Prevention and control of transboundary and emerging viral ... · patogenesis of pneumonia •...
Transcript of Prevention and control of transboundary and emerging viral ... · patogenesis of pneumonia •...
Dinko Novosel, PhD Croatian Veterinary Institute, Pathology department
Savska cesta 143, 10000 Zagreb, Croatia email: [email protected]
Prevention and control of transboundary and emerging viral infections of pigs in
Central Europe – Croatian situation
• 1. ASF – in a background of CSF control measures
• 2. PEDV, PDCoV
• 3. PPV 1-4
• 4 PRRSV
• 5. PCV2
• 6. HEV and TTSuV 1and 2
Instruction:
-all phylogeny is analyzed using MJ Phylogenetic networks
-immunohistochemistry – positive signal brown, background blue
-in situ hybridization, in situ PCR – postive signal blue, background green
Topics
African swine fewer -last outbreak of CSF in
2006, 1.5 year after
vaccination is
suspended
-last confirmed case in
2008.
-control measures
CSF
African swine fewer -two level biosecurity
measure, counties close
to Bosnia and Serbia
and others
-suspicious case-
pathology, serology and
PCR + obligatory
monitoring of all farms
serology and PCR for
CSFV and ASF ASF
PED and PRCoV
• PED – not present
• -in situ RT PCR tested, but not jet optimized
• PRCoV- never confirmed, antibodies in 2 wild boars in 2006.
• -lazy to purchase Mab-s for IHC
PED
Swine influenza
• -in 2012 health problem in one farm
• -PCV2 is identify as a „bad guy”
• -SIV is identify in background od PCV2 infection
• -actually this was the first confirmation of SIV in Croatia – ever
• -research is ongoing and we have several isolates in MDCK cells
• -2000. antibodies in wild boars, in 2006-2010. serorevalance in wild boars was 9.7%
PPV1-4
• PPV1 – very old and known disease
• -vaccination against reproductive failures
• -weak implication in reproductive disorders
• PPV3 – confirmed in fatteners
• - can be possibly attributed in patogenesis of pneumonia
• -Croatia ancestory region
• PPV2 – confirmed in fatteners and fetuses
• -can be possibly attributed in patogenesis of pneumonia
• PPV4 confirmed in fatteners and fetuses
• - can be possibly attributed in patogenesis of reproductive failures and lymphnode lesions in animals affacted with PMWS
PPV1 • -easy to recognize, 1985. reported in Croatia, seroprevalance was 38%
• -routine vaccination was introduced, seroprevalnce decline to 1%
• -involved in 2.4% of reproductive failure cases
Farm
ID
Total
no. of
fetuses
Number of
reproductive
animals
RF1
PRRSV
Status2
PMWS
status3
No. (%) of fetuses positive for indicated virus by PCR
Infections
found in
fetuses
PRRSV PCV2 TTSuV1 TTSuV2 PPV1 PPV2 PPV3 PPV4
A 14 4000 18% Category
1 Affected 3 (21.4)
14
(100) 1 (7.1) 2 (14.3) 0 (0)
PRRSV,
PCV2,
TTSuV1,
TTSuV2
B 10 2500 5% Category
1 Affected
10
(100) 0 (0)
2
(20.0) PCV2,
PPV4
C 8 12000 2% Category
2 Affected
8
(100) 0 (0)
1
(12.5) PCV2,
PPV4
D 11 800 5% Category
1 Affected
11
(100)
1
(9.1) 0 (0)
2
(18.2)
PCV2,
PPV2,
PPV4
E 2 3000 2% Category
3 -
2
(100) 0 (0) PCV2
F 33 3000 8% Category
1 Affected 3 (9.1)
33
(100) 1 (3.0) 0 (0)
1
(3.1)
PRRSV,
PCV2,
TTSuV1,
PPV4
G 2 200 2% Category
3 -
2
(100) 0 (0) PCV2
H 3 200 1,5% Category
3 -
3
(100)
2
(100) 0 (0)
PCV2,
PPV1
Total 83 6 (7.2) 83
(100) 2 (2.4) 2 (2.4)
2
(2.4)
1
(1.2) 0 (0)
6
(7.2)
Abreviations: 1 RF - Reproductive failures;
2 according to Holtkamp’s criteria;
3 according to PMWS 3 diagnostic key criteria.
