Isolation of African swine fever virus from ticks of the Ornithodoros moubata complex (Ixodoidea: Argasidae) collected within the African swine fever enzootic area of Malawi


Haresnape, JM.; Wilkinson, PJ.; Mellor, PS.

Epidemiology and Infection 101(1): 173-185

1988


Ticks of the Ornithodoros moubata complex were collected from domestic pig sties and dwelling houses, and from a warthog habitat, and tested for the presence of African swine fever (ASF) virus. Collections were made in 9 of the 24 districts of Malawi, these being primarily the districts in which O. moubata is most numerous. ASF virus was isolated from ticks collected in both domestic pig sties and houses in certain villages in Mchinji district where ASF outbreaks had recently occurred. Mchinji district is in the centre of a large ASF enzootic area which stretches into other districts of Malawi and also into Zambia and Mozambique. The high titre of virus in some of the ticks demonstrates that O. moubata can act as a virus reservoir and potential vector of disease in the field situation in Malawi.

Epidem.
Inf.
(1988),
101,
173-185
173
Printed
in
Great
Britain
Isolation
of
African
swine
fever
virus
from
ticks
of
the
Ornithodoros
moubata
complex
(Ixodoidea:
Argasidae)
collected
within
the
African
swine
fever
enzootic
area
of
Malawi
BY
J.
M.
HARESNAPE
Central
Veterinary
Laboratory,
Lilongwe,
Malawi
P.
J.
WILKINSON
AND
P.
S.
MELLOR
Institute
for
Animal
Health,
Pirbright
Laboratory,
Ash
Road,
Pirbright,
Surrey
GU24
ONF,
UK
(Accepted
8
February
1988)
SUMMARY
Ticks
of
the
Ornithodoros
moubata
complex
were
collected
from
domestic
pig
sties
and
dwelling
houses,
and
from
a
warthog
habitat,
and
tested
for
the
presence
of
African
swine
fever
(ASF)
virus.
Collections
were
made
in
9
of
the
24
districts
of
Malawi,
these
being
primarily
the
districts
in
which
0.
moubata
is
most
numerous.
ASF
virus
was
isolated
from
ticks
collected
in
both
domestic
pig
sties
and
houses
in
certain
villages
in
Mchinji
district
where
ASF
outbreaks
had
recently
occurred.
Mchinji
district
is
in
the
centre
of
a
large
ASF
enzootic
area
which
stretches
into
other
districts
of
Malawi
and
also
into
Zambia
and
Mozambique.
The
high
titre
of
virus
in
some
of
the
ticks
demonstrates
that
0.
moubata
can
act
as
a
virus
reservoir
and
potential
vector
of
disease
in
the
field
situation
in
Malawi.
INTRODUCTION
It
has
long
been
known
that
wild
pigs
such
as
warthogs
(Phacochoerus
aethiopicus)
and
possibly
bush
pigs
(Potamochoerus
porcus)
are
potential
carriers
of
African
swine
fever
(ASF)
in
Africa,
although
they
do
not
show
signs
of
disease,
and
they
have
been
considered
to
be
the
natural
hosts
and
primary
reservoirs
of
the
virus
(Montgomery,
1921).
ASF
virus
was
first
recovered
from
naturally
infected
warthogs
in
South
Africa
(Steyn,
1932)
and
later
in
Kenya
(Hammond
&
DeTray,
1955)
and
has
also
been
found
in
bush
pigs
in
South
Africa
(Thomas
&
Kolbe,
1942).
Establishing
how
the
virus
could
be
transmitted
from
wild
to
domestic
pigs
was
recognized
as
a
major
problem
since
direct
contact
between
them
is
rare,
and
attempts
to
demonstrate
transfer
of
virus
by
contact
from
'
carrier'
warthogs
to
domestic
pigs
usually
failed
(DeTray,
1963
;
Scott,
1965
;
Heuschele
&
Coggins,
1969
;
Plowright,
Parker
&
Peirce,
1969
b).
The
discovery
that
the
argasid
tick
Ornithodoros
moubata,
collected
from
warthog
burrows
in
East
Africa
contained
ASF
virus
(Plowright,
Parker
&
Peirce,
1969
a)
was
a
major
step
forward.
Following
this
it
was
established
that
ASF
virus
replicates
in
the
ticks,
that
naturally
infected
ticks
can
transmit
the
174
J.
M.
HARESNAPE,
P.
J.
WILKINSON
AND
P.
S.
MELLOR
disease
to
domestic
pigs
under
experimental
conditions,
and
that
both
transovarial
and
sexual
transmission
of
virus
are
possible
in
ticks
(Plowright,
Parker
&
Peirce,
1969
b
;
Plowright,
Perry
&
Peirce,
1970
;
Plowright
et
al.
1970
;
Plowright,
Perry
&
Greig,
1974).
It
was
therefore
proposed
that,
once
a
tick
became
infected
by
feeding
on
a
viraemic
vertebrate
it
could
maintain
virus
for
long
periods,
possibly
for
life,
without
further
exposure
to
a
viraemic
host.
The
involvement
of
ticks
of
the
Ornithodoros
moubata
complex
as
an
efficient
virus
vector
provided
an
explanation
of
how
virus
could
be
transferred
from
wild
to
domestic
pigs.
Ticks
of
the
Ornithodoros
moubata
complex
have
many
potential
hosts
beside
warthogs,
including
man
and
many
domestic
animals,
of
which
Hoogstraal
(1956)
suggests
the
domestic
pig
to
be
the
most
important.
It
seemed
likely
that
ticks
associated
with
domestic
pigs
in
Africa
may
be
more
important
in
the
epizootiology
of
ASF
in
domestic
pigs
than
those
associated
with
warthogs.
However,
no
studies
on
the
isolation
of
ASF
virus
from
0.
moubata
collected
from
domestic
pig
premises
in
Africa
had
been
done
prior
to
this
study.
Initial
investigations
in
Malawi
showed
0.
moubata
to
be
present
in
domestic
pig
pens
(kholas)
in
several
districts
with
a
more
widespread
distribution
in
dwelling
houses.
