burnum] also received hands-on training [animal handling/daily monitoring/bleeding/processing blood to generate sera/isolate cells/run ELISAs]. In addition, eleven DVM students were involved in animal handling/daily monitoring and bleeding. A Research Associate [Jianxiu Yao] received training in animal handling/daily monitoring/bleeding/processing blood to generate sera/isolate cells. Several personel [T. Kim, M. Zajac, R. Kumar, H. Sang, J. McCall, L. Burton] received training in working with a select agent (ASFV) at BSL3 and BSL3-Ag. J. McCall competed and received NBAF Scientist Training Program sponsorship from USDA-APHIS ($410,000): he will conduct his studies on ASFV. M. Zajac has also received the same sponsorship and her PhD research is also focused on ASFV subunit vaccine development. How have the results been disseminated to communities of interest?1. One abstract was accepted and presented as a virtual poster at CRWAD 2021 meeting. 2. A total of four journal articles will report comprehensive results from the studies conducted. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? 1. Objective I was completed and accomplishments were included in previous annual progress reports. 2. For objective II, safety and protective efficacy of recombinant adenovirus-encoded ASFV multivalent cocktail-II was evaluated in piglets. The adenovirus expressed ASFV antigens were shown to be authentic after validation using ASFV convalescent serum. Groups of piglets were immunized with the cocktail alone or formulated with one of two adjuvants. All the vaccine candidate formulations were safe and they were well-tolerated by piglets. Only one out of five piglets (20%) immunized with the virus cocktail without adjuvant survived until study termination 30 days post-challenge. Although this was a low protective efficacy, the outcome suggests that development of an ASFV subunit vaccine is feasible but further development and optimization is needed. 3. A total of four publications and one abstract.

Publications

  • Type: Journal Articles Status: Under Review Year Published: 2022 Citation: Zajac, M.D., N. Sangewar, S. Lokhandwala, J. Bray, R.P. Bishop, S. D. Waghela, and W. Mwangi. Adenovirus-vectored African Swine Fever Virus p220 polyprotein induces robust antibody, IFN-gamma, and CTL responses in pigs. Frontiers in Veterinary Science.
  • Type: Journal Articles Status: Awaiting Publication Year Published: 2022 Citation: Zajac, M.D., J. Yao, R. Kumar, H. Sang, N. Sangewar, S. D. Waghela, L. Burton, D. Kumar, T. Kim, and W. Mwangi. 2022. Efficacy of prototype adenovirus-vectored African swine fever virus multi-antigen vaccines. Frontiers in Veterinary Science.
  • Type: Journal Articles Status: Awaiting Publication Year Published: 2022 Citation: Zajac, M.D., J. Yao, R. Kumar, H. Sang, N. Sangewar, S. D. Waghela, T. Kim, and W. Mwangi. 2022. Granzyme B and IFN-gamma responses to putative CTL epitopes by lymphocytes from pigs immunized with adenovirus-vectored prototype multi-antigen ASFV vaccine. Frontiers in Immunology.
  • Type: Journal Articles Status: Awaiting Publication Year Published: 2022 Citation: Zajac, M.D., R. Kumar, N. Sangewar, L. Burton, S. D. Waghela, T. Kim, and W. Mwangi. 2022. Characterization of anti-porcine Granzyme B monoclonal antibodies. Frontiers in Veterinary Science.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Mwangi, W., S. Lokhandwala, N. Sangewar, J. Bray, J. Yao, H. C. Sang, and S. Waghela. Efficacy of prototype live-vectored African swine fever virus vaccines. CRWAD 2021. ID: V-P080.


