Development & validation of

confirmatory NSP antibody

ELISAs to detect infection in

vaccinated animals (DIVA)

A.Tewari, H. Ambrose, A. Di Nardo, K.G. Parekh,

T. Inoue, J. Guitian, D.J. Paton, S.Parida

Sera used to estimate diagnostic

sensitivity

A.) To detect seroconversion or infection

Group Number of animals Number of samples

1.) Unvaccinated infected recovered 30 47

a.) Contact challenged 20 31

b.) Needle challenged 10 16

2.) Vaccinated infected recovered 185 261

a.) Contact challenged 80 130

b.) Needle challenged 75 131

B.) To detect carriers

Group Number of animals Number of samples

1.) Unvaccinated infected carriers 12 22

a.) Contact challenged 8 16

b.) Needle challenged 4 6

2.) Vaccinated infected carriers 68 154

a.) Contact challenged 32 74

b.) Needle challenged 36 80

36 Bovine NSP panel sera, 159 clinically infected

field sera

991 naïve cattle sera from Italy

130 vaccinated cattle sera

Sera used to estimate diagnostic

specificity

Six in-house tests developed and

validated

2B and 3B peptide tests

3ABC, 3D, 3CD, 2C proteins

Detection of infection and/or carrier

status in unvaccinated infected cattle

Test N AUC [95% CI p Se ) Sp ) LR+ LR- κ Agreement

1) Unvaccinated infected recovered Prionics 1038 1.00[0.99-1.00] Ref 100.00 99.39 165.16 0.00 Ref Ref

