CENTRO DE INVESTIGACION EN SANIDAD ANIMAL (CISA-INIA)

European Union Reference Laboratory for ASF (URL-ASF)

African swine fever (ASF) in Africa. Main activities 2004-2011

Marisa AriasWorkshop on ASF, Nairobi, July, 2011

CISA (INIA)-ILRI Collaborative project

2004-2010ASF Collaborative Project

“Development of new diagnostic assays and epidemiological surveillance of viral pathogens of

livestock in Sub-Saharan Africa” between the International Livestock Research Institute (ILRI) and

Centro de Investigacion En Sanidad Animal (CISA-INIA)

CISA (INIA)- ASFRISK PROJECT

2008-2011

MAIN GOAL → Improvement of knowledge of the epidemiological situation of ASF in Africa

Description of the epidemiological

situation in African countries based on

epidemiological findings and samples collected in

different African countries.

INIA-ILRI

INIA-ILRI

FAOASFRISK

FAO

OBJECTIVES

To evaluate the epidemiological situation of ASF in Africa,FROM THE MOLECULAR AND BIOLOGICAL POINT OF VIEW, tocharacterize currently circulating field viruses ofepidemiological interest.

To develop, validate and apply improved, robust and/orsimple ANTIBODY AND NUCLEIC ACID DETECTION METHODSFOR ASFV for the rapid detection and differential diagnosisof suspected cases of ASF.

Technology Transfer to regional African Labs

WORK SCHEDULE

Sampling strategy → SAMPLINGCOLLECTION in collaboration with theVeterinary Services, Wildlife Services, OIE andFAOASF diagnosis at CISA-INIA using OIE-prescribed diagnostic tools.ASFV genetic characterizationASFV biological characterization

CISA (INIA)-ILRI Collaborative project

EAST AFRICA

CISA (INIA)-ILRI Collaborative project

WORK SCHEDULE

Sampling strategy → SAMPLINGCOLLECTION in collaboration with theVeterinary Services and Wildlife Services.ASF diagnosis at CISA-INIA using OIE-prescribed diagnostic tools.ASFV genetic characterizationASFV biological characterization

CISA (INIA)-ILRI Collaborative project

The last outbreak of ASF in Kenyawas reported in central Kenya(Kiambu, Nairobi and Thika) inAugust 2001.

Presence of the disease in Uganda. Theoutbreak occurred in 2003 Uganda hadspread from the Central region to the Easternregion of the country which shares a borderwith Kenya (Busia district).

Presence of the disease in Tanzania. Sporadicoutbreaks of the disease had been reportedin Northwest Tanzania since 2001.

Phase I (2004-2007)→ ASF in East Africa (background)

ASF situation in 2004

CISA (INIA)-ILRI Collaborative project

Surveillance program; Sampling collection from free-ranging pigs inWestern and Central Kenya (no apparent clinical signs related to ASF).

ASF in KENYA (SAMPLING STRATEGY)

ASF in KENYA (SAMPLING STRATEGY)

CISA (INIA)-ILRI Collaborative project

Surveillance program; Sampling collection from free-ranging pigs(no apparent clinical signs related to ASF) and bush pigs in WesternKenya districts and in neighbouring Ruma National Park

Prevalence of ASFV in bush pigs and their

role in the transmission of the

disease.

CISA (INIA)-ILRI Collaborative project

Study conducted since 2006 in 6 administrative divisions of Homabay district in Western Kenya to identify critical issues related to pig production as well as risk factors for African swine fever. The

specific study area was selected because it represents a predominantly free-range smallholder pig production system and lies in close proximity to a

national park, factors that increase the risk of ASF.

Western Kenya districts and in neighbouring Ruma National Park

ASF in KENYA (SAMPLING STRATEGY)

CISA (INIA)-ILRI Collaborative project

Surveillance program; Sampling collection from warthogs andticks in Kapiti plain state (Central Kenya) in collaboration withthe KENYAN WILD LIFE SERVICE

Prevalence of ASFV in warthogs (Phacochoerus

africanus ) and ticks and the role in the

transmission of the disease → SYLVATIC

CYCLE

ASF in KENYA (SAMPLING STRATEGY)

CISA (INIA)-ILRI Collaborative project

Surveillance program; Retrospective sampling collection fromwarthogs in Northern Kenya from 2006-2008 in collaborationwith the KENYAN WILD LIFE SERVICE