Progress 02/01/20 to 01/31/21

Outputs
Target Audience:1. Fellow NIFA National Project Directors and National Program Leaders, and CRWAD attendees during the 2020 virtual meeting. Two abstracts were accepted. One project report was presented virtually. 2. The broader audience was the scientific community which was reached through publication of our research findings. 3. Invited virtual presentation [ASFV Special symposium hosted by CRWAD] on our efforts on ASFV vaccine development were also made. Changes/Problems:In vivo studies scheduled for 2020 were hindered by constraints caused by COVID-19. The study will be conducted in 2021. What opportunities for training and professional development has the project provided?Several undergraduate and graduate students received hands-on training. Specifically, three PhD students [Huldah Sang, Rakshith Kumar, Michelle Zajac]; one DVM/PhD student [Jayden McCall], and one DVM student [Tori Matta] received hands-on training in this project [generation of adenovirus-vectored ASFV antigen expression constructs, animal bleeding, processing sera and PBMCs, determining viremia, virus neutralization assays. Four undergraduate students [Leeanna Burton, Kylynn Mallen, Kyci Sperry, Rachel Brown] also received hands-on training [animal handling/bleeding/processing blood to generate sera/isolate cells/run ELISAs]. These folks will also be involved in the pending efficacy studies. Leeanna Burton was admitted to pursue MS in this project. The project continued to train Research Associate [Jianxiu Yao] and she was involved in the execution of the studies. How have the results been disseminated to communities of interest?1. Two abstracts were accepted. One project report was presented virtually at CRWAD 2020 virtual meeting. 2. Oral virtual presentation: ASFV Special symposium hosted by CRWAD. 3. Publications; i) Cadenas-Fernández, E., J. M. Sánchez-Vizcaíno, A. Kosowska, B. Rivera, A. Rodríguez-Bertos, J. Yao, J. Bray, S. Lokhandwala, W. Mwangi, and J. A. Barasona. 2020. Adenovirus-vectored African swine fever virus antigen cocktail is not protective against virulent Arm07 isolate in Eurasian wild boar. Pathogens J. Feb 28;9(3):171. doi: 10.3390/pathogens9030171. PMID:32121082. ii) Huldah, S., G. Miller, Q. Manzil, S. Lokhandwala, N. Sangewar, S. D. Waghela, and W. Mwangi*. 2020. Progress towards development of Efficacious and Safe African Swine Fever Virus Prototype Vaccines. Front Vet Sci. 2020 Feb 21;7:84. doi: 10.3389/fvets.2020.00084. eCollection 2020. PMID: 32154279. iii) Abstract: Mwangi, W., S. Lokhandwala, N. Sangewar, J. Bray, J. Yao, H. C. Sang, S. Waghela, M. Sheahan, and R. Rowland. Efficacy of prototype live-vectored African swine fever virus vaccines. CRWAD 2020 ID:133. iv) Abstract: Mwangi, W., S. Lokhandwala, N. Sangewar, J. Bray, J. Yao, H. C. Sang, S. Waghela, M. Prospects for an African Swine Fever Virus Subunit Vaccine. Special Symposium on African Swine Fever. CRWAD 2020. What do you plan to do during the next reporting period to accomplish the goals?Conduct objective 2 animal study to evaluate safety, tolerability, and efficacy of the second experimental adenovirus-vectored ASFV subunit vaccine. This study was scheduled to be conducted in 2020 but it was not done due to constraints caused by COVID-19.

Impacts
What was accomplished under these goals? Objective 1 has been completed and outcomes were included in the previous report. Two papers and two abstracts were published as well as one oral virtual presentation. For objective 2, recombinant adenoviruses encoding ASFV multicistronic expression cassettes were generated. Protein expression was confirmed and antigens were authenticated using ASFV convalescent serum. The recombinant viruses were scaled up, quality control validated in preparation for conducting immunogenicity and efficacy studies in pigs. In addition, putative cytotoxic T lymphocyte epitopes present in the ASFV vaccine candidate antigens were identified using bioinformatic tools and peptides synthesized in readiness for conducting epitope mapping using T cells from pigs that will be immunized with the adenovirus-vectored prototype subunit vaccine.