2B 1038 0.99[0.99-1.00] 0.59 100.00 99.10 110.11 0.00 0.87 98.75

3B 1038 0.99[0.99-0.99] 0.13 100.00 98.39 61.93 0.00 0.82 98.17

3ABC 1038 0.99[0.99-1.00] 0.33 100.00 99.09 110.11 0.00 0.87 98.75

3D 1038 0.99[0.99-0.99] 0.21 97.87 97.17 34.63 0.02 0.81 96.82

3CD 1038 0.99[0.99-0.99] 0.21 97.87 98.59 64.66 0.02 0.81 98.07

2C 1038 0.85[0.79-0.89] 0.00 82.98 76.69 3.55 0.22 0.17 76.69

a.) Contact challenge

Prionics 1022 1.00[0.99-1.00] Ref 100.00 99.39 165.16 0.00 Ref Ref

2B 1022 0.99[0.99-1.00] 0.69 100.00 99.10 110.11 0.00 0.82 98.73

3B 1022 0.99[0.99-0.99] 0.13 100.00 98.39 61.93 0.00 0.85 98.14

3ABC 1022 0.99[0.99-1.00] 0.41 100.00 99.09 110.11 0.00 0.82 98.73

3D 1022 0.99[0.99-0.99] 0.32 96.77 97.17 34.25 0.03 0.82 96.77

3CD 1022 0.99[0.99-0.99] 0.23 96.77 98.59 63.93 0.03 0.83 98.04

2C 1022 0.85[0.79-0.91] 0.00 80.65 76.69 3.45 0.25 0.11 76.52

b.)Needle challenge

Prionics 1007 0.99[0.99-1.00] Ref 100.00 99.39 165.16 0 Ref Ref

2B 1007 0.99[0.99-1.00] 0.80 100.00 99.10 110.11 0 0.90 98.71

3B 1007 0.99[0.99-1.00] 1.00 100.00 98.39 61.93 0 0.81 98.11

3ABC 1007 0.99[0.99-1.00] 0.36 100.00 99.09 110.11 0 0.90 98.71

3D 1007 0.99[0.99-0.99] 0.13 100.00 97.17 35.39 0 0.88 96.82

3CD 1007 0.99[0.99-1.00] 1.00 100.00 98.59 66.06 0 0.81 98.11

2C 1007 0.85[0.74-0.96] 0.00 87.50 76.69 3.75 0.16 0.07 76.56

2.) Unvaccinated infected carrier

Prionics 1013 1.00[0.99-1.00] Ref 100.00 99.39 165.16 0.00 Ref Ref

2B 1013 0.99[0.99-1.00] 0.46 100.00 99.10 110.11 0.00 0.86 98.72

3B 1013 0.99[0.99-1.00] 0.32 100.00 98.39 61.93 0.00 0.88 98.12

3ABC 1013 0.99[0.99-0.99] 0.27 100.00 99.09 110.11 0.00 0.86 98.72

3D 1013 0.99[0.98-0.99] 0.21 95.45 97.17 33.78 0.04 0.84 96.74

3CD 1013 0.99[0.99-1.00] 0.40 95.45 98.59 63.06 0.04 0.86 98.03

2C 1013 0.77[0.68-0.87] 0.000 68.18 76.69 2.92 0.41 0.06 76.21

a.) Contact challenge

Prionics 1007 1.00[1.00-1.00] Ref 100.00 99.39 165.16 0 Ref Ref

2B 1007 0.99[0.99-1.00] 0.60 100.00 99.10 110.11 0 0.82 98.71

3B 1007 0.99[0.99-1.00] 0.43 100.00 98.39 61.93 0 0.81 98.11

3ABC 1007 0.99[0.99-1.00] 0.36 100.00 99.09 110.11 0 0.83 98.71

3D 1007 0.99[0.98-0.99] 0.10 93.75 97.17 33.18 0.06 0.86 96.72

3CD 1007 0.99[0.99-1.00] 0.62 93.75 98.59 61.93 0.06 0.89 98.01

2C 1007 0.79[0.70-0.87] 0 68.75 76.69 2.94 0.40 0.041 76.27

b.) Needle challenge

Prionics 997 0.99[0.99-1.00] Ref 100.00 99.39 165.16 0 Ref Ref

2B 997 0.99[0.99-1.00] 0.81 100.00 99.10 110.11 0 0.87 98.70

3B 997 0.99[0.99-1.00] 1.00 100.00 98.39 61.93 0 0.87 98.09

3ABC 997 0.99[0.99-1.00] 0.59 100.00 99.09 110.11 0 0.87 98.70

3D 997 0.99[0.99-1.00] 1.00 100.00 97.17 35.39 0 0.86 96.79

3CD 997 0.99[0.99-1.00] 1.00 100.00 98.59 66.06 0 0.87 98.09

2C 997 0.73[0.46-1.00] 0.35 66.67 76.69 2.86 0.43 0.012 76.33

Detection of infection and/or carrier

status in vaccinated infected cattle

Test N AUC [95% CI p Se (%) Sp (%) LR+ LR-

κ Agreement

%

1.) Vaccinated infected recovered Prionics 1252 0.92[0.90-0.95] Ref 82.49 99.39 118.55 0.17 Ref Ref