It
is
particularly
numerous
in
pig
kholas
in
several
areas
where
ASF
is
enzootic,
and
may
play
a
role
in
the
epizootiology
of
ASF
in
the
Central
Region
of
Malawi
(Haresnape
&
Mamu,
1986).
The
purpose
of
this
study
was
to
assess
the
importance
of
ticks
associated
with
domestic
animals
in
acting
as
reservoirs
of
ASF
virus
and
vectors
of
disease.
MATERIALS
AND
METHODS
Collection
of
ticks
Extensive
collections
of
0.
moubata
from
domestic
pig
kholas
and
houses
were
made
in
many
parts
of
Malawi
both
within
the
ASF
enzootic
area
and
elsewhere
(Figs.
1
and
2).
One
collection
was
made
in
Liwonde
National
Park
from
a
culvert
frequented
by
warthogs.
Collections
were
made
by
staff
of
the
Central
Veterinary
Laboratory
(CVL),
Lilongwe,
and
by
Veterinary
Assistants
stationed
in
rural
areas,
as
described
previously
(Haresnape
&
Mamu,
1986).
Ticks
were
packed
in
plastic
tubes,
keeping
those
from
different
pig
kholas
and
houses
separate,
and
the
tubes
were
packed
in
sealed
plastic
bags
and
despatched
at
ambient
temperature
to
the
Pirbright
Laboratory,
Institute
for
Animal
Health,
UK.
Collection
of
interview
data
Details
of
any
cases
of
ASF
in
the
areas
where
collections
were
made
were
obtained
from
interviews
with
pig
owners.
Some
pig owners
in
areas
where
ticks
were
found
in
houses
but
not
in
pig
kholas
were
also
interviewed.
Where
laboratory
confirmation
of
ASF
had
not
been
obtained
deaths
from
ASF
were
assessed
on
the
basis
of
the
interview
data
as
described
previously
(Haresnape,
1984
;
Haresnape,
Lungu
&
Mamu,
1985).
Virus
isolation
in
tissue
culture
Adult
ticks
were
examined
individually
or
in
pools
of
2-6.
The
nymphal
stages
were
pooled
in
groups
of
up
to
22.
Individual
ticks
or
small
pools
were
ground
up
ASF
virus
infected
0.
moubata
in
Malawi
175
Chitipa
.:karonga
10
0
TANZANIA
11
°
N
Mzimba
,-
(
Nkhata
Bay
Lake
<
Malawi
k
0
Nkhotakota
Kasungu
12
°
km
0
10
20
30
,
13
°
Ntchisi\
,
\
....
412
,
Dow
a
s
t
Mchmji
---
,
'Salim
-
a
Lilongwe
,
'Dedza
Mango
hi
Ntchet)1
9
3
34
°
MOZAMBIQUE
14
°
15
°
I
Machinga
MOZAMBIQUE
Zomba
Mwanza/
Blantyre
Chiradzuili
Mulanje
Chikwawa"
Thyol•
16
°
Nsanje
17
°
35
°
Fig.
1.
Sites
where
ticks
were
collected
from
pig
kholas
and
warthog
habitats
and
tested
for
presence
of
ASF
virus.
Number
of
ticks
from
pig
kholas
tested
for
presence
of
ASF
virus
(
0,
10-100
;
0,
100-1000;0,
more
than
1000).
Filled
in
circles
represent
sites
where
one
or
more
infected
ticks
were
found.
Number
of
ticks
from
warthog
habitats
tested
for
presence
of
ASF
virus
(0
,
more
than
1000).
—,
Regional
boundary
;
-,
District
boundary
; ,
National
Park
or
Game
Reserve
;
*,
ASF
enzootic
area.
176
J.
M.
HARESNAPE,
P.
J
.
WILKINSON
AND
P.
S.
MELLOR
hitipa
.
N
Karonga
10
°
TANZANIA
N
Rumphi
O
0
Mzimba
o
0
;Nkha
a
Bay
O
Lake
o
M
alawi
Nkhotakota
N
km
0
10
20
30
0
Kasun
u
12
°
13
°
Ntchis
MOZAMBIQUE
Dowa
I f
i
ilongwe/
Dedza
14
°
Mangochi
34
°
N
cheu
\
chin
33
°
15
°
Machinga
MOZAMBIQUE
Mwanza
Zomba
Bfarityre•..
Chiradzulu
Mulanje
16
°
Chikwawa<
Thyolo
Nsanje
17
°
35
°
Fig.
2.
Sites
where
ticks
were
collected
from
houses
and
tested
for
presence
of
ASF
virus.
Number
of
ticks
from
houses
tested
for
presence
of
ASF
virus
o,
less
than
10
;
o
,
10-100
;
0
,
100-1000
0,
more
than
1000).
Filled
in
circles
represent
sites
where
one
or
more
infected
ticks
were
found.
—,
Regional
boundary
;
,
District
boundary
;
RI,
National
Park
or
Game
Reserve
;
#
,
ASF
enzootic
area.
ASF
virus
infected
0.
moubata
in
Malawi
177
in
2
ml
of
diluent
(PBS
containing
1
%
ox
serum
and
antibiotics)
and
larger
pools
in
5
ml
of
diluent.
Suspensions
were
clarified
by
centrifugation
for
5
min
at
1000
rpm
and
the
supernatant
medium
was
either
assayed
immediately
or
stored
at
70
°C.
Primary
cultures
of
pig
bone
marrow
(PBM)
cells
were
used
for
the
detection
and
titration
of
infectious
virus.
Cultures
were
prepared
as
described
previously
(Wilkinson
et
al.
1977)
in
tubes
containing
10
7
cells
in
1.5
ml
medium
which
were
incubated
in
stationary
racks
at
37
°C
and
used
after
3-4
days.
Tick
suspensions
were
assayed
to
determine
whether
or
not
they
contained
infectious
virus
by
the
inoculation
of
0.33
ml
into
each
of
three
tubes
of
PBM
cells,
which
were
examined
daily
for
6
days
for
haemadsorption.
Positive
ticks
or
tick
pools
were
then
titrated
using
10-fold
dilutions
in
diluent
and
three
culture
tubes
per
dilution,
each
tube
being
inoculated
with
0.33
ml.
Cultures
were
examined
daily
for
haemadsorption
and
discarded
if
positive.