Publications

  • Type: Journal Articles Status: Published Year Published: 2020 Citation: Huldah, S., G. Miller, Q. Manzil, S. Lokhandwala, N. Sangewar, S. D. Waghela, and W. Mwangi*. 2020. Progress towards development of Efficacious and Safe African Swine Fever Virus Prototype Vaccines. Front Vet Sci. 2020 Feb 21;7:84. doi: 10.3389/fvets.2020.00084. eCollection 2020. PMID: 32154279.
  • Type: Journal Articles Status: Published Year Published: 2020 Citation: Cadenas-Fern�ndez, E., J. M. S�nchez-Vizca�no, A. Kosowska, B. Rivera, A. Rodr�guez-Bertos, J. Yao, J. Bray, S. Lokhandwala, W. Mwangi, and J. A. Barasona. 2020. Adenovirus-vectored African swine fever virus antigen cocktail is not protective against virulent Arm07 isolate in Eurasian wild boar. Pathogens J. Feb 28;9(3):171. doi: 10.3390/pathogens9030171. PMID: 32121082.
  • Type: Other Status: Published Year Published: 2020 Citation: Mwangi, W., S. Lokhandwala, N. Sangewar, J. Bray, J. Yao, H. C. Sang, S. Waghela, M. Sheahan, and R. Rowland. Efficacy of prototype live-vectored African swine fever virus vaccines. CRWAD 2020 ID:133.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Mwangi, W., S. Lokhandwala, N. Sangewar, J. Bray, J. Yao, H. C. Sang, S. Waghela, M. Prospects for an African Swine Fever Virus Subunit Vaccine. Special Symposium on African Swine Fever. CRWAD 2020.


Progress 02/01/19 to 01/31/20

Outputs
Target Audience:1. The target audience were fellow NIFA National Project Directors and National Program Leaders, and CRWAD attendees during the 2019 meeting in Chicago on November 4th, 2019. An abstract was submitted/accepted, and a poster was also presented. 2. The broader audience was the scientiffic community which was reached through publication of our research findings. 3. Invited presentations on our efforts on ASFV vaccine development were also made to commercial entities [Zoetis, CEVA, HUVEPHARMA, ELANCO, MDx (S. Korea), NAVETCO (Vietnam) and academic institution [University of Miyazaki, Japan]. Changes/Problems:We had proposed and got funded to conduct ASFV subunit vaccine development studies using human adenovirus-5 vector [Ad-5]. Initial studies in pigs showed that the Ad-5 vector was not powerful enough to prime and significantly expand high quality ASFV-specific immune responses in pigs [this outcoem has now been confirmed by results from other ASFV studies conducted in Pirbright, UK]. Inclusion of an adjuvant improved immune responses but the induced immune responses did not improve efficacy. For our last animal study, we modified the antigen expression platform by adding a transcription enhancer, fused in-frame, to transgene to improve protein expression and this was verified in vitro. Safety, immunogenicity, and efficacy of a cocktail of novel ASFV antigens expressed using this new platform will be evaluated in pigs in 2020. Ourcomes from this study will be included in the final project report due in Jan 31, 2021. What opportunities for training and professional development has the project provided?Several undergraduate and graduate students received hands-on training. Specifically, two PhD students [Huldah Sang, Gabrielle Miller] received hands-on training in this project [generation of adenovirus-vectored ASFV antigen expression constructs, animal bleeding, processing sera and PBMCs, determining viremia, virus neutralization assays. Four undergraduate students [Rachel Reith, Mary Markland, Bailey Fritz, Leeanna Burton] also received hands-on training [animal handling/bleeding/processing blood to generate sera/isolate cells/run ELISAs]. These folks will also be involved in the pending efficacy studies. The project continued to train Research Associate [Jianxiu Yao] and she was involved in the execution of the studies. How have the results been disseminated to communities of interest?1. Publications; Lokhandwala, S., et al. 2019. Vet Microbiol, 235: 10-20. 2. Poster Presentation: Lokhandwala, S., V. Petrovan, L. Popescu, N. Sangewar, C. Elijah, A. Stoian, M. Olcha, J. Bray, P. R. Bishop, S. D. Waghela, M. Sheahan, R. R. R. Rowland, and W. Mwangi. Efficacy of prototype live-vectored African Swine Fever Virus Multi-antigens. CRWAD. Chicago, IL, November 4th, 2019. 3. Abstract: Mwangi, W., S. Lokhandwala, N. Sangewar, J. Bray, J. Yao, H. C. Sang, S. Waghela, M. Sheahan, and R. Rowland. 2019. Efficacy of prototype live-vectored African swine fever virus vaccines. CRWAD ID: P183. 4. Invited oral presentations: Zoetis, CEVA, HUVEPHARMA, ELANCO, MDx (S. Korea), NAVETCO (Vietnam) and academic institution [University of Miyazaki, Japan. What do you plan to do during the next reporting period to accomplish the goals?One final study will be conducted to evaluate safety, tolerability, and efficacy of a novel experimental ASFV immunogen formulated using an improved adenovirus backbone. We had expected to get this study completed in 2019 [this was a 3 yr project], but this was not achieved due, in part, to challenges associated with generation of the experimental vaccine. We have requested no-cost extension and we have received approval. Data from the initial efficacy studies conducted in pigs informed our decision to develop a new formulation that is expected to confer better survival than the 5/9 obtained with one formulation that was tested.