PPV2 • -virus is in low viral load, in lungs and lymph
nodes, poorly express SLAIIDQ, almost not CD3
• -no other causative agent observed in proliferative and interstital pneumonia
• -relathionship with strains in neighbour countries
• -low prevalance in fetuses
PPV3 • -present in lungs (in bronchus) and lymph
nodes, virus is in low prevalence, in lymphocytes like cells, poorly express SLAIIDQ, CD3 even less
• -involved in pneumonia primary cause by PRRSV
• -not detected in fetuses
• -Croatia seems to be ancestor region
PPV4 • -present in lymph nodes, in lymphocyte like
cells mostly not express CD3 or SLAIIDQ antigen
• -in relationship with some lesions in lymph node in PMWS cases
• -prevalent in fetuses, some indication that it might be involved in reproductive failures and cause systemic infection in fetus
PRRSV • -introduced in 1994., two
doses of Duroc semen was smuggled from Netherland
• -devastate Croatian pig production, estimations were 17mil Euros
• -significant reproductive failures
• -one outbreak of „hot PRRSV” in 2007
• -several large pig producers close farms
• -no national control measures
• -still unknown exact prevalence
• -the first cases were confirmed by IHC
• -developed ISH, in sit RT PCR needs further optimisation
Imunohistochemistry
In situ hybridization
In situ RT PCR
PCV2
• -first outbreak in 2001.
• -devastated effect in Croatian pig production, after vaccine is introduced situation cooled down, but still unvaccinated herds suffer economical loses
• -in 2006 is reported PMWS in wild boars
Identified viruses in fetuses
Farm ID Total
samples
No. (%) positive for indicated virus
PRRSV PCV2 TTSuV1 TTSuV2 PPV1 PPV2 PPV3 PPV4
M 14 3 (21,4) 14 (100) 1 (7,1) 2 (14,3)
A 10
10 (100)
2 (20)
B 8
8 (100)
1 (12,5)
D 11
11 (100)
1 (9,1) 2 (18,2)
F 2
2 (100)
S 33 3 (9,1) 33 (100) 1 (3)
1 (3,1)
T 2
2 (100)
K 3
3 (100)
2 (100)
Total 83 6 (7,2) 83 (100) 2 (2,4) 2 (2,4) 2 (2,4) 1 (1,2) 0 (0) 6 (7,2)
PCV2-RD Diagnosis -histopathology + IHC or ISH
-common finding in almost all fetuses, but there si no specific lesion in heart and ISH or IHC are always negative -both in situ PCR methods revealed presence of PCV2 in Kupffer cells (not know!) -role in reproductive failures during subclinic infection – unknown -ongoing research...
Direct in situ PCR
Indirect in situ PCR
in situ hybridization
Imunohistochemistry
+
-
Origin of samples TTSuV1 positive
TTSuV2 positive
PCV2 RD ( Σ 63) 1 (1,58%) 2 (3,17%) Interstitial nefritis(Σ 14)
1 (7,14%) 8 (57,14%)
PCV2 SD wild boars (Σ 3)
3 (100%) 3 (100%)
Proliferative and necrotizing pneumonia (Σ 5)
1 (20%) 4 (80%)
Torque teno sus virus
-indication that TTSuV2 is involved -other researchers already made connection with TTSuV infection and glomerulonephropathy
Proliferative and necrotizing pneumonia
• High mortality, pigs per acute dies!
• Still is unresolved etiology PCV2 and PRRSV play significant role
• sever form of proliferative interstitial pneumonia
• High replication level of PCV2
• TTSuV2 identify in lymphocytes
PNP lymph node PMWS lymph node
ISH ssDNA PCV2 ISH ssDNA PCV2
ISH dsDNA PCV2 ISH dsDNA PCV2
In situ PCR TTSuV2
PCV2/PRRSV PCV2/PRRSV
no air
necrosis
TTSuV1 TTSuV2
HEV in HR
• Seroprevalence in pigs 89%
• Prevalence of virus in sera 13.3% , in liver 8.1%
Hepatitis -in PMWS affected animals -in farms with high HEV prevalence we observed outbreaks of edema gut disease without reason.
Imunohistochemistry In situ RT PCR
Take home massage:
• Many known and unknown virus circulates, real impact in health status or production effectiveness is unknown
• If you search you will find many things!
• with PCV2, everything is possible...
Acknowledgment
• PCV2 sequencing, PPVs research to Tamas Tuboly, Daniel Cadar and Attila Csagola , University in Budapest
• PCV2 phylogeny – Joaquim Segales, Marti Cortey from CReSA
• PRRSV - Tomasz Stadejek, University in Warsaw
• SIV and PED – Helena Grgic, Josepha Del Lay University in Guelph
• ISH for PRRSV – Lars Eric Larsen DTU Copenhagen
• In situ PCR – Tahar Ait Ali, Roslin Institute and Rodolfo Ocadiz-Delgado –CINVESTAV , Mexico
Thank you for your attention!