2B 1252 0.83[0.80-0.87] 0.00 59.14 99.10 66.10 0.41 0.74 93.51

3B 1252 0.89[0.86-0.91] 0.00 72.37 98.39 42.82 0.28 0.85 95.96

3ABC 1252 0.79[0.75-0.83] 0.00 59.92 99.09 60.28 0.40 0.77 94.14

3D 1252 0.92[0.90-0.94] 1.00 66.15 97.17 19.57 0.35 0.67 91.05

3CD 1252 0.92[0.90-0.94] 1.00 57.59 98.59 30.49 0.43 0.66 91.29

2C 1252 0.59[0.54-0.63] 0.00 43.19 76.69 1.872 0.73 0.18 71.65

a.) Contact challenge

Prionics 1121 0.85[0.80-0.90] Ref 69.83 99.39 116.96 0.30 Ref Ref

2B 1121 0.70[0.64-0.77] 0.00 37.93 99.10 42.35 0.62 0.61 95.45

3B 1121 0.78[0.72-0.84] 0.01 51.72 98.39 32.48 0.49 0.71 96.25

3ABC 1121 0.68[0.61-0.75] 0.00 39.66 99.09 44.28 0.60 0.63 95.63

3D 1121 0.87[0.84-0.91] 1.00 43.97 97.17 12.99 0.58 0.38 91.44

3CD 1121 0.87[0.83-0.91] 1.00 35.34 98.59 18.69 0.65 0.40 92.77

2C 1121 0.48[0.42-0.54] 0.00 29.31 76.69 1.25 0.92 0.04 73.42

b.)Needle challenge

Prionics 1122 0.99[0.98-1.00] Ref 93.89 99.39 155.08 0.06 Ref Ref

2B 1122 0.96[0.95-0.98] 0.10 82.44 99.10 102.12 0.17 0.83 96.88

3B 1122 0.98[0.96-0.99] 0.20 90.84 98.39 56.26 0.09 0.88 97.59

3ABC 1122 0.92[0.88-0.96] 0.06 80.92 99.09 89.09 0.19 0.86 97.33

3D 1122 0.97[0.96-0.98] 0.06 87.02 97.17 30.79 0.13 0.81 95.99

3CD 1122 0.97[0.96-0.98] 0.09 78.63 98.59 55.65 0.21 0.79 96.08

2C 1122 0.70[0.65-0.75] 0.00 57.25 76.69 2.47 0.55 0.20 74.42

2.) Vaccinated infected carrier

Prionics 1145 0.96[0.94-0.98] Ref 90.26 99.39 149.07 0.09 Ref Ref

2B 1145 0.90[0.87-0.93] 0.00 70.13 99.10 86.87 0.30 0.77 95.55

3B 1145 0.90[0.87-0.94] 0.00 75.97 98.39 47.05 0.24 0.81 96.07

3ABC 1145 0.89[0.85-0.93] 0.00 70.78 99.09 77.93 0.29 0.81 96.24

3D 1145 0.92[0.90-0.94] 0.01 61.04 97.17 21.60 0.40 0.64 92.75

3CD 1145 0.93[0.90-0.95] 0.06 56.49 98.59 39.98 0.44 0.64 93.19

2C 1145 0.68[0.63-0.72] 0.00 49.35 76.69 2.13 0.65 0.17 73.28

a.) Contact challenge

Prionics 1065 0.93[0.89-0.98] Ref 87.84 99.39 145.07 0.12 Ref Ref

2B 1065 0.82[0.76-0.88] 0.00 50.00 99.10 55.05 0.50 0.62 96.15

3B 1065 0.83[0.77-0.89] 0.00 58.11 98.39 35.99 0.42 0.65 96.15

3ABC 1065 0.82[0.76-0.89] 0.00 51.35 99.09 56.54 0.49 0.63 96.24

3D 1065 0.86[0.82-0.90] 0.06 35.14 97.17 12.89 0.66 0.37 93.15

3CD 1065 0.88[0.84-0.92] 0.18 31.08 98.59 20.53 0.69 0.38 94.08

2C 1065 0.59[0.52-0.65] 0.00 35.14 76.69 1.50 0.84 0.03 73.62

b.) Needle challenge

Prionics 1071 0.99[0.97-1.00] Ref 92.50 99.39 152.78 0.07 Ref Ref

2B 1071 0.97[0.95-0.99] 1.00 88.75 99.10 109.93 0.11 0.84 97.85

3B 1071 0.97[0.95-0.99] 0.49 92.50 98.39 57.29 0.07 0.85 97.85

3ABC 1071 0.95[0.91-0.99] 0.42 88.75 99.09 97.72 0.11 0.89 98.51

3D 1071 0.97[0.96-0.98] 0.25 85.00 97.17 30.08 0.15 0.73 96.08

3CD 1071 0.97[0.95-0.99] 0.70 80.00 98.59 56.62 0.20 0.76 96.83

2C 1071 0.76[0.71-0.81] 0.00 62.50 76.69 2.70 0.48 0.18 75.84

Detection of infection in vaccinated

infected cattle using Bovine NSP panel

Test N AUC [95% CI p Se (%) Sp (%) LR+ LR- κ Agreement(%)

Prionics 1027 0.99[0.99-1.00] Ref 91.67 99.39 151.40 0.08 Ref Ref

2B 1027 0.99[0.99-0.99] 1.00 91.67 99.10 113.55 0.08 0.77 98.34

3B 1027 0.96[0.91-1.00] 0.80 88.89 98.39 55.05 0.11 0.70 97.66

3ABC 1027 0.96[0.92-1.00] 0.67 75.00 99.09 92.90 0.25 0.68 97.86

3D 1027 0.91[0.84-0.98] 0.09 72.20 97.17 61.17 0.45 0.50 95.81

3CD 1027 0.94[0.89-0.98] 0.06 72.20 98.59 66.85 0.53 0.60 97.08

2C 1027 0.78[0.70-0.85] 0.00 63.80 76.69 2.97 0.40 0.10 76.24

Non-parametric ROC curve

comparison of in-house

tests & Prionics 3ABC test

Detection of infection in cattle in field

Test N AUC [95% CI p Se (%) Sp (%) LR+ LR- κ Agreement(%)

