OIE Reference Laboratory Reports
Activities in 2011
Name of disease (or topic) for
which you are a designated OIE
Reference Laboratory:
Address of laboratory:
Highly pathogenic avian influenza and low
pathogenic avian influenza (poultry)
Friedrich Loeffler Institute
Institute of Diagnostic Virology
O.I.E., FAO and National reference laboratory
for avian influenza
Suedufer 10
D-17493 Greifswald-Insel Riems
GERMANY
Tel.:
++49 38351 71152
Fax:
++49 38351 71174
e-mail address:
Timm.harder@fli.bund.de
website:
Name (including Title and
Position) of Head of Laboratory
(Responsible Official):
Prof. Dr. Timm Harder, PhD
Name(including Title and
Position) of OIE Reference
Expert:
Prof. Dr. Timm Harder, PhD
Name (including Title and
Position) of writer of this report
(if different from above):
Prof. Dr. Timm Harder, PhD
Annual reports of OIE Reference Centres, 2011
PD Dr. Christian Grund
1
Highly pathogenic avian influenza and low pathogenic avian influenza (poultry)
Part I: Summary of general activities related to the disease
1.
2.
Test(s) in use/or available for the specified disease/topic at your laboratory
Test
For
Specificity
Total
Competitive ELISA
Antibody (AIV NP)
Type (influenza A virus),
534
Hemagglutination inhibition
assay
HI antibody
Subtype
504
Embryonated hen eggs
Virus isolation, virus
amplification
Replication-competent avian
viruses
36
Cell cultures
Virus isolation/titration
Viral pathogens
25
Rapid antigen assay
Type-specific antigen
detection
Type (influenza A virus)
20
Conventional RT PCR
Amplification of AIV
nucleic acid
Group (M-segment), subtypespecific (HA, NA)
208
Real time RT PCR
Amplification and
characterization of AIV
nucleic acids
Group (M-segment), subtype
specific (HA 5, HA 7, NA 1-9)
591
Real time RT PCR
Molecular pathotyping of
AIV
HA 5, Qinghai lineage,
Eurasian low path H5
23
Nucleotide sequencing
Molecular pathotyping of
AIV, phylogenetic analysis
Fragments of HA and NA
segments
117
Immunization studies and
challenge experiments in
animals
Production of antisera,
clinical pathotyping (IPVI)
of AIV, vaccine evaluation
Broad
19
Production and distribution of diagnostic reagents
Amount supplied nationally
(including for own use)
Amount supplied to other
countries
Reference viruses
Management of virus repository with
>300 strains and isolates of all avian H/N
subtypes, human type A and B, porcine and
equine influenza viruses.
Deliveries: 4
Deliveries: 22
Virus antigen
Deliveries: 15
0
Reference RNA
Deliveries: 3
4
Primers, probes
Repository of >25 real time and
conventional RT-PCRs for influenza
diagnosis.
Deliveries: 1
Deliveries: 4
Type of reagent
2
Annual reports of OIE Reference Centres, 2011
Highly pathogenic avian influenza and low pathogenic avian influenza (poultry)
Part II: Activities specifically related to the mandate
of OIE Reference Laboratories
3.
International harmonisation and standardisation of methods for diagnostic testing or the
production and testing of vaccines
a)
Establishment and maintenance of a network with other OIE Reference Laboratories
designated for the same pathogen or disease and organisation of regular inter-laboratory
proficiency testing to ensure comparability of results
 Organizing member of the OFFLU technical group on proficiency testing; production of test vials for
PCR, sequencing and antigen characterization assays among OIE AI reference laboratories. Managing
organization by OIE reference lab at Ames, Iowa.
b)
Organisation of inter-laboratory proficiency testing with laboratories other than OIE
Reference Laboratories for the same pathogens and diseases to ensure equivalence of
results
 Participation in international ring trials organised by other laboratories:
- European Community Reference Laboratory at Weybridge, UK (PCR, serology: passed)
- Animal Health Service, Deventer, The Netherlands (serology: passed)
 Organisation by your laboratory of international ring trials:
- Egypt (PCR and serology; National Reference lab of Egypt and four regional branches thereof; all
participants passed)
- National veterinary diagnostic centers; ring trial for PCR, antigen characterization and virus isolation
 Participation in, or organisation of, interlaboratory test evaluations leading to international
harmonisation and validation (specify the tests, the organisers, participants, outcomes).
- Supply of data to the AHVLA, UK, epidemiological group for Bayesian analysis of test performances of
NP-specific influenza A virus ELISAs
 International co-operation on harmonising methods for production and quality control of vaccines.
- Contacts with vaccine producing companies for H5 and H9 vaccines to be used in Egypt and the Near
East.
4.
Preparation and supply of international reference standards for diagnostic tests or vaccines
No standard reference materials officially recognised by OIE or other international bodies are produced by this
laboratory.
Any other reference materials regarding quality control of AI-related diagnostic measures is freely available from
this laboratory (see lists under item 2).
5.
Research and development of new procedures for diagnosis and control

