flavi and pestiviruses october 12, 2010 flaviviridae pestiviruses flaviviruses hepacviruses...
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Flaviviridae
Pestiviruses
FlavivirusesHepacvirusesHepatitis C virus
BVD, Hog cholera, Border disease
Yellow feverJapanese encephalitisSt. Louis encephalitisDengueWest Nile virus
(arthropods, biological vectors)
West Nile OutbreaksWest Nile Outbreaks• Israel - 1951-1954, 1957• South Africa - 1974• Romania – 1996• Italy – 1998, 2008• Russia - 1999 (human)• United States –1999-2009 (equine, human)• Canada - 2001-2009 (equine, human)• Israel – 1998, 2000 (human)• France (Rhine delta) - 2000 (equine)
springearly summer
return from south
overwinteror
eggs
amplification in birds
late summer andfall
dead-end hosts
Saskatchewan mosquito species shown to be capable of transmitting
WNV• Aedes vexans (spring to fall)• Ochlerotatus flavescens,
spencerii(July-August)• Culex restuans*, tarsalis* (July-
August)• Culiseta inornata*• Coquillettidia perturbans
How does the virus overwinter and spread?
• migratory birds?
• overwintering mosquitos?
• bird to bird transmission?
– Komar et al. EID March 2003
in the mosquito
1. virus ingested
with blood meal
2. virus multiplies in gut epithelial cells
3. virus leaks from gut and infects salivary glands
4. virus released in saliva during feeding
(sufficient amount of virus must be ingested - > 105 infectious units/ml)
in mammalsvirus transmitted in mosquito
saliva during probing
virus deposited in extra vascular tissue
replication in skin and lymph nodes
amplification in extra-neural tissues
viremia (secretion in milk)
crosses blood/brain barrier (repl’n in vascular endothelium, exacerbated by concurrent infections)
viremia terminated by immune response
viral damage to neurons and glia or dysfunction
inflammation perivascular infiltration (plasma cells and macs), cerebral edema.
“flu” likesymptoms
“neurological”signs
IgM,
CSF IgM, pleocytosis,PCR
PCR
equine cases of WNV neurological disease
• Ataxia 86%• Depression 51%• Hind limb weakness 49%• Difficulty or inability to rise 46%• Muscle tremors 41%• Fever only 24%• Differentials:
– rabies, EHV 1, EEE, WEE, botulism
• 10% to 50% of horses with neurological signs die
clinical signs in people
• most asymptomatic• fever, “flu” like symptoms (fatigue, anorexia, nausea,
vomiting, arthralgia, rash, lymphadenopathy)• encephalitis, meningoencephalitis - ataxia, painful
eyes, seizures, change in mental status (confusion)case fatality rate in hospitalized patients - 10-12%
risk factor for severe disease (age 50-60 yr are 10 times and >80 yr are more than 40 times likely)
Petersen and Margin, WN virus: a primer for the clinician 2002. Ann Int med 137:173
unusual cases in USA
• infant infected through breast milk
• 2 people infected through blood transfusion
• 2 laboratory workers while dissecting infected animals
the Canadian experience
• 2000 - 2,288 birds examined, 185 tested - no positives
• 2001 - 2,807 bird carcasses from NF to Sask tested– 128 WNV infected birds from 12 health
dist. in Ontario– no disease in horses or humans
testing for WNV - serology
strong + cont
weak + cont
samples from 45 horses, Virden Manitoba (Aug-Sept, 2002)
IgM captureELISAPDS immunol.lab
West Nile Virus – Common Client Questions
• Should we vaccinate our horse – is it safe and does it work?
• Can I catch WNV from a horse?• What signs might a horse show early on?• Is there a treatment?• What can we do to limit the risk to our horse?• Can our other pets get it?• When should we vaccinate our horses?
