1/41

Influenza surveillance in Switzerland

Sentinel network report

Season 2008 – 2009

National Influenza Reference Centre Laboratory of Virology

University Hospitals of Geneva Geneva, Switzerland

Faculty of Medicine

University of Geneva Geneva, Switzerland

FEDERAL OFFICE OF

PUBLIC HEALTH

2/41

National Influenza Reference Centre

Laboratory of Virology, University of Geneva Hospitals

4 Rue Gabrielle Perret-Gentil, 1211 GENEVA 14 – SWITZERLAND

Dr Yves THOMAS Tel: +41/22 372 40 81 Fax: +41/22 372 40 88

� : yves.thomas@hcuge.ch

Pr Laurent KAISER Tel: +41/22 372 40 96 Fax: +41/22 372 40 97

� : laurent.kaiser@hcuge.ch

3/41

Contents 1. ACKNOWLEDGEMENTS 4

2. RESUME-SUMMARY- ZUSAMMENFASSUNG 5

2.1. Résumé 5

2.2. Summary 6

2.3. Zusammenfassung 7

4. METHOD OF DETECTION FOR INFLUENZA VIRUSES 8

4.1. Clinical identification of influenza cases 8

4.2. Detection of influenza viruses 9

5. CHARACTERIZATION OF INFLUENZA VIRUSES 10

5.1. Phenotyping and antigenic characterization 10

5.2. Genetic characterization 12

6. RESULTS FROM THE 2008-09 SEASON 14

6.1. Detection by molecular assays of influenza viruses in nasopharyngeal specimens 14

6.2 Characteristics of patients with influenza infection 16 6.2.1. Frequency of viruses detected in a particular age group 16 6.2.2. Clinical features of participating subjects 17

6.3. Antigenic and genetic characterization of influenza viruses 17 6.3.1. Influenza A (H3N2) 18 6.3.2. Influenza B 21 6.3.3. Antiviral resistance 22

6.4. Overview of influenza epidemics around the world 24 6.4.1. Influenza in Europe (www.Euroflu.org) 24 6.4.2. Influenza epidemic in North America (www.CDC.gov) 25

7. WHO RECOMMENDATION FOR THE COMPOSITION OF INFLUENZA VIRUS VACCINES FOR USE IN THE 2009-10 NORTHERN HEMISPHERE INFLUENZA SEASON. 27

8. DISCUSSION 27

9. BIBLIOGRAPHY 31

ANNEX 1 32

ANNEX 2 (A): INFLUENZA A (H3N2) HEMAGGLUTINATION 2008 33

ANNEX 2 (B): INFLUENZA A (H3N2) HEMAGGLUTINATION 2009 34

ANNEX 3: INFLUENZA B HEMAGGLUTINATION 40

4/41

1. Acknowledgements

We would like to thank:

- the Sentinel network and collaborating practitioners

- Tobias Eckert, Andreas Birrer, Patrick Mathys and Daniel Koch from the Swiss

Federal Office of Public Health (FOPH)

- Dr Maja Lièvre Wenging Zhang from the World Health Organization (WHO),

Drs Alan Hay and Vicky Gregory from the WHO Reference Laboratory (MRC)

in London, UK

- Patricia Suter and Lorena Sacco for their excellent technical assistance

- Members of the Laboratory of Virology who collaborated in the project

- Werner Wunderli for his advices

5/41

2. RESUME-SUMMARY- ZUSAMMENFASSUNG

2.1. Résumé

La saison 2008-09 de grippe s’est révélée avec une intensité parmi les plus

importante de ces huit dernières années. Le virus influenza A (H3N2) a prédominé

pendant cette saison 2008-09 et a circulé dans la première partie de l'épidémie avec

un pic observé au cours de la semaine 2/2009. Le virus influenza B a co-circulé mais

à un taux inférieur et plus tardivement dans la saison, avec un pic pendant la

semaine 13. Deux virus influenza A (H1N1) seulement ont été détectés dans des

échantillons n’appartenant pas au réseau sentinelle. Les virus influenza A (H3N2)

étaient proches de la souche vaccinale 2008-09 A/Brisbane/10/2007 et de la souche

plus récente influenza A/Finland/9/2008. La majorité des virus influenza B étaient

proches du virus influenza B/Hong Kong /45/2005, de la lignée B/Victoria/02/87.

Deux virus influenza B seulement étaient proches de la souche vaccinale 2008-09

B/Florida/4/2006 et de la souche plus récente influenza B/Barcelona/143/2008, de la

lignée B/Yamagata/16/88. Ces virus influenza B ont été détectés dans la majorité

des cas chez les personnes de moins de 30 ans. Les deux virus influenza A (H1N1)

détectés dans des échantillons non-sentinelle étaient résistants à l’oseltamivir. 37/37

virus influenza A (H3N2) testés en Suisse portaient la mutation de S31N induisant la

résistance à l’amantadine.

6/41

2.2. Summary

The 2008-09 influenza season was one of the three most important epidemics of the

past 8 years. Influenza A (H3N2) virus predominated and circulated during the first

part of the epidemic with a peak during week 2/09. Influenza B virus co-circulated,

but at a lower rate and later in the season, with a peak during week 13/09. Two

influenza A (H1N1) only were detected in non-Sentinel samples. Influenza A (H3N2)

viruses were closely related to the 2008-09 vaccine strain influenza

A/Brisbane/10/2007 and to the more recent influenza strain A/Finland/9/2008. The

vast majority of influenza B viruses were related to influenza B/Hong Kong/45/2005,

a B/Victoria/02/87-like lineage. Two influenza B viruses only were related to

B/Florida/4/2006, a B/Yamagata/16/88-like lineage and the 2008-09 vaccine strain,

and to the more recent B/Barcelona/143/2008 virus. These viruses were detected

mostly in individuals less than 30 years. The two influenza A (H1N1) viruses detected

in non-Sentinel samples were oseltamivir-resistant. 37/37 influenza A (H3N2) viruses

tested in Switzerland had the S31N mutation in the matrix gene inducing amantadine

resistance.

7/41

2.3. Zusammenfassung

Die 2008-09 Influenza Saison war eine der drei intensivsten Grippe-Epidemien der

vergangenen acht Jahre. Influenza A (H3N2) Viren dominierten die Saison. Diese

zirkulierten vor allem in der ersten Hälfte der Epidemie mit einem Maximum in der

zweiten Woche 2009. Influenza B Viren traten dazu parallel auf aber in einem

geringeren Ausmasse und später während des Winters waren sie in der 13. Woche

am häufigsten. Nur zwei Influenza A (H1N1) Viren konnten in Proben nachgewiesen

werden welche nicht aus der Sentinella Überwachung stammten.

Influenza A (H3N2) Viren waren eng verwandt mit dem im Impfstoff von 2008-09

enthaltenen Stamm Influenza A/Brisbane/10/2007 und zu der kürzlich aufgetretenen

Variante Influenza A/Finnland/9/2008. Die Mehrzahl der Influenza B Viren waren

verwandt mit Influenza B/Hong-Kong/45/2005 ein Stamm welcher aus der Influenza

B/Viktoria/02/87 Linie stammt. Nur zwei Isolate waren verwandt mit B/Florida/4/2006,

eine mit B/Yamagata/16/88 verwandte Variante. Dieser Stamm war im Impfstoff von

2008/09 enthalten. Zusätzlich waren diese Viren auch mit einem kürzlich

nachgewiesenen Isolate B/Barcelona/143/2008. verwandt. Influenza B wurde

vorwiegend in Personen unter dreissig Jahren nachgewiesen.

Die zwei Influenza A (H1N1) Viren aus den Nicht-Sentinella Proben waren gegen

Oseltamivir resistent. 37/37 Influenza A (H3N2) Viren welche in der Schweiz

nachgewiesen wurden, trugen die S31N Mutation im Matrix Gen. Eine Mutation

welche die Viren unempfindlich gegen Amantadin macht.

