欧洲疾控中心-2024年潜在人畜共患甲型流感病毒分子检测_亚型和特征的外部质量评估（英）_112页_14mb

AI报告摘要

External Quality Assessment for Molecular Detection, Subtyping and Characterisation of Potentially Zoonotic Type A Influenza Viruses, 2024

Core Content

The report presents the findings of the European External Influenza Quality Assessment Programme (EEIQAP) 2024, an external quality assessment (EQA) initiative funded by the European Centre for Disease Prevention and Control (ECDC). The programme aimed to evaluate the pandemic preparedness of national influenza centres (NICs) and reference laboratories in the European Union/European Economic Area (EU/EEA), Western Balkan countries, and Türkiye in detecting, identifying, and characterising potentially zoonotic type A influenza viruses.

Main Objectives

Assess the molecular diagnostic capabilities of laboratories for detecting and subtyping zoonotic influenza viruses.
Evaluate genetic characterisation of these viruses, including clade, subclade, and genotype determination.
Identify gaps in diagnostic accuracy and sequencing performance to improve preparedness for zoonotic influenza outbreaks.

Key Findings

Participation

30 laboratories from 27 countries participated.
One laboratory did not participate in the genetic characterisation part.
Non-participating countries were noted in the report.

Molecular Diagnostics (MD)

All laboratories detected type A influenza virus using a generic NAAT.
67% of laboratories successfully detected the lowest concentration specimen (8.11 genome copies/μl) of the H5N1 strain.
Sensitivity differences were observed among laboratories, highlighting the need for improved testing protocols.
Commercial NAATs were found capable of detecting type A influenza viruses in specimens from patients infected with zoonotic strains.

Subtyping

H-subtyping:
- 94% of laboratories correctly identified the H-subtype for human H1pdm09.
- 84% for human H3.
- 94–100% for avian H5.
- 100% for H7.
- 100% for swine H1.
- 82% for swine H3.
N-subtyping:
- 60% of laboratories used N1pdm09 or N2 NAATs.
- 33% used avian N1 NAATs.
- Only 47% of laboratories performed N-subtyping for all panel viruses.

Sequencing and Bioinformatics

70% of laboratories performed whole genome sequencing.
57% of these correctly identified H- and N-subtypes and likely host species (human, avian, swine).
8 laboratories that did not perform sequencing still obtained and uploaded correct HA/NA sequences or NAAT results.
Clade and subclade assignments were correct for 82–100% of the laboratories that reported on these.
Genotype determination for H5N1 was correct in all 5 laboratories that performed it.

Antiviral Susceptibility and Mammalian Adaptation

All sequences contained expected markers for antiviral reduced susceptibility and mammalian adaptation.
However, diverse results were observed, indicating lack of knowledge or tools for interpreting these genetic data.

Recommendations

Implement highly specific NAATs for H- and N-subtyping, and universal type A influenza sequencing.
Participate in ECDC EQA activities and WHO EQAP to enhance diagnostic and sequencing capabilities.
Training in bioinformatics and data analysis is encouraged for better interpretation of genetic data.
National influenza reference laboratories should be capable of confirming zoonotic infections identified at the local level.
If a national laboratory lacks capacity, ECDC offers technical support and training.

Critical Information

Panel composition included:
- 14 heat-inactivated type A influenza viruses with different H and N subtypes.
- 5 avian subtypes (including 3 different genotypes of H5N1).
- 3 swine subtypes (H1N1, H1N2, H3N2).
- 2 human seasonal subtypes (H1N1pdm09, H3N2).
- 1 negative control (no virus).
Data collection and analysis were coordinated by ECDC and RIVM.
Panel preparation and internal quality control were managed by RIVM.
Support from external entities included:
- WHO Collaborating Centre (WHO CC) in London for further characterisation of positive specimens.
- QCMD in Glasgow for panel distribution and data services.

Conclusion

The EEIQAP 2024 provided a comprehensive assessment of the diagnostic and molecular characterisation capabilities of laboratories across the EU/EEA, Western Balkans, and Türkiye. While all 30 laboratories detected all virus strains, there were variations in subtyping accuracy and sequencing performance. The results highlight the importance of specific NAATs and universal sequencing for accurate identification of zoonotic influenza viruses. The need for training and technical support is emphasized to improve diagnostic accuracy and response capacity.

References

[1-3] Outbreaks of HPAI H5N1 in wild birds and poultry.
[4,5] Human cases reported to WHO.
[6] Importance of diagnostic capacity in reference laboratories.

Annexes Overview

Annex 1: List of participating laboratories.
Annex 2: Overview of challenge types.
Annex 3: Equipment and reagents used.
Annex 4-15: Detailed results for molecular detection, subtyping, and sequence analysis.

Figures Overview

Figure 1: Map of participating countries.
Figure 2-12: Correlation of Ct values, sensitivity, and specificity for various subtypes.
Figure 13-15: Performance of laboratories in clade, subclade, and genotype determination.