THE FIRST YEAR OF OBLIGATORY BVD
CONTROL IN GERMANY –
DIAGNOSTIC STRATEGIES, RESULTS AND
EXPERIENCES
Horst Schirrmeier, Günter Strebelow, Martin Beer
2nd Congress of the Eurorean Association of Veterinary Laboratory Diagnosticians (EAVLD),
Kazimierz Dolny, Poland , 1-4 July 2012
Institute of Diagnostic Virology
Bovine Virusdiarrhoe/Mucosal Disease
BDV
BDV
Gifhorn
Moredun
Virus: 40-60 nm small, unveloped
pestivirus, with a single strained RNA
with positive polarity of 12,5 kbp in size.
CSFV
CSFV
Reindeer-1
CSFV 39
99
100
• 2 biotypes (cp, ncp)
100
• 2 genotypes with numerous subtypes
Brescia
Alfort 187
100
99
HoBi
Giraffe
Giraffe
H. GranzowGiraffe-1
90
Further members of the Genus:
100
• Classical swine fever virus (CSFV)
• Border disease virus (BDV 1 bis 4 (7))
100
93
US890
Osloss
Giessen-3
• atypical pestiviruses (Giraffe, Hobi
Pronghorn, Bungowannah)
NADL
BVDV-1
BVDV-1
0.1
PT810
BVDV-2
BVDV-2
Acute infection of non pregnant seronegative animals
transient viremia
scarely clinical signs
seroconversion
special form: severe
haemorrahagic disease
– unclear pathogenesis
infection of pregnant seronegative cattle
•transient viremic dam
• seroconversion
•infection of fetus
•abort
•PI (ag- pos, abnegativ)
•normal (ag-negativ,
ab-positiv
1st stage of gestation
fetus retention
2nd. stage of gestation
(30-90 d)
re-heat
Abort
Reproduction
failure
PI-animal
3th stage of gestation
immune competent calves
(congenital damages, abort,
stillbirth)
Control target
Persistent infected animal (pi-animal)
wasting animals („Kümmerer“ ) (50 %) or normal development
livelong virus shedding (virus perpetutation, reinfection of herds)
Hiding of viremia by maternal antibodies („Diagnostic gap“ !!)
Biggest challenge of BVDV diagnosis - shortening of
the diagnostic gap
Notifiable disease since November 2004
Voluntary control programs in some federal states basing on a
federal guidline,
Case definition:
detection of a persistent viremic animal
diagnosis: Mucosal disease (with or without virus detection)
Persistent viremic animal
two times ag/genome-positive with methods according to
officially approved methods (interval depends on methods, 3-12
weeks)
ones positive without second checkup
(NRL recommendation; pi-cut off for PCR: > 104 genome copies/well)
BVDV control strategy in Germany
1. Legal basics:
•
Official Federal BVD regulation 2008 (BVD-Verordnung), in force
since January 2011
•
Collection of approved methods (prescribed)
•
Reduction of PI-prevalence
•
Establishing of unsuspicious (virus free) animals and farms
(certified)
•
Obligatory investigation of all calves up to the 6th month of life for
BVDV antigen/genome)
•
Elimination of pi-animals (immediately)
•
Only trade with certifed unsuspious animals
•
Prevention of reinfections by qualified measures and vaccination
(voluntarely)
2. Aim:
3. Way:
Obligatory testing of calves for
virus/genome
negative
positive
Consideration to cull
Methods applied depend
on age of animals (diagnostic gap)
or
Retesting (3-12
weeks)
Notified case
positive
cull
negative
Certificate:
BVDV-unsuspicious
animal (livelong status)
Approved methods for virus/genome testing considering
the diagnostic gap
method
sample
Diagnostic gap
E RNS –Ag-capture
ELISA
serum, plasma, EDTA-blood < 60. day
organes, ear notches
no gap
NS3 Ag- capture
ELISA
blood leukocytes
3. -90. day
Fluorescence activated
cell sorting (Facs)
blood leukocytes
3. - 90. day
Virus isolation
blood leukocytes
7. – 40. day
RT-PCR
serum, plasma, EDTAblood, leukocytes,
pooled sample: 7.- 40. Tag
individual sample: no
dagnostic gap
organes, milk, ear notches
no diagnostic gap
Ear notches
- the final solution ??
☺ very early investigation (< 7 day post partum)
☺ clear identification of specimens
☺ simple sampling by the farmer
☺ no relevant influence of maternal antibodies
Qualified trial period in the diagnostic routine is essential !!!
