Source: UNIV OF CALIFORNIA (VET-MED) submitted to NRP
UNRAVELING THE EFFECT OF CONTACT NETWORKS & SOCIO-ECONOMIC FACTORS IN THE EMERGENCE OF INFECTIOUS DISEASES AT THE WILD-DOMESTIC INTERFACE
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
COMPLETE
Funding Source
AFRI COMPETITIVE GRANT
Reporting Frequency
Annual
Accession No.
1017060
Grant No.
2019-67015-28981
Cumulative Award Amt.
$2,500,000.00
Proposal No.
2018-04788
Multistate No.
(N/A)
Project Start Date
Dec 1, 2018
Project End Date
Nov 30, 2024
Grant Year
2019
Program Code
[A1222]- Ecology and Evolution of Infectious Diseases
Recipient Organization
UNIV OF CALIFORNIA (VET-MED)
(N/A)
DAVIS,CA 95616
Performing Department
Vm: Medicine & Epidemiology
Non Technical Summary
The emergence of African Swine Fever (ASF) in Africa, Caucasus region and Eastern Europe in the last decade has increase the concerns about the global spread and potential impact in the swine industry, particularly if central EU-countries, USA or China become infected. The proposed interdisciplinary, multi-institution, research project aims to understand the evolution of ASF at the very source of the problem to better prevent its global spread. The Southern African Development Community is the unique area in the world where i) ASFV is believed to have originated/emerged and continues being an active source of long-distance ASFV spread (e.g., to Europe in 2007), ii) 23 out of the 24 ASFV genotypes are present, iii) both the domestic and sylvatic transmission cycles (involving domestic pigs, wild pigs and ticks) coexist.Our aims are to 1) assess the pig contact networks, pig management and socio-economic factors, tick involvement in ASFV transmission and viral diversity in the sylvatic and domestic cycles, 2) model ASFV transmission dynamics, economic impact and risk of introduction/spread into free areas in different eco-epidemiological settings using multi-scale simulation models, 3) integrate genomic-to population level data and modeling methods into a comprehensive, open-access, user-friendly, long-term, analytical platform referred to as ASF-BioPortal and develop interactive educational and training materials. All these will contribute to the vertical advancement in ASF research and will provide a modeling framework and generalizable principles useful for the study of the emergence, prevention and control of many other transboundary diseases at a local, continental and global scale.
Animal Health Component
60%
Research Effort Categories
Basic
10%
Applied
60%
Developmental
30%
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
3113599117030%
3114030108020%
3073599209030%
3113120107020%
Knowledge Area
311 - Animal Diseases; 307 - Animal Management Systems;

Subject Of Investigation
3120 - Spiders, mites, ticks, and other arthropods; 3599 - Swine, general/other; 4030 - Viruses;

Field Of Science
2090 - Statistics, econometrics, and biometrics; 1070 - Ecology; 1080 - Genetics; 1170 - Epidemiology;
Goals / Objectives
Our long-term goal is to better understand the transmission dynamics of emerging transboundary animal diseases (TADs) that hamper food security and limit the socio-economic development of human communities. Thisstudy will focus to improve the understanding of the complex pig contact networks that contribute to African Swine Fever virus (ASFV)transmission and evolution in theSouthern African Development Community (SADC) region and its impact on global spread. We have selected this region as this is a unique area in the world where i) ASFV is believed to have originated/emerged and continues being an active sources of long-distance ASFV spread (e.g., to Europe in 2007), ii) 23 out of the 24 difference ASFV genotypes are present, iii) both the domestic and sylvatic transmission cycles (involving domestic pigs, wild pigs and ticks) coexist.Our specific goals are to 1) assess the pig contact networks, pig management and socio-economic factors, tick involvement in ASFV transmission and viral diversity in the sylvatic and domestic cycles, 2) model ASFV transmission dynamics, economic impact and risk of introduction into free areas in different eco-epidemiological settings using multi-scale simulation models, 3) integrate genomic-to population level data and modeling methods into a comprehensive, open-access, user-friendly, long-term, analytical platform referred to as ASF-BioPortal and develop interactive educational and training materials.
Project Methods
Aim 1. Comprehensively assess the pig contact networks, pig management and socio-economic factors, tick involvement in ASFV transmission, ASF seroprevalence and viral diversity in the sylvatic and domestic cyclesFor this aim we will use a combination of field work (sampling and surveys), diagnostic methods, full genome sequencing and experimental infections to gather necesary data. Data will be then analyzed using value chain analysis, network analysis and exponential random graph models.Aim 2. Model ASFV transmission dynamics, economic impact and risk of introduction into free areas in different eco-epidemiological settings.We will use a hybrid spatial-explicit stochastic disease spread and economic model and risk assessmentto understand the transmission dynamics of ASF and evaluate the risk of ASF introduction/spread into new territories, respectively.Aim 3. Integrate data and modeling methods into a comprehensive, accessible, operational, user-friendly, long-term, analytical platform and develop interactive educational and training materials.We will develop a user-friendly dedicated site referred to as ASF-BioPortal as well as guidelines and training materials to facilitate data access, analysis and visualization by stakeholders, policy makers and the general public

Progress 12/01/18 to 11/30/24

Outputs
Target Audience:Our target audience is stakeholders related with the pig industry and wild pig management such as farmers, animal health officials, hunters, park rangers, extension specialists and other policy makers working in animal health Changes/Problems:There were few challenges/problems during this last year of the project: ·CIRAD -The PhD work from Cynthia Mapendere at the University of Pretoria experienced some delays but will be finalized before June 2025. ·IIAM -No technical challenges and problems arising during the reporting period. The administrative and financial constraints related to the SAM Registration Validation for work with UC-Davis was a significant problem that was not possible to be resolved despite trying for more than one year and despite the official translations from Portuguese to English languages submitted to FSD.com and SAM.Gov. ·UP -We experienced some problems with section 20 approvals.These were overcome by processing samples in an accredited laboratory in the Kruger National Park, but this did result in delays. ·OVI -Domestic pig numbers are low in some of the areas investigated. It seems that either a sylvatic cycle or domestic cycle exist. Direct contact at interface was not seen to date. ·UEM -None ·UCM -None ·UCDavis-The calibration of the models was delayed as we were waiting to incorporate the last data collected in the field. ASF spread modelisexpected to be finalized during 2025 by PhD student O. Pretorius. Despite those challenges the support provided by this project has been instrumental for building capacity for ASF and disease ecology research in a young cohort of scientists from South Africa, Zimbabwe, Madagascar and Mozambique.In addition to the professional development benefits, the results of the research conducted have advanced our understanding of virus evolution and the role of both warthogs andOrnithodorossoft ticks in the eco-epidemiology of the disease in southern Africa.It has also resulted in a strong network of ASF researchers in this sub-region and globally. We aim to continue our work with future focus more in the Americas. What opportunities for training and professional development has the project provided?The project has provided excellent opportunities for training the next generation of TADs and ASF researchers and strengthening diagnostic and epidemiology capacity. During the last trimester of 2024, two PhD students of the project successfully finalized their PhD and three others are expecting to graduate in 2025. Florian Taraveaufrom University of Montpellier defended his PhD thesis "Multipartite interactions (pathogens, symbionts, vectors) within vector pathosystems: the example of the tick Ornithodoros moubata and the African swine fever viru"s on 19th September 2024 at CIRAD, Montpellier. Rianja Rakotoarivonyfrom University of Antananarivo submitted his PhD manuscript at the University of Montpellier in November 2024 and defended his PhD thesis in December 2024. Cynthia Mapendere, registered at the University of Pretoria has made substantial progress and will defend her PhD thesis at the University of Pretoria during the first semester 2025. Elsa Lameirafrom Mozambique registered at the PhD program at the University of Pretoria, Department of Veterinary Tropical Diseases is expected to finalize her dissertation by the end of 2025. Her project title is ?The dynamics of Ornithodoros ticks and ASFV interactions at the wild-domestic suid interface of Macossa Hunting Reserve-Central Mozambique? Iolanda Vieira Anahory Monjanefrom Mozambique registered at the PhD Program at the University of Pretoria, Department of Animal Production Studies is expected to finalize her thesis in the second semester of 2025. Her project title is: ?Molecular characterization of ASFV in domestic pigs, wild suids and ticks from south region of Mozambique to improve husbandry systems, biosecurity measures and disease control? Antonia Andradefrom Mozambique is conducting her MSc Program at the Eduardo Mondlane University - Faculty of Veterinary Medicine in Mozambique and is expected to finalize in Fall 2025. Her project title is ?The ocurrence of ASFV at Macossa Game Reserve and its effect on pig production in Macossa District ? Stuart Neilon(MSc student at the University of Pretoria) attended the African Bioacoustics Community Conference in Cape Town (1-6 September, 2024). He will graduate the first semester of 2025. Charlene Clarke(postdoctoral fellow at the University of Pretoria) attended Oxford Nanopore Technologies workshop (11-13 September 2024) presented by Diplomics and AfricaBio Genome Initiative SA and a basic qPCR workshop (25-26 March 2024), presented by Diplomics in Pretoria, South Africa. Keaton Rea, MSc studentandCynthia Mapendere, PhD studentat the University of Pretoria, both received training on molecular techniques at a BLS3 facility, the Onderstepoort Veterinary institution and at the Hans Hoheisen Wildlife Research Station in the Kruger National Park, where they processed tick and warthog samples in compliance with the section 20 approvals granted for each project. Training was provided toMs T Rametseon ASF ELISA, DNA extraction both manual and automated, Real-time PCR. Two PhD students from the University Complutense of Madrid in Spain, have benefit from the training offered in this project and have successfully graduated:Carolina Muñoz-PérezandCecilia Aguilar-Vega. One post doc was working in research and mentoring activities in this project(Jaime Bosch) UCDavis: A South African student,Oonagh Pretorius, has been actively working in this project and it is expected to graduate from UCDavis at the end of 2025. Other two students were involved during the project duration and already graduated from UCDavis (Kathleen O'Hara, PhD; Narjara Veras, MPVM; Kyuyoung Lee, PhD). Several Post Doctoral researchers were also participating in this project and contributing to both research and mentoring activities within UCDavis and in collaboration with our international partners (Carlos Gonzalez Crespo, Jose Pablo Gomez, Jose Manuel Diaz Cao) How have the results been disseminated to communities of interest?All researchers and students have had to opportunity to present their results in different conferences during the project (see Products) as well as during the Annual Project Meetings. Our Final Project meeting was held in Dinokeng, South Africa in October 2024. We have generated dozens of publications in high impact journals (see Products) and several more are expected during 2025. We generated also several training courses and capacity building activities. All our training, outputs, data and results are made available and posted in our project website:https://www.asf-nifnaf.organd through the ASF-BioPortal platform:https://bioportal.ucdavis.edu The reports of different field trips were shared to the relevant stakeholders in Mozambique (District Agriculture and Game Reserve Authorities), Madagascar and South Africa (Veterinary Services). Additional presentations are planned for 2025 (including attendance and presentations in GARA in Italy as well as in stakeholder meetings with veterinary services and pig producers during 2025 in South Africa) We are actively searching for funding and opportunities to continue and expand our work and apply all our knowledge, risk assessment and modeling tools and "lessons learnt" with primary focus in the Americas. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? During this project we have collected huge amount of samples and data which have already generated groundbreaking results, trained new researchers in ASF eco-epidemiology and generated dozens of impact publications and will produce many more in the months to come. In this reporting period, although we continued with the field work, most of achievements were associated with the laboratory-based and modeling activities leading to the completion of the MSc and PhD projects. AFV transmission dynamics and viral diversity in the sylvatic and domestic cycle (aim 1): Two field missions were carried out in Macossa and Gorongosa Districts in Mozambique: The first one was in Mar 2024 to collect ticks from the warthog burrows and Granite boulders at Coutada-9 Game Reserve. ASFV positives were detected through diagnostic test and 14 ASFV positives samples were used for multi-locus analysis (p54). The second mission was in Oct 2024 to collect O. ticks is the warthog burrows and decks in the Gorongosa National Park and expand the retrospective survey of these vectors (the first done between 2006 and 2008). A total of 278 O. ticks were collected and DNA was extracted to complete the tick population genetic studies. To assess the genetic diversity of ASFV circulating in wild suids and ticks at the Limpopo National Park (LNP), 719 O. ticks and 228 wild pig blood samples were collected during the field trip to LNP in Nov 2023. The ticks were analyzed by PCR and the results were all negative.The blood samples were analyzed for anti-ASFV and anti-tick antibodies by ELISA. All the samples were negative for the presence of anti-ASFV antibodies and 68 out of 228 (~30%) samples were weak positive for the presence of anti-tick antibodies showing a medium probability of tick presence. Microbiome analysis from the ticks collected in Mozambique was performed by F. Taraveau and H. Jourdan team. They also studied the impact of microbiota on vector competence. A total of 199 domestic pig samples collected from outbreaks occurring from 2012 to 2021 in different Mozambican provinces, were sent to IAEA Seibersdorff Animal Production Laboratory, in Vienna Austria for sequencing. The phylogenetic analysis of the p72, revealed ASFV genotype II. Sequences were submitted to GenBank, a paper is in progress and will be part of I. Monjane dissertation. MSc student, A. Andrade analyzed the data produced in the cross-sectional study conducted between Nov and Dec 2023 to describe the smallholder pig-production system and the economic impact related with ASF in Macossa district. We conducted a validation of an Ornithodoros salivary antigen tick ELISA and tested pig samples from Mozambique and Zimbabwe. This test will facilitate the identification of pigs exposed to O. ticks. We conducted a pig smallholder survey analysis in South Africa: a total of 151 pigs were bled from 23 farms in 5 areas. Biosecurity in Ekurhuleni continues to be of a serious concern, with free roaming pigs, co-existing with cattle, sheep, goats, dogs, and chickens. Bloodmeal analysis in O. ticks from central Mozambique revealed that warthogs and tortoises are the dominant sources of blood meals. ASFV diversity in O. ticks from South Africa and Mozambique using a multi-gene approach revealed that despite a relatively restricted sampling area in Coutada 9, that virus diversity is high with two p72 genotypes and multiple variants of each being identified. Evaluation of the haemoparasite infection status of warthogs and O. ticks sampled from Kruger National Park (an ASF-positive region of South Africa) revealed high infection rates of both warthogs and ticks with an Anaplasma species that is specific to these two species, implicating the O. tick as the vector. The impact of Anaplasma infection status on ASF infection in O. ticks remains to be investigated. Genetic characterization of warthogs was performed to gain insight into historical dispersal patterns that underlie the substantially expanded geographical range of this host species within the last few decades. Similarly, ASF outbreaks are no longer restricted to the ASF control region in the northern parts of South Africa, and both the warthog, which acts as an amplification host for the ASFV, as well as O. ticks species occur throughout vast tracts of South Africa. These range expansions of warthogs and virus beyond their traditional ranges a very real threat that new sylvatic cycles may become established outside of the ASF control zone posing new threats for ASF spread locally and globally. Establishment of a carbon dioxide trap method for improved sampling of O. ticks from warthog habitats (natural and man-made) was developed by C. Mapendere (paper under revision). This practical advance is significant for ASF researchers investigating virus-tick dynamics, distribution and diversity. Full genome sequencing of 16 ASF viruses, representative of the two genotypes that have attained pandemic status, viz. genotype I and II. All viruses are from countries within the Southern African Development Community (SADC) region, an under-represented region of the world and provide valuable insights into the evolution of the virus and the likelihood of multiple genotype I incursions into previously ASF-free regions of South Africa, over a very narrow time period (less than a decade). A PCR assay that provides a better assessment of DNA integrity and Ornithodoros host diversity was developed by C. Clarke. This will substantially benefit future research projects. Genomic analysis of putative hybridized individuals between domestic pigs and two different species of Potamochoerus spp, the red river hog (P. porcus) and the bushpig (P. larvatus) suggested both species are genetically very distant and that putative hybrids are just atypical phenotypes of domestic pigs appearing in areas of sympatry. Modeling, risk analysis and decision-support tools (aim 2 and 3): We finalized the distribution maps for four wild suid species involved in the ASF epidemiology:Phacochoerus africanus,Potamochoerus larvatus,Potamochoerus porcus, andHylochoerus meinertzhageni. Spatial analyses confirmed that species occurrence data closely match with high quality of available habitats (QAH) areas, especially forP. larvatus, and correlate with many ASF cases. This information will be integrated in the ASF agent-based disease spread models. UCDavis PhD student, O. Pretorious trained 87 Animal Health Technicians and dip tank assistants from Mpumalanga Province, South Africa, to conduct 770 semi-structured electronic questionnaires and evaluate the knowledge, attitudes, and practices of smallholder pig farmers. The questionnaire was offered in eight official local languages. Descriptive and multivariate analyses are being used to evaluate relationships between management practices and the risk of introduction or spread of ASF as well as to define farm profiles based on their distinguishing characteristics. All these data will inform our ASF agent-based disease spread model to improve our understanding of ASF transmission dynamics and risk factors in Mpumalanga and provide decision-makers with tools to support more cost-effective ASF preventive and control strategieslocally and globally. Dr Pretorius presented these results at the ISVEE17 in Australia (Nov 24). She will continue with the calibration and validation of the spatial-explicit ASF disease spread model and the integration of all information and beta-testing of the ASF-BioPortal platform under the supervision of Dr. Martinez-Lopez. She will be communicating her results through conferences (e.g. GARA meeting in Italy, Apr 2025; CRWAD 2026, etc.), and stakeholder meetings and publications during 2025 and early 2026. She is expected to graduate by the end of 2025.