Prionics 159 0.99[0.98-0.00] Ref 96.86 99.39 159.97 0.03 Ref Ref

2B 159 0.98[0.97-1.00] 1.00 96.23 99.10 105.95 0.03 0.93 98.43

3B 159 0.98[0.97-0.99] 1.00 97.48 98.39 60.37 0.02 0.92 98.26

3ABC 159 0.98[0.97-0.99] 0.73 96.23 99.09 105.95 0.03 0.94 98.61

3D 159 0.87[0.83-0.92] 0.00 75.47 97.17 26.71 0.25 0.74 94.09

3CD 159 0.84[0.79-0.89] 0.00 71.07 98.59 46.95 0.29 0.75 94.61

2C 159 0.76[0.69-0.83] 0.00 81.76 76.69 3.50 0.23 0.37 77.39

Non-parametric ROC curve

comparison of in-house

tests & Prionics 3ABC test

Performance of joint-simultaneous testing with the

PrioCHECK® FMDV NS test for Unvaccinated infected

animals

Serial Testing Parallel Testing

Test Se Sp Se Sp

1.) Unvaccinated infected recovered

Prionics Ref Ref Ref Ref 2B 0.93[0.82-0.99] 0.99[0.99-1.00] 0.99[0.99-1.00] 0.97[0.95-0.99]

3B 0.93[0.82-0.99] 0.99[0.99-1.00] 0.99[0.99-1.00] 0.97[0.99-1.00]

3ABC 0.93[0.82-0.99] 0.99[0.99-1.00] 0.99[0.99-1.00] 0.99[0.99-1.00] 3D 0.92[0.82-0.98] 0.99[0.99-1.00] 0.99[0.99-1.00] 0.95[0.93-0.97]

3CD 0.92[0.82-0.98] 0.99[0.99-1.00] 0.99[0.99-1.00] 0.97[0.94-0.98]

2C 0.80[0.72-0.84] 0.99[0.99-0.99] 0.99[0.97-0.99] 0.75[0.73-0.78]

a.) Contact challenge

Prionics Ref Ref Ref Ref

2B 0.93[0.81-0.99] 0.99[0.99-1.00] 0.99[0.99-1.00] 0.97[0.96-0.99] 3B 0.93[0.81-0.99] 0.99[0.99-1.00] 0.99[0.99-1.00] 0.97[0.95-0.98]

3ABC 0.93[0.81-0.99] 0.99[0.99-1.00] 0.99[0.99-1.00] 0.97[0.96-0.99]

3D 0.92[0.81-0.97] 0.99[0.99-0.99] 0.99[0.99-1.00] 0.96[0.94-0.97] 3CD 0.92[0.81-0.97] 0.99[0.99-1.00] 0.99[0.99-1.00] 0.97[0.95-0.98]

2C 0.77[0.69-0.84] 0.99[0.99-0.99] 0.99[0.97-0.98] 0.76[0.73-0.78]

b.) Needle challenge Prionics Ref Ref Ref Ref

2B 0.93[0.81-0.99] 0.99[0.99-1.00] 0.99[0.99-1.00] 0.98[0.96-0.98]

3B 0.93[0.81-0.99] 0.99[0.99-0.99] 0.99[0.99-1.00] 0.97[0.95-0.98] 3ABC 0.93[0.81-0.99] 0.99[0.99-1.00] 0.99[0.99-1.00] 0.98[0.96-0.98]

3D 0.93[0.81-0.99] 0.99[0.99-0.99] 0.99[0.99-1.00] 0.96[0.94-0.97]

3CD 0.93[0.81-0.99] 0.99[0.99-1.00] 0.99[0.99-1.00] 0.97[0.95-0.98] 2C 0.83[0.73-0.91] 0.99[0.99-0.99] 0.99[0.97-0.99] 0.76[0.73-0.78]