Prevalence of ASFV in warthogs

(Phacochoerus aethiopicus )

in Northern Kenya

ASF in KENYA (SAMPLING STRATEGY)

CISA (INIA)-ILRI Collaborative project

Sampling collection from ASF outbreaks occurred in Kenya 2006-2007 (OIE report 04/05/2007) and in 2010-2011 (OIE report 04/03/2011 )

2006-2007 2010-2011

Kenyan Dept. Vet. Service, (Joseph Macharia)

ASF in UGANDA (SAMPLING STRATEGY)

CISA (INIA)-ILRI Collaborative project

→ Sampling collection from ASF outbreaks occurred in Uganda in 2007

→ Sampling collection from National Parks in Western Uganda to determinethe prevalence of ASFV virus in warthogs and their role in the transmission ofthe disease. In Collaboration with Conservation Tthrough Public Health(CTPH)(Gladis)

Uganda, Ministry of Agriculture (Rose Ademun)

ASF in TANZANIA (SAMPLING STRATEGY)

CISA (INIA)-ILRI Collaborative project

Surveillance program;Sampling collection fromfree-ranging pigs in Northand Eastern Tanzania werelast ASF outbreaks werereported (no apparent clinicalsigns related to ASF).

2005

ASF in TANZANIA (SAMPLING STRATEGY)

CISA (INIA)-ILRI Collaborative project

→ Sampling collection in the Serengeti National Park todetermine the prevalence of ASFV virus in warthogs and theirrole in the transmission of the disease.

Collaboration with Tanzania Wild Life Service –TAWIRI-

WEST AFRICA

CISA (INIA)-ASFRISK

Improve understanding of virus spread and

maintenance in West African countries

Sampling and characterisation of currently circulating West

Africa field strains

CISA (INIA)-ASFRISK

2,446 samples from…. ASF free country → Côte d'Ivoire

468 serum samples(surveillance program)

2009

ASF in WEST AFRICA (SAMPLING STRATEGY)

CISA (INIA)-ASFRISK

Study area and sampling collection

2,446 samples from…. ASF endemic country → Burkina Faso

261 samples (serum, tissue, whole blood)

collected from 15 provinces out of 45 during ASF outbreks

2007-2009

ASF endemic country → Ghana

31 tissues collected from domestic

pigs during ASF outbreaks

2002-2008

ASF endemic country →Togo

31 samples (serum and tissues)

collected from domestic pigs during ASF outbreak

2009

ASF endemic country → Benin

57 samples (serum and tissues) collected

from domestic pigs during ASF outbreaks

2009

ASF in WEST AFRICA (SAMPLING STRATEGY)

CISA (INIA)-ASFRISK

2,446 samples from…. ASF endemic country → Nigeria

1,598 samples (serum and tissues) collected

from domestic pigs during ASF outbreaks and endemic regions

2006-2008

ASF in WEST AFRICA (SAMPLING STRATEGY)

CISA (INIA)-ASFRISK

ASF endemic country → Republic of the Congo

ASF in WEST AFRICA (SAMPLING STRATEGY)

WORK SCHEDULE

Sampling strategy → SAMPLING COLLECTION in collaborationwith the Veterinary Services, Wildlife Services, OIE and FAO

ASF diagnosis at CISA-INIA using OIE-prescribeddiagnostic tools.ASFV genetic characterization to determine;Source of outbreaksRole of sylvatic cycle in the transmission of ASF

ASFV biological characterization

WORK SCHEDULE1. ASF infection status on samples collected.

1. Ab detection (ELISA+IB)

2. Virus detection (PCR + virus isolation)

2. ASF molecular characterization on selected positivesamples using ASF genotyping standarizedprocedures1. P72 genotyping (C-terminal end)2. P54 genotyping (full gene)3. CVR subtyping

MAIN FINDINGS → EAST AFRICA

Sylvatic cycle → Existence an endemically stableepidemiological situation involving domestic, wild pigs(bushpigs and warthogs) and ticks.

Presence of ASF virus in absence of antibody response in healthydomestic pigs in Central and Western Kenya.Presence of ASF virus in absence of antibody response in bushpigs inWestern Kenya.Presence of ASF virus and specific antibody response in warthogs andASFV in ticks in Kenya, Uganda and Tanzania.

MAIN FINDINGS → EAST AFRICA

ASFV isolates obtained in Kenya from ticks and domesticpigs collected in surveillance program without ASF reportedoutbreaks clustered in P72 genotype X → SYLVATIC CYCLE

ASF viruses genotyped directly from warthogs at samephysical locality clustered within p72 genotype IX and weretherefore genetically similar to viruses responsible forrecent disease outbreaks in East Africa (Kenya and Uganda).

