Figure 5-1. Survival curves of IFNAR
isolates. Each ten mice per group were inoculated subcutaneously with 100 TCID50 of WNV lineage 1 NY, DA, WNV lineage 2 UG and GO. Lineage 1 infected WT mice had 20
WT survived. In contrast all IFNAR was used.
Survival curves of IFNAR -/- and wild type (C57/Bl6) mice infected with four different WNV Each ten mice per group were inoculated subcutaneously with 100 TCID50 of WNV lineage 1 NY, DA, WNV lineage 2 UG and GO. Lineage 1 infected WT mice had 20-30% lethality, whereas all lineage 2 infected WT survived. In contrast all IFNAR -/- mice succumbed rapidly after infection regardless which WNV lineage
33
and wild type (C57/Bl6) mice infected with four different WNV Each ten mice per group were inoculated subcutaneously with 100 TCID50 of WNV lineage 1 NY, DA, 30% lethality, whereas all lineage 2 infected apidly after infection regardless which WNV lineage
34
Figure 5-2. Viral loads in the brains WNV loads in brains of IFNAR -/-
control mice (IFNAR -/- and wild type) inoculated with dilution medium. Deceased NY and DA infected WT mice had medium viral loads in brain, whereas UG and
mice had high viral loads in this organ with values of UG infected IFNAR loads were determined with qRT-PCR and given in Ct values (Figure (Figure 5-2B) or with virus titration given in TCID50 per mg brain (Figure
A
B
C
in the brains of different mouse groups determined by qRT
and wild type mice infected with WNV NY, DA, GO and UG and negative and wild type) inoculated with dilution medium. Deceased NY and DA infected WT mice had medium viral loads in brain, whereas UG and GO infected WT mice were negative. All IFNAR mice had high viral loads in this organ with values of UG infected IFNAR -/- mice being slightly lower. WNV
PCR and given in Ct values (Figure 5-2A) and in genome copies per m 2B) or with virus titration given in TCID50 per mg brain (Figure 5-2C).
Manuscript IV
determined by qRT-PCR or cell culture.
and wild type mice infected with WNV NY, DA, GO and UG and negative and wild type) inoculated with dilution medium. Deceased NY and DA infected WT GO infected WT mice were negative. All IFNAR -/- mice being slightly lower. WNV 2A) and in genome copies per mg brain
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Figure 5-3. Viral loads in the hearts WNV loads in hearts of IFNAR -/-
control mice (IFNAR -/- and wild type) inoculated with dilution medium. In deceased NY and DA infected WT mice low viral loads were detected in heart. In I
infected with lineage 1 and 2 isolates, with UG infected mice revealing slight lower values. WNV loads were determined with qRT-PCR and given in Ct values (Figure
3B) or with virus titration given in TCID50 per mg heart (Figure
A
B
C
in the hearts of different mouse groups determined by qRT
and wild type mice infected with WNV NY, DA, GO and UG and negative and wild type) inoculated with dilution medium. In deceased NY and DA infected WT mice low viral loads were detected in heart. In IFNAR -/- mice high viral loads were detected in hearts of mice infected with lineage 1 and 2 isolates, with UG infected mice revealing slight lower values. WNV loads were
PCR and given in Ct values (Figure 5-3A) and in genome copies per 3B) or with virus titration given in TCID50 per mg heart (Figure 5-3C).
35
groups determined by qRT-PCR or cell culture.
and wild type mice infected with WNV NY, DA, GO and UG and negative and wild type) inoculated with dilution medium. In deceased NY and DA infected WT mice high viral loads were detected in hearts of mice infected with lineage 1 and 2 isolates, with UG infected mice revealing slight lower values. WNV loads were 3A) and in genome copies per mg heart (Figure
5-36
Figure 5-4. Viral loads in the spleens WNV loads in spleens of IFNAR
-/-control mice (IFNAR -/- and wild type) inoculated with dilution medium. Deceased WT mice infected with NY and DA had low viral loads. In contrast, very hi
lower viral loads in single animals. WNV loads were determined with qRT 5-4A) and in genome copies per mg spleen (Figure
(Figure 5-4C).
A
B
C
in the spleens of different mouse groups determined by qRT
- and wild type mice infected with WNV NY, DA, GO and UG and negative and wild type) inoculated with dilution medium. Deceased WT mice infected with NY and DA had low viral loads. In contrast, very high viral loads were detected in IFNAR
-/-lower viral loads in single animals. WNV loads were determined with qRT-PCR and given in Ct values (Figure 4A) and in genome copies per mg spleen (Figure 5-4B) or with virus titration given in TCID50 per mg spleen
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groups determined by qRT-PCR or cell culture.
and wild type mice infected with WNV NY, DA, GO and UG and negative and wild type) inoculated with dilution medium. Deceased WT mice infected with NY - mice, with UG having PCR and given in Ct values (Figure n TCID50 per mg spleen
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Figure 5-5. Immunohistochemistry score for brain (5
groups. For IHC the whole slide was evaluated and a score for each brain region, for splenic follicles and sinus and for liver Kupffer cells and hepatocytes was given, dependent of the percentage of positive antigen stained tissue (brain: IHC score 0 = negative, I
IHC score 3 = ≥ 10 % positive tissue; spleen: IHC score 0 = negative, IHC score 1 = ≤ 9 % positive tissue; IHC score 2 = 10 – 24 % positive tissue; IHC score 3 =
definable; liver: IHC score 0 = negative, IHC score 1 = <1 % positive tissue; IHC score 2 = 1
A
B
C
. Immunohistochemistry score for brain (5-5A), spleen (5-5B) and liver (5
For IHC the whole slide was evaluated and a score for each brain region, for splenic follicles and sinus and for liver Kupffer cells and hepatocytes was given, dependent of the percentage of positive antigen stained tissue (brain: IHC score 0 = negative, IHC score 1 = ≤ 1 % positive tissue; IHC score 2 = 2
≥ 10 % positive tissue; spleen: IHC score 0 = negative, IHC score 1 = ≤ 9 % positive tissue; IHC 24 % positive tissue; IHC score 3 = ≥ 25 % positive tissue; pos = positive, but proportion not definable; liver: IHC score 0 = negative, IHC score 1 = <1 % positive tissue; IHC score 2 = 1
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B) and liver (5-5C) of all mouse For IHC the whole slide was evaluated and a score for each brain region, for splenic follicles and sinus and for liver Kupffer cells and hepatocytes was given, dependent of the percentage of positive antigen stained
≤ 1 % positive tissue; IHC score 2 = 2 - 9 % positive tissue;
≥ 10 % positive tissue; spleen: IHC score 0 = negative, IHC score 1 = ≤ 9 % positive tissue; IHC sue; pos = positive, but proportion not definable; liver: IHC score 0 = negative, IHC score 1 = <1 % positive tissue; IHC score 2 = 1 – 4 % positive
38 Manuscript IV
tissue; IHC score 3 = ≥ 5 % positive tissue; inc = inconclusive). The highest given score for one region/cell type gives the total organ score. Positive cells were found in brain in cerebrum, endbrain, hippocampus, diencephalon, brainstem, and cerebellum of lineage 1 infected WT mice. In IFNAR -/- mice no antigen was detected in brain. Spleens of WT mice remained negative, whereas in IFNAR-/- mice massive antigen was detected regardless of WNV lineage used. All WT livers remained negative for WNV antigen, whereas IFNAR -/- livers had medium amounts of antigen, with UG infected IFNAR --/- mice revealing lower scores in this organ than the other three isolates.
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