Publications

  • Type: Conference Papers and Presentations Status: Published Year Published: 2024 Citation: Rakotoarivony, R. Assessing interactions between domestic and wild pigs and implications for the transmission of African swine fever and other pathogens in rural habitats of Madagascar. 3rd joint conference of the AITVM-STVM, 21st-24th May 2024, Montpellier.
  • Type: Peer Reviewed Journal Articles Status: Published Year Published: 2024 Citation: Kabuuka, T., Mulindwa, H., Bastos, A.D.S., van Heerden, J., Heath, L. & Fasina, F.O. 2024. Retrospective multi-locus sequence analysis of African swine fever viruses by PACT confirms co-circulation of multiple outbreak strains in Uganda. Animals, 14(1), 71. https://doi.org/10.3390/ani14010071
  • Type: Peer Reviewed Journal Articles Status: Published Year Published: 2024 Citation: Goatley, L.C., Freimanis, G.L., Tennakoon, C., Bastos, A., Heath, L. & Netherton, C.L., 2024. African swine fever virus NAM P1/95 is a mixture of genotype I and genotype VIII viruses. Microbiology Resource Announcements, pp. e00067-24. https://doi.org/10.1128/mra.00067-24
  • Type: Peer Reviewed Journal Articles Status: Published Year Published: 2024 Citation: Spinard, E., Dinhobl, M., Erdelyan, C.N.G., O'Dwyer, J., Fenster, J., Birtley, H., Tesler, N., Calvelage, S., Leijon, M., Steinaa, L., ODonnell, V., Blome, S., Bastos, A.D., Ramirez-Medina, E., Lacasta, A., St�hl, K., Qiu, H.-J., Tennakoon, C., Nilubol, D., Masembe, C., Faburay, B., Ambagala, A., Williams, D.T., Ribeca, P., Borca, M.V. & Gladue, D.P. 2024. A standardized pipeline for assembly and annotation of the African swine fever virus genome. Viruses, 16(8), 1293 https://doi.org/10.3390/v16081293
  • Type: Other Journal Articles Status: Submitted Year Published: 2025 Citation: Mapendere, C., Bastos, A., Etter, E., Heath, L & Jori, F. A comparison of manual and carbon dioxide trap sampling of Ornithodoros soft ticks from warthog resting sites in South Africa. Parasites and Vectors (under revision)
  • Type: Conference Papers and Presentations Status: Published Year Published: 2024 Citation: Bastos, A.D. Closing the One Health circle to the benefit of conservation and communities (Invited Keynote Address), 2nd Joint WDA-AME/KVA Wildlife Branch Scientific Conference, Nairobi, Kenya (1-3 October 2024)
  • Type: Peer Reviewed Journal Articles Status: Published Year Published: 2023 Citation: Kabuuka J, Mulindwa H, Van Heerden J, Heath L, Bastos AD, Fasina F. Retrospective multi-locus typing of African swine fever viruses by PACT confirms co-circulation of multiple strains. Animals. 2023 Dec 24;14(1):71. doi: 10.3390/ani14010071.
  • Type: Peer Reviewed Journal Articles Status: Published Year Published: 2024 Citation: Penrith, M.-L.; van Emmenes, J.; Hakizimana, J.N.; Heath, L.; Kabuuka, T.; Misinzo, G.; Odoom, T.; Wade, A.; Zerbo, H.L.; Luka, P.D. African Swine Fever Diagnosis in Africa: Challenges and Opportunities. Pathogens 2024, 13, 296. https://doi.org/10.3390/pathogens13040296.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2024 Citation: Juanita van Emmenes, Puseletso Johnston, Refiloe Malesa & Keneiloe Montsu. Biobanking for animal health: storing natural treasures. SAAVT congress 2024, Oral presentation.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2024 Citation: Anthony F. Craig, Refiloe Malesa, Livio E. Heath, Jan E. Crafford, J�rgen A. Richt and Robert Swanepoel. Prevalence of African swine fever virus in selected warthog (Phacochoerus africanus) tissues in South Africa. 2nd Joint WDA-AME/KVA Wildlife Branch Scientific Conference. Oral Presentation.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2024 Citation: Juanita van Emmenes, Kerstin Malan, Refiloe Malesa, Keneiloe Montsu, Eduard Roos, Maruping Mangena and Anthony F. Craig. Importance of biobanking for animal health. 2nd Joint WDA-AME/KVA Wildlife Branch Scientific Conference. Oral Presentation.
  • Type: Theses/Dissertations Status: Published Year Published: 2024 Citation: Mthombeni, R.F. 2024. Molecular epidemiology of African swine fever genotype I and II viruses: insights from genome sequencing. PhD thesis, University of Pretoria.
  • Type: Other Journal Articles Status: Published Year Published: 2024 Citation: Special issue on African swine fever virus transmission and control: the role of wild and domestic suids in the MDPI journal Animals (Leade editor: Dr Juanita van Heerden (ARC) & co-editor: Armanda Bastos). https://www.mdpi.com/journal/animals/special_issues/9787K02RMA
  • Type: Conference Papers and Presentations Status: Awaiting Publication Year Published: 2025 Citation: Pretorius O., Gomez-Vazquez JP, Mart�nez-L�pez B. Enhancing African swine fever control strategies in South Africa: Contributions to disease introduction and spread by smallholder pig farmers. Global African Swine Fever Research Alliance (GARA) Scientific Meeting in Rome, Italy from 28-30 April, 2025.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2024 Citation: Pretorius O., Gomez-Vazquez JP, Mart�nez-L�pez B. New Modeling Approaches To Improve Surveillance And Control Of African Swine Fever In South Africa. 17th International symposium on veterinary epidemiology and economics (ISVEE17), Sydney, 11-15 November 2024.
  • Type: Peer Reviewed Journal Articles Status: Submitted Year Published: 2025 Citation: Florian Taraveau, David Bru, Carlos Quembo et al. Development of microsatellite markers for Afrotropical Ornithodoros ticks, 25 January 2024, PREPRINT (Version 1) available at Research Square [https://doi.org/10.21203/rs.3.rs-3891482/v1]
  • Type: Peer Reviewed Journal Articles Status: Published Year Published: 2024 Citation: Taraveau, F., Bru, D., Quembo, C.J. and Jourdan-Pineau H. Development of microsatellite markers for the soft tick Ornithodoros phacochoerus. Parasites and Vectors 17, 301 (2024). https://doi.org/10.1186/s13071-024-06382-7
  • Type: Peer Reviewed Journal Articles Status: Published Year Published: 2024 Citation: Rakotoarivony, R.; Kassi�, D.; Andriamahefa, A.; Andria-Mananjara, D.; Rakotoarinoro, M.; Ramaroson, H.S.; Raliniaina, M.; Rasamoelina, M; Gomez-Vazquez, P and Jori, P. Assessment of domestic pigbushpig (Potamochoerus larvatus) interactions through local knowledge in rural areas of Madagascar. Sci Rep 14, 16310 (2024). https://doi.org/10.1038/s41598-024-67208-1
  • Type: Other Journal Articles Status: Submitted Year Published: 2025 Citation: Mapendere, C.; Bastos, A. ; Etter, E. Heath, L and Jori, F. Evaluating the efficacy of carbon dioxide traps to investigate the sylvatic cycle of African Swine Fever in Sub-saharan Africa. Submitted to Parasites and Vectors.


Progress 12/01/22 to 11/30/23

Outputs
Target Audience:Our main target audience is stakeholders related with the pig industry and wild pig management such as farmers, animal health officials, hunters, park rangers, extension specialists and other policy makers working in animal health. Changes/Problems:No technical challenges and problems arising during the reporting period. The administrative and financial constraints related to the SAM Registration Validation for our partners at IIAM and UEM to be able to workwith UC-Davis still to be solved. The incident created as result of this problems is being resolved as the official translations from Portuguese to English languages were done and submitted to FSD.com and SAM.Gov. There have been continued delays with securing the necessary approvals to secure wild suid samples for the PhD project of Cynthia Mapendere. Cynthia has distributed sampling kits to professional bushpig hunters and has secured section 20 approval, but the final approval from the animal ethics committee of the University of Pretoria is still pending. A new collaboration has been established with the Faculty of Engineering to develop new approaches for monitoring the internal warthog burrow environment. This is a key component of tick distribution modelling and prediction of tick and virus movement. What opportunities for training and professional development has the project provided? Our strategy to train the next generation of researchers includes the involvement of a diverse pool of students and young researchers with different experiences and focus areas to work collaboratively in our consortium of partners. There have been several training and professional development opportunities provided by the project both in the field and laboratory work as well as the modeling and other quantitative methods. We have been able to meet and share knowledge among ASF experts, domestic pigs and wild pigs, as well as more knowledge about ticks involved in the ASF. It has been possible to discuss the methodology and the different distribution models of the most suitable species for the three study areas of the project. Knowledge, experience, and data on wildlife and environmental variables (necessary for the development of these models) have been shared. Students have been involved, and a webinar on different geographical information systems and statistical software with model techniques will be developed in the following year for the project participants. We also incorporate a new study about the patterns of activity of warthog using a GPS radiotracking collars. This helpful information obtained will be shared with colleagues in our project. These techniques will help us to understand the ecology and spatiotemporal patterns of the warthog activity in the year and seasons and obtain data to improve the modeling of this species. The specific students and young researchers involved and activities conducted are summarized below: Keaton Rea obtained experience in working under BSL3 laboratory conditions at the Onderstepoort Veterinary Institute in South Africa Stuart Neilon developed skills in AI processing of camera trap data. Elsa Lameira received on-site training at the laboratory in Chimoio Mozambique when Armanda Bastos spent a week there in March 2023. Dr Anthony Craig, new researcher at ARC, was trained on validation principles for an ELISA and practical testing of samples. Three PhD studens from Mozambique, one in the south and two other in central part of Mozambique presented their projects and advanced in their research. One MSc student was also involved in the project activities in central part of Mozambique. The students working in the project are: Antónia Andrade, MSc Student registered at the Universidade Eduardo Mondlane in Maputo-Mozambique. Her project title is: ?The ocurrence of ASFV at Macossa Game Reserve and its effect on pig production in Macossa District ?. Elsa Lameira and Iolanda Monjane, PhD students all registered at the University of Pretoria in South Africa. Elsa project title is: ?The dynamics of Ornithodoros ticks and ASFV interactions at the wild-domestic suid interface of Macossa Hunting Reserve-Central Mozambique?. Iolanda project title is: ?Molecular characterization of ASFV in domestic pigs, wild suids and ticks from south region of Mozambique to improve husbandry systems, biosecurity measures and disease control? One undergraduate student from computer science (Bi Nguyen), two PhD students (Kathleen O'Hara, Oonagh Pretorius) and two post docs (Jose Pablo Gomez and Carlos Gonzalez) from UCDavis were involved in the development of the risk assessment framework and the modeling activities. How have the results been disseminated to communities of interest? Reports of different field trips were shared to the relevant stakeholders (District Agriculture and Game Reserve Authorities). The PCR optimization of different gene regions and 16S rRNA PCRs for all soft tick collected in the field are still ongoing. It is planned to share the results of serology from upcoming domestic pig sampling and this will done in conjunction with some data interpretation and explanations. The result of field missions as well as the laboratory and modeling work has been shared during in the annual project meeting held in Dinokeng - Pretoria - RSA in 2022 and in Antananarivo-Madagascar in October 2023. Cynthia Mapendere and Livio Heath presented talks on ASF and Cynthia's project to stakeholders in the Marloth Park area in Mpumulanga Province in November 2023. Keaton Rea presented the results of his MSc study at the Annual General Meeting of the Department of Zoology and Entomology in December 2023. Armanda Bastos presented a talk at the GARA meeting in Uganda in February 2023. Cynthia Mapendere and Livio Heath travelled to Malelane, South Africa toreporton African swine fever virus and the work that has been conducted to date in the study area. Results of the modeling work was presented in the GeoVet 2023 international conference. What do you plan to do during the next reporting period to accomplish the goals? The serology results, DNA sequencing and the first data analysis as well as the first scientific articles to be derived from the completed works. Morphological characterization and sorting the recently collected soft ticks Refinement the student's protocols (with the respective co-supervisors from CIRAD, UEM and UP) and the two PhD candidates to present their protocols at the University of Pretoria (UP) in October and November 2022 Domestic pigs soft tick sampling using the CO2 traps in Macossa Gorongos and Massingir Districts, Sharing the already existing data to the other Nif Naf team involved in the modeling Finish the sampling of all domestic pigs in the study areas in South Africa, testing and evalution of those samples and share the data for modeling and risk assessment. Field trip to collect soft ticks (the proposed areas are Zingane and Decada da Vitoria, located in Massingir district close to the Limpopo National Reserve); Field trip for domestic pig sampling (the proposed areas are Sabie Game Park and Massintoto Reserve (in the greater transfrontier conservation area Maputo Province). To develop baiting sites to test the non-invasive methods in conservation areas in the South of Mozambique and in the Macossa District; Adapt to the Mozambican context the questionnaires that will be used in the Project; Manuscript preparation and submission; To attend the monthly meetings and other specific group meetings. Publication of a number of components, including: A comparison of manual versus CO2 tick trapping success The use of oral baits for non-invasive sampling of wild and domestic suids Genetic diversity of warthogs from South Africa Seasonal variation in tick infection rates in the Kruger National Park, South Africa Genome sequencing of an atypical genotype II virus from the SADC region Genome sequencing of genotype I viruses from Haemoparasite prevalence and diversity in warthogs andOrnithodorossoft ticks Drivers of warthog burrow activity patterns Haemoparasite prevalence and diversity in soft ticks from South Africa