8/41

3. INTRODUCTION

Since 1986, annual influenza epidemics are monitored by the Swiss Sentinel

network, the National Reference Centre of Influenza (NRCI), and the Federal Office

of Public Health (FOPH). This collaboration allows an epidemiological, clinical, and

virological surveillance of the epidemic. Volunteer practioners from different Swiss

regions enrol subjects presenting with an influenza-like illness (ILI) during the winter

season. Cases are screened for the presence of influenza in upper respiratory tract

specimens. Two different viruses predominated during this season: first, an influenza

A virus, followed by an influenza B virus that appeared during the second part of the

winter. The duration and period of viral circulation in the population was similar to

many other past seasons. Influenza viruses were characterized by phenotypic testing

(inhibition of the hemagglutination [IHA]) and by sequencing at the genotypic level to

characterize the hemagglutinin (HA) and to screen for antiviral resistance by

neuraminidase (NA) sequencing.

4. METHOD OF DETECTION FOR INFLUENZA VIRUSES

4.1. Clinical identification of influenza cases

During the 2008-09 season, a network of 181 practioners participated actively to the

clinical surveillance of influenza cases. This surveillance is based on a weekly count

of medical consultations for an influenza-like illness (MC-ILI). The case definition

used is the presence of fever of >38°C with or with out a feeling of sickness, myalgia,

or an alteration of the general status. In addition to fever, acute respiratory symptoms

such as cough and/or rhinorhea must be present. The geographic distribution of the

participating general practitioners is shown in Figure 1.

A subgroup of 90 Sentinel participants (50%) are chosen to provide clinical

specimens from selected patients in addition to clinical surveillance. Combined

nasopharyngeal and pharyngeal specimens are sent in transport medium by regular

mail to the NRCI in Geneva for subsequent viral detection and characterization.

The sampling selection procedure of specimens is adapted to the epidemic phases

as follows.

9/41

1) Pre- and post-epidemic phase: the number of MC-ILI by Sentinel practitioners

remains below the threshold level of 58 cases per 100,000 inhabitants. During

this phase, respiratory screening is performed in all cases.

2) Epidemic phase: the number of persons presenting ILI increases. The MC-ILI

is over the threshold of 58 cases of MC-ILI per 100,000 inhabitants. During this

phase, respiratory screening is performed in a subgroup of cases according to

predefined rules and only 1:5 ILI cases are systematically screened.

Figure 1 : Geographical distribution of the 181 participants of the Sentinel network

Each practitioner is represented as follows: red square = participants conducting both clinical surveillance and specimen collection (n=90); blue triangle = participants conducting only clinical surveillance (n=90)

4.2. Detection of influenza viruses

The presence of influenza virus in samples is determined first by two reverse

transcription polymerase chain reaction (RT-PCR) assays. This allows rapid

detection of the genome of influenza A and B types, even if the whole virus has been

inactivated by the transport. In positive cases, the nature of the NA is also

determined by subtype-specific N1 or N2 RT-PCR assays (Schweiger et al, 2000).

Positive samples are then cultivated on appropriate cell lines for antigenic

characterization by IHA to assess the antigenic property of circulating strains and to

10/41

determine the potential efficiency of recognition by influenza vaccine-induced

antibodies (Figure 2). Cell lines used for the surveillance are specific for influenza

isolation, i.e., MDCK and SIAT cells. The latter are modified MDCK cells enriched

with Sialic acid-coupled protein, which is the cellular receptor used by influenza virus

to enter the cell. These cells are assumed to provide a higher efficiency of influenza

virus multiplication by cell culture.

During the first and last weeks of surveillance, a random sampling of negative

specimens are regularly inoculated on cells for virus culture. The goal of this strategy

is to detect influenza strains that could escape RT-PCR detection. This could be the

case in presence of a drifted mutant in the regions of the viral genome targeted by

the RT-PCR primers and probes. A virus of animal origin could also escape RT-PCR

detection as the method is intended for human viruses only. Cell culture is coupled

with immunofluorescence detection using viral nucleoprotein-specific monoclonal

antibodies. This sensitive method allows for the detection of viral antigen, even in low

yields of viral culture cases (see Figure 3).

5. CHARACTERIZATION OF INFLUENZA VIRUSES

5.1. Phenotype and antigenic characterization

In the presence of a positive cell culture, the cell supernatant containing the viral

isolate is phenotyped with IHA reaction. In this latter reaction, the ability of the virus

to link to the red blood cell receptor is tested in the presence or absence of subtype-

specific antisera from immunized ferrets. A specific recognition of the HA by a given

antiserum inhibits the interaction between this HA and the red blood cell receptor. In

the present season, guinea pig red blood cells were used for this reaction. Results

are interpreted according to an antigenic table adapted to circulating strains and

established at the beginning of the season. The 2008-09 reference antigenic table

comprised 5 reference ferret antisera/strains for influenza A (H1N1), A (H3N2) and B

viruses, respectively (Table 1).

11/41

Figure 2 : Procedure used for the detection of influenza vi ruses by Sentinel surveillance

Green arrows: if the clinical sample contains sufficient virus particle, sequencing analysis is performed directly on the original specimen

Negative

Random sampling of

RT-PCR negative

specimens RT-PCR : N1, N2 Influenza A Influenza B

Genotyping characterization by sequencing of the

HA and NA genes

Selection of representative

specimens throughout the season

Nasopharyngeal or pharyngeal specimen

RT-PCR for influenza A RT-PCR for influenza B

Positive

Hemagglutination titration

Culture

Phenotypic characterization by IHA using an antigenic table

adapted to each season

Positive

Negative

12/41

Figure 3 : SIAT cells infected by influenza A/Solomon Island s/03/06 (H1N1)

a) Negative control b) In green, influenza viruses detected with monoclonal anti-influenza A primary antibody and monoclonal FITC conjugate revealing the presence of viral antigen in cells (Chemicon®,, USA)

In this procedure, the titres obtained with each strain are identified and compared

with reference antisera adapted to available antisera and circulating strains. This

allows a standardized identification of the antigenic characteristics of the HA of a

given strain. The ratio between the homologous titres and the observed titres

obtained with the circulating influenza strains define the antigenic relationship to the

reference strains. In turn, this could identify any significant antigenic drift of the HA,

the major target of the immune response.

5.2. Genetic characterization

Genetic characterization is performed by sequencing analysis target genes. Genes

like the HA, are more variable than any others and provide indication on the

phylogeny origin of the virus. Another important task that revealed to be increasingly

important over the last few years is the monitoring of antiviral resistance. Resistance

of influenza viruses to antivirals has been observed with amantadine, oseltamivir,

and zanamivir. Influenza viruses have developed a resistance against these antivirals

with key mutations inducing a modified protein that is no more recognised by the

antiviral. Each antiviral resistance is associated with specific amino acid mutation on

a specific gene, respectively NA gene for neuraminidase inhibitors and Matrix gene

for amantadanes. All known mutations of influenza genes and their corresponding

antiviral resistance detected in human patients are summarised in Table 2.

a) b)

13/41

Table 1 : Hemagglutination inhibition (IHA) titres of refer ence influenza strains tested with the 2008-09 reference antisera

The value obtained in the reaction of the reference strain with the corresponding antiserum represents the homologous titre (HT). The titre obtained with the clinical isolate from a Sentinel sample (SenS) is then compared with HT. If the ratio SenS/HT is ≤4, Sentinel sample is considered as antigenically related to the reference strain. If the ratio is >4, Sentinel sample is considered as antigenically different from the reference strain.

a) Influenza A (H3N2 ) :

b) Influenza A (H1N1)

c) Influenza B

14/41

Table 2 : Key mutations conferring antiviral mutation to influenza viruses

.