Five
days
after
inoculation
0.2
ml
of
a
1
%
suspension
of
pig
erythrocytes
was
added
to
each
culture
and
final
readings
were
made
on
day
6.
Titres
were
expressed
as
50
%
haemadsorbing
doses
(HAD
50
)
per
tick
or
per
pool.
Virulence
tests
Cross-bred
Large
White
x
Landrace
pigs
were
infected
with
the
original
suspension
of
tick
(isolate
TIK/82),
with
virus
isolated
at
Pirbright
from
pig
bone
marrow
cells
infected
with
tick
suspension
(LIL
18/2,
LIL
20/1,
LIL
31/20),
with
tissues
from
a
pig
inoculated
with
tick
suspension
(KON/83),
or
with
virus
isolated
at
CVL,
Lilongwe
from
a
pig
which
died
of
ASF
(BONGERA/83).
Between
two
and
four
pigs
were
used
for
each
isolate,
and
were
infected
by
the
inoculation
of
between
10
06
and
10
36
HAD
50
of
ASF
virus
by
the
intramuscular
route
or
by
keeping
them
in
contact
with
pigs
infected
by
inoculation
and
observed
until
death.
Blood
samples
were
taken
at
intervals
and
viraemia
measured
using
pig
bone
marrow
cells.
RESULTS
Virus
isolation
from
ticks
A
total
of
17
405
ticks
consisting
of
8335
from pig
kholas,
7670
from
houses
and
1400
from
a
single
warthog
habitat,
were
collected
in
Malawi
and
tested
at
Pirbright
for
the
presence
of
ASF
virus.
Of
these
7183
three
were
from
pig
kholas
and
5146
were
from
houses
within
the
ASF
enzootic
area
(Fig.
1).
Virus
assays
were
carried
out
on
a
total
of
5581
ticks
from
pig
kholas
and
3034
ticks
from
houses
in
Mchinji
district,
which
is
within
the
enzootic
area.
Virus
was
found
in
181
pools
from
the
pig
kholas
and
48
pools
from
the
houses.
No
virus
was
isolated
from
ticks
collected
in
pig
kholas
or
houses
in
any
other
district
or
from
the
ticks
from
the
culvert
used
by
warthog
in
Liwonde
National
Park
in
Machinga
district
(Tables
1
and
2,
Figs.
1
and
2).
Enzootic
area
(1)
Mchinji
district
(a)
Chalaswa
area.
The
vast
majority
of
infected
ticks
were
collected
from
three
closely
associated
villages,
Chalaswa,
Guluza
and
Kamende,
in
the
south-eastern
178
J.
M.
HARESNAPE,
P.
J.
WILKINSON
AND
P.
S.
MELLOR
Table
1.
0.
moubata
collection
sites
within
the
ASF
enzootic
area
and
results
of
ASF
virus
isolation
tests
No.
of
pools
positive
Map
Date
Total
no.
of
ticks
tested
A
District
Locality
square
collected
Pig
khola
House
Ntchisi
Malomo
1333B2
Nov.
84—Sep.
85
0/565
0/1348
Khuuwi
1333B4
Nov.
1984
0/9
Total
0/565
0/1357
Lilongwe
Mkoko
1333C4
Jun.
83—Dec.
84
0/153
Kamandauna
1433A2
Apr.
1983
0/3
Chilinda
1433B1
Jan—Jun.
84
0/1037
0/599
Total
0/1037
0/755
Mchinji
Molosiyo
1332D2
Mar.
82,
Nov.
83
0/23
0/117
Ndawambe
1332D2
Mar.
1982
1/323
May.
83,
Nov.
83
0/83
0/314
Sitolo
1332D2
Mar.
82,
Nov.
83
0/84
0/51
Mkanda
1332D2
Feb.
85,
May.
85
0/38
0/45
Tikoliwe
1332D4
Apr.
1982
1/33
May
83—May
84
0/81
0/178
Kondoole
II
1332D4
June
1983
1/8
0/189
Nov.
83—May
84
0/462
0/63
Kondoole
I
1332D4
Apr.—May
84
0/304
0/47
Menyani
1332D4
May
82—Oct.
83
0/37
0/109
Chamani
1433A2
June
1983
0/30
0/3
Chalaswa
1433A2
July
1983
0/297
Nov.
1983
141/1190
31/255
Jan.
1984
13/114
10/96
Mar.
1984
16/432
5/133
Apr.
1984
6/295
1/124
July
1984
2/217
1/89
Aug.
1984
0/424
0/377
Nov.
1984
0/604
0/228
Sep.
1985
0/502
0/616
Total
181/5581
48/3034
Find
Total
181/7183
48/5146
part
of
Mchinji
district
close
to
the
Mozambique
border,
hereafter
referred
to
as
Chalaswa
(Table
1,
Figs.
1
and
2).
These
villages
were
affected
by
an
ASF
outbreak
early
in
November
1983
in
which
58
pigs
died
and
the
estimated
mortality
in
infected
pigs
was
77-92
%
(Haresnape,
Lungu
&
Mamu,
1985).
Tick
collections
were
made
before
the
outbreak,
3
weeks
following
the
outbreak,
and
at
intervals
afterwards.
Those
collected
in
November
1983
were
assayed
in
pools
of
between
1
and
22
ticks,
and
141
out
of
1190
pools
of
ticks
from
pig
kholas
and
31
out
of
255
pools
of
ticks
from
houses
contained
virus
(Table
1).
The
actual
number
of
infected
ticks
was
undoubtedly
much
larger
.
than
this
but
was
not
ascertained
exactly.
Of
the
11
kholas
in
which
ASF-infected
ticks
were
found,
7
had
suffered
losses
from
the
disease.
The
proportion
of
infected
ticks
from
houses
in Chalaswa
was
not
significantly
different
from
that
in
pig
kholas
at
any
time
during
the
study.
The
overall
proportion
of
adult
females,
adult
males,
larger
nymphae
and
smaller
ASF
virus
infected
0.
moubata
in
Malawi
179
Table
2.