Coexistence in Kenya ofdistinct ASFV genotypes inwarthog- burrow associatedticks and in adult wildwarthogs and the apparenttransfer of both genotypes todomestic pigs.

MAIN FINDINGS → EAST AFRICAPresence of a domestic pig-associated genotype IXcausing ASF outbreaks in the border region betweenKenya and Uganda occurred in 2006 and 2007, 2010 and2011.•The rapid spread of the virus among pigs and the acute forms of thedisease suggest that the disease may have been maintained in theborder regions either in contaminated pork products, or live pigs thatbecame carrier-pigs, surviving the first outbreak.•The evidence of trans-boundary transmission between thesecountries indicates that a regional approach to disease control would bemore efficient.

MAIN FINDINGS → WEST AFRICA

The results obtained from the ASFdiagnosis on samples collected fromdomestic pigs in target West Africancountries confirms the evidence of ahigh incidence of the disease in Ghana,Benin, Burkina Faso, Togo, Nigeria andRepublic of Congo.

MAIN FINDINGS → WEST AFRICA

Domestic pig-cycle associated genotype I.

Cross-virus circulation between neighbouringcountries Togo, Ghana, Benin and Nigeria in Maritime areas, as well as in bordering areas among Ghana and Burkina Faso.

P72 genotype I

P54 genotype Ic

West Congo districts → related historical West Africa viruses

MAIN FINDINGS → WEST AFRICA

P72 genotype IXP54 genotype IX

East Congo districts → related recent East Africa viruses

MAIN FINDINGS → WEST AFRICA

Movement of a virus genotype previously associated with virulent ASF in easternAfrica to western Africa, where the viruses have hitherto always been classified in p72genotype I. there would be the possibility of future outbreaks of disease caused by novelviruses in western Africa. This study confirms the continuing spread of ASFV.

KENYA outbreaks 2006-2007

KENYA sylvatic cycle

UGANDA outbreaks 2007

Surveillance in Nigeria

Congo outbreaks 2009

TITLE: African Swine Fever Virus p72 Genotype IX in Domestic Pigs,Congo, 2009Carmina Gallardo, Raquel Anchuelo,Virginia Pelayo, Frédéric Poudevigne, Tati Leon,Jacques Nzoussi, Richard Bishop,Covadonga Pérez, Alejandro Soler, Raquel Nieto,Hilario Martín, and Marisa Arias

Emerging Infectious Diseases • www.cdc.gov/eid • Vol. 17, No. 8, August 2011DOI: 10.3201/eid1708.101877

WORK SCHEDULE

Sampling strategy → SAMPLING COLLECTION incollaboration with the Veterinary Services, WildlifeServices, OIE and FAOASF diagnostic at CISA-INIA using OIE-prescribed diagnostic tools.ASFV genetic characterizationASFV biological characterization.

Clinical, biological and immunological characteristics of ASF recently investigated using

European pig breeds by inoculation with three ASFV

Kenyan isolates belonging to the most variable defined

genotypes IX and X.

MAIN FINDINGSAcute to subacute forms of the disease showing typicalclinical signs and lesions associated to ASFV moderate strains.

Viremia detectable by OIE- prescribed virological diagnostictechniques at early times post infection and was maintainedduring the whole infection.

Antibody response detectable by OIE- prescribedserological diagnostic techniques developed in thesecond week of infection.

Complex epidemiological situation in eastern regions of Africa

ASFRISK: Why Non evident ASF clinical signs in ASF outbreaks, co-existing with a high viral

load and a significant lack of antibody response

Increase of the risk of the endemicity of ASF and virus spreading → Increased difficulty for the control of the disease.

Are the current ASF diagnostic tools adapted to all epidemiological situations?

Existence Different Transmission cyclesASFV genetic and antigenic variability

The current ASF serological diagnostic tools ARE ADAPTED TO ALL

EPIDEMIOLOGICAL SITUATIONS

The results obtained using new Ags based on current and variable circulatingASFV strains were 100%according to those obtained using OIE prescribedantibody detection techniques.

Immunogenetics and genetic characteristics of theEast Africa indigenous pig populations?

OBJECTIVEComparative in vivo study of the clinical,

pathological and immune response against the ASFV infection in Kenyan

“indigenous pigs” and European domestic pigs using Kenya ASFV strain.