2.) Unvaccinated infected carriers

Prionics Ref Ref Ref Ref 2B 0.93[0.81-0.99] 0.99[0.99-1.00] 0.99[0.99-1.00] 0.97[0.95-0.98]

3B 0.93[0.81-0.99] 0.99[0.99-1.00] 0.99[0.99-1.00] 0.97[0.95-0.98]

3ABC 0.93[0.81-0.99] 0.99[0.99-1.00] 0.99[0.99-1.00] 0.97[0.95-0.98] 3D 0.91[0.81-0.96] 0.99[0.99-0.99] 0.99[0.98-1.00] 0.95[0.93--.97]

3CD 0.91[0.81-0.96] 0.99[0.99-1.00] 0.99[0.98-1.00] 0.97[0.95-0.98]

2C 0.65[0.54-0.73] 0.99[0.99-0.99] 0.99[0.95-0.99] 0.76[0.73-0.78]

a.) Contact challenge

Prionics Ref Ref Ref Ref

2B 0.93[0.81-0.99] 0.99[0.99-1.00] 0.99[0.99-1.00] 0.98[0.96-0.99] 3B 0.93[0.81-0.99] 0.99[0.99-1.00] 0.99[0.99-1.00] 0.97[0.95-0.98]

3ABC 0.93[0.81-0.99] 0.99[0.99-1.00] 0.99[0.99-1.00] 0.98[0.96-0.99]

3D 0.90[0.81-0.95] 0.99[0.99-0.99] 0.99[0.98-0.99] 0.96[0.94-0.97] 3CD 0.90[0.81-0.95] 0.99[0.99-1.00] 0.99[0.98-0.99] 0.97[0.95-0.98]

2C 0.65[0.54-0.74] 0.99[0.99-0.97] 0.99[0.95-0.99] 0.76[0.73-0.78]

b.) Needle challenge Prionics Ref Ref Ref Ref

2B 0.94[0.81-0.99] 0.999[0.99-1.00] 0.99[0.99-1.00] 0.98[0.96-0.98]

3B 0.94[0.81-0.99] 0.99[0.99-0.99] 0.99[0.99-1.00] 0.97[0.95-0.98]

3ABC 0.94[0.81-0.99] 0.99[0.99-1.00] 0.99[0.99-1.00] 0.98[0.96-0.98]

3D 0.93[0.81-0.99] 0.99[0.99-0.99] 0.99[0.99-1.00] 0.96[0.94-0.97]

3CD 0.93[0.81-0.99] 0.99[0.99-1.00] 0.99[0.99-1.00] 0.97[0.96-0.98] 2C 0.64[0.55-0.71] 0.99[0.99-0.97] 0.99[0.95-0.99] 0.76[0.73-0.78]

Performance of joint-simultaneous testing with

the PrioCHECK® FMDV NS test for vaccinated

infected animals

Serial Testing Parallel Testing

Test Se Sp Se Sp

1.) vaccinated infected recovered

Prionics Ref Ref Ref Ref 2B 0.50[0.45-0.56] 0.99[0.99-1.00] 0.91[0.89-0.93] 0.97[0.94-0.99]

3B 0.61[0.58-0.64] 0.99[0.99-1.00] 0.93[0.92-0.94] 0.97[0.93-0.99]

3ABC 0.51[0.46-0.57] 0.99[0.99-1.00] 0.91[0.89-0.93] 0.97[0.94-0.99] 3D 0.55[0.50-0.60] 0.99[0.99-1.00] 0.93[0.90-0.95] 0.95[0.91-0.97]

3CD 0.48[0.43-0.54] 0.99[0.99-1.00] 0.91[0.89-0.94] 0.96[0.93-0.98]

2C 0.36[0.33-0.39] 0.99[0.99-0.99] 0.89[0.87-0.91] 0.75[0.72-0.79]

a.) Contact challenge

Prionics Ref Ref Ref Ref

2B 0.28[0.21-0.36] 0.99[0.99-1.00] 0.79[0.71-0.86] 0.97[0.95-0.99] 3B 0.38[0.31-0.45] 0.99[0.99-1.00] 0.82[0.75-0.88] 0.97[0.94-0.99]

3ABC 0.30[0.22-0.38] 0.99[0.99-1.00] 0.79[0.71-0.87] 0.97[0.94-0.99]