0.
moubata
collection
sites
outside
the
ASF
enzootic
area
and
results
of
ASF
virus
isolation
tests
No.
positive/No.
tested
A
Map
Date
Pig
Warthog
District
Locality
square
collected
khola
House
habitat
Mzimba
Luzi
1033D4
Sep.
1983
0/17
Various*
1133
Sep.—Nov.
85
0/270
Various*
1233
Oct.
1983
0/197
Kasungu
Various*
1233
Aug.
83—May
85
0/152
0/300
Ntchisi
Chintembwe
1333B4
Apr.—May
83
0/727
Dowa
Mponela*
1333D
Sep.
83—Aug.
85
0/692
0/481
Chimangamasa*1334C
Aug.
83,
Nov.
85
0/34
Lilongwe
Jumpha*
1333D4
Aug.
83,
Aug.
85
0/241
0/364
Chinsapo
1433B1
Oct.
1983
0/5
Dedza
Nyama
1434A2
June—Aug.
83
0/129
Machinga
Liwonde
1535A2
July
1985
0/1200
Nov.
1985
0/200
Mulanje
Mpasa
1535D3
May
1985
0/67
Total
0/1152
0/2524
0/1400
*
Collections
made
over
a
wide
area
covering
several
localities
(see
Figs.
1
and
2).
nymphae
infected
were
not
significantly
different.
The
proportion
of
infected
ticks
gradually
decreased
following
the
outbreak
and
virus-infected
ticks
were
still
present
in
July
1984,
8
months
after
the
outbreak.
The
virus
titres
in
the
three
single
infected
ticks
collected
on
this
occasion
were
high
(1046,
10
4.6
,
10
31
HAD50
per
tick)
although
no
deaths
from
ASF
had
occurred
in
Chalaswa
after
November
1983.
ASF
isolates
LIL
18/2,
LIL
20/1
and
LIL
31/20
were
made
in
pig
bone
marrow
cultures
from
pools
of
3
adult
females,
4
adult
males
and
5
adult
males
respectively,
collected
from
pig
kholas
in
Chalaswa
in
November
1983.
(b)
Ndawambe.
Ndawambe
village
is
in
the
western
part
of
Mchinji
district
about
1
km
from
the
Mozambique
border.
Three
hundred
and
twenty
three
ticks
were
collected
from
11
pig
kholas
in
March
1982,
and
ASF
virus
was
isolated
from
a
single
adult
tick
in
a
batch
of
82
adults
and
29
nymphae
collected
in
one
khola
containing
20
pigs.
The
titre
of
virus
in
the
tick
was
not
measured.
There
had
been
suspected
cases
of
ASF
in
the
village
in
January
and
February
1982,
just
prior
to
the
time
of
collection,
but
these
20
pigs
were
apparently
unaffected.
More
cases
of
ASF
were
reported
between
October
1982
and
January
1983
and
a
further
collection
of
ticks
including
67
from
this
khola,
was
made
in
May
1983
but
all
were
negative.
(c)
Tikoliwe.
Tikoliwe
village
is
also
in
western
Mchinji
district
close
to
the
Mozambique
border.
Thirty-three
ticks
were
collected
from
two
pigs
kholas
on
29
April
1982,
and
ASF
virus
was
isolated
from
one
adult
female
of
31
ticks
from
one
khola.
The
titre
of
virus
in
this
tick
was
10"
HAD
50
(isolate
TIK/82).
There
had
been
an
outbreak
of
ASF
in
March
1982
just
prior
to
the
time
of
collection
but
the
four
pigs
in
the
khola
were
apparently
unaffected.
Further
collections
of
ticks
made
in
May
1983,
November
1983
and
May
1984
were
not
infected
with
virus
(Table
1).
(d)
Kondoole.
Kondoole
consists
of
two
villages,
Kondoole
I
and
Kondoole
II,
180
J.
M.
HARESNAPE,
P.
J.
WILKINSON
AND
P.
S.
MELLOR
situated
0.5
km
apart
between
Ndawambe
and
Tikoliwe.
Both
villages
had
been
affected
by
ASF
in
early
June
1983
and
10
out
of
15
pigs
died
of
the
disease
in
two
kholas
in
Kondoole
II.
Eight
ticks
were
collected
from
one
khola
in
Kondoole
II
on
10
June
1983,
and
ASF
virus
was
isolated
from
one
N4/N5
nymph.
The
virus
titre
in
the
tick
was
not
measured
but
an
isolate
was
made
following
passage
in
a
pig
(KON/83).
The
five
surviving
pigs
were
still
alive
and
apparently
healthy
in
November
1983.
The
estimated
percentage
mortality
in
infected
pigs
in
Kondoole
II
was
thus
67
%.
In
November
1983
a
further
383
ticks
were
collected
from
five
kholas
in
Kondoole
II,
including
186
from
a
khola
in
which
pigs
had
died,
but
no
virus
was
isolated
from
any
of
them.
Further
losses
from
ASF
occurred
in
both
Kondoole
I
and
Kondoole
II
in
March
and
April
1984,
but
no
virus
was
isolated
from
a
further
383
ticks
collected
in
April
and
May
from
12
kholas
which
included
4
of
those
affected
by
ASF.
No
samples
were
sent
to
CVL
from
pigs
in
any
of
the
villages
in
which
infected
ticks
were
found,
so
laboratory
confirmation
of
ASF
as
the
cause
of
death
of
pigs
in
these
villages
was
not
obtained,
but
pig
owners
variously
described
symptoms
consistent
with
ASF
such
as
lack
of
appetite,
immobility,
weakness,
refusal
to
walk,
loss
of
balance,
huddling
together,
coughing,
shivering,
diarrhoea
and
loss
of
body
condition
with
death
following
3-7
days
after
onset
of
sickness.
Some
internal
organs,
particularly
the
spleen,
were
reported
to
be
unusually
dark
in
colour.
However,
laboratory
confirmation
was
obtained
from
an
ASF
outbreak
which
occurred
in
September
1983
in
Bongera,
which
is
also
in
Mchinji
district,
situated
approximately
halfway
between
Chalaswa
and
Ndawambe
and
about
40
km
from
each
(Haresnape,
Lungu
&
Mamu,
1985).
An
isolate
from
this
outbreak
was
made
in
pig
leucocyte
cultures
at
CVL,
Lilongwe
(BONGERA/
83).