Production of recombinant baculovirus-expressed influenza A virus hemagglutinins for use in competitive
ELISA (continued and extended from last year; 2 publications released); advanced validation level.
Performance characteristics variable with species and subtype. Extensive evaluation of cross-subtype
reactivity published.

Development of a multiplex RTqPCR for the detection and lineage-specific characterization of swine
influenza viruses in Europe (1 publication submitted); advanced validation level.
Annual reports of OIE Reference Centres, 2011
3
Highly pathogenic avian influenza and low pathogenic avian influenza (poultry)
6.
Collection, analysis and dissemination of epizootiological data relevant to international disease
control
 Epidemiological surveys: Continuation of wild bird and poultry monitoring programs at national level.
Continuing presence of a broad spectrum of different LP AIV subtypes in wild water birds in Germany.
Subtype H5 viruses are still moderately prevalent in wild birds.

Scientific projects: None
 Notifications sent to OIE: None
7.
Maintenance of a system of quality assurance, biosafety and biosecurity relevant to the
pathogen and the disease concerned
Fully ISO-certified QM system is in place since 2008. Certifying body changed to company AKS. Regular audits
(internal and external) ensure tight update schedules of QM measures.
8.
Provision of consultant expertise to OIE or to OIE Member Countries
 Member of several OFFLU technical groups: Standard PCR controls; Proficiency testing.
 International missions: None
 Writing: A short note of the Twinning project between our laboratory and the Egyptian NLQP institute at Giza
has been compiled for publication in the OIE bulletin.
 Consultation: Frequent e-mail exchange with scientists from India, South Korea, Singapore, and Sudan on AIV
related matters.
 Participation in expert panel for the validation of candidate “Anigen Antigen detection kit” for inscription on
the OIE register.
9.
Provision of scientific and technical training to personnel from other OIE Member Countries
The laboratory is partner in a twinning project with the Egyptian National Reference Institute for Avian Influenza
and Newcastle (NLQP) at Giza, Egypt. In 2011, the project went into its third year. Several research projects, ring
trials and further consultancies have been successfully carried out. A financially neutral prolongation for the third
year had been granted by OIE. In this last year (2011) Newcastle-related matters have been tackled predominantly
but also AI-related diagnostic training had been carried out. Four trainees of the NLQP visited the laboratory for
eight weeks.
The project closed with a final work shop held in October in Giza, Egypt. Apart from members of the twinning
partner institutes, representatives of the OIE regional offices and headquarters were present. In addition,
invitations were issued to FAO, WHO as well as to members of Egyptian veterinary faculties and CVOs of
countries in the Near and Middle East. The emphasis of this meeting was put on facilitating contacts between the
different parties tackling the HPAI problem of Egypt. In a separate session for NLQP and FLI members an
evaluation of the achievements of the project was attempted. A sketch of lines on future co-work has been drawn.
10. Provision of diagnostic testing facilities to other OIE Member Countries
None.
11. Organisation of international scientific meetings on behalf of OIE or other international bodies
Joint Symposium on Avian Influenza and Newcastle Disease in Northern Africa and the Middle East,
28–29 September 2011, Safir Hotel – Dokki – Giza, Egypt
Organized in co-operation with NLQP, Giza.
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Annual reports of OIE Reference Centres, 2011
Highly pathogenic avian influenza and low pathogenic avian influenza (poultry)
No proceedings etc. will be published from this meeting. For a brief summary, see item 9. For a detailed account
on the contents and achievements of this meeting pls. see the final report on the Twinning project between FLI and
NLQP.
12. Participation in international scientific collaborative studies

FluLabNet – Development and maintenance of a diagnostic network of AIV-specialized reference
laboratories (closed in 2011).

