African swine fever –

all you must know

FLI / February 2019

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Outline

• The causative agent and its characteristics

• Host species and transmission pathways
• The disease in domestic pigs and wild boar
• Laboratory diagnosis Picture: The Pirbright Institute, UK

• Distribution and current situation

• Disease control (general)
• Lessons learned from the current epidemic
• Summary

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The causative agent
• Large, complex, envolped DNA-Virus, replicates
primarily in macrophages
– Why is this important?
• Large and complex: Multiple factors that help the virus to modulate and evade the
host immune system → one of the difficulties in vaccine production
• Many components are uncharacterized in location and function
• Replication in macrophages: antibodies can be detrimental, replication in
designated phagocytes
• Enveloped: important for the choice of disinfectants
• DNA: stable, rather slow evolution (in general)
• ASFV is the only known ARBO virus with a DNA genome
– ARBO: arthropod borne virus
• ASFV has a soft tick vector (replication in the vector, transmission)
• So far no relevance of soft ticks for Central Europe and several other parts of the
world, Asia?
• Vector has to be included in control measures
„ASFAR“
• For the completeness: African
Swine
– Genus Asfivirus in the Asfarviridae family Fever
And
Related viruses
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 Stability of the virus
Tenacity:
• 3 h at 50°C
• Up to 10 days in feces, even longer periods mentioned in old
publications (>3 month under favourable conditions)
• Up to 70 days in blood at room temperature
• Up to 15 weeks in cooled pork
• Up to 6 months in conserved ham
• Up to 18 months in cooled blood (fridge)
• Years or even decades in frozen carcasses and pork

pH stability:
Inactivation at pH <3,9 and >11.5
Serum (proteins) has a strong stabilizing effect: 21 h at pH 13,4!

Disinfection:
• Commercial disinfectants for enveloped viruses, e.g. Vennovet
• Peracetic acid, formic acid
• Hypochlorite, iodine, quarternary ammonium compounds
• Lime slurry, (caustic lime)

Heat treatment: 70°C für 20 min; 60°C 30 min…

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Host
Wirte species

- Domestic pigs

- Wild pigs, e.g. European wild boar

- Warthogs

- Soft ticks of the Ornithodoros genus

Ornithodoros moubata
Picture: R. Klein
X X
X
ASF is not a zoonosis… X
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 Transmission pathways
Introduction into the Cycle in domestic
Sylvatic cycle in Africa domestic pig population pigs and wild boar

Persistently infected adult warthogs do not show

high level viremia but may carry infected ticks Direct contact,
swill feeding

Trans-stadial and trans-

ovary transmission

Indirect contact

O. erraticus played a
European wild boar are as
Young warthogs in the burrow – role as vector on the
susceptible as domestic pigs
High viremia, source of infection for soft ticks Iberian peninsula
and played a role on Sardinia
(Ornithodoroos moubata) and in the Caucasus/Russia
Source of warthog pictures: Natur-Lexikon.com, Frank Stober und http://www.imagesofafrica.co.za/
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Clinical signs in domestic pigs

• First signs are observed app. four days post infection: High fever, reluctance to move, inappetence,
huddling, it is „too quiet“ when you enter the stable
• Some animals develop conjunctivitis and gastro-intestinal signs (vomiting, diarrhea)
• With progression of the disease, animals become somnolent, appear desorientated, and show dyspnea
• In the final phase, affected animals may show petechiae or map-like bleedings and cyanoses as well as
seizures and haemorrhages (epistaxis, bleedings from the anus)
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Clinical signs in wild boar

• In general no differences observed between domestic pigs and wild boar (skin lesions cannot be seen)
• No age dependence of clinical signs with highly virulent strains
• In the field: desorientation, lack of fear towards humans and dogs
• General observation from lab and field: young animals survive more often
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Differentiation of African and classical swine fever is impossible based on clinical
signs alone! The same is true for several other differential diagnoses!