6. RESULTS FROM THE 2008-09 SEASON

6.1. Detection by molecular assays of influenza vi ruses in nasopharyngeal specimens

The active surveillance phase began on 27 September 2008 and ended on 17 April

2009 after a period of 29 weeks. Nine hundred and sixteen samples from 66 different

Sentinel practitioners were analysed. Of these 916 cases, 516 were detected

influenza positive by RT-PCR, representing an average positive rate of 56% over 29

weeks surveyed (Figure 4a).

Among the 516 positive cases, 422 (82%) were influenza type A viruses and 94

(18%) influenza type B (Figure 4b). Among influenza A viruses, 305 (72%) were from

the H3N2 subtype and the remaining ones were influenza A viruses that could not be

subtyped.

Influenza A viruses were dominant during the 2008-09 season and represented 82%

of all influenza viruses detected (Figure 4b). Of these type A viruses, 72% could be

subtyped and were from the A (H3N2) subtype; 23% of influenza A viruses could not

be subtyped due to insufficient viral titre. No influenza A (H1N1) viruses have been

detected in the Sentinel network during the 2008-09 season. However, two influenza

15/41

A (H1N1) viruses were detected in hospitalised patients from Geneva (data not

shown). Finally, 18% of influenza viruses detected were of the B subtype.

a)

b)

Figure 4 : Nasopharyngeal specimens positive for any influen za virus during the 2008-09 season (n=916)

a) Number of RT-PCR-positive versus -negative specimens; b) distribution of the different types and subtypes of influenza viruses detected.

Influenza A and B viruses co-circulated in a sequential manner during the season

(Figure 5). Influenza A (H3N2) virus predominated during the first part of the season

with a peak during week 2. Influenza B virus predominated during the second part of

the season with a peak during week 13. The MC-ILI kinetic as reported in our system

well paralleled that observed for influenza A virus detection (Figure 5). However, the

influenza B detection rate did not parallel the kinetic of MC-ILI cases. This

observation suggests that the circulation of influenza A viruses are more often

associated with influenza-like illness symptoms.

516 RT-PCR Positive (56%)

400 RT-PCR Negative (44%)

94 B (18%)

117 A non subtyped

(23%)

306 A (H3N2) (59%)

16/41

0

20

40

60

80

100

120

40 41 42 43 44 45 46 47 48 49 50 51 52 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16

Weeks

Sam

ple

rece

ived

(n)

0

10

20

30

40

50

60

70

80

90

100

MC

-ILI

08-

09 (

‰)

& R

T-P

CR

pos

itive

spe

cim

en (

%)

Influenza A

Influenza B

‰ MC-ILI 08-09

Number of samples tested

2008 2009

Figure 5 : MC-ILI, positivity rate, and distribution type of RT-PCR-positive cases

Proportion of influenza A and B viruses (%), sample received and MC-ILI (%) 2008-09 distributed per week.

6.2 Characteristics of patients with influenza infe ction

6.2.1. Frequency of viruses detected in a particular age group

The proportion of influenza viruses detected in Sentinel specimens is shown

according to different age groups and virus types (Figure 6). The detection rate over

the whole season averaged 56%. The detection rate was relatively similar across

each age groups but the relative contribution of each subtype seemed to vary

significantly. The highest positive rates were observed in patients less than 30 years

old and those 50 to 69 years old. The positive rate was not higher in those over 70

years old (50%).

Influenza A (H3N2) virus rates were the dominant type detected in all age groups.

Between 30% and 40% of samples received in each group contained an influenza A

(H3N2) virus. In contrast, 79% of influenza B viruses were detected in the less than

30 years old group and only 21% in the over 30 years old.

17/41

0

10

20

30

40

50

60

70

80

90

100

≤1 2-4 5-9 10-19 20-29 30-39 40-49 50-59 60-69 70≤

Age group

RT

-PC

R p

ositi

ve s

peci

men

(%

)

A not sub.

A (H3N2)

inf. B

Figure 6 : Percentage of viruses detected on samples accordi ng to age groups

A not sub: influenza A virus not subtyped; A (H3N2): influenza A (H3N2) virus; inf. B: influenza B virus

6.2.2. Clinical features of participating subjects

Forty-six per cent (237/516) of patients infected with influenza virus were female and

54% (279/516) were male. Patient age ranged between three weeks to 86 years old.

Five main symptoms associated with influenza infection have been documented. The

respective proportion of each of these symptoms observed in the case of a

laboratory-confirmed influenza infection is shown in Figure 7. The most frequent

symptoms observed were fever (99%) and cough (96%).

6.3. Antigenic and genetic characterization of inf luenza viruses

Of all influenza viruses detected by RT-PCR, 318 of 516 influenza viruses (62%)

revealed to be culture-positive (Figure 8). Sixty-seven influenza B viruses and 262

influenza A viruses have been phenotyped by IHA assay. All influenza A viruses

appeared to be influenza A (H3N2) viruses (Figure 8). A sample of these viruses

have also been submitted to sequencing analysis. The HA sequence of 35 influenza

A (H3N2) and 13 influenza B viruses were obtained and analysed. Finally, the matrix

18/41

gene of 37 influenza A (H3N2) viruses have also been sequenced to determine the

amantadine resistance status of these influenza viruses.

80

9996

75 73

0

10

20

30

40

50

60

70

80

90

100

Acute onsetof illness

Fever Cough Myalgia Headache

Symptoms

Sym

ptom

s in

pos

itive

sam

ples

(%)

Figure 7 : Symptoms recorded in laboratory confirmed influen za samples

6.3.1. Influenza A (H3N2)

Antigenic characterisation was conducted thanks to IHA reactions, as described

previously. Two different series of influenza A (H3N2) antisera have been selected,

one from 2008 and one from 2009. IHA results are shown in two different tables, one

with 2008 antisera results (Annex 2a) and one with 2009 antisera results (Annex 2b).

Almost all influenza A (H3N2) viruses were antigenically related to the 2008-09

vaccine strain influenza A/Brisbane/10/2007 (H3N2). Half of these influenza A

(H3N2) viruses were also recognised by influenza A/Brisbane/10/2007 antiserum and

showed IHA titres closer to the more recent influenza strain A/Finland/9/2008 (H3N2)

(Annex 2). Four influenza A (H3N2) viruses were also recognised by influenza

A/Brisbane/10/2007 antiserum, although with a reduced titre.

Sequencing analysis of a subgroup of 35 of these culture-positive strains confirmed

the nature of the HA and was used for phylogenetic analysis (Figure 9). A non

significant divergence of HA sequence can be observed on the phylogenetic tree (low

bootstrap values) and these sequences did not form distinct clusters. Bootstrap

19/41

values were not statistically different and were genetically closely related to the

reference 2008-09 vaccine strain A/Brisbane/10/2007 (H3N2).

Figure 8 : Summary of the analysis performed on Sentinel sa mples

IHA: inhibition of the hemagglutination; B/Vic: B/Victoria-like sublineage; B/Yama: B/Yamagata sublineage; HA: hemagglutinin; NA: neuraminidase.

20/41

The matrix gene of 37 influenza A (H3N2) strains have been sequenced for detection

of the mutation related to amantadine resistance. Results are presented in the

section on antiviral resistance (6.3.4.).

Figure 9 : Phylogenetic analysis of the influenza A (H3N2) h emagglutinin

Vaccine strains are labelled in red. Year of viral detection is mentioned at the end of the sample name.

Surprisingly, no influenza A (H1N1) virus was detected in the 516 influenza viruses

detected in the Sentinel network. However, two influenza A (H1N1) were detected in

immuno-compromised patients hospitalised at the University of Geneva Hospitals.

Both viruses could be cultivated and were subtyped by IHA, which revealed that they

were antigenically related to the vaccine strain influenza A/Brisbane/59/2007 (H1N1).

21/41

These isolates have also been analysed for antiviral resistance by NA gene

sequencing. Results will be discussed in the section on antiviral resistance (6.3.4.).