FluTest – Standardization and improvement of AIV-specific diagnostic tests. Particular task of our laboratory
is the evaluation of internal controls for RRT PCRs (closed in 2011)

O.I.E. Twinning Project Egypt (closed in 2011)

ESNIP 3 – Surveillance of swine influenza in Europe
13. Publication and dissemination of information relevant to the work of OIE (including list of
scientific publications, internet publishing activities, presentations at international conferences)

Presentations at international conferences and meetings
Harder, T., Overview on Twinnning program between NLQP / FLI, Twinning work shop, Giza
Harder, T., Overview on molecular diagnostic of influenza A viruses, Twinning work shop, Giza

Scientific publications in peer-reviewed journals
1: Abdelwhab EM, Grund C, Aly MM, Beer M, Harder TC, Hafez HM. Influence of maternal immunity on
vaccine efficacy and susceptibility of one day old chicks against Egyptian highly pathogenic avian influenza
H5N1. Vet Microbiol. 2011 [Epub ahead of print] PubMed PMID: 21920677.
2: Starick E, Fereidouni SR, Lange E, Grund C, Vahlenkamp T, Beer M, Harder TC. Analysis of influenza A
viruses of subtype H1 from wild birds, turkeys and pigs in Germany reveals interspecies transmission events.
Influenza Other Respi Viruses. 2011; 5: 276-84.
3: Abdelwhab EM, Grund C, Aly MM, Beer M, Harder TC, Hafez HM. Multiple dose vaccination with
heterologous H5N2 vaccine: immune response and protection against variant clade 2.2.1 highly pathogenic avian
influenza H5N1 in broiler breeder chickens. Vaccine. 2011; 29: 6219-25.
4: Kohls A, Hafez HM, Harder T, Jansen A, Lierz P, Lüschow D, Schweiger B, Lierz M. Avian influenza virus
risk assessment in falconry. Virol J. 2011; 8: 187.
5: Abdelwhab EM, Lüschow D, Harder TC, Hafez HM. The use of FTA® filter papers for diagnosis of avian
influenza virus. J Virol Methods. 2011; 174: 120-2.
6: Starick E, Lange E, Fereidouni S, Bunzenthal C, Höveler R, Kuczka A, grosse Beilage E, Hamann HP,
Klingelhöfer I, Steinhauer D, Vahlenkamp T, Beer M, Harder T. Reassorted pandemic (H1N1) 2009 influenza A
virus discovered from pigs in Germany. J Gen Virol. 2011; 92: 1184-8.
7: Grund C, Abdelwhab el-SM, Arafa AS, Ziller M, Hassan MK, Aly MM, Hafez HM, Harder TC, Beer M.
Highly pathogenic avian influenza virus H5N1 from Egypt escapes vaccine-induced immunity but confers clinical
protection against a heterologous clade 2.2.1 Egyptian isolate. Vaccine. 2011; 29: 5567-73.
8: Postel A, Ziller M, Rudolf M, Letzel T, Ehricht R, Pourquier P, Dauber M, Grund C, Beer M, Harder TC. Broad
spectrum reactivity versus subtype specificity-trade-offs in serodiagnosis of influenza A virus infections by
competitive ELISA. J Virol Methods. 2011; 173: 49-59.
Annual reports of OIE Reference Centres, 2011
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Highly pathogenic avian influenza and low pathogenic avian influenza (poultry)
9: Postel A, Letzel T, Müller F, Ehricht R, Pourquier P, Dauber M, Grund C, Beer M, Harder TC. In vivo
biotinylated recombinant influenza A virus hemagglutinin for use in subtype-specific serodiagnostic assays. Anal
Biochem. 2011; 411: 22-31.

Other communications
1. Siebert U, Gulland F, Harder T, Jauniaux T, Seibel H, Wohlsein P and Baumgärtner W. Epizootics in harbour
seals: Clinical aspects and epidemiology. NAMMCO Sci Publ. 2011; 8: 265-274.
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Annual reports of OIE Reference Centres, 2011