Domestic fattening pigs at the seventh day post Domestic pigs and wild boar at the seventh day
inoculation with a highly virulent ASFV strain post inoculation with a highly virulent CSFV
(„Armenia08“) strain („Koslov“)

Laboratory confirmation is mandatory!

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Pathology

Animals that die from ASF show lesions in lymphnodes, lungs, kidney, and spleen:
• Ebony-colored, bloody and enlarged lymphnodes, especially in the gastro-hepatic area, throat,
and kidney
• Froth in lungs and trachea (lung edema), lung haemorrhages
• Petechia in the kidneys, kidney infarctions
• Haemorrhages in other organs
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 Laboratory diagnosis

• Neither clinical signs nor pathological lesions allow a reliable

diagnosis of ASF!
• Swift and reliable diagnostic methods are of paramount
importance
• Direct (pathogen detection) and indirect methods (antibody
detection) are available
• A growing number of ELISA and PCR kits is commercially
available and licensed in Europe
• ASF is still an exotic disease with severe impact: high
biosafety standards (containment, e.g. level L3+) are
mandatory for the work with ASFV
• In some EU Member States, screening methods are carried out
at regional level (PCR and ELISA, early detection is passive)
• Quality of diagnosis is ensured through national and
international ring trials

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Some words on sample quality…
0-Wert 1 Monat 2 Monate 3 Monate
4°C 20°C 37°C 4°C 20°C 37°C 4°C 20°C 37°C
cq cq cq cq cq cq cq cq cq cq
Blut 24,78 23,57 26,15 20,94 28,51 25,41 20,58 27,74 25,49 19,97
Milz 20,09 18,33 30,77 23,43 23,71 25,17 21 20,67 28,12 27,6
Bache 3
Muskel 25,55 25,94 27,02 25,78 27,53 27,4 29,76 29,26 41,41 27,3
Kot 34,4 32,95 33,54 34,27 29,26 28,86 31,3 33,62 33,9 33,57
Blut 27,5 27,8 29,65 26,47 27,66 32,69 29,37 31,07 31,54 27,68
Milz 21,75 20,41 24,18 27,02 26,17 31,81 28,14 25,27 26,94 27,28
Paula
Muskel 27,39 26,29 28,25 25,88 28,44 32,59 30,11 27,15 32,75 28,41
Kot 40,32 35,55 42,48 35,46 30,86 31,87 no cq 40,7 37,81 no cq
Blut 22,55 24,09 21,16 18 28,06 26,33 20,72 27,75 24,27 18,66
Milz 18,85 18,62 18,93 27,52 28,45 29,31 20,46 19,85 19,78 24,28
Hannibal
Muskel 26,05 25,65 27,63 22,94 26,9 26,05 27,47 28,6 24,45 25,19
Kot 40,44 36,38 no ct 30,9 32,19 30,33 31,45 42,24 38,8 41,74

„Sample too bad“ is not a good excuse…

… at least for most PCR applications…

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Targets in the EU: carcasses…

Source: Food and Veterinary Service of Latvia

• Sampling is not easy
• Rotten carcasses are smelly and disgusting…
• Some organs are not available anymore
• Blood (decent fluid) is not available
• Some carcasses are just skeletons
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Alternative Sampling

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“Point-of-care“ Diagnostics

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Distribution
• First description in Kenya 1921
• Reason was probably the introduction of domestic pigs through
colonists → unbalancement, new susceptible host
• 1957 first occurrence outside Africa → Portugal
• Thereafter, again in Portugal and subsequently in Spain, France, Italy,
Malta, Belgium, and The Netherlands
• Endemic on Sardinia, eradicated since the late 1990s in all other EU
countries
• 1980ies: Cuba, Brazil, Dominican Republic, and Haiti (eradicated)
• Over the last decade (since 2007), ASF has spread from Georgia over
several Trans-Caucasian countries and Russia to the Baltic states and
Poland
• Recent outbreaks in Hungary, Belgium, Czech Republic, Romania,
Bulgaria, Ukraine, and Moldova
• China, Mongolia, Viet Nam
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„Historical“ ASF outbreaks within the EU

Country Outbreaks

Portugal 1957, 1960-1993; 1999

Spain 1960-1995

Italy 1967, 1969, 1993

Sardinia: since 1978
France 1964, 1967, 1977

Malta 1978

The Netherlands 1986

Belgium 1985

Successful eradication with the

exception of Sardinia!