Impacts
What was accomplished under these goals? 1. Data from two animal studies in which experimental adenovirus-vectored ASFV immunogens were tested in two pig studies was published [Lokhandwala S, Petrovan V, Popescu L, Sangewar N, Elijah C, Stoian A, Olcha M, Ennen L, Bray J, Bishop RP, Waghela SD, Sheahan M, Rowland RRR, and Mwangi W. Adenovirus-vectored African Swine Fever Virus antigen cocktails are immunogenic but not protective against intranasal challenge with Georgia 2007/1 isolate. 2019. Vet Microbiol, 235: 10-20]. Following challenge, all pigs immunized with one formulation, as well as negative controls, succumbed to intranasal challenge with wildtype virus. Five out of nine pigs [5/9] immunized with an alternate formulation survived challenge and cleared viremia by day 17 post-challenge. 2. We also conducted another efficacy study and used infected animals to challenge test and negative controls by contact: the model mimics actual disease transmission during outbreaks. All animals succumbed to the challenge. A manuscript to report study outcomes is currently under review [Cadenas-Fernández, E., et al., 2020. Pathogens]. 3. Continued to develop a highly sensitive and superior lateral flow diagnostic device in partnership with a commercial entity [MDx, S. Korea].

Publications

  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Lokhandwala S, Petrovan V, Popescu L, Sangewar N, Elijah C, Stoian A, Olcha M, Ennen L, Bray J, Bishop RP, Waghela SD, Sheahan M, Rowland RRR, and Mwangi W. Adenovirus-vectored African Swine Fever Virus antigen cocktails are immunogenic but not protective against intranasal challenge with Georgia 2007/1 isolate. 2019. Vet Microbiol, 235: 10-20.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Mwangi, W., S. Lokhandwala, N. Sangewar, J. Bray, J. Yao, H. C. Sang, S. Waghela, M. Sheahan, and R. Rowland. 2019. Efficacy of prototype live-vectored African swine fever virus vaccines. CRWAD ID: P183.
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2019 Citation: Poster Presentation: Lokhandwala, S., V. Petrovan, L. Popescu, N. Sangewar, C. Elijah, A. Stoian, M. Olcha, J. Bray, P. R. Bishop, S. D. Waghela, M. Sheahan, R. R. R. Rowland, and W. Mwangi. Efficacy of prototype live-vectored African Swine Fever Virus Multi-antigens. CRWAD. Chicago, IL, November 4th, 2019.