(2)
Other
districts
Although
no
virus
infected
ticks
were
found
in
other
districts
(Table
1),
it
is
interesting
to
note
that
ASF
outbreaks
had
occurred
in
some
villages
prior
to
the
time
of
collection.
For
example
ASF
outbreaks
occurred
between
August
and
September
1983
in
Chilinda
and
neighbouring
villages
in
Lilongwe
district,
and
in
September
1984
in
Malomo
in
Ntchisi
district
(Haresnape,
Lungu
&
Mamu,
1985).
Substantial
collections
of
ticks
from
affected
pig
kholas
were
made
in
January,
April
and
June
1984
in
the
Chilinda
area,
and
in
November
and
December
1984
and
January
1985
from
Malomo,
but
none
were
infected.
These
parts
of
Lilongwe
and
Ntchisi
districts
are
within
the
ASF
enzootic
area
(Haresnape,
Lungu
&
Mamu,
1985).
Non-enzootic
area
Collections
of
ticks
were
also
made
from
areas
outside
the
enzootic
area
in
which
ASF
outbreaks
had
occurred
(Table
2),
such
as
Mwera
Hills
in
Ntchisi
district,
Dzoole
in
Dowa
district
and
Chikuse
in
Lilongwe
district,
but
these
collections
were
all
from
houses
no
ticks
were
found
in
pig
kholas
in
these
localities
despite
thorough
searches
(Haresnape
&
Mamu,
1986).
Most
of
the
other
localities
from
which
the
ticks
originated
were
well
outside
the
ASF
enzootic
area
(Table
2),
and
in
parts
of
Malawi
where
ASF
has
never
been
reported
(Mzimba,
northern
Kasungu,
north-eastern
Dedza
and
Machinga
districts).
Virus
isolate
TIK/82
(tick
isolate)
BON/83
(pig
tissue)
KON/83
(tick
isolate)
LIL
18/2
(tick
isolate)
LIL
20/1
(tick
isolate)
Route
of
infection
(titre
of
inoculum)*
Intramuscular
(10
3-3
)
Contact
Intramuscular
(10
3-3
)
Intramuscular
(10
36
)
Contact
Intramuscular
(10
16
)
Contact
Intramuscular
(10
1.8
)
LIL
31/20
Intramuscular
(tick
isolate)
(10
0
'
6
)
Contact
ASF
virus
infected
0.
moubata
in
Malawi
181
Table
3.
Virulence
of
ASF
virus
isolates
from
the
enzootic
area
of
Malawi
in
European
pigs
No.
of
days
to
:
Virus
titre
A
(log
in
HAD„)
Onset
of
fever
Death
Pig
no.
(days
after
infection)
Blood
Spleen
OP
38
3
9
NT
8.3
OP
39
3
7
8.3
8.0
OP
40
9
dpet
13
dpe
7.5
7.3
PA
10
4
7
7.3
NT
PA
11
4
7
7.3
NT
PA
39
3
7
7.2
8.8
PA
40
6
dpe
13
dpe
8.0
8.1
PA
43
4
9
8.2
8.3
PA
44
12
dpe
14
dpe
8.0
8.0
PB
40
4
7
8.0
9.1
PB
41
4
8
7.6
7.8
PB
42
3
8
7.3
9.3
PB
43
3
7
9.5
9.8
PA
41
4
9
8.0
7.6
PA
42
13
dpe
14
dpe
7.0
8.3
*
HAD
50
.
t
dpe,
Days
post
exposure.
NT,
not
tested.
ASF
virus
was
not
isolated
from
any
ticks
collected
in
the
non-enzootic
areas.
Virulence
tests
Five
isolates
from
ticks
(KON/83,
TIK/82,
LIL
18/2,
LIL
20/1
and
LIL
31/
20)
and
one
isolate
from
a
pig
(BONGERA/83)
were
obtained
from
Mchinji
district
and
were
all
found
to
be
very
virulent
in
European
cross-bred
pigs.
The
incubation
period
was
3-4
days
in
all
the
inoculated
pigs
which
died
between
7
and
9
days
after
infection.
The
pigs
infected
by
contact
had
a
longer
incubation
period
(9-13
days)
and
died
13-14
days
after
exposure
(Table
3).
All
pigs
had
high
titres
of
virus
in
the
blood
and
tissues
(tonsil,
parotid
and
gastrohepatic
lymph
nodes,
kidney,
spleen
and
heart).
DISCUSSION
ASF
virus
infected
ticks
were
found
in
pig
kholas
in
Mchinji
district
which
is
in
the
centre
of
a
large
ASF
enzootic
area.
In
all
villages
where
virus
infected
ticks
were
found,
deaths
of
pigs
from
ASF
had
been
reported
shortly
beforehand
and
in
some
villages,
such
as
Kondoole
and
possibly
also
Ndawambe
and
Tikoliwe,
deaths
from
ASF
also
occurred
in
kholas
after
infected
ticks
were
collected
from
them,
possibly
as
the
result
of
a
bite
from
an
infected
tick.
High
titres
of
ASF
virus
were
found
in
some
ticks
notably
that
from
Tikoliwe
(10
6.4
HAD
50
),
collected
on
29
April
1982,
which
compares
with
the
peak
virus
titre
182
J.
M.
HARESNAPE,
P.
J.
WILKINSON
AND
P.
S.
MELLOR
of
10
6
*
3
HAD
50
recorded
by
Greig
(1972)
from
a
tick
approximately
100
days
after
infection.
In
Chalaswa,
the
mean
titre
of
virus
in
infected
ticks
was
at
its
highest
about
8
months
after
the
ASF
outbreak
in
November
1983
which
demonstrates
that
virus
had
replicated
in
the
ticks
in
the
field
situation.
A
new
outbreak
of
ASF
could
start
following
a
bite
from
an
infected
tick
even
many
months
after
the
last
cases
of
ASF.
Thus
domestic
pig
ticks
of
the
Ornithodoros
moubata
complex
are
important
reservoirs
of
ASF
virus
and
vectors
of
disease
in
rural
Malawi.