Impacts
What was accomplished under these goals? Field work and laboratory analyses (aim 1): Two field missions were planned and carried out in Macossa and Gorongosa Districts-Central Mozambique for the project to better understand the transmission dynamics of emerging TADs: The first mission, occurred between 28thMay and 03thJune 2023 aimed to collect soft tick samples from the warthog burrows at Coutada 9-Macossa and the respective geo-referencing as well as the deployment of Data Loggers inside and outside of the wild suids burrows.Additional work was for oral fluids collection in domestic pigs aiming for ASFV PCR screening. There was also sampling of warthog burrows and bedrock: a total of 40 sites (mainly warthog burrow, bedrocks and under the trees) were checked for tick presence (in addition to the 98 already checked in the 4 previous field missions). Burrows were inspected manually and 311 specimens of soft ticks of different developmental stages were collected and counted. In the PNG a total of 750 soft ticks in the 15 of these natural habitats and under the tourist buildings also known as decks. Data loggers were used inside and outside of those sites (warthog burrows) for the temperature and humidity measurement, the data analysis still in progress. The second mission was from 11thto June 17th2023. This mission was for soft tick sampling, geo-referencing different locations and the Data Loggers in the warthog burrows and decks in the Gorongosa National Park (PNG). The laboratory component of this mission was to extract DNA of the sampled ticks (and shared in vials, one for IIAM, another one for CIRAD, and The University of Pretoria). All details of these activities are detailed bellow and the remaining component will be reported further. These two field activities were followed by the laboratory work: DNA extraction of ticks sampled in Coutada 9 and Gorongosa. Soft tick DNA were screed with 16S rRNA for Soft tick Phylogeny. Those testing positive to 16S rRNA were also tested using TK protocol. A second sampling activity in Limpopo National Park, (Gaza Province, south of Mozambique), was conducted in November 2023 to collect Ornithodorus ticks. PhD student Iolanda Monjane led the field mission carried out by researchers from UEM, CIRAD and IIAM. A total of 199 domestic pigs samples were collected from outbreaks (from 2012 to 2021), from several Mozambican provinces, were sent to IAEA Seibersdorff Animal Production Laboratory, in Vienna Austria for sequencing. The phylogenetic analysis of the p72, revealed ASFV genotype II. The results of this work will be included in a thesis chapter of Iolanda. Part of the 143 blood samples collected in the previous field trip to Limpopo National Park in April 2022, were sent to South Africa for the detection of specific ASFV and anti-tick antibody. Iolanda PhD project research was submitted to Animal Ethic for approval. MSc protocol defense by the student Antónia Andrade. Virus genome sequence data were generated for genotype II viruses from the SADC region in Africa. The results helped to refine the likely origin of the Georgia2007/1 strain that was introduced to the Caucasus in 2007 and which now poses a global threat to pig production. The most likely origin of the pandemic strain was shown to be a virus circulating in Mozambique in 2005, based on near identical genome sequences. The results were published in Pathogens: Stuart Neilon, registered for an MSc degree in 2023 and successfully presented and defended his project proposal in 2023 at UP Protocols to assess prevalence and diversity of three haemoparasite genera that may serve as proxies for exposure of domestic ticks to the Ornithodoros soft tick vector of ASF virus were established. A manuscript co-authored by Vincent Simbizi, Rebone Moerane, Juanita van Heerden and Bruce Gummow titled "The role of smallholder pig farmers in the biosecurity of pig diseases in the Eastern Cape Province of South Africa using African swine fever as a model", was submitted to the journal "Transboundary and Emerging Diseases". The article was accepted and is undergoing some in-house review before publication. Non-invasive sampling, including CO2 traps for ticks and ropes for warthog and bush pigs, was evaluated in the Dinokeng and Malelane area of South Africa. A manuscript on the effectiveness of sampling ticks with CO2 traps vs manual collection of ticks co-authored by Cynthia Mapendere, Livio Heath, Armanda Bastos and Ferran Jori, is in preparation for submission to Experimental and Applied Acarology. Extraction methods including automated, kit based, and Silica were compared. Currently, all ticks collected will be extracted using the Silica method. The ticks will be identified using 16S rRNA sequencing and p72 sequencing for genotyping of the viruses isolated from ticks. Full genome sequence analysis is being done and articles are in draft. Several farmer questionnaires were completed in Gauteng (21), South Africa. Farmer questionnaires were done in Nyanga (68) and Mt Darwin (74), Zimbabwe. Serum (215) was collected from domestic pigs in Mt Darwin but all tested negative on the ASF antibody ELISA. Modeling, risk analysis and platform development (aim 2 and 3): We continued working in the development, calibrationand validation of the risk assessment and disease spread models with the field data collected in all the participant countries. We have recruit a new PhD student to focus fully on this for her thesis. We have built a data lake within AWS to host the ASF-BioPortal and facilitate the ingestion, integration and sharing of ASF (and other diseases) related data. Users can upload their own propietary data. Different mapping, sequencing and other visualization tools areavailable at:https://bioportal.ucdavis.edu/. We have developed a new R-language package and its associated shiny dashboardto facilitate the development of rapid risk assessments for ASF (and other diseases). We are developing the documentation, associated paper and beta testing the dashboard to be launchedsoon. We have developed a Delphi expert opinion on wild African suidsto assess the distribution of the 4 most representative host species related to ASF (i.e. Phacocheorus africanus (warthog), Potamochoerus larvatus (bushpig), Potamochoerus porcus and Hylochoerus meinertzhangeni). The objective was tocalculate the value of land cover mostly associated with the presence of wild suidpopulations in Africa andobtain the distribution map of the Quality of available habitat of Wild Suids (QAHS). The species distribution models developed in Madagascar and African continent, are acrucial step to map the distribution of the ASF wild hosts and the potential contact areas between wild hosts and domestic animals. The questionnaires we are conducting with experts will collect very valuable information to characterize and quantify the distribution of these animals and can be applied to different epidemiological scenarios. We use several data sources to build the models and for the training and evaluation of the results. However, more data is needed to be able to more accurately evaluate the results obtained. Unfortunately, these species do not have extensive and good-quality databases. Therefore, during 2022, data has been obtained from different sources, institutions, and researchers (other projects, publications, local researchers, etc.). Expert opinion and land cover, GBIF - Presences/Occurrences (X, Y coordinates), IUCN - historical distributions of the animals (spatial data), Other African Projects: Snapshot Project South Africa (coordinates of the presence of wild African suids). In addition, we incorporated collected data from other studies and colleagues within our consortium during the year 2022. The phototrap coordinates (x, y) of the field studies for warthog and bushpig, samples of oral fluid, other samples in the field, baiting, other sources, studies, publications, etc.

Publications

  • Type: Peer Reviewed Journal Articles Status: Published Year Published: 2023 Citation: Jori, F.; Bastos, A.; Boinas, F.; Van Heerden, J.; Heath, L.; Jourdan-Pineau, H.; Martinez-Lopez, B.; Pereira de Oliveira, R.; Pollet, T.; Quembo, C.; et al. An Updated Review of Ornithodoros Ticks as Reservoirs of African Swine Fever in Sub-Saharan Africa and Madagascar. Pathogens 2023, 12, 469. https://doi.org/10.3390/pathogens12030469
  • Type: Peer Reviewed Journal Articles Status: Published Year Published: 2023 Citation: Mthombeni, R.F.; Bastos, A.D.; van Schalkwyk, A.; van Emmenes, J.; Heath, L. Phylogenomic Comparison of Seven African Swine Fever Genotype II Outbreak Viruses (19982019) Reveals the Likely African Origin of Georgia 2007/1. Pathogens 2023, 12, 1129. https://doi.org/10.3390/pathogens12091129
  • Type: Peer Reviewed Journal Articles Status: Published Year Published: 2023 Citation: Penrith, M.-L.; van Heerden, J.; Pfeiffer, D.U.; O?aevskis, E.; Depner, K.; Chenais, E. Innovative Research Offers New Hope for Managing African Swine Fever Better in Resource-Limited Smallholder Farming Settings: A Timely Update. Pathogens 2023, 12,355. https://doi.org/10.3390/pathogens12020355
  • Type: Conference Papers and Presentations Status: Published Year Published: 2023 Citation: Bosch, J., Aguilar-Vega, C., Ivorra, B., Goodman, S. M., Penrith, M. L., Jori, F., ... & Mart�nez-L�pez, B. (2023, September). P. 8.3 Eco-Epi cartographic tool for managing ASF in Africa: mapping African wild suids distribution based on the quality of available habitat. In GeoVet 2023 International Conference, Italy.