6.3.2. Influenza B

Since the 2002-03 season, two distinct influenza B lineages have been circulating in

Europe, either both during the same season (2004-05, 2005-06), or alternatively in

successive seasons. The first lineage is represented by the B/Victoria/02/87 strain

and the second by the influenza B/Yamagata/16/88 strain. Antisera directed against

the strain of one lineage showed no cross-reactivity (or limited cross-reactivity) in the

IHA reaction against strains of the other lineage.

Antigenic analysis by IHA showed that both influenza B lineages were present in

Switzerland: a Victoria-like one and a Yamagata-like lineage (Annex 3). The former

was predominant representing 65/67 (97%) of the influenza B strains detected

(Figure 8, Annex 3). Most were antigenically related to influenza B/Hong Kong/45/05.

Ten influenza B strains were recognised by B/Malaysia/2506/2004 and

B/Victoria/304/2006 antisera, but with a reduced titre (Annex 3). These strains have

been sent to the European WHO Coordinating Centre (MRC, London, UK) for further

analysis. They appeared to be closely related to the recent strain influenza

A/England/393/2008, a Victoria-like lineage.

Two Yamagata-like lineage strains were detected in Switzerland. Both were

antigenically related to B/Barcelona/143/2008 (Figure 8, Annex 3). Both influenza

viruses were analysed genetically and the HA sequence confirmed the homology

with influenza B/Barcelona/143/2008 and the vaccine strain influenza

B/Florida/4/2006 (Figure 10). A high homology between HA sequences of influenza B

viruses detected during the 2008-09 season was observed with influenza B viruses

detected during the previous season in Switzerland (sequences labelled with a blue

circle, Figure 10).

22/41

6.3.3. Antiviral resistance

The resistance of influenza viruses to antiviral drugs has already occurred in past

years. Table 3 provides an update on resistant viruses detected in European

countries as established by EISS/EuroFlu, the European Network of National

Influenza Centres. Resistant viruses have been detected in many European

countries in recent years.

Figure 10 : Phylogenetic comparison sequences of influenza B viral hemagglutinin

Vaccine strains are labelled in red. Both sub-lineages observed in influenza B viruses are defined by orange and green brackets. Blue circles indicate HA sequences of influenza strains detected in Switzerland during the 2007-08 season

B/Yamagata/16/88 sublineage

B/Victoria/02/87 sublineage

23/41

During the 2007-08 season, influenza A (H1N1) viruses circulated in Europe

(Lackenby et al. 2008), particularly in Switzerland (Thomas et al, 2008). Almost 20%

of these viruses harboured the H275Y mutation in the NA gene that confer

oseltamivir resistance, the main antiviral used specifically for influenza treatment.

This resistance was not promoted by oseltamivir overuse in the population and

emerged among influenza A (H1N1) viruses in absence of selective pressure. The

resistance rate among influenza A (H1N1) viruses was heterogeneous in European

countries and varied from 0% in many Eastern European countries to 68%, as

observed in Norway (Lackenby et al. 2008). During the 2008-09 season, 230/234

influenza A (H1N1) viruses displayed the NA mutation (Euroflu.org). In Switzerland,

no influenza A (H1N1) virus was detected in the Sentinel network. However, two

influenza A (H1N1) viruses were detected in immunocompromised patients

hospitalised at the University of Geneva Hospitals. Sequencing analysis of the NA

gene confirmed that both viruses possess the H275Y mutation that confers

oseltamivir resistance. None of the influenza A (H3N2) and influenza B viruses tested

displayed oseltamivir and zanamivir resistance.

Resistance to amantadine has been shown in influenza A (H3N2) viruses in USA and

Europe (Bright et al., 2006). In 2008, 5/5 of the few influenza A (H3N2) viruses

detected in Switzerland were resistant to amantadine. During the 2008-09 season,

influenza A (H3N2) viruses were predominant in Europe and 552/552 A (H3N2)

viruses tested in European national influenza centres showed the S31N mutation in

the matrix gene due to a G to A mutation that induce to amantadine resistance

(Euroflu.org). In Switzerland, sequencing analysis of the matrix gene of 37 influenza

A (H3N2) viruses have been conducted and all displayed the G to A mutation,

leading to the S31N mutation conferring resistance to amantadine (Figure 11).

24/41

Figure 11 : Partial sequences of the matrix gene of influenza A (H3N2) viruses detected in Switzerland during the 2008-09 season

In red, the nucleotide position that confers the resistance to amantadine.

6.4. Overview of influenza epidemics around the wor ld

6.4.1. Influenza in Europe (www.Euroflu.org)

As observed in Switzerland, both types of influenza A and B viruses circulated in

Europe during the influenza epidemic 2008-09. Influenza A viruses were predominant

and started to circulate early in the season (Figure 12a) with the peak during week 4.

Influenza B viruses started to circulate later and culminated during weeks 10 and 11,

but at a lower rate than influenza A viruses.

Among influenza A viruses, influenza A (H3N2) subtype was predominant (Figure

12b). These influenza A viruses were antigenically and genetically related to the

G to A

25/41

vaccine strain influenza A/Brisbane/10/2007 (H3N2) viruses (Figure 12c). Influenza A

(H1N1) viruses were also detected in some countries, but in a sporadic manner

(Figure 12b). These strains were antigenically related to influenza

A/Brisbane/59/2007 (H1N1) viruses (Figure 12c).

Influenza B viruses of the two lineages were detected in European countries.

However, influenza B viruses of the B/Victoria lineage were predominant and

antigenically related to influenza B/Malaysia/2506/2004 strain. A very low ratio of

influenza B viruses was related to the Yamagata lineage and were related to

influenza B/Florida/4/2006.

6.4.2. Influenza epidemic in North America (www.CDC.gov)

In the USA, the influenza epidemic seems to have had quite different characteristics

than in Europe. Influenza A and influenza B viruses co-circulated during the 2008-09

winter epidemic (MMWR 2009). Sixty-seven per cent of circulating influenza viruses

were influenza A viruses, while 33% were influenza B viruses. Influenza A viruses

predominated in the first part of the epidemic, and influenza B viruses were

predominant in the second part. Influenza A (H1N1) viruses represented 90% of all

influenza A viruses subtyped; 10% were influenza A (H3N2) viruses. Antigenic

properties of influenza A viruses that circulated in USA were related to the vaccine

strains influenza A/Brisbane/57/2007 (H1N1) and A/Brisbane/10/2007 (H3N2).

Influenza B viruses were of the two different lineages: a minority (19%) was related to

the 2008-09 vaccine strain influenza B/Florida/04/2006 and the predominant group

was more related to influenza B/Victoria-lineage (MMWR, 2009).

Of 699 influenza A (H1N1) viruses tested for NA inhibitors resistance (zanamivir and

oseltamivir), 694 (99.3%) were resistant to oseltamivir and all were sensitive to

zanamivir. One hundred and three influenza A (H3N2) and 274 influenza B viruses

were also analysed for resistance to NA inhibitors: none of these two type of viruses

was resistant to NA inhibitors.

One hundred influenza A (H3N2) viruses were also tested for amantadine resistance.

As previously observed, all were resistant (Bright et al. 2006).

26/41

a)

Figure 12 : Influenza viruses detected in the Sentinel networ ks in European countries

a) Type of influenza viruses by week. b) Subtype of influenza viruses detected during the 2008-09 season. c) Antigenic characterization of influenza viruses. Data provided by EISS (http://www.eiss.org)

b)

c)

27/41

7. WHO RECOMMENDATION FOR THE COMPOSITION OF INFLUENZA VIRUS VACCINES FOR USE IN THE 2009-10 NORTHERN HEMISPHERE INFLUENZA SEASON.

The annual meeting for the composition of the influenza vaccine took place on 12

February 2009 at WHO headquarters in Geneva. Based on the epidemiological data

available at that time, recommendations were issued for the composition of the

influenza vaccine for the 2009-10 season (WHO, 2009) (Table 4).