In
Chalaswa,
it
is
perhaps
surprising
that
no
further
deaths
from
ASF
occurred
after
the
loss
of
58
pigs
in
November
1983,
despite
the
presence
of
large
numbers
of
infected
ticks
in
the
kholas
and
it
seems
unlikely
that
the
pigs
in
infected
kholas
escaped
being
bitten
by
them.
The
six
Malawi
isolates
of
ASF
virus
from
ticks
in
the
enzootic
area
all
produced
acute,
fatal
ASF
in
European
pigs
but
the
mortality
rate
in
ASF
outbreaks
in
indigenous
village
pigs
in
Mchinji
district
is
often
much
lower
than
100
%
(Haresnape,
Lungu
&
Mamu,
1985).
One
explanation
for
this
difference
is
that
field
isolates
are
less
virulent
for
indigenous
pigs
in
Malawi
than
they
are
for
European
breeds
of
pig.
Recovered
pigs
may
be
resistant
to
experimental
challenge
with
homologus
virus
although
this
virus
may
replicate
without
producing
clinical
signs
(Wilkinson,
1984).
A
similar
situation
may
also
occur
in
the
field
in
Malawi
where
recovered
pigs
could
become
infected
under
natural
conditions
with
local
virus
strains
which
replicate
and
produce
mild
or
unnoticed
clinical
signs.
The
interaction
between
recovered
pigs
and
infected
ticks
may
thus
be
important
in
the
epizootiology
of
ASF
in
the
enzootic
area.
The
overall
infection
rate
in
ticks
collected
from
pig
kholas
in
the
Mchinji
district
was
approximately
3
%
(Table
1).
However,
in
Chalaswa,
11
%
of
the
ticks
collected
from
pig
kholas
during
the
3
months
following
the
outbreak
of
ASF
were
infected
and
the
high
infection
rate
in
ticks
collected
in
November
1983
was
presumably
a
result
of
their
having
fed
on
viraemic
pigs
in
early
November.
By
July
to
August
1984,
the
infection
rate
was
0.3
%,
which
is
similar
to
the
overall
infection
rate
elsewhere
in
Mchinji
district.
The
estimated
tick
infection
rates
were
1
%
(1
in
102),
3
%
(1
in
31)
and
12
%
(1
in
8)
respectively,
in
kholas
in
Ndawambe,
Tikoliwe
and
Kondoole
in
each
of
which
one
infected
tick
was
found,
although
these
cannot
be
regarded
as
accurate
estimates
because
of
the
small
sample
sizes.
The
collections
from
pig
kholas
in
localities
in
Ntchisi
and
Lilongwe
districts
which
are
within
the
ASF
enzootic
area
and
had
recent
cases
of
ASF
(Table
1)
may
have
been
too
small
to
detect
virus-infected
ticks.
It
is
somewhat
surprising
that
in
Chalaswa
equally
high
infection
rates
were
found
in
ticks
collected
from
houses
or
from
pig
kholas.
The
common
practice
in
Malawi
of
airing
blankets
by
spreading
them
over
pig
kholas
could
be
the
explanation
for
the
movement
of
ticks
between
pig
sties
and
human
dwellings.
This
seems
a
more
likely
route
of
transfer
than
on
people,
animals
and
poultry,
as
the
ticks
are
generally
only
found
on
their
hosts
at
night
(Haresnape
&
Mamu,
1986).
No
infected
ticks
were
found
in
houses
elsewhere,
even
though
3096
ticks
from
houses
within
the
enzootic
area
was
tested.
In
most
areas
where
ticks
were
found
in
kholas
they
were
easier
to
find,
and
therefore
presumably
more
numerous,
in
houses
(Haresnape
&
Mamu,
1986)
and
Chalaswa
was
the
only
area
where
they
appeared
more
numerous
in
kholas.
ASF
virus
infected
0.
moubata
in
Malawi
183
The
ASF
virus
infection
rates
reported
here
for
naturally
infected
ticks
collected
from
domestic
pigs
compare
with
infection
rates
in
0.
moubata
from
warthog
burrows
of
between
0.017
and
1.35
%
in
different
localities
in
East
Africa
(Plowright,
1977
a,
b)
and
between
0.06
and
1.4
%
in
South
Africa
and
Namibia
(Thomson
et
al.
1983).
In
ticks
collected
from
warthog
burrows
the
infection
rate
was
higher
in
adults
than
in
nymphae,
but
this
was
not
the
case
in
the
large
numbers
of
infected
ticks
found
in
Chalaswa
immediately
following
the
cases
of
ASF
there.
If
a
larger
number
of
persistently
infected
ticks
had
been
found,
the
same
trend
might
have
been
observed.
While
warthogs
and
ticks
in
warthog
burrows
must
have
been
the
original
hosts
of
ASF
virus
before
domestic
pigs
were
introduced
to
Africa,
they
may
play
only
a
minor
role
in
the
epizootiology
in
some
parts
of
Africa.
0.
moubata,
being
a
multiple
host
tick,
is
ideally
suited
as
a
potential
vector
of
ASF
virus
(Heuschele
&
Coggins,
1965)
and
it
seems
likely
that
in
Malawi
ticks
from
domestic
pig
premises
may
be
important
reservoirs
of
the
virus.
Viraemias
of
10'
or
10
8
HAD
50
/m1
are
very
common
in
domestic
pigs
shortly
before
death
from
ASF,
and
these
are
the
likely
source
of
virus
for
the
persistently
infected
ticks
in
Malawi.
Plowright,
Parker
&
Peirce
(1969
b)
found
that
the
Tengani
strain
of
ASF
virus,
which
originated
from
Nsanje
district
in
the
Southern
Region
of
Malawi
(Matson,
1960
;
Cox
&
Hess,
1962)
caused
persistent
infection
in
only
about
5
%
of
East
African
ticks
fed
on
viraemic
pigs
and
this
might
also
be
true
of
other
Malawi
strains
of
ASF
virus
in
Malawi
ticks.
This
would
then
explain
the
comparatively
low
numbers
of
persistently
infected
ticks,
as
was
found
in
Chalaswa
8
months
after
the
outbreak
of
ASF,
even
though
large
numbers
of
ticks
were
infected
immediately
after
the
outbreak
of
ASF.