Progress 12/01/21 to 11/30/22

Outputs
Target Audience:Our main target audience is stakeholders related with the pig industry and wild pig management such as farmers, animal health officials, hunters, park rangers, extension specialists and other policy makers working in animal health. Changes/Problems: UCD - no changes or problems found during this reporting period. CIRAD- Our team in Madagascar suffered a car accident and one of our FOFIFA researchers (Fleurette, rest in peace) died in the accident. Other threehad several broken bones and went through multiple surgeriesand are stillrecovering from their injuries. FOFIFA hadto rearrenge the team and contract a new person to cover up the lab testing in Madagascar that was conducted by Fleurette. IIAM-No technical challenges and problems arising during the reporting period. The administrative and financial constraints related to the SAM Registration Validation for work with UC-Davis still to be solved as the systems is not running smoothly and need assistance on this. UP- There have been some delays with securing the necessary approvals and samples for the warthog and bushpig samples needed to conduct the genetic component included in the MSc and PhD studies, respectively of Keaton Rea and Cynthia Mapendere. Cynthia has distributed sampling kits to professional bushpig hunters. OVI- Domestic pig numbers are low in areas investigated. It seems that either a sylvatic cycle or domestic cycle exist. Direct contact at interface was not seen to date. UEM- None UCM- There is no change or problem that has arisen during this reporting period. What opportunities for training and professional development has the project provided?CIRAD: Participation of the two PhD students from the project to the International Wild Boar Symposium in Barcelona from 6th-9th September 2022 Ferran Jori, was a keynote speaker, opening this conference with the presentation: "African swine fever and wild pigs. A moving target." Rianja Rakotoarivony presented his PhD in the framework of the NIF-NAF project entitled "Health implications of interactions between bushpigs, Potamochoerus larvatus and domestic pigs in Madagascar". Cynthia Mapendere presented some of her previous work on African swine fever in South Africa and some of her ongoing activities within the NIF-NF project on the presentation entitled "Do wild suids from Ndumo Game Reserve, South Africa, play a role in the maintenance and transmission of African Swine Fever to domestic pigs?" IIAM: Three students' candidates from Mozambique have been identified as a strategy of contributing to the training of next generation of researchers. These students are: 1. Elsa Lameira- PhD program at the University of Pretoria - Department of Veterinary Tropical Diseases Project title:The dynamics of Ornithodoros ticks and ASFV interactions at the wild-domestic suid interface of Macossa Hunting Reserve-Central Mozambique Supervisory team: Prof Armanda D. S. Bastos (UP, Main supervisor),Dr. Hélène Jourdan (CIRAD, Co supervisor),Prof. Mary Louise Penrith (UP, Co supervisor), Dr. Carlos João Quembo (IIAM,Co supervisor) 2. Iolanda Vieira Anahory Monjane- PhD Program at the University of Pretoria - Department of Animal Production Studies Project title:Molecular characterization of ASFV in domestic pigs, wild suids and ticks from south region of Mozambique to improve husbandry systems, biosecurity measures and disease control Supervisory team: Prof. Geoffrey Fosgate (UP, Main supervisor), Dr Livio Edward Heath (ARC,Co supervisor),Dr. Carlos João Quembo (IIAM, Co supervisor),Prof. José Manuel Fafetine (UEM,Co supervisor). This candidate presented her proposal at the UP-DPAS on the 27thOctober 2022 3. Antonia Andrade*-MSc Program at the Eduardo Mondlane University - Faculty of Veterinary Medicine Project title:The ocurrence of ASFV at Macossa Game Reserve and its effect on pig production in Macossa District Supervisory team: Prof. José Manuel Fafetine (UEM,Main supervisor),Dr. Carlos João Quembo (IIAM,Co supervisor), Dr. Ferran Jori (CIRAD, Co supervisor). *Ms Antonia Andrade replaces the late Manuel Popinsky which unfortunately passed away in July 2021 UP: The project has provided photogrammetry and coding training opportunities to Nathan Mordaunt, a BSc(Hons) student as well as field and desktop training opportunities to a third-year BSc student, Micaela Venter. These students had the opportunity to go into the field with research assistant Dewald Kleynhans, where they assisted with downloading data from camera traps and replacing batteries as well as with the sorting of photographic images and videos. Cynthia Mapendere received training in ASF molecular detection methods at the Onderstepoort Veterinary Institute, BSL3 laboratory and is currently sorting ticks at this biocontainment facility. Two PhD candidates from Mozambique, Elsa Lameira and Iolanda Vieira, registered for their PhD degrees early in 2022, and Keaton Rea, registered for an MSc degree. Iolanda Viera, Cynthia Mapendere and Keaton Rea successfully presented and defended their project proposals in 2022. OVI: Dr. van Heerden and Ms. Mthombeni provided Laboratory training on DNA extraction, ASF p72 PCR and ELISA for Dr. Spargo, Ms. Ravaomanana and Ms. Mapendere from 24-26 October 2022. UEM: Last year, three PhD student were recruited to work in the Project in the south and central Mozambique. However, one dropped out due to personal issues. Currently two PhD and one MSc students are involved in the project activities. The students working in the project are: Antónia Andrade, MSc Student registered at the Universidade Eduardo Mondlane in Maputo-Mozambique Elsa Lameira and Iolanda Monjane, PhD students all registered at the University of Pretoria in South Africa. As previously mentioned, during the reporting period the students worked in their research proposals. UCM: There are several training and professional development opportunities provided by the project: we have been able to meet and share knowledge among ASF experts, domestic pigs and wild Suidae, as well as more knowledge about ticks involved in the ASF. It has been possible to discuss the methodology and the different distribution models of the most suitable species for the three study areas of the project. Knowledge, experience and data on wildlife and environmental variables (necessary for the development of these models) have been shared. Students have been involved, and a webinar on different geographical information systems and statistical software with model techniques will be developed in the following year for the project participants. UCDAVIS: - Two PhD students (Kathleen O'Hara, Oonangh Pretorius) and threepost docs (Carlos Gonzalez, Jose Pablo Gomez, Jose Manuel Diaz Cao) have beenworking in the risk assessment and modeling activities in collaboration with UCM, CIRAD and other partners. How have the results been disseminated to communities of interest?CIRAD: Results have been disseminated in the form of publications and participation to an international conference (International Wild Boar and Other Suids Symposium, Barcelona 4-6thSeptember 2022) Diaz Cao, et al., (2022). A maximum entropy species distribution model to estimate the distribution of bushpigs on Madagascar and its implications for African swine fever". Transboundary and Emerging Diseases (submitted) One paper published byRakotoarivony, R., Molia, S., Rakotomalala, E., Ramy-Ratiarison, R., Jori, F., Pedrono, M., 2022. Bushpig (Potamochoerus larvatus) Hunting in Rural Areas of Madagascar and Its Health and Socioeconomic Implications. Frontiers in Conservation Science 3:732626., doi: 10.3389/fcosc.2022.732626 One publication submitted Mapendere et al. 2022. "Husbandry practices and trade networks between subsistence pig farmers: A potential risk for African swine fever transmission in Northern KwaZulu Natal, South Africa". Transboundary and Emerging Diseases IIAM: No results suitable for dissemination so far outside of the project. However, the reports of different field trips were shared to the relevant stakeholders (District Agriculture and Game Reserve Authorities). The PCR optimization of different gene regions and 16S rRNA PCRs for all soft tick collected in the field are still ongoing. It is planned to share the results of serology from upcoming domestic pig sampling and this will done in conjunction with some data interpretation and explanations. All the field information collected will be used to parameterize the risk assessment and modeling activities. The result of field missions as well as the laboratory works as been shared during in the presence Nif Naf Meeting held in Dinokeng - Pretoria - RSA. UP Dewald Kleynhans, Livio Heath and Armanda Bastos presented an overview of the study to Dinokeng management and to field rangers in March 2022. In addition, Cynthia Mapendere presented her project to stakeholders in the Marloth Park area in Mpumulanga Province in July 2022. OVI Information has not yet been disseminated to communities UEM No result were produced during the reporting period that were suitable for dissemination.However, field work reports were shared with the authorities of the visited conservation areas. UCM 13th International wild boar symposium on wild boar and other suids 6-9 September 2022 - A review of the important role of wildlife in African Swine Fever.Jaime Bosch, Teresa Goicolea, Pablo Cisneros, María Cruz Mateo, Satoshi Ito, Cecilia Aguilar-Vega, Carolina Muñoz, Jose Manuel Sánchez-Vizcaíno.Oral presentation at Seva, Barcelona, Spain. The annual meeting celebrated in South Africa in October 2022. UCDAVIS We presented our risk assessment and modeling results in the Global African swine fever research alliance (GARA) Scientific Meeting (24-27 May 2022, Punta Cana, Dominican Republic), theISVEE, August 7-12 2022. Halifax, Canada,the 13th International wild boar symposium on wild boar and other suids 6-9 September 2022 in Spain and the annual meeting celebrated in South Africa in October 2022 Several publications were generated in collaboration with our other team members What do you plan to do during the next reporting period to accomplish the goals?CIRAD: Preparation of a review onOrnithodorosticks in Africa Ticks: Jori et al., (2022)Ornithodorosticks as reservoirs of African swine fever in Sub-Saharan Africa and Madagascar.An updated review. Pathogens (to be submitted in January 2022). Preparation of a manuscript entitledRakotoarivony et al. (2022) A combination of participatory methods to assess the occurrence and frequence of interactions between bushpig and domestic pig in Madagascar" to be submissted to Plos One In South Africa: Preparation of a publication reporting the results of the use of CO2 traps for Ornithodoros tick collection in warthog burrows Analysis of the ticks collected in warthog burrows. Local Organization of collections of blood and tissue samples from wild pigs in Southern Kruger NP Additional collection and analysis of oral fluid samples in buhspigs and warthogs, laboratory validation and publication of the results. Finalization and analysis of the questionnaires Participation to the GARA gap analysis in Uganda (February 2023) In Madagascar: Data Analysis of the interface characterization activities Analysis of the samples and information collected to characterize which will be synthesized in the form of a publication to be submitted in the first months of 2023 Analysis of the camera trap pictures to determine how bushpigs and domestic pigs share the same habitat and get quantifiable information on the nature, frequency and intensity of those interactions. Those results will be prepared to published and presented in international conferences. Serological analysis The collected samples from buhspigs and domestic pigs in two different areas will be tested with ELISA and molecular methods to monitor the potential sharing of ASF and CSF. Genetic analysis Samples collected from buhspigs, domestic pigs and hybrids will be analyzed with SNP method to detect potential levels of genetic introgression in suspected hybrid individuals. The results will be analyzed by Rianja Rakotarivony, with the help of Hélène Jourdan (CIRAD) IIAM: Culling some warthogs for research purposes (Diagnostic tests comparison) as part of a MSc student candidate work Possibility to develop baiting sites to test the non-invasive methods in the Macossa District and other two conservation areas in the South of Mozambique Morphological characterization and sorting the recently collected soft ticks Refinement the student's protocols (with the respective co-supervisiors from CIRAD, UEM and UP) and the two PhD candidates to present their protocols at the University of Pretoria (UP) in October and November 2022 Domestic pigs soft tick sampling using the CO2 traps in Macossa District Sharing the already existing data to the other Nif Naf team involved in the modeling UP: Publication of the use of oral baits for non-invasive sampling of wild and domestic suids and test of saliva for ASF antibodies: This component has been presented at two international conferences and a manuscript is in preparation for submission to an appropriate international journal. Sylvatic cycle risk assessment (ongoing): Using a combination of questionnaires, camera traps and genetics, an in-depth investigation into the role of wild suids, and bushpigs in particular, will be undertaken by PhD candidate Cynthia Mapendere. In addition, warthog burrows will be sampled forOrnithodorosticks and tested for ASF virus genome presence. Role of warthogs in disease transmission and resistance: Keaton Rea will undertake a genetic investigation into the role of warthogs in pathogen maintenance and transmission (2022-2023) using banked warthog samples from a broad range of sampling sites throughout South Africa. OVI: Sampling from all domestic pigs in study areas. Complete sampling to provide data for modeling, value chain analysis and risk assessment. Testing and evaluation of samples from the study areas. UEM: The activities planned for the next reporting period are mainly related to Aim 1"Collect information about pig contact networks, pig management and socio-economic factors, ticks, ASF seroprevalence and viral diversity in the sylvatic and domestic cycles", namely: Field trip to Sabie Game Park to collect soft ticks; To develop baiting sites to test the non-invasive methods in conservation areas in the South of Mozambique and in the Macossa District; Adapt to the Mozambican context the questionnaires that will be used in the Project; Manuscript preparation and submission; To attend the monthly meetings and other specific group meetings. UCM: All tasks have been started and are being carried out within the scheduled times. And the next steps will be: Regarding to the QAHS models for African wild suids will be to improve consensus among experts and to provide a map for future ASF eco-epidemiological and conservation studies for wildlife and domestic animals for all partners of the project. But also this models will using as a base to implement and develop more complex models of ecological niche and species distribution models in wild suids. In the annual meeting of the project celebrated the last October in South Africa, we discussed and we start to work all partners to share ecological data on tick and wild suid studies developed in the project to include and fed all distribution models for all African wild suids. This synergy will improve studies, not only in African wild suid distribution models or tick studies, but also help to better understand the transmission cycle of ASF in Africa. UCDAVIS Finalize the calibration and validation of the risk assessment and disease spread models with the field data collected in all the countries Finalize the wild suid and tick distribution models for global Africa (andthe participant countries at more fine scale resolution) Conduct a training course in rapid risk assessment and dashboard generation Launch the ASF Disease BioPortal new dashboards and user friendly tools

Impacts
What was accomplished under these goals? In South Africa, a PhD student started her first year of field work with very exciting preliminary results: ·Proof of concept of tick collection with CO2 traps in warthog burrows in the Kruger NP interface: results suggest that CO2 traps aremore efficient than the previously used manual collection method to collect ticks from warthog burrows, should dry ice be available. This method also shows very good prospect to be used to collect soft ticks in domestic pig premises. ·Baiting and use of ropes to collect saliva from wild pigs: this innovative system is working well to collect saliva from both, warthogs and bushpigs in 4 sites.Resultswillallow to describe a new and practical non-invasive method to monitor ASF exposure in wild African pigs. ·Assessing the interface (questionnaires and fence monitoring): The goal wasto evaluate wild pig movements, crop damage from wild pigs to understand and estimate presence and abundance of wild pigs at the interface of park and the potential of these territories to host ticks and ASF. There was a significant difference in the number of holes observed in areas bordering agricultural crops (7 holes/Km) vs areas bordering non-agricultural land (3.74 holes/Km). We also deployed non-invasive sampling methods that were established in the previous reporting period (2021) to new sites in South Africa. PhD student, Cynthia Mapendere, demonstrated that our prediction that warthogs that were habituated to human presence and structures would readily take the oral baits was proven when she deployed baits at a new site in Mpumulanga in 2022. Warthogs at this Mpumulanga site and at a new site in Dinokeng readily took the oral baits. Refinement of laboratory protocol for processing the oral baits for testing (spearheaded by Dr Livio Heath) confirmed that the collecting saliva from warthogs using oral baits is a viable means of confirming the ASF status of warthogs. In addition to successfully deployed baits and collecting saliva from wild suids in South Africa for the first time, methodologies for the trapping of ticks were also refined. Using a modified CO2trap and double-sided sticky tape, Cynthia and Ferran were able to demonstrate high levels of success with this method. Cynthia has established good contacts at the Mpumulanga sampling sites and has collected in excess of 1,000Ornithodorosticks. Other activities leaded by OVI were: ·Completed testing of eastern Cape serum samples and paper submission to Prev Vet Med. ·Non-invasive sampling, including CO2 traps for ticks and ropes for warthog and bush pigs, was evaluated in the Dinokeng area ·A field trip to Marloth area was undertaken from 13-23 Feb 2022 to sample pigs and collect ticks. ·Census of animals in the Marloth area and in Zimbabwe is almost completed. ·Dr. van Heerden and Ms. Rametse visited farmers in Gauteng North to investigate pig numbers. In Madagascar, Rianja Raokotoarivony continued his field work during the 2ndyear of his PhD with very promising results: Characterization on the interface betweenbushpigsand free-ranging domestic pigs in Menabé and Boeny. These two sites are prone to raise free ranging pigs in areas where bushpigs are also abundant. We used a combination of sociological (questionnaires, focus groups and participative mapping) and ecological approaches (camera traps). Most 14% of the people interviewed hasobserved interactions, which were mainly indirect (10%).Presence of crops, water sources and forest cover seems to facilitate inter-specific interaction. Collection and analyses of samples (filter papers & tissues) from bushpigs and domestic pigs in areas with different gradient of interactions: atotal of 85 hunters were trained to collect blood samples on filter papers and 120 bushpig samples and 73 domestic pig samples have already been collected and analyzed to identify antibodies or presence of ASF and CSF virus. So far, all the results were negative for bushpig and only 4 domestic pigs were positive to ASFV antibodies. Currently, a larger amount of domestic pig samples is being collected in the Menabé region to reach a sample of 120 individuals in contrasted areas of suspected presence and absence of interaction with bushpigs. Collection of genetic samples: Genetic samples were collected from bushpig x domestic pig hybrids (n=15), pure bushpigs (n=29) and local domestic pigs (n=15). Those samples will be analyzed genetically using Single Nucleotide Polymorphism to confirm the potential existence of these hybrids. These results would allow to confirm the occurrence of sexual interactions but also open new research questions about the presence of resistance genes against ASFV in domestic pig populations. In Mozambique,three field missions were organized in Macossa District to better understand the transmission dynamics of emerging TADs: The first mission (17-23Apr 2022) collected blood samples from domestic pigs in different locations of the district;The second mission (20-25 Jun2022) was for domestic pig blood sampling within human settlements at the outskirts of Coutada 9-Macossa; The third mission (23-29Oct 2022)involved Drs Jourdan and Jeanneau from CIRAD and recorded temperature and soil humidity in warthog burrows in Coutada 9, collectedsoft ticks in pigsties in villages around Coutada 9 with CO2 traps with dry ice;sampled ticks in warthog burrows in Gorongosafor population genetic analyses and extracted DNA of the sampled ticks. As a result919 soft ticks were collected in the 12 of these areas and one burrow had no soft ticks.These missions also resulted in sampling of ticks in 11 inspected pig pens in villages closed to Coutada 9-Macossa withCO2 traps with dry ice and manually. No ticks were found. The sampling was supplemented with the short interview to the pig farmers who's reported no occurrence of warthog near their settlements. The husbandry is mostly free-ranging pigs or pigs penned in the building made of wall mud, wood poles and covered by grass in the roof. The sampling resulted in 353 pigs and 7 warthog samples (5 sera and 2 tissue) for epidemiological studies (serology and the interviews through structured questionnaires) as part of Ms Andrade Msc project. In the lab 919 soft ticks form the burrows and bedrocks were sorted. Dead (152 ticks) were eliminated. Life stages of the remaining 767 ticks were recorded. 258 ticks were individually extracted with Qiagen DNeasy blood and tissue kit. This represents 4 sites with 30 ticks per site in Coutada 9 (2 were already sampled in 2020) and 3 sites with 42, 45 and 51 ticks in Gorongosa. About 352 sera collected from domestic pigs were negative, only one sera was positive. On the other hand, 3 of the 5 warthog sera tested ASFV positive.PCR in pools of soft ticks were also performed by Elsa (PhD student) and Mélanie (the CIRAD scientist) tested PCR (p72) on ticks pooled before extraction (3 pools from Coutada9 site 77) and after extraction (and dilution) (pools from Gorongosa). 4 pools from Gorongosa were positive for ASFV. This confirms the ability to detect ASFV with this protocol and the use of pooling (at least after extraction). Horizontal activities (all the countries involved) UCD has developed a rapid risk assessment and disease spread modeling framework for ASF that is now in the process of calibration, validation and sensitivity analysis using the field data collected by our field teams. The ASF Disease BioPortal will be launch in 2023 UCM and UCD teams worked in thespecies distribution models for wild suids (Phacocheorus africanus(warthog), Potamochoerus larvatus(bushpig), Potamochoerus porcus and Hylochoerus meinertzhangeni)in Madagascar and global Africa using Maximum enthropy and a expert-opinion based Quality of available habitat (QAHS) approach. These maps are valuable to design studies targeting ticks, borrows, baiting, epidemiological studies, for domestic pigs, etc.