Table 4 : Recommended composition of influenza vaccine for the 2009-10 and 2008-09 seasons

1. B/Brisbane/60/2008-like virus is a B/Victoria-lineage. 2. B/ Florida/4/2006 is a B/Yamagata-lineage

Influenza A strains recommended for the 2009-10 vaccine remained unchanged in

comparison with the 2008-09 vaccine. The influenza B/Brisbane/60/2008 is a

B/Victoria-like strain and replaces the influenza B/Florida/4/2006 of the 2008-09

vaccine, which is a B/Yamagata-like virus.

8. DISCUSSION

During the 2008-09 season, two different influenza viruses circulated: first, an

influenza A (H3N2) that was predominant and then an influenza B virus. The first

influenza virus was at the origin of an epidemic wave that culminated during three to

four weeks early during the year (between weeks 2 and 5). A second wave of

Vaccine strain 2009/2010

Vaccine strain 2008/2009

A

(H1N1) A/Brisbane/59/2007 A/Brisbane/59/2007

A

(H3N2) A/Brisbane/10/2007 A/Brisbane/10/2007

B B/Brisbane/60/2008 1 B/Florida/4/2006 2

28/41

influenza B then circulated in the population and culminated during week 13,

although at a very moderate level.

In comparison with the previous influenza season, the MC-ILI curve registered during

the 2008-09 epidemic showed a peak of around 50%. This value is one of the three

highest observed during the past eight years (Figure 13), with all three resulting from

an influenza A virus of the H3N2 subtype (2003-04, 2004-05, and 2008-09).

However, the present one remains below the most important epidemic observed

during the 1997-98 season and which was associated with the emergence of the

influenza A/Sydney/5/97 (H3N2) strain (Thomas et al., 1997). Since 2006, the

number of influenza viruses detected has increased for the following reasons: first,

RT-PCR screening was introduced and increased the sensitivity of viral detection;

and second, there has been an increase of specimens sent for viral screening in the

Sentinel network.

Figure 13 : Number of influenza viruses and medical consultat ions for influenza-like illness registered from 2001 to 2009 in Switzerland

IHA analysis showed that most influenza A (H3N2) viruses detected during this

season were antigenically related to the vaccine strain influenza A/Brisbane/10/2007

(H3N2), but half of these viruses showed a low decrease in IHA titre. HA sequence

29/41

obtained by sequencing analysis did not show any significant difference or clustering

between these influenza A (H3N2) viruses.

Very surprisingly, no influenza A (H1N1) virus was detected in the Sentinel network.

However, two influenza A (H1N1) viruses were detected by our laboratory in

hospitalised patients at the University of Geneva Hospitals. Both were related to the

2008-09 vaccine strain influenza A/Brisbane/59/2007 (H1N1). This suggest that

adding hospital surveillance to community surveillance might reveal different pattern

of influenza circulation and that susceptible population might be expected to different

strains. Also this reveals that H1N1 was circulating at least at a very low level. In

European countries, only sporadic detection of influenza A (H1N1) viruses was

reported. IHA analysis showed that they were antigenically related to the vaccine

strain influenza A/Brisbane/10/2007 (H1N1).

IHA studies showed that the large majority of influenza B viruses detected were

antigenically related to influenza B/Hong Kong/45/05, a Victoria-like lineage. Two

influenza B strains only were antigenically related to influenza B/Barcelona/143/2008

and the 2008-09 vaccine strain B/Florida/4/2006 of the Yamagata-like lineage. Thus,

predominant influenza B viruses circulating this season were unlikely to be covered

by the 2008-09 vaccine. Interestingly, the rate of influenza B detection did not

correlate with the kinetic of ILI. Based on available data, it could not be determined

whether this reflects a selection bias or a different disease pattern associated with

influenza B. Of note, the same observation was also made during the 2007-08

season (Thomas et al., 2008).

Influenza A viruses circulated among all age groups. In contrast, influenza B viruses

were detected mostly in subjects younger than 30 years old. This has been also

observed in previous seasons. It might be that older people have experienced

influenza B illnesses during their life that have conferred cross-protection as the

influenza B viral genome is more stable than influenza A viruses.

The survey of antiviral resistance is now a routine part of influenza surveillance.

Amantadine-resistant strains emerged spontaneously in influenza strains circulating

during the 2005-06 epidemic in the USA. In Switzerland, the first resistant strains

30/41

were detected during the 2007-08 epidemic (Thomas et al, 2007). During the 2008-

09 season, 100% of influenza strains tested were amantadine- resistant. The same

observation was made in Europe where 100% of 532 strains were resistant to

amantadine (ECDC, 2009). Another resistance that emerged spontaneously was the

resistance to oseltamivir observed in seasonal influenza A (H1N1) viruses. First

observed in Norway during the previous 2007-08 season, this resistance was then

detected in other European countries including Switzerland, where 18% of influenza

A (H1N1) viruses were resistant. Two seasonal influenza A (H1N1) viruses detected

had the H275Y mutation in NA gene conferring the oseltamivir resistance. In Europe,

98% of influenza A (H1N1) viruses were resistant to oseltamivir. The present pattern

of H3N2 and H1N1 virus resistance, respectively, imposes an appropriate laboratory

strategy when antiviral treatment needs to be prescribed. Rapid influenza subtyping

has to be considered at least for hospital-based laboratories.

31/41

9. BIBLIOGRAPHY

Bright, R. A., D. K. Shay, B. Shu, N. J. Cox, A. I. Klimov, R. A. Bright, D. K. Shay, B. Shu, N. J. Cox, and A. I. Klimov . Adamantane resistance among influenza A viruses isolated early during the 2005-2006 influenza season in the United States. JAMA 2006; 295:891-894.

ECDC, “Monitoring of Influenza antiviral resistance in EU during 2008-09 season”, (http://ecdc.europa.eu/en/Health_Topics/influenza/antivirals.aspx). Euroflu.org, update 22/04/2009. Lackenby A, Hungnes O, Dudman SG, Meijer A, Paget W J, Hay AJ, Zambon MC. Emergence of resistance to Oseltamivir among influenza A(H1N1) viruses in Europe. Euro Surveill. 2008; 13: issue 5. MMWR Weekly , CDC, Update: Influenza Activity --- United States, September 28, 2008--April 4, 2009, and Composition of the 2009--10 Influenza Vaccine 2009; 58;369-374. (http://www.cdc.gov/mmwr/preview/mmwrhtml/mm5814a4.htm) Schweiger B, Zadow I, Heckler R, Timm H, G. Pauli G . Application of a fluorogenic PCR assay for typing and subtyping of influenza viruses in respiratory samples. J Clin Microbiol 2000; 38:1552-1558. Thomas Y, Kaiser L. Influenza surveillance in Switzerland: Sentinella study, Winter 2007-08. 2008, Annual report. Thomas Y, Kaiser L, Wunderli W. Influenza surveillance in Switzerland: Sentinella study, Winter 2006-07. 2007, Annual report. Thomas Y, Kaiser L, Wunderli W. Influenza surveillance in Switzerland: Sentinella study, Winter 1996-97. 1997, Annual report. World Health Organization, « Composition recommandée des vaccins antigrippaux pour la saison 2009-2010 (prochain hiver dans l’hémisphère Nord) ». n°9, 2009, 84, p 72. (http://www.who.int/wer/2009/wer8409.pdf)

32/41

ANNEX 1

Sentinel surveillance - Season 2008-09

Total Inf. A A (H3N2) Influenza B Total Weeks Dates ‰ MC-ILI

08-09 sample Undet. Undet. A/Bris Total Undet. Malaysia Florida Total virus (n) Pos (%) ‰ MC-ILI

