Download Characterization of opsonizing antibodies against FMD virus, A. Summerfield

Appendix 18
Hypothesis
Characterization of opsonizing antibodies
against foot-and-mouth disease virus
Artur Summerfield
Institute of Virology and
ophylaxis, Switzerland
The immunological value of antibodies may not be
sufficiently determined in a simple VNT.
Protection against FMDV could involve antibodies
opsonizing the virus for transfer on to Fc receptors (FcR):
 virus destruction by phagocytes (McCullough et al 1992
J. Virol. 66:1835.)
 IFN-a responses by plasmacytoid DC (Guzylack-Piriou
et al. 2006 Eur. J. Immunol. 36:1674)
 efficient antigen presentation to DC for T-cell activation
In vivo evidence:
In a mouse model, depletion of macrophages abrogates
the protective effect of antibody transfer, and nonneutralizing can mediate protection (McCullough et al.
1988. Immunology 65:187).
FMDV stimulates plasmacytoid DC only in the
presence of anti-FMDV antibodies
AIMS
1. Determine the relationship between the neutralising
capacity and the opsonising activity of antibodies
2. Cross-reactivity of opsonizing antibodies
3. Use of a murine macrophage cell line expressing bovine
Fc g
RII to quantify opsonizing antibodies in cattle sera
after vaccination
Immune complexed FMDV
FMDV
anti-FMDV Ig
FcR
FcR
DNA
ssRNA
TLR7
TLR9
„plasmacytoid DC“
No/weak response
ssRNA
DNA
TLR9
TLR7
„plasmacytoid DC“
IFN-a
Guzylack et al., Eur. J. Immunol, 2006
Serotype-crossreactivity of porcine opsonizing
antibodies using pDC cultures
Relationship between neutralization and
opsonization titres of porcine sera.
anti-O Manisa serum
Viral Strain
NT in log10 OT* in log10
O UKG 2001
4.1
4
Type-O
O VietNam 7/97
3.1
3
A Brazil 10/93
<1.5
4
Type-A
<1.5
A Turkey /99
2
<1.5
Type Asia-1 Asia-1 Turkey /99
2
<1.5
Type-C
C1-Noville
0
* Opsonizing titres representing minimum serum dilution
able to enhance IFN-α responses by pDC
Serotype
Serum: O Manisa-vaccinated pigs
 Reacts also with A Brazil 10/93
Lannes et al., 2012. Vet Res. 43:64
Open Session of the EuFMD: 2012, Jerez de la Frontera, Spain
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Appendix 18
RAW264.7 murine macrophage cell line to
characterize opsonizing antibodies
Anti-O Mab D9: against linear epitope near RGD of VP1
RAW_bovCD32 cells: murine macrophage cell line
engineered to express bovine CD32
RAW264.7
Characterisitics:
1. Only (with exceptions) in presence of reactive mAb
infection by FMDV
2. This can be detected by intracellular staining of the
mAb/FMDV complex or by measurement of CPE
1st Vaccination challenge experiment in
cattle
Merial Aftovaxpur: inactivated O3039, C
Noville, Asia1 Shamir, A Iran96
Adjuvant: aluminium hydroxide and saponin
After 21 days infection with FMDV O BULGARIA
(10000 ID50)
=> Cannot neutralize O Turkey, O Brazil, A serotype but still opsonize these viruses
VOT/VNT @ 21 days p.v. and protection
full
dose
¼ dose
1/20
dose
100 / 13
10 / 10
10 / 11
100 / 27 100 / 32
10 / 7
100 / 10
10 / 8
10 / 5
100 / 11 100 / 16
-/-
10 / 13
Group 1: 4 animals full dose vaccine
Group 2: 5 animals 1/4 dose
Group 3: 5 animals 1/20 dose
Red: not protected!
No obvious relationship to protection
2nd Vaccination challenge experiment in
cattle
But relationship to vaccination dose?
VOT/VNT @ 21 days p.v. and protection
O3039 O 3039
and
Manisa
O
Manisa
Merial vaccine
Adjuvant: aluminium hydroxide and saponin
After 21 days infection with FMDV O BULGARIA
(10000 ID50)
10 / 19 10 / 27
100 / 108
-/7
10 / 45
10 / 10 100 / 64
100 / 23
Group 1: 4 animals O3039
Group 1: 5 animals O3039 and O Manisa
Group 3: 5 animals O Manisa
10 / 13 100 / 38
100 / 32
10 / 19
100 / 54
100/45
Higher levels of OT and NT against O Bulgaria after vaccination with O Manisa
Open Session of the EuFMD: 2012, Jerez de la Frontera, Spain
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Appendix 18
Conclusions and recommendations
1. Opsonizing antibodies against FMDV can be broadly
cross-reactive even amongst serotype.They enhance
virus infection of macrophages and dendritic cells
resulting in virus destruction and IFNa
responses.
2. Often mAbs which efficiently neutralize are also efficient
at opsonization. Non-neutralizing mAbs (against
homologous virus) can probably not opsonize FMDV.
However, opsonization of non-neutralized virus isolates is
often seen.
3. Measurement of opsonizing antibody may represent a
supplement to other immunological readouts to
determine correlates of protection.
4. O Manisa protects against O Bulgaria 2001
Acknowledgements
Heidi Gerber
Trix Zumkehr
Monika Gsell Albert
Nils Lannes
Lukas Bruckner
Raffael Fricker
Dr. Satya Parida
Dr. Santina Grazioli and Dr. Emiliana
Brocchi. IZSLER, Brescia, Italy
Funding
Open Session of the EuFMD: 2012, Jerez de la Frontera, Spain
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