Publications

  • Type: Journal Articles Status: Under Review Year Published: 2022 Citation: Cynthia Mapendere, Jan Helenus Ferguson, Ferran Jori, et al. Husbandry practices and trade networks between subsistence pig farmers: A potential risk for African swine fever transmission in Northern Kwa Zulu Natal, South Africa. Authorea. April 10, 2022. DOI: 10.22541/au.164961543.38493108/v1
  • Type: Journal Articles Status: Accepted Year Published: 2022 Citation: Jos� Manuel D�az-Cao, N�rjara Grossmann, Steven M. Goodman, Jaime Bosch, Helene Guis, Miatrana Rasamoelina, Rianja Rakotoarivony, Ferran Jori, Beatriz Mart�nez-L�pez, "A Maximum Entropy Species Distribution Model to Estimate the Distribution of Bushpigs on Madagascar and Its Implications for African Swine Fever", Transboundary and Emerging Diseases, vol. 2023, Article ID 7976252, 10 pages, 2023. https://doi.org/10.1155/2023/7976252
  • Type: Journal Articles Status: Published Year Published: 2022 Citation: Rakotoarivony, R., Molia, S., Rakotomalala, E., Ramy-Ratiarison, R., Jori, F., Pedrono, M., 2022. Bushpig (Potamochoerus larvatus) Hunting in Rural Areas of Madagascar and Its Health and Socioeconomic Implications. Frontiers in Conservation Science 3:732626., doi: 10.3389/fcosc.2022.732626
  • Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: Jori, F. African swine fever and wild suids. A moving target. Proceedings of the International Wild Boar Symposium in Barcelona from 6th-9th September 2022.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: Rakotoarivony, R.Health implications of interactions between bushpigs, Potamochoerus larvatus and domestic pigs in Madagascar. Proceedings of the International Wild Boar Symposium in Barcelona from 6th-9th September 2022
  • Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: Mapendere, C. Do wild suids from Ndumo Game Reserve, South Africa, play a role in the maintenance and transmission of African Swine Fever to domestic pigs? Proceedings of the International Wild Boar Symposium in Barcelona from 6th-9th September 2022.
  • Type: Book Chapters Status: Published Year Published: 2022 Citation: Bastos, A. & Boinas, F. 2022. Ornithodoros tick vectors and African swine fever virus (Chapter 58). In: Climate, Ticks and Disease, Edited by P. Nuttall. CAB International, pp. 405-412
  • Type: Journal Articles Status: Published Year Published: 2022 Citation: Penrith, M.L., van Heerden. J., Heath, L., Abworo, E.O. & Bastos, A.D.S. 2022. Review of the pig-adapted African swine fever viruses in and outside Africa. Pathogens, 11(10), 1190. https://doi/org/10.3390/pathogens11101190
  • Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: Kleynhans, D., Heath, L., Martinez-Lopez, B., & Bastos, A. 2022. Rope-based oral fluid sampling of warthogs (Phacochoerus africanus): lessons learned from South Africa. Global African swine fever research alliance (GARA) Scientific Meeting (24-27 May 2022), Punta Cana, Dominican Republic (poster presentation).
  • Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: Jacquier, M., Malherbe, G., Harrison, T.A., Heath, L., Bengis, R.G., Thomson, G.R., Maree, S., Retief, L. & Bastos, A.D. 2022. Molecular detection and characterisation of an African swine fever p72 genotype XXV virus in Ornithodoros ticks from the Kruger National Park, South Africa. Global African swine fever research alliance (GARA) Scientific Meeting (24-27 May 2022), Punta Cana, Dominican Republic (poster presentation).
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Kleynhans, D., Heath, L., Martinez-Lopez, B and Bastos, A. 2021. African swine fever surveillance: challenges and opportunities of non-invasive sampling of warthogs  Abstract was submitted and accepted for CRWAD (3-7 December 2021), Chicago, USA (Poster presentation).
  • Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: Mthombeni R, Bastos A, Van Schalkwyk A, Van Heerden J and Heath L. Phylogenomics of genotupe II African swine fever viruses from outbreaks in southern Africa (1993-2019), GARA Scientific Meeting 24-27 May 2022 (Poster presentation).
  • Type: Journal Articles Status: Published Year Published: 2022 Citation: Rakotoarivony R, Molia S, Rakotomalala E, Ramy-Ratiarison R, Jori F and Pedrono M (2022) Bushpig (Potamochoerus larvatus) Hunting in Rural Areas of Madagascar and Its Health and Socioeconomic Implications. Front. Conserv. Sci. 3:732626. doi: 10.3389/fcosc.2022.732626
  • Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: Gonz�lez-Crespo C, G�mez-Vazquez JP, Kathleen O'H, Patterson L, Mart�nez-L�pez B. Assessment of management strategies to control potential African Swine fever outbreaks in California using an Agent-based epidemiological model. Proceedings of the International Wild Boar Symposium in Barcelona from 6th-9th September 2022.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: Gonz�lez-Crespo C, Kyuyoung L, Patterson L, Belkhiria J, Pires AFA, Mart�nez-L�pez B. Suitability modelling of Ornithodoros spp. in California and overlapping with domestic and feral pig populations to identify ASFV high-risk areas under current and future climatic conditions. Global African swine fever research alliance (GARA) Scientific Meeting (24-27 May 2022), Punta Cana, Dominican Republic (poster presentation).
  • Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: Gonz�lez-Crespo C, Kyuyoung L, Patterson L, Belkhiria J, Pires AFA, Mart�nez-L�pez B. Suitability modelling of Ornithodoros spp. in California and overlapping with domestic and feral pig populations to identify ASFV high-risk areas under current and future climatic conditions. ISVEE, August 7-12 2022. Halifax, Canada.