Progress 02/01/18 to 01/31/19

Outputs
Target Audience:1. Presented a poster at the American Association of Immunologist Annual Meeting, Austin, TX, May 4-8, 2018 2. Invited speaker [by USDA-NIFA National Program Leaders]: 122nd USAHA Annual Meeting, Kansas City, MO, Oct. 21, 2018 3. Invited Speaker [KeyNote Talks] North American PRRS Symposium. Chicago, IL, Dec. 2, 2018. 4. Presented two posters at the the CRWAD Meeting, Chicago, IL, Dec. 3, 2018. Target audience were stakeholders from academia, industry, and federal government. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?One Postdoctoral fellow [Dr. Shehnaz Lokhandwala] was involved in the planning of in vitro and in vivo experiments, hands-on execution [generation of the prototype vaccines/immunizations/immune readouts/data procesing and analysis], and training graduate and undergraduate students. Thus, she received training that will enable her to become an independent investigator in the area of transboundary animal diseases and working in high biocontainment facility. One DVM-PhD trainee [Luka Popescu] was also involved in study execution during the immunization phase [pig immunizations/bleeding] and the ASFV challenge study [challenge dose inoculations/monitoring clinical outcomes/euthanization] in high biocontainment facility. Four PhD gradute students [Neha Sangewar, Vlad Petrovan, Annah Stoian, Mathew Olcha] received hands-on training in this project [vaccine formulation/immunization/bleeding post-immunization/monitoring clinical-viremia outcomes post-challenge]. In addition, one DVM student [Catherine Elijah] received hands-on training in handling pigs, bleeding, monitor and evaluation post-immunizations, and treatment of minor issues under the guidance of the attending veterinarian. Three undergraduate students [Lyndsey Ennen, Rachel Reith, and Mary Markland] also received hands-on training [animal handling/bleeding/processing blood to generate sera/isolate cells/run ELISAs]. These folks will also be involved in the pending efficacy studies. The project also recruited and trained a Research Associate [Jianxiu Yao] and she will be involved in the execution of pending studies. How have the results been disseminated to communities of interest?Data presentations at meetings [invited speaker], publication of abstracts, and poster presentations as indicated below. What do you plan to do during the next reporting period to accomplish the goals? We plan to evaluate protective efficacy of the second prototype ASFV experimental immunogen [Objective 2].

Impacts
What was accomplished under these goals? 1. A manuscript was generated using data generated from the previously concluded three animal studies in which we evaluated the protective efficacy of one prototype adenovirus-vectored ASFV antigen cocktail formulated in two adjuvants. Lokhandwala, S., V. Petrovan, L. Popescu, N. Sangewar, C. Elijah, A. Stoian, M. Olcha, J. Bray, P. R. Bishop, S. D. Waghela, M. Sheahan, R. R. R. Rowland, and W. Mwangi. 2018. Efficacy of adenovirus-vectored African Swine Fever Virus antigen cocktails. Scientific Reports Journal [under review]. 2. We conducted a fourth animal study to evaluate protective efficacy of the experimental ASFV subunit vaccine using contact [infected naive animals] to challenge treatment and control animals. This challenge model mimics the actual disease transmission during disease outbreak. All animals succumbed to the challenge. Data from this study is being analyzed and a manuscript will be written to report the outcome. 3. Identified novel ASFV antigen for development of a highly sensitive and superior lateral flow based diagnostic device that will be a suitable DIVA component of the subunit vaccine under development.

Publications

  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Abstracts: 1. Mwangi, Waithaka*, Shehnaz Lokhandwala, Luca Popescu, Neha Sangewar, Catherine Elijah, Vlad Petrovan, Ana Stoian, Mathew Olcha, Jocelyn Bray, Surya Waghela, Maureen Kerrigan and Raymond R. R. Rowland. Efficacy of a Prototype Live-vectored Multi-antigen African Swine Fever Virus Vaccine. J. Immunol May 1, 2018, 200 (1 Supplement) 59.10. 2. Lokhandwala, S#., N. Sangewar, J. Bray, S. D. Waghela, and W. Mwangi*. 2018. Immunogenicity of live-vectored African Swine Fever Virus pp220 antigen. CRWAD ID: 42588. 3. Lokhandwala, S#., V. Petrovan, L. Popescu, N. Sangewar, C. Elijah, A. Stoian, M. Olcha, J. Bray, P. R. Bishop, S. D. Waghela, M. Sheahan, R. R. R. Rowland, and W. Mwangi*. 2018. Efficacy of prototype live-vectored African Swine Fever Virus vaccines. CRWAD ID: 42477. Posters: 1. Lokhandwala, S#., V. Petrovan, L. Popescu, N. Sangewar, C. Elijah, A. Stoian, M. Olcha, J. Bray, P. R. Bishop, S. D. Waghela, M. Sheahan, R. R. R. Rowland, and W. Mwangi*. 2018. Efficacy of prototype live-vectored African Swine Fever Virus vaccines. CRWAD. Chicago, IL, December 3rd, 2018. 2. Lokhandwala, S#., N. Sangewar, J. Bray, S. D. Waghela, and W. Mwangi*. 2018. Immunogenicity of live-vectored African Swine Fever Virus pp220 antigen. CRWAD. Chicago, IL, December 2rd, 2018. 3. Lokhandwala, S., Luca Popescu, Neha Sangewar, Catherine Elijah, Vlad Petrovan, Ana Stoian, Mathew Olcha, Jocelyn Bray, Surya Waghela, Maureen Kerrigan, Raymond R. R. Rowland and W. Mwangi. Efficacy of a Prototype Live-vectored Multi-antigen African Swine Fever Virus Vaccine. American Association of Immunologists International Annual Meeting. Austin, TX, May 4-8, 2018.


