Title page
Contents
Abbreviations 6
Executive summary 7
1. Introduction 9
Objectives 10
2. Study design 11
2.1. Organisation 11
2.2. Panel composition, preparation and validation 11
2.3. Participation 12
2.4. Testing 12
2.5. Data reporting 13
2.6. Data analysis 14
3. Results 15
3.1. Participating laboratories 15
3.2. Molecular detection 15
3.3. Virus isolation and antigenic and genetic characterisation 18
3.4. Antiviral susceptibility 24
3.5. Accreditation requirements 31
4. Discussion 34
5. Conclusions 36
6. Recommendations 37
Molecular detection, typing, type A H- and N-subtyping and type B lineage determination 37
Virus isolation 37
Strain characterisation 37
Antigenic characterisation 37
Genetic characterisation 38
Antiviral susceptibility testing 38
Genetic antiviral susceptibility testing 38
Phenotypic antiviral susceptibility testing 38
Accreditation 39
References 40
Annexes: extra tables and figures 43
Table 1. Panel composition, 2018 EEQIAP 11
Table 2. Expected results panel, EEQIAP 2018 molecular detection, virus isolation and antigenic and genetic characterisation 13
Table 3. Expected results panel, EEQIAP 2018 antiviral susceptibility 14
Table 4. Breakdown of number of participants by challenge type 15
Table 5. Overview of molecular detection, typing, type A H- and N-subtype and type B lineage determination results by specimen code 16
Table 6. Overview of molecular detection, typing, type A H- and N-subtype and type B lineage determination results by challenge type 18
Table 7. Overview of virus isolation results by specimen code 19
Table 8. Overview summarising reported genetic characterisation categories by specimen code 24
Table 9. Overview summarising the reported identified amino acid substitutions associated with reduced antiviral susceptibility by specimen code 25
Table 10. Overview summarising reported interpretation of amino acid substitution identification associated with reduced antiviral susceptibility by specimen code 25
Table 11. Overview of phenotypic antiviral susceptibility testing results by specimen code 27
Table 12. Methodologies used by laboratories to determine and evaluate IC50 values 28
Table 13. Summary of survey on laboratory accreditation and participation in external quality assessment programmes 31
Figure 1. Calendar days between starting molecular testing and return of results for molecular detection, typing and subtyping/lineage determination 15
Figure 2. Overview of cumulative performance scores for molecular detection, typing (A/B) and type A H-subtyping 17
Figure 3. Overview of cumulative performance scores for molecular detection, typing (A/B), type A H-and N-subtyping and type B lineage determination 17
Figure 4. Calendar days used for virus isolation and antigenic and genetic characterisation 19
Figure 5. Overview of cumulative performance scores for virus isolation 20
Figure 6. Overview summarising reported antigenic characterisation categories by specimen code 22
Figure 7. Overview of cumulative performance scores for genetic characterisation 23
Figure 8. Calendar days used for genetic and phenotypic antiviral susceptibility testing 24
Figure 9. Overview of cumulative performance scores for genetic antiviral susceptibility determination, amino acid substitution analysis and interpretation of this analysis 26
Figure 10. Overview of reported IC50 values by specimen code 29
Figure 11. Overview of calculated IC50 fold-change values for EISN_AV18-01 and 02 specimens 30
Figure 12. Overview of cumulative performance scores for phenotypic antiviral susceptibility determination 31
Annex Tables
Annex Table 1. List of participants 49
Annex Table 2. Overview in which challenge type each participating laboratory participated 51
Annex Table 3. Overview of molecular detection and typing and type A H-subtype, type A N-subtype and type B lineage determination results by participant with performance score and used methodology 52
Annex Table 4. Overview of virus isolation results with performance score and used methodology 55
Annex Table 5. Overview of virus antigenic characterisation results with used methodology 58
Annex Table 6. Overview of genetic characterisation results with performance score and used methodology 63
Annex Table 7. Overview of genetic antiviral susceptibility testing results with performance score and used methodology 67
Annex Table 8. Overview of phenotypic antiviral susceptibility testing results with performance score and used methodology(assay type only) 77
Annex Figures
Annex Figure 1. Antigenic cartography maps created based on HI-assay data for A(H1N1)pdm09 (A), B/Victoria (C) and B/Yamagata influenza viruses (D) and based on virus neutralisation data for A(H3N2) influenza virus (B) 43
Annex Figure 2A. Phylogenetic tree of full HA of A(H1N1)pdm09 influenza viruses with common amino acid changes for viruses at indicated branch 44
Annex Figure 2B. Phylogenetic tree of full HA of A(H3N2) influenza viruses with common amino acid changes for viruses at indicated branch 45
Annex Figure 2C. Phylogenetic tree of full HA of B/Victoria influenza viruses with common amino acid changes for viruses at indicated branch 46
Annex Figure 2D. Phylogenetic tree of full HA of B/Yamagata influenza viruses with common amino acid changes for viruses at indicated branch 47
Annex Figure 3. Neuraminidase inhibition curves for determination of IC50 for oseltamivir of B/Victoria NA- E105K mutant (A) and A(H1N1)pdm09 N1-H275Y(mixH/Y) mutant (B) using native virus and inactivated by Triton X-100 or formalin 48
Annex Figure 4. Molecular methodologies used by 55 laboratories in detection of influenza virus types A and B (A), type A H-subtyping (B), type A N-subtyping (C) and type B lineage determination (D) 54
Annex Figure 5. Methodologies used by 44 laboratories in virus isolation; type of cells or eggs for A(H1N1)pdm09 (A), A(H3N2) (B) and type B viruses (C); assay type used for confirmation of virus growth (D), type of red blood cells used in haemagglutination assay (E) 57
Annex Figure 6. Source and species of sera used for antigenic characterisation in HI assay and virus neutralisation 62
Annex Figure 7. Methods used for genetic antiviral susceptibility determination 70
Annex Figure 8. Overview of reported IC50 values by method and participant ID of laboratories participating in the phenotypic antiviral susceptibility determination challenge 71
Annex Figure 9. Overview of calculated IC50 fold-change values by method and participant ID of EISN_AV18-01 and 02 specimens 76