Progress 12/01/20 to 11/30/21

Outputs
Target Audience:Farmers, private and official veterinarians, state parks managers, hunters, dealers, District Agriculture and Game ReserveAuthorities Changes/Problems:UCDAVIS- we continue with COVID19 restrictions which prevented UCDavis students to participate in field work incollaborationwithourprojectpartners.Therefore,studentshavebeenworkinginriskassessmentanddiseasespreadmodeldevelopment. Their field work part has been removed from their dissertation plan. Unfortunately, some of our partners gotCOVID(mostlyin Mozambique)andwelostManuelPopinsky,whounfortunatelypassedawayinJuly2021. CIRAD- COVID19 has delayed some of the field activities and there were delays to gather ASF testing reagents andcameratraps(flightswerecancelledanditwasnotpossibletoaccesssomeareasinMadagascar).Thoseactivitieswerere-scheduledlaterintheyearandweresuccesfullyconducted. IIAM-Notechnicalchallengesandproblemsarisingduringthereportingperiod.Theadministrativeandfinancialconstraintswere alreadysortedout. UP-RegistrationofthetwoPhDcandidatesfromMozambique(ElsaandIolanda)wasdelayeduntil2022forarangeoflogistical reasons. OVI-COVIDrestrictionsinSouthAfricamadetravelingwithinSouthAfricadifficultanddelayedthestartofsampling. UEM-No technical challenges, but COVID affected some of our team members (ManuelPopinsky unfortunately passed in July 2021) UCM-Thereisnochangeorproblemthathasarisenduringthisreportingperiod What opportunities for training and professional development has the project provided? UCDAVIS: A Master student (Narjara Veras) and two PhD students (Kathleen O'Hara and Pablo Gomez) at UC Davis havebeeninvolvedinthisproject.Apostdoctoralstudent(JoseManuelDiaz)wasalsoinvolvedinseveralactivitiesofthisproject.We developed and conducted a workshop on ASF network analysis and network based modeling in which 40 researchersandstudentsparticipated(seeotherproducts). CIRAD:TwoPhDstudentsthatareinvolvedintheMadagascaractivitieswillbeco-mentoredbyCIRADandFofifaresearchers:RianjaandHerilanto. IIAM:Fourstudents'candidatesfromMozambiquehavebeenidentifiedasastrategyofcontributingtothetrainingofnextgenerationofresearchers.Thesestudentsare: ElsaLameira-PhDprogramattheUniversityofPretoria-DepartmentofVeterinaryTropicalDiseases Projecttitle:ThedynamicsofOrnithodorosticksandASFVinteractionsatthewild-domesticsuidinterfaceofMacossaHuntingReserve-CentralMozambique Supervisoryteam: ProfArmandaD.S.Bastos(UP)-Mainsupervisor Dr.HélèneJourdan(CIRAD)-Cosupervisor Prof.MaryLouisePenrith(UP)-Cosupervisor Dr.CarlosJoãoQuembo(IIAM)-Cosupervisor ClaudioAfonsoGule-PhDProgramattheUniversityofPretoria-DepartmentofAnimalProductionStudies Projecttitle:AfricanSuineFeverinMacossaDistrict,CentralMozambique:Seroprevalenceandriskfactorstodomestic pigs Supervisoryteam: Prof.EricMarcelCharlesEtter(CIRADandUP)-Themainsupervisor Prof.GeoffreyFostgate(UP)-Cosupervisor Prof.BeatrizMartinezLopez-Cosupervisor DrCarlosJoãoQuembo(IIAM)-Cosupervisor IolandaVieiraAnahoryMonjane-PhDProgramattheUniversityofPretoria-DepartmentofAnimalProductionStudies Projecttitle:MolecularcharacterizationofASFVindomesticpigs,wildsuidsandticksfromsouthregionofMozambiqueto improvehusbandrysystems,biosecuritymeasuresanddiseasecontrol Supervisoryteam: Prof.GeoffreyFosgate(UP)-Mainsupervisor DrLivioEdwardHeath(ARC)-Cosupervisor Prof.JoséManuelFafetine(UEM)-Cosupervisor AntoniaAndrade*-MScProgramattheEduardoMondlaneUniversity-FacultyofVeterinaryMedicine Projecttitle:ThePenside-testevaluationforantibodydetectionofASFinwildlife Supervisoryteam: Prof.JoséManuelFafetine(UEM)-Mainsupervisor Dr.CarlosJoãoQuembo(IIAM)-Cosupervisor Dr.FerranJori(CIRAD)-Cosupervisor *MsAntoniaAndradereplacesthelateManuelPopinskywhichunfortunatelypassedawayinJuly2021 UP:The project has provided field and desktop training opportunities to eight of third-year BSc student interns. Thesestudents had the opportunity to go into the field with research assistant Dewald Kleynhans, where they assisted withdownloading data from camera traps and replacing batteries as well as with the sorting of photographic images and videos. Inaddition, two PhD students, Cynthia Mapendere and Thapelo Rametse, whose projects will focus of the sylvatic and domesticcycles of ASF, respectively, accompanied Dewald Kleynhans in the field and assisted with tick sampling and with camera trapmaintenance (data download, resetting and battery replacement). Cynthia Mapendere is also currently completing an onlinethird-year module, ZEN 362 Molecular Evolution and Phylogeny, which will be valuable for the genetic component of her PhDstudy. Finally, third-year intern, Stuart Neilon, gained extensive experience with preparing and deploying baits, processingcameratrap dataandiscurrentlyinvestigatingartificialintelligence(AI)approachesforimageclassificationusingopensourcesoftware. Onceestablished,thiswillpermitrapidbinningandsortingofcameratrapphotographs OVI:Duringthisperiodnotrainingandprofessionaldevelopmentwereobtainedbyteammembers.TheprojectdidinformaltrainingforDrVincentSimbizionuseoftheASFELISAandOmTSGP1ELISAforusetoevaluatepigserums. UEM:ThethreePhDstudentrecruitedtoworkintheProjectinthesouthandcentralMozambiquearecurrentlyinvolvedinthe projectactivities.OtherMScstudentwasrecruitedtoreplacethedeceasedstudentthatwasworkingintheproject. Currently,thestudentsinvolvedintheprojectare: AntóniaAndrade,MScStudentregisteredattheUniversidadeEduardoMondlane(UEM)inMaputo-Mozambique Cláudio Gule, Elsa Lameira and Yolanda Anahori, PhD students candidates all to be registered at the University ofPretoria(UP)inSouthAfrica. Aspreviouslymentioned,duringthereportingperiodthestudentsworkedontheirresearchproposals. UCM:Thereareseveraltrainingandprofessionaldevelopmentopportunitiesprovidedbytheproject:wehavebeenabletomeet and share knowledge among ASF experts, domestic pigs and wild suidae, as well as more knowledge about ticksinvolvedintheASF.Ithasbeenpossibletodiscussthemethodologyandthedifferentdistributionmodelsofthemostsuitablespecies for the three study areas of the project. Information have been shared for the elaboration of baits in the field for wildsuids. Knowledge, experience and data on wildlife and environmental variables (necessary for the development of thesemodels) have been shared. One student at UCM have been involved, and one post doctoral researcher (Jaime Bosh) and atraining course on network analysis and network based modeling given by the UCDavis team has benefited three students atUCM. How have the results been disseminated to communities of interest?UCDAVIS: We participated in several conferences and several publications are in preparation (to be submitted in 2022).ExperienceandknowledgegainedinthisprojectwasusedtobuildcollaborationsandsupportresearchactivitiesinothercountriesatriskofASFintroductionornewlyASFinfected(e.g.NorthMacedonia) CIRAD:Multiple training activities were conducted to train hunters about ASF and engage them to collect samples usingblottingpaper.ElevenPRAwereconductedinwhichwewereabletoincreaseawarenessaboutASFandprovideinformationabouthowtomitigateriskpracticestoreduceASFinfection IIAM:No results suitable for dissemination so far outside of the project. However, the reports of different field trips wereshared to the relevant stakeholders (District Agriculture and Game Reserve Authorities). The PCR optmisation of differentgeneregionsarestillongoing.Itisplannedtosharetheresultsofserologyfromupcomingdomesticpigsamplingandthiswilldoneinconjunctionwithsomedatainterpretationandexplanations. UP:Dewald Kleynhans presented an overview of the study to Dinokeng management and to field rangers. Livio Heath,Cynthia Mapendere and Thapelo Rametse undertook a site visit to the Marloth Park area in Mpumulanga Province andpresentedanoverviewoftheprojecttokeycollaboratorswhowillprovidefieldassistancewithboththedomesticandsylvaticcyclecomponentsofthetwoPhDstudies. OVI:Informationhasnotyetbeendisseminatedtocommunities.Itisplannedaspartofthesampling. UEM:Noresultswereproducedduringthereportingperiodthatweresuitablefordissemination.However,fieldworkreportsweresharedwiththeauthoritiesofthevisitedconservationareas. UCM:- Congress: CRWAD 2021 - December -3rdto 7th 2021 - Chicago IL: Estimation of the distribution of bushpigs inMadagascaranditsimplicationforAfricanswinefeverrisk.Althoughnextyeartheywillbepresentedseveralresultsatdifferentinternationalcongresses. What do you plan to do during the next reporting period to accomplish the goals? UCDAVIS: Testandvalidatetheriskassessmentanddiseasespreadmodelsusingtheavailabledataobtainedinthefieldworkfor SouthAfrica,MozambiqueandMadagascarincollaborationwithprojectpartners Conduct1or2trainingworkshopsinadvancedepitoolsincludingnetworkanalysis(followupoftheworkshopconducted thisyear),agent-basedmodelingandriskassessment OrganizeaASFsymposiumandin-personmeetinginSouthAfricaattheendof2022invitingprojectparticipantsand stakeholderstopresentresultofthestudyandfacilitatefurthercollaborationsforbetterASFpreventionandcontrol Continueexpandingandadaptingtheriskassessmentsandmodelingmethodsdevelopedinthisprojecttoothernon-AfricanASFendemicsettingsandpotentialcountriesatriskofASFintroduction(e.g.US) CIRAD: FinalizethebushpigriskmappingforMadagascar ExpandthesamplingandquestionnairestobetterunderstandthepigvaluechainandASFpresenceinMadagascarand identify locationswheredomesticpigshavemoreinteractionwithbushpigsusingSNA ContributetotheadaptationoftheriskassessmentanddiseasespreadmodelsforMadagascarincollaborationwith UCDavisandUCMteams IIAM: Cullingsomewarthogsforresearchpurposes(Diagnostictestscomparison)aspartofaMScstudent candidate Possibilitytodevelopbaitingsitestotestthenon-invasivemethodsintheMacossaDistrictandothertwoconservation areasintheSouthofMozambique Morphologicalcharacterizationandsortingtherecentlycollectedsoftticks Refinementthequestionnaires(withcolleaguesfromUC-DavisandUP)anddomesticpigsbleedingin MacossaDistrict UP: Non-invasivesamplingofwildanddomesticsuids:Ropebaitscollectedtodateandstoredat-20°Cwillbeprocessedin the BSL 3 laboratory at ARC-OVR and tested for ASF antibodies. The focus will shift away from natural sites to interface areas where transmission risk is highest. These areas will be identified following the pig census in and around the Dinokengarea which is scheduled for early 2022. In addition, domestic and wild suid sampling using the baiting approach will be undertakenatthesecondfieldsiteintheMarlothParkarea,MpumulangaProvince. Capacity building: Two PhD candidates from Mozambique, Elsa Lameira and Iolanda Vieira, will register for their PhD degreesearlyin2022andallthreePhDstudents(Elsa,IolandaandCynthia)willpresentanddefendtheirprojectproposalsinthefirsthalfof2022.ThapeloRametsewillalsoregisterforaPhDin2022. Sylvatic cycle risk assessment: Using a combination of questionnaires, camera traps and genetics, an in-depth investigationintotheroleofwildsuids,andbushpigsinparticular,willbeundertakenbyPhDcandidateCynthiaMapendere.In addition,warthogburrowswillbesampledforOrnithodorosticksandtestedforASFvirusgenomepresence. Role of warthogs in disease transmission and resistance: An MSc student will register in 2022 and will undertake a geneticinvestigationintotheroleofwarthogsinpathogenmaintenanceandtransmission(2022-2023)usingbankedwarthog samplesfromabroadrangeofsamplingsitesthroughoutSouthAfrica. OVI: Samplingfromalldomesticpigsinstudyareas. Testingandevaluationofsamplesfromthestudyareas. RegistrationofstudentsandobtainingapprovalforresearchproposalsfromUniversity. UEM: FieldtriptoSabieGameParktocollectsoftticks Todevelopbaitingsitestotestthenon-invasivemethodsinconservationareasintheSouthofMozambiqueandinthe MacossaDistrict AdapttotheMozambicancontextthequestionnairesthatwillbeusedintheProject ToregisterthePhDstudentsthatwillbeinvolvedintheProjectactivities Discusstheproposedtopicsandobjectivesofdifferentstudentcandidates Toattendthemonthlymeetingsandotherspecificgroupmeetings. UCM: Nowthatmoredataiscollected,wecanadvancewiththeparameterizationoftheriskassessmentanddiseasespread modelsforthedifferentstudyregions. Finalizethesuitabilitymappingforwildsuidsintheparticipantcountries ExplorethepossibilitytouseRadiotracckingofwildsuidae(eartag,pilotstudy)forwarthogsinSouthAfrica,possible expansiontoothercountries.

Impacts
What was accomplished under these goals? UCDAVIS: Adaptationofquestionnairestobedeployedinallstudyareasincollaborationwithprojectpartners ExpansionoftheASFagent-basedmodeltoincludethesylvaticcycle(wildsuids&ticks)andswillfeedingtransmission pathways.ModelisnowbeingadaptedtootherEuropeanandAmericancontext(i.e.NorthMacedoniaandUSA) Coordinationoftheworkinggroupsandmonthlymeetings Updateoftheprojectwebsite UpdateoftheASFDiseaseBioPortaldashboard(availableathttps://bioportal.ucdavis.edu) EstablishcollaborationsbeyondthepartnersofthisprojecttosupportrecentlyASFinfectedareasandbetterpreparefor potential incursion of ASF into the US (starting the adaptation of the risk assessment and disease spread models for US) CIRAD: IncollaborationwithFofifa&LNDV,weworkedinthebushpigMCDAforMadagascar(leadbyNarjara).Bushpig expertshelped to validate thesuitabilitymap Implementationofphonequestionnairestodescribetheinteractionbetweendomesticpigsandbushpigsindifferent zones and to identify areas with potential ASF circulation. We targeted vets (62 responses from 155 contacted) and park managers(72responsesfrom98contacted) Farmer,hunter,dealer,butcher,etc.questionnairestounderstandthepigvaluechainandhigh-riskpracticesadaptedto Madagascar Test46samplesfromdifferentregions:20sampleswerePCRpositive,confirmingthatASFwasresponsibleformultiple outbreaksinthecountry. PhDstudentRianjastartedfieldworkandworkon: Contacthunters(Menabeandother12huntingcommunities)andtrainthemtocollectsamplesusingblottingpaper. InvestigationandsamplinginArivonimamoonMar17-18(vetreportedhighpigmortality):seraandtissue collection:4/10werepositive Conduct12PRAexercisestounderstandhuntingstrategies,tradeandcarcassdisposal Organizevalidationtestforfilterpaper Testcameratrapslookingforbushpigs Investigationofplacesfrequentedbybushpigsanddomesticpigs:maizefields,naturalwatersources,etc. Surveyofmarketingchannelsforbushpigmeatandpigmovements(collectors,pigfarmer,butcher) Sample29bushpigs:alltestednegativetoPCR Boeny'smission(Oct18-27):nodirectinteractionbetweendomesticandbushpigsfound SNAandpigfarming:85interviews(10pigbreeders,49hunters,16farmers).Ongoinganalysis. IIAM: ThreefieldmissionswereorganizedinMacossaDistrict-CentralMozambique: 1stmission(29Mar-02Apr)tocollectbloodsamplesfromdomesticpigs. 2ndmission (31Aug-4 Sept) for manual ticks collection in the warthog burrows and the "resting yard" at the edge of the rocksandatthevicinityofhumansettlementsofCoutada9. 3rdmission (10-17 Oct) with partners from CIRAD to collect ticks in the natural habitats in that conservation area.These missions resulted in blood sampling (145 domestic pigs from 6 villages) and surveys of pig farmers in Macossa.Additionally, 6 sites of warthog burrows near human settlements of Coutada 9 were inspected and georeferenced (3 sites werepositiveforticks).Inthelastmission,40warthogburrowswereinspected,in25ofthesewecollected1053ticks.The ticksamplingwascomplementedbytestingofCO2(yeast)cameratraps Labworkandonjobtrainings? Countingandsortingallsampledburrows.Lifestageswererecordedforallindividuals(only10%wereadultticks). DNAextractionoftickssampledinCoutada9:atotalof585tickswereextractedwithQiagenDNeasybloodandtissuekit.Thisrepresents20siteswith30individualspersite.DNAextractsweredividedinto2 parts,oneforIIAMandoneforCIRAD. Establishment of tick colony in IIAM: training for tick rearing (identification of life stages, sexing, blood feeding) was providedtostudentsinthelab.BloodfeedingprotocolusedinCIRADwassharedwithIIAMandappropriatedeviceswere provided.Bloodfeedingwassuccessfullyperformedwithtickssampledoneyearago,onfreshpigblood. 120ticksfrom7sitesweresenttoCIRADtoestablishatickcolony. UP: We focused on non-invasive sampling methodologies for warthogs. First, our activities were limited to the Dinokeng GameReserve which is situated near UP. Informed by results of oral baiting of wild boars in Europe, we deployed a range of oralbaits withtheaimofcollectingsalivafromwarthogstobetestedtoASFantibodies.Resultsindicatedthatnon-targetspecies preferentially interacted with the baits and that warthog interaction was minimal. We subsequently expanded our baiting activitiestoincluderesidenceswithinandadjacenttoDinokeng,whicharefrequentedbywarthogs.Oncethenecessaryapprovals were secured from private landowners, several baits were deployed simultaneously at these "warthog-habituated" sitesandatthe"natural"siteswithinDinokeng.Allbaitingsitesweremonitoredusingcameratrapsandconsumedbaitswere collected and replaced daily. This allowed us to assess bait preference and to evaluate warthog behavior and bait interaction within the natural and residential (warthog-habituated) areas. Our results indicate that warthogs in natural areas appear toavoid baits, whereas warthogs at transformed sites are more likely to interact with the baits, particularly if non-target speciesshow an interest in the baits. These results indicate that non-invasive sampling of warthogs is likely ineffective in naturalareas, but effective in areas where warthogs have become habituated to human presence, confirming the utility of thisapproach for assessing the ASF-status of warthogs at interface areas where the risk of disease transmission between wildanddomestic suidsishighest. OVI: CompletetestingofEasternCapeserumsamplesandASFELISAresultsreported.Sampleswerescreenedon OmTSGP1ELISAandfinalreportisbeingprepared.Anarticleisplannedforpublicationofthesefindings Non-invasivesamplinginDinokengandCO2trapsfortickswereevaluated Studentsbusypreparingresearchproposalsandbudgetswillbeallocatedperstudentfortheplannedactivities SitevisittoMarlothtodiscusstheworkwiththestatevets.ThenumberofdomesticpigskeptintheKomatipoortand Mjejaneandthesuitabilityoftheseareastodeploynon-invasivesamplingmethodswereevaluated CensusofanimalsintheMarlotharea(SouthAfrica)andinZimbabweiscompleted.Thiswillbeusedforplanningof sampling EthicsapplicationsweresubmittedtoUPforstudiesinMarlothandDinokengareas UEM: Visit to Maputo Special Reserve in Nov 2021 to collect ticks. We deploy 10 camera traps in locations close to warthogburrowstoconfirmtheidealplacesfortickcollection.Weusedamanualtickcollectionmethodthatincludedtheuseofyeastovernight in the warthog burrows. In total, 14 warthog burrows located in 12 sites were examined, and none had ticks.Camera traps records were analyzed to confirm the presence of the wild suids and in two of the warthog burrows revealedtheir presence. Data describing the study area around the warthog burrows were collected. This mission was done withcolleaguesofIIAM Sequencingof199ASFVsamplesfrom9ofthe10Mozambicanprovinces.ThesequenceswillbeusedbyonePhD studenttoassessthegeneticdiversityoftheASFVindomesticpigsinMozambiqueasathesischapter Refinementoftheresearchtopicsandobjectivesforeachofthestudent'sprojectproposalandidentificationofthe studentsupervisors UCM: WorkinriskmappingactivitieswithourMalagasypartnersandUCDavis.Wehavecontributedtothecreationof environmentalvariablesforthedistributionofthebushpiginMadagascarandsharedthemodelcode. Shareandapplymethodologiestoimprovethespeciesdistributionmodels,specifically,toreducespatialautocorrelation andpseudo-replicationbetweenoccurrencerecordsofbushpigs Helpinthedesignofquestionnairesforallthestudyregions Shareprotocolsforcollectingandstoringsalivafromropesamples