07-08 40 29-sept-06 05-oct-06 1.3 9 0 0 0 0 0.7 41 06-oct-06 12-oct-06 1.3 7 0 0 0 0 0.9 42 13-oct-06 19-oct-06 1.1 6 0 0 0 0 0.6 43 20-oct-06 26-oct-06 0.7 5 0 0 0 0 1.4 44 27-oct-06 02-nov-06 0.7 4 0 0 0 0 1.5 45 03-nov-06 09-nov-06 1.4 11 2 0 0 2 18 1.4 46 10-nov-06 16-nov-06 0.8 11 3 1 1 0 4 36 0 47 17-nov-06 23-nov-06 1.5 14 1 1 0 1 7 2 48 24-nov-06 30-nov-06 1.3 19 0 0 0 0 2.9 49 01-dec-06 07-dec-06 2.2 20 0 1 1 1 5 3.1 50 08-dec-06 14-dec-06 2.6 26 5 5 0 5 19 4.1 51 15-dec-06 21-dec-06 3.2 39 6 6 0 6 15 4.6 52 22-dec-06 28-dec-06 4.8 24 4 8 8 2 2 14 58 11.4 1 29-dec-06 04-jan-07 9.1 25 3 4 8 12 0 15 60 15.2 2 05-jan-07 11-jan-07 24 120 21 11 61 72 3 1 4 97 81 21.9 3 12-jan-07 18-jan-07 37.6 125 16 8 67 75 1 1 92 74 31.2 4 19-jan-07 25-jan-07 54.4 91 20 11 38 49 0 69 76 29 5 26-jan-07 01-feb-07 52.1 71 8 3 39 42 2 2 52 73 28.5 6 02-feb-07 08-feb-07 43.4 51 15 1 17 18 1 1 34 67 25.2 7 09-feb-07 15-feb-07 28.5 42 11 2 6 8 5 3 8 27 64 20.1 8 16-feb-07 21-feb-07 18.4 28 3 2 2 4 1 7 8 15 54 18.8 9 23-feb-07 29-feb-08 13.7 31 2 3 3 2 7 9 14 45 12.3

10 01-march-07 07-march-08 9.1 32 4 1 1 2 10 12 17 53 7.8 11 08-march-07 14-march-08 7.3 33 3 0 8 6 14 17 52 7.5 12 15-march-07 21-march-08 4.7 27 0 1 11 12 12 44 5.6 13 22-march-07 28-march-08 3.7 18 1 0 5 4 9 10 56 4.6 14 29-march-07 04-apr-08 2.4 18 0 4 2 6 6 33 3.2 15 05-apr-07 11-apr-08 2.6 8 1 0 4 4 5 63 2.6 16 12-apr-07 18-apr-08 0.9 1 0 1 1 1 100 1.1 43 262 38 54 2

916 117 305 94

516

33/41

ANNEX 2 (a): INFLUENZA A (H3N2) HEMAGGLUTINATION 2008 A (H3N2) 2008 Antisera A/Brib07 A/Tries07 A/Wisc05 A/Finl08 A/Nep06 A/Johan08 A/Bribane/10/2007 128 128 32 128 32 32 A/Trieste/25c/07 128 64 32 32 16 32 A/Wisconsin/67/05 256 512 1024 256 512 256 A/Finland/9/08 64 32 8 64 16 32 A/Nepal/921/06 512 1024 256 1024 4096 2048 A/Johannesburg/15/08 256 256 64 256 64 256

Date HA sample n° HA titre Cell/Passage A/Brib07 A/Tries07 A/Wisc05 A/Finl08 A/Nep06 A/Johan08 Result IHA 27.11.2008 1285490 64 ZH/3 256 128 32 64 32 64 A/Bribane/10/2007 02.01.2009 1370948 16 ZH/2 128 32 32 32 A/Bribane/10/2007 09.01.2009 1373613 64 ZH/3 128 64 128 A/Bribane/10/2007 02.01.2009 1373630 64 S/2 256 128 64 128 32 128 A/Bribane/10/2007 02.01.2009 1373732 32 ZH/2 256 128 64 128 32 32 A/Bribane/10/2007 09.01.2009 1380121 16 ZH/3 128 32 32 A/Bribane/10/2007 09.01.2009 1389056 16 ZH/2 32 16 16 A/Finland/9/08 09.01.2009 1397477 32 ZH/2 64 32 32 A/Finland/9/08 09.01.2009 1397481 64 ZH/2 128 64 64 A/Bribane/10/2007 09.01.2009 1397487 64 ZH/2 256 64 128 A/Bribane/10/2007 13.01.2009 1409256 64 ZH/1 128 32 64 A/Bribane/10/2007 13.01.2009 1409274 16 ZH/1 128 32 64 A/Bribane/10/2007 15.01.2009 1409276 64 ZH/2 64 64 64 A/Finland/9/08 15.01.2009 1409281 64 ZH/2 64 32 64 A/Finland/9/08 15.01.2009 1409294 64 ZH/2 64 32 64 A/Finland/9/08 15.01.2009 1420151 128 ZH/1 128 128 128 A/Bribane/10/2007 15.01.2009 1420239 16 ZH/1 64 32 32 A/Finland/9/08 15.01.2009 1420255 128 ZH/1 512 256 256 A/Bribane/10/2007 15.01.2009 1425478 8 ZH/1 128 128 128 A/Bribane/10/2007 22.01.2009 1426767 64 ZH/2 64 64 32 64 32 A/Finland/9/08 15.01.2009 1426769 64 ZH/1 256 256 256 A/Bribane/10/2007 22.01.2009 1426772 32 ZH/2 64 16 16 32 16 A/Finland/9/08 15.01.2009 1426835 32 ZH/1 64 64 64 A/Finland/9/08 15.01.2009 1429309 32 ZH/1 64 64 32 A/Finland/9/08 15.01.2009 1429331 16 ZH/1 64 128 64 A/Finland/9/08 15.01.2009 1431863 64 ZH/1 64 64 64 A/Finland/9/08 15.01.2009 1432042 32 ZH/1 128 128 A/Bribane/10/2007 15.01.2009 1432102 32 ZH/1 256 256 128 A/Bribane/10/2007 22.01.2009 1432521 64 ZH/2 64 64 64 64 32 A/Finland/9/08 22.01.2009 1432561 64 ZH/2 64 64 32 64 16 A/Finland/9/08 22.01.2009 1432789 16 ZH/1 64 32 32 64 16 A/Finland/9/08 19.01.2009 1435736 16 ZH/1 32 16 16 16 16 16 A/Finland/9/08 19.01.2009 1442659 16 ZH/1 128 64 <16 64 <16 64 A/Bribane/10/2007 22.01.2009 1447419 16 ZH/1 64 64 32 64 16 A/Finland/9/08 22.01.2009 1447470 16 ZH/1 64 32 16 32 16 A/Finland/9/08 22.01.2009 1448039 32 ZH/1 32 32 16 32 16 A/Finland/9/08 22.01.2009 1448046 8 ZH/1 32 16 <16 <16 A/Finland/9/08

34/41

ANNEX 2 (b): INFLUENZA A (H3N2) HEMAGGLUTINATION 2009 Antiserum 2009 A/Brib07 A/Tries07 A/Wisc05 A/Finl08 A/Nep06 A/Bribane/10/2007 256 512 512 2048 1024 A/Trieste/25c/07 512 1024 512 512 512 A/Wisconsin/67/05 1024 20248 4096 1024 512 A/Finland/9/08 64 128 32 256 64 A/Nepal/921/06 128 512 256 256 2048