Pestiviruses
• Systemic haemorrhagic disease - pigs (USA)
• Enteric disease - calves (USA)
• Congenital, neurological “hairy-shakers” (England/Wales)
1940sHCV
BVDV
BDV
FlavivirusesFlaviviruses
Pestiviruses Hepatitis CLouping ill
Yellow feverDengue
Pestiviruses of Artydactyla
bovine viral diarrhoea(BVDV)
hog cholera(HCV)border disease
(BDV)
Japaneseencephalitis
Murray ValleySt. Louisencephalitis
West Nile
Wide host range (camelids, deer etc)
BVDV genome and gene products
UTR structural nonstructural
Regions that show the most variation
gp53 p125(p54+p80)
Biotypes of BVDV
NON-cytopathic(natural state)
Cytopathic(mutant)
Based on effect on cells in tissue-culture
Biotypes - implications
• Non-cytopathic– Implications for vaccines and research
• Cytopathic– Mucosal disease
BVDV “types” Not different serotypes!!
• BVDV-1
• BVDV-2
Based on:1. sequence differences In non-translated region of genomeDoes not imply differences in pathogenicity2. Antigenic differences
UTR structural nonstructural
Antigenic differences between BVDV-1 and BVDV-2
Serum against
BVDV-1strains
BVDV-2strains
VN titre againstBVDV-1strains
BVDV-2strains
800->12,800 100->3,200
50->400 3,200->51,200
Pellerin et al.(1994) Virology 203:260-268
Pathogenesis - infection
• Intra-species– PI carriers
• Inter species• Vaccine related?• Artificial breeding programs• Blood• Persistence in acutely infected animals
Pathogenesis - disease in imm.competent, non-pregnant
animals• Sub-clinical• Mild fever, leukopenia, decreased milk
production• Mild BVD - mild erosive lesions, ulcerative
stomatitis, diarrhoea, respiratory• Severe disease - lesions mimic MD,
thrombocytopenia, haemorrhagic syndrome, hyphemia
Pathogenesis - pregnant animals
• All syndromes described above
• Embryonic death
• Abortions
• Birth defects
• Persistently infected calves
consequences of having a PI animal
• Loneragan et al. JAVMA, Feb 15, 2005– PI animals more likely to be ill, require
treatment or die– Incontact animals more likely to be sick,
require treatment
• Dieguez et al. Res Vet Sc, Aug 2009• Correlation BVD status and respiratory
disorders, mortality
Mucosal disease
• NCP->CP (p80)• Infection with antigenically related CP virus
(mutation, herd-mate, vaccination?)• Low morbidity high mortality• High fever, depression, anorexia, diarrhoea• Ulcerative mucosal lesions• Death 2 days -> 3 weeks
Diagnostic procedures• Virus isolation
– Small numbers ($31.50)– Herd screening - microtitre ($10 -2->10, $6 >10)
• Antigen capture ELISA– Herd screening
• Serology– VN ($14)– ELISA ($5/animal)
• PCR– genotyping ($62)– pooled samples ($30)
Diagnostic procedures
• PCR (BVDV-1 vs BVDV-2)• Immunohistochemistry or IFA
– Abortions ($45)– PI animals (approx. $6/animal) BVDV-2
BVDV-1a
BVDV-1b
Immunohistochemistry - PI vs acute
Persistently infected calf:Antigen in hair follicle epithelium
Acutely infected calf:Antigen in superficial layerof epidermis (foci)
From Brad Njaa et al. 2000. J. Vet. Diag. Invest. 12:393-399.
Diagnostic Parameters (acute infection)
infectionincubation period
clinical disease
virus
infect. virus detectable in serum
infect. virus detectable in WBC
Virus det.by PCR
antigens in biopsy
antibody5-7 days
Diagnostic parameters (abortion)
• Fetus– Often no infectious virus– Submit liver, kidney, spleen, thymus for IH
or IFA– Fetal antibody if late term
Diagnostic parameters (PI)
• Large amounts of virus in blood, serum and secretions (103-107 TCID50/ml)
• Maternal antibody interferes with isolation• <3 months - submit blood• >3 months - serum • Repeat isolation in 3 months• Immunohistochemistry - follicle epithelium
Management of BVD
• Test and remove PI animals
• Test all new born calves for 9 months
• For 9-12 months segregate age groups
• Quarantine replacements
• Vaccination with MLV BVDV
• BVD infections may persist for some time after removal of PI animals (Collins et al. 2009)