Publications

  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Kleynhans, D., Heath, L., Martinez-Lopez, B and Bastos, A. 2021. African swine fever surveillance: challenges and opportunities of non-invasive sampling of warthogs  Abstract was submitted and accepted for CRWAD (3-7 December), Chicago (Poster presentation).
  • Type: Books Status: Published Year Published: 2021 Citation: Bastos, A. & Boinas, F. 2021. Ornithodoros tick vectors and African swine fever virus (Chapter 58). In: Climate, Ticks and Disease, Edited by P. Nuttall. CAB International, pp. 405-412.
  • Type: Journal Articles Status: Published Year Published: 2021 Citation: Mushagalusa, C.A., Etter, E., Penrith, M.L. 2021. Review of African swine fever outbreaks history in South Africa: From 1926 to 2018. Journal of Veterinary Research, 88(1). doi: 10.4102/ojvr.v88i1.1919
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: J. Diaz-Cao, H. Guis, N. Veras Grossman, M. Rasamoelina, D. Mananjara, H. Ramaroson, D. Kassie, J. Bosch, F. Jori, S. Goodman, B. Martinez-Lopez 2021. Estimation of the distribution of bushpigs in Madagascar and its implication for African swine fever risk. CRWAD 2021. Oral presentation.
  • Type: Journal Articles Status: Published Year Published: 2021 Citation: O'Hara KC, Beltran-Alcrudo D, Hovari M, Tabakovski B, Martinez-Lopez B. Descriptive and Multivariate Analysis of the Pig Sector in North Macedonia and Its Implications for African Swine Fever Transmission. Front Vet Sci. 2021 Nov 30;8:733157. doi: 10.3389/fvets.2021.733157.


Progress 12/01/19 to 11/30/20

Outputs
Target Audience:Pig farmers, official and private veterinarinas, hunters, state parks and game reservemanagers Changes/Problems: UCDAVIS - COVID19 did not allowed to send students from UC Davis to participate in field activities in collaboration with our African partners. Many conferences were cancelled or postponed to 2021 so we were not able to get those abstracts published (e.g. GARA 2020).We were able to continue working remotely in the risk assessmnet and model development activities, but some of the field activities in collaboration with our partners inSouth Africa, Mozambique and Madagascarwere significantly delayed. CIRAD -Covid-19 crisis blocked the countries: it was not possible to carry out field missions, to recruit students,and to enter the country from abroad between March and September 2020. It also greatly delayed theshipmentoflaboratoryconsumablesandreagents(listsoforderssentsinceNovember) IIAM-No technical challenges and problems arising during the reporting period. The only concern is the administrative matters related to the disbursementrate that isvery low which some how hampersthe acquisition of equipment, materials and reagents and at same time conduct the field work withthe sameallocated budget. UP -We were unable to initiate the study due to the hard lockdown that was imposed at the start of the pandemic and which was kept in place until the end of April. Despite moving to a lower lockdown level, inter-provincial travel was restricted making it impossible to initiate studies at the three sampling sites identified at the start of the project which all fall outside of the Gauteng Province. In addition, we could not enroll any students on the project from March - July due to the severity of the restrictions. As our academic year runs from January - December, and as registrations close in the first half of the year, we could not proceed with registering students once the lockdown level was eased sufficiently to allow for the 33% occupancy of campus facilities. To ensure project continuity, we therefore opted to appoint a part-time research assistant on the project, to work at a field site within the Gauteng Province. OVI -COVID restrictions in South Africa made traveling within South Africa difficult and delayed the start of sampling. UEM - None.In the reporting period,all the administrative procedure for the first disbursement of funds were carried out and the funds transferred in March 2020. Simultaneously, the procurement of reagents and consumables started. UCM-There is no change or problem that has arisen during this reporting period. What opportunities for training and professional development has the project provided? Thisprojectprovides outstandingprofessionaldevelopmentforseveralstudentswhowilluseresearchproject activitiestoobtaintheirMScorPhDdegrees. UCDAVIS: 1. Kathleen O'Hara: her PhD is conducted at the Gradaute Group of Epidemiolgyat UC Davis on the risk assessment and modeling of ASF in endemic settings. Director: Beatriz Martinez Lopez (UCDavis),co-directorEricEtter(CIRAD) 2. Jose Manuel Diaz Cao: Post Doctoral researcher at UC Davis that is involved in the risk mapping of ASF-susceptible populations (bushpigs, domestic pigs, ticks and warthogs) in Africa CIRAD: Rianja Rakotoarivony, Madagascar: His PhD is registered at the University of Antananarivo andtheUniversityofMontpellier.Title:Roleofthebushpig,PotamochoeruslarvatusinthedynamicsoftransmissionofAfricanswinefevertodomesticsuidaeDirector:FerranJori CynthiaMapendere,SouthAfrica.HerPhDwillberegisteredattheUniversityofPretoria.Director(AramandaBastos(UP),co-directorFerranJori(CIRAD),co-directorEricEtter(CIRAD) ManuelPopinsky,Mozambique.HisMScisentitled"EvaluationofpensidetestsforASFantibodydetection in samples of bushpigs ansd warthogs in Central Mozambique". Co-director Ferran Jori(CIRAD) Florian Taraveau: this student did his MSc internship with CIRAD in Montpellier working onO.moubatamicrobiome,hewillregisterforaPhDUniversityofMontpellier:Multipartyinteractions(pathogens, symbionts, vectors) within vector pathosystems: the example of the OrnithodorosmoubatatickandAfricanSwineFevervirus.Directors:ThomasPollet&HeleneJourdan(CIRADresearchers) Elsa Lameira Candidate for the PhD program at the University of Pretoria -Department ofVeterinary Tropical Diseases. Project title:The dynamics of Ornithodoros ticks and ASFVinteractions at the wild-domestic suid interface of Macossa Hunting Reserve-CentralMozambique.Supervisory team: Prof Armanda D. S. Bastos (UP) - Main supervisor; Dr.Hélène Jourdan (CIRAD) - Co supervisor; Prof. Mary Louise Penrith (UP) - Co supervisor;Dr.Carlos JoãoQuembo(IIAM) -Cosupervisor Claudio Afonso Gule. Candidate for the PhD Program at the University of Pretoria -Departmentof Animal Production Studies. Project title: African Suine Fever in Macossa District, CentralMozambique: Seroprevalence and risk factors to domestic pigs.Supervisory team: Prof. EricMarcel Charles Etter (CIRAD and UP) - The main supervisor; Prof. Geoffrey Fostgate (UP) - Cosupervisor; Prof. Beatriz Martinez Lopez- Co supervisor; Dr Carlos João Quembo (IIAM) - Cosupervisor TopicsforveterinaryorMScstudentswerealsodiscussedforanimplementationduringthe2021fieldwork. IIAM: Four students' candidates from Mozambique have been identified as a strategy of contributing to the trainingofnextgenerationofresearchers.Thesestudentsare: ManuelPopinsky,the MScStudent Candidateto beregistered at the Universidade EduardoMondlane(UEM)inMaputo-Mozambique ClaudioGule,Elsa LameiraandYolandaAnahori,all the PhD students candidates to beregistered UniversityofPretoria(UP)inPretoria-RSA UP: Limited field-based opportunities due to the COVID-19 regulations. Dewald Kleynhans, did however, receive training on sampling ticks using the traditional wall and floor scraping and sand sieving method. He also assisted with making Zoom recordings of the Global African swine fever research alliance (GARA) webinar available to attendees and via a Youtube link, thereby facilitating his own professional development and that of ASF researchers globally. OVI: During this period no training and professional development were obtained by Dr van Heerden (PI) and Dr Heath (co-worker). UEM: One MSc student and three PhD student were recruited to work in the Project in the south and central Mozambique. These students are: Manuel Popinsky, MSc Student candidate to be registered at the Universidade Eduardo Mondlane (UEM) in Maputo-Mozambique Claudio Gule, Elsa Lameira and Yolanda Anahori,PhD students candidates all to be registered at the University of Pretoria (UP) in South Africa. UCM: There are several training and professional development opportunities provided by the project: we have been able to meet and share knowledge among ASF experts, domestic pigs and wild Suidae, as well as more knowledge about ticks involved in the ASF. It has been possible to discuss the methodology and the different distribution models of the most suitable species for the three study areas of the project. Knowledge, experience and data on wildlife and environmental variables (necessary for the development of these models) have been shared. Two students have been involved and received training onnetwork analysis and modeling. Different lines have been shared for the elaboration of baits in the field for wild suids. etc. How have the results been disseminated to communities of interest?UCDAVIS: - We have participated in different conferences and share some of the project results and ASF expertise with collaborators,veterinary servicesand FAO collaborators outside the project in newly ASF infected areas (e.g. China, North Macedonia, South Korea)and here in the US (in informal working groups and meetings with CDFA, USDA-APHIS, Departmnet of Interior, etc.) CIRAD: Wild pig sampling Hunting community - In the project, contacts with the hunting community are required in order to have access to samples of hunted animals. We can distinguish two categories of hunters: Sport hunters practicing leisure hunting on weekends- They have been informed by telephone, by e-mail and in some cases by personal meetings. Subsistence hunters - They have been informed through focus group meetings with local communities during field missions in order to develop participative mapping of the study areas. In the different study sites, contacts with hunters have been established Madagascar: Contacts have been established with sport hunters in Madagascar and Mahajanga. In addition, a first field visits allowed to organize focus groups with 11 subsistence hunting communities in Menabé study site.Awareness on ASF risk was raised following the PRA with the traditional hunters and the interviews of the leisure hunters Mozambique: Contacts have been established with the owners of a private hunting area (Coutada 9) in Central Mozambique. Hunting activities are expected to start in July 2020. South Africa: Contacts with hunters are planned in some private hunting reserves but have not yet been established. Domestic pigs sampling Report to the Veterinary Services on the outbreak mission Website Shared working space on google drive Tick competence analysis Soft ticks life history traits: Oral presentation at the "Tiques et Maladies à Tiques" annual meeting. (http://tmt.sfecologie.org/) Tick microbiome/tick immunityand ASFV infection:PhD report of RemiPereira (http://www.theses.fr/2020MONTG013)and publications IIAM:Noresultssuitablefordisseminationsofaroutsideoftheproject.However,thereportsofdifferent fieldtripsweresharedtotherelevantstakeholders(DistrictAgricultureandGameReserveAuthorities).Itis planned toshare the results of serology from upcoming domestic pig sampling and this will done in conjunction with some data interpretation and explanations. UP:No. Our efforts in the final four months of the reporting period were focused on securing the necessary approvals, designing baiting and tick collection protocols, training of the newly appointed research assistant and identifying active warthog burrows. OVI:None to date. UEM:No relevant results were produced during the reporting period that were appropriate for dissemination. Nevertheless, the field work reports were shared with the authorities of the visited conservation areas. UCM:At the moment, they have not been able to spread any results. Although nextyear they will be presented several results at different international congresses. What do you plan to do during the next reporting period to accomplish the goals?UCDAVIS: 1. Continue with the development of the risk assessment and disease spread models to get them ready for implementation and validation in the participating countries when we finish the field data collection. 2. Support the survey implementation using Qualtrics 3. Conduct workshops and training activities in network analysis and modeling for researchers, students and collaborators in Africa 4. Continue with the coordination of the working groups and montly meetings to achieve project goals. CIRAD: Wild pig sampling: Exploring the wild-domestic animal interface: This will be done following a two-stage process: Implementation of questionnaires among pig farmers to identify hot spots of interaction. Deployment of camera traps in hot spots of interaction in order and quantify to identify the frequency and occurrence of potential direct and indirect interactions. Madagascar-Ménabé and Marovoay South Africa- Marloth Park Tick competence analysis Importance of soft tick microbiome on life history traits (Master's traineeship): impact of antibiotic treatment on life historytraits. Characterizationoflifehistorytraits:followup. DesignmicrosatelliteprimersonOrnithodorosmoubatatostudytickspatialdynamicsusingpopulationgenetics. IIAM: Cullingsomewarthogsforresearchpurposes(Diagnostictestscomparison)aspartofaMScstudent candidate Possibilitytodevelopbaitingsitestotestthenon-invasivemethodsintheMacossaDistrictandothertwoconservationareasintheSouthofMozambique Discusstheproposedtopicsandobjectivesofdifferentstudentcandidates Discusswith CIRAD-France the MTA (to transfersoftticksfrom Mozambiqueto Montpellierand back)andpossible establishment of aSoft tick colony in Mozambique (either at IIAMorUEMfacilities) Morphologicalcharacterizationandsortingtherecentlycollectedsoftticks Refinementthequestionnaires(withcolleaguesfromUC-DavisandUP)anddomesticpigsbleedinginMacossaDistrict UP: Optimise bait deployment to ensure successful sampling of warthog saliva, finalise the selection of the three study sites, enroll students in postgraduate degree programmes, apply for ethical clearance to conduct the research, secure section 20 approval, draft the questionnaires that will be used in the study and approach game park managers to secure approval to conduct the studies in the selected sites. OVI: Complete testing of Eastern Cape serum samples and report results. Non-invasive sampling in the Dinokeng area of South Africa was delayed because of recent rains, which made tick collection impossible. We plan to collect ticks when weather improves, and to start non-invasive sampling of warthog and setting up CO2traps for ticks. Students will prepare research proposals and budgets will be allocated per student for the planned activities. A site visit to Marloth is planned, to discuss the work plan with the state veterinarians. Census of animals in the Marloth area (South Africa) and in Zimbabwe is planned to identify where animals are within these areas, which will help with planning for sampling Ethics applications were submitted to University of Pretoria for studies in Marloth and Dinokeng areas. UEM:The activities planned for the next reporting period are mainly related to Aim 1"Collect information about pig contact networks, pig management and socio-economic factors, ticks, ASF seroprevalence and viral diversity in the sylvatic and domestic cycles" which are: Field trip to Maputo Special Reserve to collect soft ticks using CO2trap protocol; Field trip to Sabie Game Park to collect soft ticks; To develop baiting sites to test the non-invasive methods in conservation areas in the South of Mozambique and in the Macossa District; Adapt to the Mozambican context the questionnaires that will be used in the project; Discuss the proposed topics and objectives of different student candidates; To register at different universities the post-graduate students that will be involved in the project activities; To attend the monthly meetings and other specific group meetings. UCM 1. Continue supporting the risk mapping activities in the study countries 2. Provide technical support for the oral fluid and camera trap activities 3. Help in the development and implementation of the risk assessment and disease spread models for the different study areas