Date HA sample n° HA titre Cell/Passage A/Brib07 A/Tries07 A/Wisc05 A/Finl08 A/Nep06 Result IHA 27.01.2003 1380108 16 ZH/3 32 16 <16 16 <16 reduced 27.01.2009 1425412 64 ZH/2 64 64 16 16 16 A/Bribane/10/2007 27.01.2009 1425437 16 ZH/2 256 256 64 128 128 A/Finland/9/08 27.01.2009 1428524 64 ZH/2 128 128 64 128 32 A/Finland/9/08 27.01.2009 1428528 64 ZH/2 64 64 64 64 32 A/Finland/9/08 10.02.2009 1428538 32 ZH/2 256 512 128 256 A/Bribane/10/2007 10.02.2009 1428545 16 ZH/2 256 512 128 256 A/Bribane/10/2007 27.01.2009 1429349 32 ZH/2 512 512 512 256 256 A/Bribane/10/2007 27.01.2009 1429432 64 ZH/2 128 128 64 128 16 A/Finland/9/08 26.01.2009 1429477 64 ZH/2 256 128 64 64 32 A/Finland/9/08 26.01.2009 1431768 32 ZH/2 64 32 32 32 32 A/Bribane/10/2007 26.01.2009 1431914 128 ZH/2 256 64 64 128 32 A/Finland/9/08 26.01.2009 1432026 32 ZH/2 256 512 256 256 256 A/Bribane/10/2007 26.01.2009 1432074 512 ZH/2 128 128 32 64 64 A/Finland/9/08 27.01.2009 1432111 64 ZH/2 64 64 16 64 16 A/Finland/9/08 26.01.2009 1432111 256 ZH/2 512 1024 1024 1024 1024 A/Bribane/10/2007 26.01.2009 1432126 32 ZH/2 64 64 16 32 16 A/Bribane/10/2007 27.01.2009 1432140 64 ZH/2 128 128 128 128 128 A/Finland/9/08 27.01.2009 1432197 128 ZH/2 128 256 256 256 128 A/Finland/9/08 27.01.2009 1432538 16 ZH/2 64 64 16 64 16 A/Finland/9/08 27.01.2009 1432601 16 ZH/2 256 512 128 256 256 A/Bribane/10/2007 02.03.2009 1432643 32 ZH/2 512 512 512 256 512 A/Bribane/10/2007 27.01.2009 1432709 128 ZH/2 512 512 512 2048 256 A/Bribane/10/2007 27.01.2009 1435593 64 ZH/2 64 32 16 16 16 A/Bribane/10/2007 27.01.2009 1435619 64 ZH/2 128 64 64 64 64 A/Finland/9/08 29.01.2009 1435711 16 ZH/2 64 64 16 32 16 A/Bribane/10/2007 27.01.2009 1435769 64 ZH/2 64 64 32 64 32 A/Finland/9/08 27.01.2009 1435811 32 ZH/2 512 128 256 256 128 A/Bribane/10/2007 27.01.2009 1435849 128 ZH/2 512 2048 512 512 256 A/Bribane/10/2007 03.02.2009 1435906 128 ZH/2 64 128 64 128 64 A/Finland/9/08 27.01.2009 1436004 128 ZH/2 512 1024 256 512 256 A/Bribane/10/2007 29.01.2009 1436422 32 ZH/2 64 32 32 32 32 A/Bribane/10/2007 27.01.2009 1442706 64 ZH/2 256 128 128 128 128 A/Finland/9/08 29.01.2009 1442720 16 ZH/2 64 64 16 64 32 A/Finland/9/08 27.01.2009 1442732 128 ZH/2 256 512 1024 512 128 A/Bribane/10/2007 27.01.2009 1442771 64 ZH/2 256 512 512 512 256 A/Bribane/10/2007 27.01.2009 1442782 256 ZH/2 512 2048 1024 1024 512 A/Bribane/10/2007 27.01.2009 1442804 256 ZH/2 1024 1024 512 128 128 A/Bribane/10/2007 12.02.2009 1442840 64 ZH/3 256 512 128 128 A/Finland/9/08

35/41

Antiserum 2009 A/Brib07 A/Tries07 A/Wisc05 A/Finl08 A/Nep06 A/Bribane/10/2007 256 512 512 2048 1024 A/Trieste/25c/07 512 1024 512 512 512 A/Wisconsin/67/05 1024 20248 4096 1024 512 A/Finland/9/08 64 128 32 256 64 A/Nepal/921/06 128 512 256 256 2048

01.04.2009 1442854 128 ZH/2 256 512 256 256 256 A/Bribane/10/2007 12.02.2009 1447380 64 ZH/3 128 128 64 128 A/Finland/9/08 27.01.2009 1447408 64 ZH/2 256 128 64 128 64 A/Finland/9/08 27.01.2009 1447434 32 ZH/2 32 32 32 32 32 reduced 09.04.2009 1447548 128 ZH/2 64 128 64 64 32 A/Finland/9/08 29.01.2009 1447593 16 ZH/2 64 32 <16 32 16 A/Bribane/10/2007 27.01.2009 1447645 128 ZH/2 128 256 256 256 128 A/Finland/9/08 27.01.2009 1447655 64 ZH/2 64 64 64 64 16 A/Finland/9/08 29.01.2009 1447965 128 ZH/2 64 64 32 64 32 A/Finland/9/08 18.03.2009 1447971 128 ZH/2 128 512 128 256 128 A/Finland/9/08 16.02.2009 1447975 32 ZH/2 512 1024 256 512 A/Bribane/10/2007 29.01.2009 1447994 64 ZH/2 64 64 64 64 32 A/Finland/9/08 09.04.2009 1448012 256 ZH/2 64 64 32 64 64 A/Finland/9/08 03.02.2009 1448017 8 ZH/2 128 64 128 32 A/Finland/9/08 29.01.2009 1448166 64 ZH/2 128 128 32 128 64 A/Finland/9/08 03.02.2009 1448186 256 ZH/2 128 256 256 128 128 A/Finland/9/08 29.01.2009 1448192 32 ZH/2 64 64 32 32 32 A/Bribane/10/2007 16.02.2009 1450547 32 ZH/2 128 64 16 64 A/Finland/9/08 29.01.2009 1451165 32 ZH/2 64 64 32 64 16 A/Finland/9/08 02.03.2009 1451181 512 ZH/2 512 1024 1024 1024 512 A/Bribane/10/2007 29.01.2009 1451213 32 ZH/2 64 32 32 64 32 A/Finland/9/08 02.03.2009 1451238 256 ZH/2 1024 512 512 1024 512 A/Bribane/10/2007 03.02.2009 1451339 128 ZH/2 128 128 256 128 128 A/Finland/9/08 29.01.2009 1451362 32 ZH/2 64 32 16 64 32 A/Finland/9/08 29.01.2009 1451383 128 ZH/2 128 128 64 128 32 A/Finland/9/08 29.01.2009 1451393 16 ZH/2 128 256 <16 128 64 A/Finland/9/08 03.02.2009 1451409 256 ZH/2 256 512 256 512 128 A/Bribane/10/2007 03.02.2009 1451442 32 ZH/2 256 512 128 256 128 A/Bribane/10/2007 29.01.2009 1451463 128 ZH/2 256 512 512 1024 256 A/Bribane/10/2007 29.01.2009 1451474 32 ZH/2 64 64 32 64 16 A/Finland/9/08 03.02.2009 1451499 8 ZH/2 256 64 128 32 A/Finland/9/08 26.01.2009 1455181 128 ZH/2 128 256 128 256 64 A/Finland/9/08 29.01.2009 1455245 32 ZH/2 64 32 32 32 16 A/Bribane/10/2007 03.02.2009 1455295 256 ZH/2 128 64 32 32 32 A/Bribane/10/2007 29.01.2009 1455315 32 ZH/2 128 32 32 64 16 A/Finland/9/08 02.03.2009 1455334 256 ZH/2 512 1024 512 256 256 A/Bribane/10/2007 17.02.2009 1455355 16 ZH/2 1024 1024 256 512 A/Bribane/10/2007 05.02.2009 1455365 32 ZH/2 256 512 256 512 256 A/Bribane/10/2007 04.02.2009 1455374 16 ZH/2 256 256 64 128 64 A/Finland/9/08