3D 0.30[0.24-0.35] 0.99[0.99-1.00] 0.83[0.76-0.89] 0.95[0.92-0.97] 3CD 0.24[0.18-0.30] 0.99[0.99-1.00] 0.80[0.72-0.87] 0.96[0.93-0.98]

2C 0.20[0.13-0.29] 0.99[0.99-0.98] 0.79[0.69-0.87] 0.76[0.72-0.79]

b.) Needle challenge Prionics Ref Ref Ref Ref

2B 0.77[0.73-0.80] 0.99[0.99-1.00] 0.98[0.97-0.99] 0.97[0.94-0.99]

3B 0.84[0.78-0.89] 0.99[0.99-1.00] 0.99[0.98-0.99] 0.97[0.93-0.98] 3ABC 0.76[0.70-0.81] 0.99[0.99-1.00] 0.98[0.97-0.99] 0.98[0.94-0.99]

3D 0.81[0.74-0.86] 0.99[0.99-1.00] 0.98[0.97-0.99] 0.95[0.92-0.97]

3CD 0.73[0.66-0.80] 0.99[0.99-1.00] 0.98[0.96-0.99] 0.97[0.93-0.98] 2C 0.53[0.46-0.60] 0.99[0.99-0.98] 0.97[0.95-0.98] 0.75[0.72-0.78]

2.) Vaccinated infected carriers

Prionics Ref Ref Ref Ref 2B 0.63[0.57-0.69] 0.99[0.99-1.00] 0.96[0.94-0.98] 0.97[0.94-0.99]

3B 0.69[0.62-0.74] 0.99[0.99-1.00] 0.97[0.94-0.98] 0.97[0.93-0.99]

3ABC 0.64[0.58-0.70] 0.99[0.99-1.00] 0.96[0.93-0.98] 0.97[0.94-0.99] 3D 0.55[0.49-0.61] 0.99[0.99-1.00] 0.95[0.92-0.97] 0.95[0.92-0.98]

3CD 0.51[0.44-0.57] 0.99[0.99-1.00] 0.95[0.92-0.97] 0.97[0.93-0.99]

2C 0.44[0.37-0.52] 0.99[0.99-0.99] 0.94[0.91-0.97] 0.75[0.72-0.78]

a.) Contact challenge

Prionics Ref Ref Ref Ref

2B 0.44[0.37-0.50] 0.99[0.99-1.00] 0.93[0.88-0.96] 0.98[0.95-0.99] 3B 0.51[0.42-0.59] 0.99[0.99-1.00] 0.94[0.89-0.97] 0.97[0.94-0.98]

3ABC 0.45[0.38-0.52] 0.99[0.99-1.00] 0.93[0.88-0.96] 0.98[0.95-0.99]

3D 0.30[0.24--.36] 0.99[0.99-1.00] 0.92[0.86-0.95] 0.96[0.93-0.97] 3CD 0.26[0.20-0.33] 0.99[0.99-1.00] 0.91[0.85-0.95] 0.97[0.95-0.99]

2C 0.29[0.24-0.35] 0.99[0.99-0.98] 0.92[0.86-0.95] 0.75[0.73-0.78]

b.) Needle challenge Prionics Ref Ref Ref Ref

2B 0.80[0.73-0.87] 0.99[0.99-1.00] 0.98[0.97-0.99] 0.97[0.94-0.99]

3B 0.85[0.81-0.88] 0.99[0.99-1.00] 0.99[0.98-0.99] 0.97[0.94-0.98]

3ABC 0.81[0.75-0.87] 0.99[0.99-1.00] 0.98[0.97--.99] 0.98[0.94-0.99]

3D 0.78[0.72-0.82] 0.99[0.99-1.00] 0.98[0.97-0.99] 0.95[0.92-0.97]

3CD 0.73[0.68-0.78] 0.99[0.99-1.00] 0.98[0.97-0.98] 0.97[0.94-0.98] 2C 0.57[0.54-0.60] 0.99[0.99-0.99] 0.97[0.95-0.98] 0.76[0.72-0.78]