Impacts
What was accomplished under these goals? Main accomplishments byeach participant institution: UCDAVIS: 1. Development of the surveys to conduct the field work using Qualtricsin collaboraiton with CIRAD, FOFIFA, UEM, IIAM and UP 2. Development of the foundations for the ASF spatial-explicitagent based model for Africa. Data, parameters and validationof the modelwill be based on the field work collected by project partners. 3. Coordination of working groups andmonthly meetings 4. Mantainance/updateof the project website. 5. Development of the ASF Disease BioPortal Dashboard(to be launchedin thehttps://bioportal.ucdavis.edu next year) 6. Expansion of collaborations in ASF beyond the parthers of this project to support recently ASF infected areas andwork in Eastern Europe (i.e. North Macedonia) and Asia (i.e. China). One paper published (see products), others pending submission. CIRAD: Attended with Malagasy partners (Fofifa, LNDV)the remote kick-off meeting in Jun2020 and all monthly meetings Wild pig sampling: Identification of sampling areas:Coutada 9,Maputo Elephant Reserve, Ménabé,Morovoay,Dinokeng andMarloth Park Authorization requests submitted Material purchase(diagnostic kits andcamera traps) Protocol preparation for testing non-invasive saliva collection in warthogs Bushpig, pig and ASFV presence phone questionnaires (Madagascar):Two questionnaires were designed, one for National Parks staffand the second onefor vets. 64 responses were collected MCDA bushpig distribution map completed Identification of hunters for further field work:In Sep and Oct 2020, on the way to Menabe region, two leisure bushpig hunters were interviewed to obtain information on their hunting practices and on the existence of hunting camps, the community of leisure hunters to engaged them inbushpig sample collections. Bushpig blood samples on filter paper were obtained from one of the hunters 4. Domestic pigs sampling: Field mission in Ambanja and Ambilobe outbreak area (Madagascar):After themission of Nov2019 where ASFV circulation wasclinically observed, a 8-day field mission (17-24th December 2019) was carried outin Ambilobe and Ambaja districts to collect samples in infected pig farmsto confirm that the pathogen responsible of these outbreaks was ASFV. Overall, 56 samples from 12 sick and 11 dead animals were collected from 5 sites in Ambilobe district and 1 site in Ambanja district. These samples are waiting for lab reagents for PCR to be shipped Farmer, butcher and collector, abattoir, rural subsistence hunter questionnaires were developed and adapted to the Malagasy context Field mission to Menabe (Oct2020) as a suitable study site, to test questionnaires and to investigate an outbreak and collect samples. Three localities were visited.A total of 35 pig samples were collected: 30 from pigs slaughtered at Ambalanomby slaughterhouseand 5 from surviving pigs.SNAquestionnairesweretestedbyinterviewingpigfarmers,collectorsandbutchers Oneparticipatoryruralappraisal (PRA)wascarriedoutamongtraditionalhunterstocompletethosedoneinNov2019intheNorthofMadagascarandduringthemissioninSouthWesterninOct2020. The mission in Oct2020 confirmed that these regions weregood field sites toinvestigate as ASF circulation. The network analysisshowed that pigs raised inBefasyandManjawerecollected bycollectorsandbutchersinAmbalanomby.Smallerpigs(<60kg)wereslaughteredforlocalconsumptionwhereasbigger onesweresoldinAntananarivo.BefasyandManjawereareaswithadomesticpig-bushpiginterface. 5. Tick competence analysis: Characterization of soft ticks (Ornithodoros moubata) life history traits: reproduction and larval development. We used a quantitative genetics design to allow estimating heritabilities required to evaluate the evolutionary potential of those traits Interaction between tick microbiome and ASFV infection. Antibiotic treatments modified the microbiome. Potential tick immunity pathway was also investigated based on expression of candidate genes. Acquisition of genomic data for O. moubata thanks to collaboration with FLI (Jan Forth) IIAM: Two visits were organized inpotential study sitestounderstand the transmission dynamics of emerging TADs: One visit was done jointly on Mar 20th 2020 with UEMin the Especial Reserve of Maputo The other visitwas done by the IIAM team on May 11th 2020 in a potential study site in the Macossa District Followingthat field trip, the first soft tick sampling was conducted at Coutada 9in order to provide the first evidence of the existence of a sylvatic cyclein that area of the country which could be a permanent source of virus threatening the domestic pig value chain in Central Mozambique. In this regard, 40 sites throughout of Coutada 9 were inspected and georeferenced. Thirty eight sites were positive for the presence of soft ticks and 2 negative sites TheCo-PIfromIIAMwasinvolved,inNov2020,inthefirstsamplingactivitydoneinSouthMozambique (seeUEM's report) UP: Due to COVID-19 travel restrictions we identified an alternative study site north or Pretoria to trial camera traps and baits for non-invasive sampling of warthogs. Dinokeng was selected as the study site as previous surveys have confirmed the presence of ASF-infectedOrnithodorosticks. Most of the fieldwork was undertaken by a technical assistant instead of postgraduate students due to the hard lockdown enforced in South Africa in Mar2020 which made fieldwork and access to labs impossible until Jul2020, by which time postgraduate student registrations had closed. With the assistance of Dewald Kleynhans,we secured the approval from park management to undertake the study, we submitted the necessary applications for approval, researched approaches to rope baits (this is unchartered territory as it has not been attempted with warthogs), designed a range of carbon dioxide traps to test in active warthog burrows and conducted a field scouting trip to identify warthog burrows at which we could deploy camera traps, and paths to water which would be "high-traffic" areas suitable for deploying baits. OVI: Section 20 approval received for sampling and testing for ASF in Eastern Cape and Marloth areas as well as non-invasive sampling in sylvatic cycle. ARC ethical approval received for Marloth area. UP ethical approval obtained for Eastern Cape area. Planningfor serology of 1000 samples from domestic pigs in Eastern Cape Planning of sampling in Zimbabwe Planning tick and non-invasive sampling in Dinokeng Kick-off meeting 17-19 Jun2020 Approval received from state vet services for sample collection in the Eastern Cape. Domestic pig samples (660) were collected and tested, and 3/660 (0.45%) positives found. All positive samples were from the reported ASF outbreak areas in the province. Attendance to all project meetings during 2020. UEM: Data describing the study area (hydrography, geomorphology, proximity with human buildings and type of habitat) around the warthog burrows and bush pig nests were collected Joint visit to Maputo Special Reservewith IIAM partners on 20thMar2020, to present the project tolocal vets and conservation areas authorities. After authorization by these authorities a sampling activity to Maputo Special Reserve was organized in Nov2020to collect the tick vectors of ASFV. The manual soft ticks collection method in the warthog burrows, around lagoons and along the Futi River was used. Thirty onewarthog burrows and bush pig nests located in 17 sitesthroughout the Maputo Special Reserve was examined for the presence of soft ticks, and none was positive UCM: Contribution to develop species distribution modelsasthebasic step to detect potential contact areas between wildlife and domestic animals. Help designing the questionnaires that will collect very valuable information to characterize and quantify the different epidemiological scenarios existing in the three study areas. Supported the pilot study on bait in the field, which may help to collect non-invasively the health status of wild swine in the field.

Publications

  • Type: Conference Papers and Presentations Status: Awaiting Publication Year Published: 2020 Citation: Randriamparany T., Rasamoelina M., Ravaomanana F., Guis H. Threat of food security and Outbreak of African Swine Fever Virus in Ambilobe district, north of Madagascar. Abstract submitted to GARA.
  • Type: Conference Papers and Presentations Status: Awaiting Publication Year Published: 2020 Citation: Etter et al. 2. Nif-Naf project: from complex pig contact networks that contribute to ASF transmission to dynamics of emerging TADs. Abstract submitted to GARA.
  • Type: Conference Papers and Presentations Status: Awaiting Publication Year Published: 2020 Citation: Veras Grossmann N., Rasamoelina M., Ravaomanana F., Guis H., Mart�nez-L�pez B. Participatory Rural appraisals with hunting communities in Madagascar on African Swine Fever awareness and bushpig hunting and trade practices. Abstract submitted to GARA
  • Type: Journal Articles Status: Submitted Year Published: 2020 Citation: Rakotoarivony, R. Molia, S., Rakotomalala, E., Jori, F. and Perdrono, M. 2020. Health and socioeconomic aspects implications of bushpig (Potamochoerus larvatus) hunting in rural areas of Madagascar. EcoHealth (in preparation)
  • Type: Journal Articles Status: Published Year Published: 2020 Citation: Pereira de Oliveira R, Hutet E, Paboeuf F, Duhayon M, Paboeuf F, Le Potier M, Vial L. 2020. No Experimental Evidence of Co-Feeding Transmission of African Swine Fever Virus between Ornithodoros Soft Ticks. Pathogens, 9(3): 168. doi: 10.3390/pathogens9030168
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Mart�nez-L�pez B, O'Hara KC, Caron A, Bastos A, Quembo C, Kassie D, Mananjara DE, Jori F, Ravaomanana F, Fosgate G, Guis H, Jourdan H, Ramaroson H, Fafetine J, S�nchez Vizca�no JM, van Heerden J, Vial L, Heath L, Penrith ML, Rasamoelina M, Rakotoarinoro M, Raliniaina M, de Oliveira RP, Molia S, Randriamparany T, Pollet T, Rakotoharinome VM, Porphyre V, Bosch J, D�az JM, Barasona JN, Etter E. Tracing African swine fever: viral evolution and disease transmission in the Southern African Development Community. Conference Presentation #134.
  • Type: Journal Articles Status: Published Year Published: 2020 Citation: O'Hara K, Zhang R, Jung YS, Zhou X, Qian Y, Mart�nez-L�pez B. Network Analysis of Swine Shipments in China: The First Step to Inform Disease Surveillance and Risk Mitigation Strategies. Front Vet Sci. 2020 Apr 28;7:189. doi: 10.3389/fvets.2020.00189. PMID: 32411733; PMCID: PMC7198701.


Progress 12/01/18 to 11/30/19

Outputs
Target Audience:Our main target audience is stakeholders related with the pig industry and wild pig management such as farmers, animal health officials, hunters, park rangers, extension specialists and other policy makers working in animal health. Changes/Problems:The main problem in this reporting period was the delay in the release of funds. We were not able to receive the project funds during 2019 from USDA-NIFA, which basically has delayed the initiation of the field work activities one year. What opportunities for training and professional development has the project provided?We initiated the recruitment of Master, PhD and Post Doctoral students that will be involved in this project. How have the results been disseminated to communities of interest?Methods and results proposed to be conducted under the project activitiesweredisseminated through our websitehttps://www.asf-nifnaf.org, presentation in conferences (e.g., CRWAD 2018; GeoVet 2019; CRWAD 2019) as well as through other partnerships and databases (e.g. International Research Consortium on Animal Health databasehttps://roadmaps-public.star-idaz.net/#/mAOhYand through partnership with the Global Alliance for African Swine Fever (GARA)https://www.ars.usda.gov/gara/. What do you plan to do during the next reporting period to accomplish the goals?We will continuethe data collection through the field work (sampling and surveys)andcontinue with the advancement of the risk assessment and modeling activities.

Impacts
What was accomplished under these goals? During this year, unfortunately we were not able to initiate the field work due to the significant delay in the release of funds from USDA-NIFA. However, we advanced with the planning, logistics and organization of the field work (including the selection of sampling areas, the request ofsampling permits for some national parks, IRBs and otherethical approvals, etc), initiated the collection of secondary data that is already available and will be necesary to conduct part of the modeling and risk assessment activities, initiated the recruitment of students and staff to be involved in the project and built the project website.

Publications

  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: B. Mart�nez-L�pez, C. Quembo, J. Van Heerden, A. Bastos, ML. Penrith, JM Sanchez-Vizcaino, F Jori, J Fafetine, F Fasina, VM Rakotoharinome, H Guis, L Heath, S Molia, V Porphyre, P Thompson, L Vial, A Caron, E Etter. Unraveling the role of pig trade and socio-economic factors in the spread of African Swine Fever in endemic areas. Poster Presentation. CRWAD 2018. Available at: https://crwad.org/wp-content/uploads/CRWAD-2018.FINAL-Author-Index-and-Abstracts.pdf
  • Type: Websites Status: Published Year Published: 2019 Citation: A special session on African Swine Fever was organized thanks to this project within the GeoVet 2019 international conference. The GeoVet 2019 website is: https://geovet2019.ucdavis.edu
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: A total of 9 abstracts were published under the Special topic of African Swine Fever within the GeoVet 2019. All these abstracts are available in the Open Access Journal of Frontiers in Veterinary Science: https://www.frontiersin.org/events/GeoVet_2019_Novel_spatio-temporal_approaches_in_the_era_of_Big_Data/6796/topic
  • Type: Websites Status: Published Year Published: 2019 Citation: The project website has been published at: https://www.asf-nifnaf.org
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: B. Mart�nez-L�pez, A. Bastos, K. O'Hara, M. Pedrono, H. Guis, L. Vial, L. Heath, F. Folorunso Oludayo, J. Fafetine, F. Jori, J.M. S�nchez-Vizca�no, M.L. Penrith, J. van Heerden, C. Quembo, A. Caron, E. Etter. Evaluation of the evolution and main pathways of African Swine Fever virus (ASFV) transmission in endemic areas. Poster Presentation. CRWAD 2019. Available at: https://crwad.org/wp-content/uploads/CRWAD-2019-Author-Index-and-Abstracts.FINAL_.pdf