In
Malawi,
ASF
is
enzootic
over
an
area
of
approximately
8000
km
2
in
the
Central
Region,
but
there
is
virtually
no
contact
between
domestic
pigs
and
warthogs
in
this
area
as
warthogs
are
largely
confined
to
the
National
Parks
and
Game
Reserves
(Hough,
1982),
where
there
are
no
domestic
pigs.
Although
it
has
been
pointed
out
that
the
distribution
of
ASF
in
Africa
as
a
whole
closely
resembles
the
distribution
of
warthogs
(Anon,
1962),
this
is
not
the
case
in
Malawi.
The
warthog
habitat
in
which
0.
moubata
were
found
during
this
study
was
in
Liwonde
National
Park
in
the
Southern
Region
of
Malawi,
which
is
well
outside
the
area
in
which
ASF
is
enzootic
for
domestic
pigs,
and
none
of
the
ticks
collected
there
were
infected,
although
further
south
in
Lengwe
National
Park
in
Chikwawa
district,
five
of
six
warthog
sera
tested
were
seropositive
(Haresnape,
Lungu
&
Mamu,
1985).
Domestic
pigs
come
into
contact
with
bush
pigs
much
more
frequently,
as
bush
pigs
are
widely
distributed
throughout
Malawi
(Hough,
1982).
No
evidence
was
found
of
ASF
infection
in
11
bush
pigs
examined
(Haresnape,
Lungu
&
Mamu,
1985)
and
a
brief
search
for
ticks
in
bush-pig
sleeping
places
in
Lilongwe
district
was
unsuccessful.
Whether
or
not
they
play
a
role
in
the
epizootiology
of
ASF
in
Malawi
is
therefore
not
clear.
The
findings
presented
here
show
that
control
of
ASF
will
be
more
difficult
than
previously
envisaged,
because
of
the
important
role
played
by
ticks
feeding
on
domestic
pigs
as
reservoirs
of
virus,
because
these
ticks are
extremely
difficult
to
eradicate.
Acaricides
can
reduce
the
tick
population
but
cannot
eliminate
it
entirely
as
ticks
can
hide
deep
in
cracks
in
the
wood
or
mud
of
kholas
and
houses
184
J.
M.
HARESNAPE,
P.
J.
WILKINSON
AND
P.
S.
MELLOR
which
are
inaccessible.
Evacuation
of
infested
premises
is
not
likely
to
be
effective
as
0.
moubata
is
extremely
resistant
to
starvation
and
some
ticks
of
the
0.
moubata
complex
may
survive
for
5
years
without
food
(Walton,
1964).
Burning
infested
premises,
although
possible
for
pig
kholas,
would
clearly
not
be
acceptable
for
houses
and,
since
the
ticks
in
houses
may
be
infected
and
could
reinfest
new
pig
kholas,
this
would
not
be
an
effective
means
of
eradication.
These
ticks
occur
over
a
much
wider
area
than
the
present
ASF
enzootic
area
and
there
is
a
danger
that
the
enzootic
area
could
become
larger
in
future
(Haresnape
&
Mamu,
1986
;
Haresnape,
Lungu
&
Mamu,
1987).
It
is
difficult
to
see
any
way
in
which
ASF
could
be
eradicated
altogether
but
it
is
possible
that
it
could
be
prevented
from
becoming
enzootic
over
a
wider
area
through
imposition
of
strict
control
of
pig
movements
from
the
enzootic
area
and
special
efforts
to
detect
the
spread
of
ASF
outbreaks
in
those
areas
outside
the
enzootic
area
where
0.
moubata
occur.
At
particular
risk
are
those
parts
of
Dowa,
Lilongwe
and
Kasungu
districts
which
are
at
present
outside
the
ASF
enzootic
area,
but
where
0.
moubata
are
common
in
pig
kholas.
Burning
affected
kholas
might
be
advisable
in
the
event
of
an
ASF
outbreak
in
these
areas
in
order
to
prevent
the
virus
from
becoming
established
in
the
0.
moubata
population.
We
would
like
to
thank
the
Malawi
Secretary
for
Agriculture
for
permission
to
submit
this
work
for
publication,
the
Malawi
Chief
Veterinary
Officer
and
his
field
and
laboratory
staff
for
their
support,
and
the
Department
of
Parks
and
Wildlife
for
their
co-operation.
Particular
thanks
to
Mr
S.
Lungu
and
Mr
F.
Mamu
of
the
Central
Veterinary
Laboratory,
Lilongwe,
Mr
G.
Hutchings
and
Mr
S.
Williams,
Institute
for
Animal
Health,
Pirbright
Laboratory,
Pirbright,
Woking,
for
technical
assistance
and
Mrs
Beryl
Hutchinson
for
typing
the
manuscript.
O.D.A.
(UK)
kindly
provided
financial
support
for
Janet
M.
Haresnape
during
the
preparation
of
the
manuscript.
REFERENCES
ANON
(1962).
The
geographic
distribution
of
the
warthog
and
domestic
pigs
in
Africa.
Bulletin
of
Epizootic
Diseases
of
Africa
10,
91-92.
Cox,
B.
F.
&
HESS,
W.
R.
(1962).
Note
on
an
African
swine
fever
investigation
in
Nyasaland.
Bulletin
of
Epizootic
Diseases
of
Africa
10,
439-440.
DETRAY,
D.
E.
(1963).
African
swine
fever.
Advances
in
Veterinary
Science
8,
299-333.
GREIG,
A.
(1972).
The
localization
of
African
swine
fever
virus
in
the
tick
Ornithodoros
moubata
porcinus.
Archiv
far
die
gesamte
Virusforschung
39,
240-247.
HAMMOND,
R.
A.
&
DETRAY,
D.
E.
(1955).
A
recent
case
of
African
swine
fever
in
Kenya,
East
Africa.
Journal
of
the
American
Veterinary
Medicine
Association
126,
389-391.
HARESNAPE,
J.
M.
(1984).
African
swine
fever
in
Malawi.
Tropical
Animal
Health
and
Production
16,
123-125.
-
HARESNAPE,
J.
M.,
LUNGU,
S.
A.
M.
&
MAMU,
F.
D.
(1985).
A
four-year
survey
of
African
swine
fever
in
Malawi.