Progress 02/01/17 to 01/31/18

Outputs
Target Audience:Presented progress report at the Annual Project Director's meeting in Chicago on December 01, 2017. The target audience were fellow NIFA National ProjectDirectors and National Program Leaders. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?One post doctoral fellow [Shehnaz Lokhandwala] was involved in the planning of in vitro and in vivo experiments, hands-on execution [generation of the prototype vaccines/immuizations/immune readouts/data procesing and analysis], and training graduate and undergraduate students. Thus, she received training that will enable her to become an independent investigator in the area of transboundary animal diseases and working in high biocontainment facility. One DVM-PhD trainee [Luka Popescu] was also involved in study execution during the immunization phase [pig immunizations/bleeding] and the ASFV challenge study [challenge dose inoculations/monitoring clinical outcomes/euthanization] in high biocontainment facility. Four PhD gradute students [Neha Sangewar, Vlad Petrovan, Annah Stoian, MathewOlcha] receivedhands-on training in this project [vaccine formulation/immunization/bleeding post-immunization/monitoring clinical-viremia outcomes post-challenge]. In addition, one DVM student [Catherine Elijah] received hands-on training in handling pigs, bleeding, monitor and evaluation post-immunizations, and treatment of minor issues under the guidance of the attending veterinarian. An undergraduate student [Lyndsey Ennen] also received hands-on training [animal handling/bleeding/processing blood to generate sera/isolate cells/run ELISAs]. These folkswill also be involved in the pending efficacy studies. How have the results been disseminated to communities of interest?Progress report was present at the annual Project Director's meeting in Chicago on December 01, 2017. Abstract accepted: American Association of Immunologists [AAI] annual meeting in Austin TX on May 2018. A poster will also be presented to report the data from the first eficacy study. What do you plan to do during the next reporting period to accomplish the goals?Execute Specific Aim 2: Evaluate protective efficacy of the second prototype ASFV experimental immunogen.

Impacts
What was accomplished under these goals? In objective one, we have conducted one efficacy study in which we evaluated the protective efficacy of one prototype adenovirus-vectored ASFV antigen cocktail formulated in two adjuvants.Notably, both formulations elicited strong ASFV-specific immune responses. Interestingly, animmunogen formulated using one adjuvant, but not the alternate, conferred protection in 5/9 (56% survival) upon oral challenge with the ASFV (Georgia 2007/1). Surprisingly, the alternate adjuvant elicited non-protective responses that significantly enhanced disease after challenge. On day 17 post-challenge when the study was terminated, the 5/9 survivors mentioned above were healthy and had low clinical scores. In comparison, 2/10 pigs from the group that received the alternate adjuvant were alive on day 17, but they had high clinical scores andhad to be euthanized for animal welfare reasons.

Publications

  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2018 Citation: Abstract and Poster: Efficacy of a Prototype Live-vectored Multi-antigen African Swine Fever Virus Vaccine. American Association of Immunologists (AAI) 2018 Annual Meeting, May 5, 2018.