difficulty of retesting (confirmation of positive result, material,
time point, which test, diagnostic gap)
contamination problems (licking one another,) esp. by using PCR
handling problems
fitness for mass investigation
Non PI , transient or
false positive
Ct 35
PI
Ct 25
Approved diagnostics (antigene detection)
Herdchek BVDV Ag/Serum Plus (IDEXX)
serum, plasma, ear notches
ERNS-based, sensitivity better than specificity
SERELISA BVD/MD Antigen, Synbiotics
p80-based, not for serum/plasma and ear notches
suitable for cell cultures and leukocytes
AK-sensitive
BVDV-antigen mix (formerly Pourquier, now IDEXX)
serum, plasma, ear notches, leukocytes
ERNS/p80 based, sensitivity sufficient, specificity exellent
approved, but not available in Germany, unfortunately
PrioCheck BVDV AG PI-focus/plus (Prionics) –
ear notches, antibody control
p80-based, no approved batch on the German market
not state of the art,
SNAP test – failed to pass the licence procedure in Germany
Approved diagnostics (antigene detection)
To be left:
Herdchek BVDV Ag/Serum Plus (IDEXX GmbH)
serum, plasma, ear notches
ERNS-based,
sensitivity better than specificity (sera)
problem arising: low OD values around cut off but high
genome loads in PCR in ear notches- cut off has to be
optimised
Approved diagnostics (genome detection)
ADIAVET®BVD REAL TIME, (ADIAVET BVD/MD) AES Chemunex
BoVir-SL®BVDV TaqMan RT-PCR, AnDiaTec GmbH & Co KG
VIROTYPE®BVDV, Labordiagnostik Leipzig GmbH
Cador BVDV RT-PCR, Qiagen GmbH
Cador BVDV Type 1/2 RT-PCR Kit , Qiagen GmbH
PrioCHECK® BVDV RT-PCR, Prionics Deutschland GmbH
Virella BVD, Gerbion
Approved diagnostics (antibody detection)
Indirect ELISA:
HerdChek BVDV Ab, IDEXX
SVANOVIR BVDV-Ak Screening and conformation,
Svanova
NS3-blocking ELISA:
BVDV p80, Synbiotics
BVD/MD P80 antibody, IDEXX
PrioCheck BVD Ab, Prionics
Gold standard: SNT
Serology in German program: checking the efficacy of PI removal, and not
to detect farms with PI-animals primarely - in contrast to the
Scandinavian model
Supervisor or detective
German pi-statistics 2011
number of detected pi-animals:
17410
percentages rel. to entire cattle population: 0,14 %
percentages rel. to calves born in 2011:
0,36 %
number of farms with at least one pi-animal: 6016
percentages rel. to entire farms:
4,11 %
Monthly distribution of pi-animals in
Germany 2011
2500
2000
1500
1000
2001
1798
1422
1944
1930
1577
1570
1382
1375
1315
500
936
490
De
ze
mb
er
ve
mb
er
No
er
Ok
tob
Se
pt e
mb
er
gu
st
Au
Ju
li
i
Ju
n
Ma
i
ril
Ap
Mä
rz
Fe
br
ua
r
Ja
nu
ar
0
?
?
Monthly distribution of pi-animals in
Germany 2011 /2012
2500
2000
1500
1000
1798
1422 1382
1570
2001 1944 1930
1577
1375
1315
500
1036
936
869
836 734
490
pr
il
A
M
är
z
ug
us
Se
t
pt
em
be
r
O
kt
ob
er
N
ov
em
be
r
D
ez
em
be
r
Ja
nu
ar
Fe
br
ua
r
A
Ju
li
i
Ju
n
M
ai
pr
il
A
M
är
z
Ja
nu
ar
Fe
br
ua
r
0
ai
Ap
ril
är
z
900
800
700
600
Ap
ril
Au
gu
Se
st
pt
em
be
r
O
kt
ob
er
No
ve
m
be
De
r
ze
m
be
r
Ja
nu
a
Fe r
br
ua
r
M
är
z
Ju
li
30
Ju
ni
M
Ap
ril
M
300
Ju
li
Au
gu
Se
st
pt
em
be
r
O
kt
ob
er
No
ve
m
be
De
r
ze
m
be
r
Ja
nu
a
Fe r
br
ua
r
400
Ja
nu
a
Fe r
br
ua
r
M
är
z
ai
Ju
ni
M
Ap
ril
Ja
nu
a
Fe r
br
ua
r
M
är
z
Bavaria
Differences in course
and frequency between
German lander
500
8814
200
100
0
Saxony Anhalt
100
90
80
70
60
50
40
20
250
10
0
Cumulative pi-prevalence in German laender
(percentages relating to the number of birth)
0,70
0,58
0,60
0,54
0,50
0,44
0,39
0,40
0,30
0,35
0,34
0,29
0,26
0,24
0,21
0,20
0,19
0,20
0,16
0,10
0,10
0,02
TH
ST
SN
V
M
BB
Be
rli
n
SL
BY
BW
RP
HE
NW
HB
NI
HH
SH
0,00
5‘ ntr-phylogeny: 445 German BVD-isolates
2b
2c
1k
2a
0,7%
0,4%
1h 0,2%
4,2%
1a
2,9%
1g
2,2%
1f
0,7%
8,8%
1e
3,5%
1b
46,2%
1d
30,1%
76,2 % of all isolates belong to BVDV-1b bzw. 1d
2,2% BVDV-1a
5,1% BVDV-2
Summary:
the obligatory BVD control strategy started in 2011 after long
lasting voluntary programs in the German lander
key point: testing of all newborn calves (< 6 month of age) for BVDV
antigene/genome
major test matrix: ear notches
major methods: real time RT-PCR and ERNS ELISA in equal parts
About 6 millions tests were performed
In 2011, 17 410 pi-animals (0,36 % of born calves) were detected
and removed
virus pressure has been decreased substantially
Summary:
with the increasing of the number of BVDV free herds, the
danger of reinfection as well as the economic losses to
expect are increasing too
Therefore flanking measures (i.e. vaccination) and
serologiocal monitoring as well have to be implemented.
Phylogenetic analysis have shown BVDV-1b and BVDV-1d as
the most frequent subtypes
Many Thanks
Foto: C. Illing