Performance of joint-simultaneous testing with

the PrioCHECK® FMDV NS test for panel and

field samples

Serial Testing Parallel Testing

Test Se Sp Se Sp

Panel

Prionics Ref Ref Ref Ref

2B 0.84[0.81-0.86] 0.99[0.99-1.00] 0.99[0.98-0.99] 0.97[0.95-0.98] 3B 0.79[0.70-0.86] 0.99[0.99-1.00] 0.98[0.97-0.99] 0.97[0.95-0.98]

3ABC 0.70[0.62-0.73] 0.99[0.99-1.00] 0.97[0.96-0.98] 0.98[0.96-0.99]

3D 0.66[0.63-0.68] 0.99[0.99-1.00] 0.97[0.96-0.98] 0.96[0.94-0.97] 3CD 0.66[0.63-0.68] 0.99[0.99-1.00] 0.97[0.96-0.98] 0.97[0.95-0.98]

2C 0.58[0.54-0.62] 0.99[0.99-0.99] 0.96[0.95-0.97] 0.76[0.73-0.78]

Field outbreak sera Prionics Ref Ref Ref Ref

2B 0.90[0.83-0.95] 0.99[0.99-1.00] 0.99[0.98-0.99] 0.98[0.95-0.99]

3B 0.91[0.83-0.96] 0.99[0.99-1.00] 0.99[0.99-0.99] 0.97[0.94-0.98] 3ABC 0.90[0.83-0.95] 0.99[0.99-1.00] 0.99[0.98-0.99] 0.98[0.95-0.99]

3D 0.71[0.64-0.76] 0.99[0.99-1.00] 0.98[0.96-0.99] 0.96[0.93-0.97]

3CD 0.66[0.60-0.72] 0.99[0.99-1.00] 0.98[0.96-0.99] 0.97[0.94-0.98] 2C 0.77[0.69-0.83] 0.99[0.99-0.99] 0.99[0.97-0.99] 0.75[0.72-0.77]

• Six in-house NSP antibody ELISAs were developed and validated.

• Bovine sera extracted from naïve animals, experimental vaccine challenged animals,a well-established bovine serum panel and sera field outbreak were used

• In-house tests were further evaluated to detect infection and carrier status in vaccinated and/or unvaccinated FMD infected and subsequently recovered and carrier animals, respectively

• In-house tests were used as confirmatory tests to the PrioCHECK® FMDV NS screening test and the increased sensitivity and specificity have been calculated using a Bayesian approach

Summary

• Preliminarily evaluated ID Vet c-ELISA NSP kits (Over night incubation and 2 hour incubation)

• 36 Bovine serum panel and 91 naïve serum samples were used and all were found negative

• Similar results as Prionics 33/36 (91.6%) were positive

• Only one common sample was found negative by both the tests

• Joining both the commercial tests 35/36 (97.2%) samples were seen positive

Good news

Test N AUC p Se (%) Sp (%) LR+ LR- κ Agreement %

1) Panel ID Vet short 128 0.99 Ref 91.67 99.39 151.40 0.08 Ref Ref

2B 128 1 1 91.67 99.10 113.55 0.08 0.89 96.09

3B 128 0.96 0.67 88.89 98.39 55.05 0.11 0.81 92.90

3ABC 128 0.94 0.26 75.00 99.09 92.90 0.25 0.65 87.50

3D 128 0.89 0.08 72.20 97.17 61.17 0.45 0.65 87.50

3CD 128 0.91 0.08 72.20 98.59 66.85 0.53 0.68 89.06

2C 128 0.84 0.00 63.80 76.69 2.97 0.40 0.60 85.06

2.) Panel

ID Vet o/n 128 1.00 Ref 91.67 99.39 151.40 0.08 Ref Ref

2B 128 1 1 91.67 99.10 113.55 0.08 0.91 96.88

3B 128 0.96 0.65 88.89 98.39 55.05 0.11 0.83 93.75

3ABC 128 0.94 0.25 75.00 99.09 92.90 0.25 0.67 88.28

3D 128 0.89 0.08 72.20 97.17 61.17 0.45 0.67 88.28

3CD 128 0.91 0.08 72.20 98.59 66.85 0.53 0.71 8980

2C 128 0.84 0.00 63.80 76.69 2.97 0.40 0.63 85.90

Funding

ICAR IF, New DelhDefra, UK

Defra, UK

EU DISCONVAC

Acknowledgements