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Current situation in Europe

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August 2018:

Die ASP hat den

größten
Schweineproduzenten
der Welt erreicht…

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News Jan 7, 2019 (Pig progress):
ASF China: Infected pig carcass found at Taiwanese beach
A dead pig, infected with African Swine Fever (ASF), has been found on the
shores of an islet that belongs to Taiwan. Most probably it washed ashore from
mainland China.

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August 2018:

ASF reached the

world`s largest pork
producer, China …
… swill feeding …
… detections in food
and feed…

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ASF in Sardinia - … since 1978

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A variant virus…

Zani et al., 2018

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Biological characterization of the variant

Moderate/rather low virulence in potbelly mini pigs and domestic pigs

18 dpi

No transmission to fetuses of infected sows EM-TH-19-0052

 Dr. Jekyll and Mr. Hyde?
The plan: use the variant to answer the question
whether the variant can protect against the co-
circulating virulent virus strains
Studien Design:
• Inokulation of five wild boar of different age calsses
with the variant ASFV „Estonia P2“
• Challenge with ASFV „Armenia08“ (the „Killer“)
Infection with the Challenge with Necropsy
attenuated phenotype ASPV Armenia

Photo by Henry Van der Weyde (1838-1924; London, England) - http://www.photography-museum.com/jekyll.html /

EM-TH-19-0052 Originally uploaded to en.wikipedia
 Plans can be changed…
120

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survival rate [%]

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days post infection

• Highly virulent phenotype with severe • Typical lesions…

clinical signs already in the first week

What is the moral of the story? → we still do not know the virus…

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Control measures (general)
• Suspicions and outbreaks have to be notified!
• Strict and legal binding control measures
• So far, no vaccine or treatment!
• Integral part of all control measures is an early detection
system (detection of clinical signs and a rapid laboratory
diagnosis)
• The affected holding and the contact holdings must be
culled
• Stand-still
• Restriction zones are implemented (3 km control zone, 10
km surveillance zone)
• Trade with pigs, pork and pork products comes to a rest
• ASF in wild boar: control plan taking into account the local
situation, measures also in domestic pigs
• Prevention: biosecurity and awareness (do not feed swill
and do not let wild boar have access to it)
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 ASF in Czech Republic

500 km

Graphik: FLI, IfE

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Hunting rest Intensify
hunting

Removal of
carcasses

Use harvest
Incentives time

Fencing
Repell
Military

Reduce public
Infrared/ Snipers
night vision

Poison

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Picture: IfE, A. Globig / C. Probst

Gary Kramer - National Digital Library of the United States Fish and Wildlife Service

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Mysterious Transmission

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Five grams of gamma-irradiated ingredients (in 50 mL mini bioreactor tubes) were spiked
with 100 μL of MEM (minimum essential media, Gibco, ThermoFisher Scientific, Waltham,
MA, US) containing 1 x 105 tissue culture infectious dose 50 (TCID50) of each virus [12].