EXPERIMENTAL DESIGN

Selected DOMESTIC PIG BREEDS29 Indigenous domestic swine (local breed) from Homa Bay

district (6-month old)

10 Exotic domestic swine (Landrace) from Kitengela

(6-month old)

EXPERIMENTAL DESIGN

Selected DOMESTIC PIG LOCAL BREEDSHoma Bay

Homa Bay district (Western Kenya)

represents a predominantly free-

range smallholder pig production system

Selected ASFV isolate → Ken05/K2

PRELIMINARY CONCLUSIONS

Different behaviour in ASFV infection, in ASF clinical signs.

Delay of onset of ASF in “indigenous pigs”

No external clinical signs related to ASF → the disease couldbe easily unrecognized in field conditions. Several animalseven died without fever.

High variety of pathological findings. Further investigation isrequired .

The laboratory was essential to confirm the presence of ASF

Slight delay in the seroconversion in indigenous pigs.March-May 2011. STUDIES ON GOING

COMPARATIVE RESULTS

Comparative Tª (average)

TECHNOLOGY TRANSFER

•Delivery of ASF reference reagents and Standard

operating procedures (SOP).

•Training courses on African Swine Fever (ASF)

diagnostic techniques in Africa → Transfer the ASF OIE-

prescribed diagnostic tools to participants from Department of

Veterinary Services (DVS) and National systems staff (regional labs) from

Eastern and Southern African countries and Nigeria.

European Union Reference Laboratory

TRANSFER OF TECHNOLOGY ON ASF diagnostic techniques

-Venue: Nigeria, Rwanda, Kenya -(funds: INIA-ILRI)

Mean: 10 days Training Course, on Diagnostic techniques

-Venue: Uganda : 25 attendances from Vet Services of Kenya, Tanzania and Uganda. (funds: ASFRISK RTD,EC, INIA-ILRI)

2007-2011

-Venue: Tanzania, Tanzania, Kenya, Rwandaand Uganda .30 attendances. (funds: ASFRISK RTD,EC INIA-ILRI)

5 days Training Course,

-Venue: OVI, Onderstepoort, South Africa. May 2011. DIAGNOSIS- More than 20 delegates coming from south and south East African countries : Malawi, Botswana, Angola, Zambia, Tanzania, Mauritius, Seychelles, Mozambique, Swaziland, Zimbawe, DR Congo, SouthAfrica.

Collaboration in TRAINING Courses ON ASF diagnostic techniques

MAIN CONCLUSIONS

• Complex epidemiological situation in easternregions of Africa with the presence of endemicallystable situation involving domestic, wild pigs and tickswhich difficult the control of the disease.

• Multiple genotypes in countries with the sylvatic cycle.

• Widespread of ASFV genotypes from eastern towestern Africa → evidence of trans-boundarytransmission between neighboring countries related tomovement of domestic pigs and pork products.

MAIN CONCLUSIONS

• Improvement of knowledge of epidemiology ofASF. Map distribution of pig density as well as natural

hosts/vectors. Prevalence of the disease trough and appropriate

sampling strategy and the application of prescribedASF diagnostic tools.

Molecular studies of virus from outbreaks and naturalhosts.

Examine mechanisms of natural resistance in domesticpigs.

MAIN CONCLUSIONS

Transfer technology at regional laboratory level

FOLLOW UP

FUTURE ACTIVITIESKey Areas of Research with implications forcontrol will mainly include:

Further investigation of the importance of thewarthog/tick sylvatic cycle in causing disease outbreaksthrough genotyping of viruses from these species frommultiple sites within the region. Kenya and Uganda.

Focus on in-depth surveillance and outbreakmonitoring of ASFV in Tanzania

Link virulence phenotype of viruses determinedthrough experimental infection of indigenous and exoticdomestic pigs in Spain and Kenya with completegenome sequences of three Kenyan isolates.

AKNOWLEDGMENTS

Cote d`Ivoire

Congo RepTanzania

AKNOWLEDGMENTS

EU Reference Laboratory CISA-INIA

INTERNATIONAL LIVESTOCK RESEARCH INSTITUTE

Special thanks to the coordinator at ILRI: Dr. Richard Bishop

Our Thanks/Ahsante Sana

TO OUR HOST COUNTRY:

SCIRO sponsor of this Event

AND

MY SPECIAL THANKS

Dr. Carmina Gallardo Frontaura