Journal
of
Hygiene
95,
309-323.
-
HARESNAPE,
J.
M.
&
MAMU,
F.
D.
(1986).
The
distribution
of
ticks
of
the
Ornithodoros
moubata
complex
(Ixodoidea
:
Argasidae)
in
Malawi
and
its
relation
to
African
swine
fever
epizootiology.
Journal
of
Hygiene
96,
535-544.
-
HARESNAPE,
H.
M.,
LUNGU,
S.
A.
M.
&
MAMU,
F.
D.
(1987).
An
updated
survey
of
African
swine
fever
in
Malawi.
Epidemiology
and
Infection
99,
723-732.
HEUSCHELE,
W.
P.
&
COGGINS,
L.
(1965).
Studies
on
the
transmission
of
African
swine
fever
virus
by
arthropods.
Proceedings
of
the
United
States
Livestock
Sanitary
Association
60,
94-100.
ASF
virus
infected
0.
moubata
in
Malawi
185
HEUSCHELE,
W.
P.
&
COGGINS,
L.
(1969).
Epizootiology
of
African
swine
fever
in
warthogs.
Bulletin
of
Epizootic
Diseases
of
Africa
17,
179-183.
HOOGSTRAAL,
H.
(1956).
African
Ixodidea.
I.
Ticks
of
the
Sudan.
United
States
Naval
Medical
Research
Report
N.M.
005.050.29.07,pp.
121-190.
HOUGH,
J.
(1982).
Mammals
of
Malawi,
pp.
27,30.
Lilongwe
:
Environmental
Unit,
Department
of
National
Parks
and
Wildlife.
MATSON,
B.
A.
(1960).
An
outbreak
of
African
swine
fever
in
Nyasaland.
Bulletin
of
Epizootic
Diseases
of
Africa
8,
305-308.
MONTGOMERY,
R.
E.
(1921).
On
a
form
of
swine
fever
occurring
in
British
East
Africa
(Kenya
Colony).
Journal
of
Comparative
Pathology
34,
159-191
and
243-262.
PLOWRIGHT,
W.
(1977a).
Vector
transmission
of
African
swine
fever
virus.
In
Hog
Cholera/
Classical
Swine
Fever
and
African
Swine
Fever.
EEC
publication
EUR
5904
EN,
pp.
575-587.
PLOWRIGHT,
W.
(1977
b).
African
swine
fever.
In
Inf
ectious
Diseases
of
Wild
Mammals,
pp.
178-190.
Ames,
Iowa,
USA
:
Iowa
State
University
Press.
PLOWRIGHT,
W.,
PARKER,
J.
&
PEIRCE,
M.
A.
(1969a).
African
swine
fever
virus
in
ticks
(Ornithodoros
moubata.
Murray)
collected
from
animal
burrows
in
Tanzania.
Nature
221,
1071-1073.
PLOWRIGHT,
W.,
PARKER,
J.
&
PEIRCE,
M.
A.
(1969
b).
The
epizootiology
of
African
swine
fever
in
Africa.
Veterinary
Record
85,
668-674.
PLOWRIGHT,
W.,
PERRY,
C.
T.
&
GREIG,
A.
(1974).
Sexual
transmission
of
African
swine
fever
virus
in
the
tick
Ornithodoros
moubata
porcinus,
Walton.
Research
in
Veterinary
Science
17,
106-113.
PLOWRIGHT,
W.,
PERRY,
C.
T.
&
PEIRCE,
M.
A.
(1970).
Transovarial
infection
with
African
swine
fever
virus
in
the
argasid
tick,
Ornithodoros
moubata
porcinus,
Walton.
Research
in
Veterinary
Science
11,582-584.
PLOWRIGHT,
W.,
PERRY,
C.
T.,
PEIRCE,
M.
A.
&
PARKER,
J.
(1970).
Experimental
infection
of
the
argasid
tick,
Ornithodoros
moubata
porcinus,
with
African
swine
fever
virus.
Archly
fiir
die
gesamte
V
irusforschung
31,
33-50.
SCOTT,
G.
R.
(1965).
The
virus
of
African
swine
fever
and
its
transmission.
Bulletin
de
l'Office
International
des
Epizooties
63,
645-677.
STEYN,
D.
G.
(1932).
East
African
virus
disease
in
pigs.
Eighteenth
Report
by
the
Director
of
Veterinary
Services
and
Animal
Industry,
vol.
1,
pp.
99-109.
Onderstepoort
(South
Africa).
THOMAS,
A.
D.
&
KOLBE,
F.
F.
(1942).
The
wild
pigs
of
South
Africa
:
Their
distribution
and
habits,
and
their
significance
as
agricultural
pests
and
carriers
of
disease.
Journal
of
the
South
African
Veterinary
Medical
Association
13,
1
-
11.
THOMSON,
G.,
GAINARU,
M.,
LEWIS,
A.,
BIGGS,
H.,
NEVILLE,
E.,
VAN
DER
PYPEKAMP,
H.,
GERBER,
L.,
ESTERHUYSEN,
J.,
BENGIS,
R.,
BEZUIDENHOUT,
D.
&
CONDY,
J.
(1983).
The
relationship
between
African
swine
fever
virus,
the
warthog
and
Ornithodoros
species
in
South
Africa.
In
African
Swine
Fever,
CEC/FAO
Research
Seminar,
Sardinia,
1981
(ed.
P.
J.
Wilkinson),
pp.
85-100.
CEC
Publication
EUR
8466
EN.
WALTON,
G.
A.
(1964).
The
Ornithodoros
moubata
group
of
ticks
in
Africa.
Control
problems
and
implications.
Journal
of
Medical
Entomology
1,
43-64.
WILKINSON,
P.
J.,
DONALDSON,
A.
I.,
GREIG,
A.
&
BRUCE,
W.
(1977).
Transmission
studies
with
African
swine
fever
virus.
Infection
of
pigs
by
airborne
virus.
Journal
of
Comparative
Pathology
87,
475-495.
WILKINSON,
P.
J.
(1984).
The
persistence
of
African
swine
fever
in
Africa
and
the
Mediterranean.
Preventive
Veterinary
Medicine
2,
71-82.