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What ASF did not do after its re-introduction into the EU:

XX
• No explosive spread to the West
• No self-termination of outbreaks
Lessons learned:
• Contagiosity was overestimated
• Endemic cycles were established in the wild boar population, independently from
domestic pigs
• The contaminated habitat plays a crucial role
• Transmission routes are often „golden bullet events“ with very low probability but
high impact, risk assessments and predictions are most difficult
• Remember transmission in the natural cycle: one droplet of blood can start an
epidemic
• Lethality is still high but mortality can be low, especially at the beginning of an
epidemic
• There are some virus variants with lower (moderate) virulence; these viruses seem
to have a disadvantage in wild boar (disappeared quickly)
• Surviving animals carry the virus for a long time and genome even longer; however,
the percentage of animals that do not eliminate the virus in the end is very low
(not existent in our long-term experimental studies), no transmission from true
convalescent animals to sentinels (surviviors are detected by ELISA)
• Antibody detection does not have a predictive value towards the outcome of the
infection
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 „Group dynamics“ of ASF

• Oral infection is rather inefficient (remember that the virus is optimized for tick transmission)
• In the majority of cases, rather high virus doses are needed for oral infection and not all inoculations
result in infection (~10.000 HAU)
• An infected animal dies with high probability, but not all animals in a stable/pen have to be infected
• The virus is found in high amounts in blood of diseased animals, much less in feces and saliva
• Contagiosity can be moderate or even low if no blood shedding occurs!
• Even with close contact, some animals may go uninfected (see pictures above)
• In units with separate pens, the disease may stop after one or some pens
• Transmission after an isolated introduction can be sluggish
• Fulminant courses are seen in breeding farms where abortions and thus blood contact is frequent

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➢ African swine fever is caused by a large, complex DNA virus
➢ The virus modulates the immune system
➢ The sylvatic cycle involves a soft tick vector, related soft ticks can
complicate control
➢ The virus is highly stable and under cool conditions carcasses, meat
products etc. can be infectious for months (if frozen, years)
➢ Despite its high stability, it can be easily inactivated using heat or
disinfectants for enveloped viruses
➢ Contagiosity is moderate (!) under our conditions

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➢ African swine fever virus affects only Suids and soft ticks of the genus
Ornithodoros
➢ ASF is not a zoonosis and no other competent vectors are described
➢ The sylvatic cycle in sub-Saharan Africa involves O. moubata soft ticks
and warthogs (ASFV is an ARBO virus)
➢ Warthogs do not show overt disease (inapparent infection)
➢ When introduced into the domestic pig sector, ASF is usually a highly
lethal disease and is no longer dependent on vector transmission
➢ ASF can be transmitted directly from pig to pig and indirectly through
fomites
➢ Wild boar are as susceptible as domestic pigs

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➢ ASF is a severe multi-systemic disease that can be accompanied by signs of
a viral haemorrhagic fever
➢ First unspecific signs occur app. 4 days after oral infection
➢ Signs include high fever, inappetence, huddling, and somnolence
➢ In the final stages, haemorrhages and neurological signs can be present
➢ A reliable clinical diagnosis is impossible and needs laboratory confirmation
➢ Lethality upon infection with highly virulent strains is very high
➢ Pathomorphological lesions depend on the course of infection and include
enlarged and haemorrhagic lymphnodes, bleedings in different organs, and
lung edema
➢ Signs in wild boar are similar
➢ Direct (virus detection) and indirect (antibody detection) diagnostic
methods exist
➢ Both antibody ELISAs and real-time PCR kits are commercially
available and licensed at European level
➢ High containment levels are needed to work with African swine
fever virus
➢ Special conditions require pragmatic approaches: wild boar
carcasses can be sampled using dry swabs
➢ Blood swab samples can be tested for antibodies and viral genome
➢ PCR is suitable over a long period of time and for samples of limited
quality

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➢ African swine fever was first described in Kenya 1921
➢ Sporadic outreaks occurred in Europe and on the American
continent after 1957
➢ Spain and Portugal eradicated the disease after intensive control
measures
➢ ASF is endemic on the island of Sardinia since 1978
➢ Large epidemic in Eastern Europe since 2007
➢ Started in Georgia and reached the European Union in 2014
➢ Recently, ASF was reported from several European Union Member
States
➢ Control measures have to rely on strict veterinary hygiene,
biosecurity, and timely diagnosis
➢ Neither vaccine nor treatment exist

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Thanks for your attention!

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