36/41

Antiserum 2009 A/Brib07 A/Tries07 A/Wisc05 A/Finl08 A/Nep06 A/Bribane/10/2007 256 512 512 2048 1024 A/Trieste/25c/07 512 1024 512 512 512 A/Wisconsin/67/05 1024 20248 4096 1024 512 A/Finland/9/08 64 128 32 256 64 A/Nepal/921/06 128 512 256 256 2048

Date HA sample n° HA titre Cell/Passage A/Brib07 A/Tries07 A/Wisc05 A/Finl08 A/Nep06 Result IHA 26.01.2009 1455385 32 ZH/1 256 512 256 512 128 A/Bribane/10/2007 05.02.2009 1455396 128 ZH/2 256 1024 512 512 256 A/Bribane/10/2007 05.02.2009 1455405 16 ZH/2 256 512 256 512 1024 A/Bribane/10/2007 04.02.2009 1455425 8 ZH/2 128 128 64 64 64 A/Finland/9/08 05.02.2009 1455430 64 ZH/2 128 128 32 64 64 A/Finland/9/08 10.02.2009 1455440 32 ZH/2 64 32 32 64 A/Finland/9/08 04.02.2009 1455475 8 ZH/2 256 256 64 128 64 A/Finland/9/08 02.03.2009 1456355 128 ZH/2 1024 512 512 256 A/Bribane/10/2007 05.02.2009 1456435 64 ZH/2 128 128 32 128 A/Finland/9/08 26.03.2009 1456441 128 ZH/1 256 512 64 256 128 A/Bribane/10/2007 02.04.2009 1456445 128 ZH/2 64 64 64 64 64 A/Finland/9/08 05.02.2009 1456445 128 ZH/1 128 128 64 128 A/Finland/9/08 04.02.2009 1456462 16 ZH/2 256 256 64 256 32 A/Bribane/10/2007 12.02.2009 1456481 128 ZH/2 128 128 64 128 A/Finland/9/08 17.02.2009 1456505 16 ZH/2 128 256 128 256 A/Finland/9/08 17.02.2009 1456506 8 ZH/2 128 256 128 512 A/Bribane/10/2007 11.02.2009 1456514 128 ZH/2 256 256 128 128 A/Finland/9/08 12.02.2009 1456525 128 ZH/2 256 256 64 256 A/Bribane/10/2007 11.02.2009 1456528 64 ZH/2 256 256 128 128 A/Finland/9/08 10.02.2009 1459746 8 ZH/2 128 128 64 64 A/Finland/9/08 27.01.2009 1459772 16 ZH/1 128 32 32 128 32 A/Finland/9/08 12.02.2009 1459789 128 ZH/2 256 512 256 512 A/Bribane/10/2007 10.02.2009 1459857 32 ZH/2 256 256 64 128 A/Finland/9/08 19.03.2009 1459895 8 ZH/2 512 512 2048 512 A/Bribane/10/2007 01.04.2009 1459912 128 ZH/2 256 1024 256 1024 256 A/Bribane/10/2007 12.02.2009 1459920 256 ZH/2 512 2048 2048 2048 A/Bribane/10/2007 12.02.2009 1459942 256 ZH/2 1024 1024 1024 1024 A/Bribane/10/2007 12.02.2009 1459960 256 ZH/2 2048 1024 1024 1024 A/Bribane/10/2007 04.02.2009 1459971 16 ZH/2 64 32 32 64 16 A/Finland/9/08 04.02.2009 1460005 16 ZH/2 256 512 128 512 128 A/Bribane/10/2007 05.02.2009 1466500 128 ZH/2 256 256 256 256 256 A/Bribane/10/2007 05.02.2009 1470589 128 ZH/2 256 256 64 256 64 A/Bribane/10/2007 05.02.2009 1470613 64 ZH/2 128 128 64 64 64 A/Finland/9/08 05.02.2009 1470643 256 ZH/2 128 256 256 256 256 A/Finland/9/08 05.02.2009 1470693 128 ZH/2 256 512 128 512 128 A/Bribane/10/2007 05.02.2009 1470708 128 ZH/2 256 1024 512 512 512 A/Bribane/10/2007 05.02.2009 1470721 512 ZH/2 256 256 256 512 128 A/Bribane/10/2007 04.02.2009 1470763 8 ZH/2 128 128 32 128 32 A/Finland/9/08

37/41

Antiserum 2009 A/Brib07 A/Tries07 A/Wisc05 A/Finl08 A/Nep06 A/Bribane/10/2007 256 512 512 2048 1024 A/Trieste/25c/07 512 1024 512 512 512 A/Wisconsin/67/05 1024 20248 4096 1024 512 A/Finland/9/08 64 128 32 256 64 A/Nepal/921/06 128 512 256 256 2048

Date HA sample n° HA titre Cell/Passage A/Brib07 A/Tries07 A/Wisc05 A/Finl08 A/Nep06 Result IHA 17.02.2009 1470784 32 ZH/2 256 256 64 128 A/Finland/9/08 05.02.2009 1470803 32 ZH/2 128 128 64 128 128 A/Finland/9/08 05.02.2009 1470830 32 ZH/2 128 128 64 128 128 A/Finland/9/08 05.02.2009 1470856 256 ZH/2 256 256 256 256 256 A/Bribane/10/2007 05.02.2009 1470868 32 ZH/2 128 128 64 128 64 A/Finland/9/08 17.02.2009 1470888 8 ZH/2 256 128 128 256 A/Bribane/10/2007 17.02.2009 1470930 16 ZH/2 256 256 128 256 A/Bribane/10/2007 05.02.2009 1470944 16 ZH/2 256 256 64 128 A/Finland/9/08 03.02.2009 1474833 32 ZH/1 64 32 31 32 16 A/Bribane/10/2007 10.02.2009 1479911 32 ZH/2 256 1024 512 512 A/Bribane/10/2007 02.03.2009 1479951 64 ZH/2 256 512 128 512 256 A/Bribane/10/2007 10.02.2009 1479985 32 ZH/2 256 128 16 16 A/Bribane/10/2007 19.03.2009 1480025 512 ZH/2 256 1024 512 256 256 A/Bribane/10/2007 19.03.2009 1482505 512 ZH/2 128 256 64 256 256 A/Finland/9/08 03.02.2009 1482515 32 ZH/1 64 16 16 16 16 A/Bribane/10/2007 20.03.2009 1482525 128 ZH/2 128 256 64 128 128 A/Finland/9/08 10.02.2009 1482540 8 ZH/2 128 128 64 64 A/Finland/9/08 19.03.2009 1482556 128 ZH/2 128 256 128 256 256 A/Finland/9/08 10.02.2009 1482616 16 ZH/2 64 32 <16 16 A/Bribane/10/2007 16.02.2009 1482621 32 ZH/2 256 1024 256 512 A/Bribane/10/2007 16.02.2009 1484117 32 ZH/2 128 128 256 256 A/Finland/9/08 16.02.2009 1484159 32 ZH/2 256 256 64 256 A/Bribane/10/2007 16.02.2009 1484307 64 ZH/2 256 256 64 256 A/Bribane/10/2007 16.02.2009 1484346 64 ZH/2 256 256 128 256 A/Bribane/10/2007 16.02.2009 1484351 8 ZH/2 256 256 64 128 A/Finland/9/08 01.04.2009 1491091 128 ZH/1 128 512 256 512 512 A/Bribane/10/2007 20.03.2009 1491500 128 ZH/2 256 256 128 256 256 A/Bribane/10/2007 16.02.2009 1491512 64 ZH/2 256 256 64 128 A/Finland/9/08 16.02.2009 1491539 8 ZH/2 256 1024 128 128 A/Finland/9/08 16.02.2009 1491558 64 ZH/2 256 128 64 256 A/Bribane/10/2007 16.02.2009 1491736 8 ZH/2 256 128 64 128 A/Finland/9/08 05.02.2009 1498532 8 ZH/2 128 128 64 64 A/Finland/9/08 16.02.2009 1498543 64 ZH/2 128 64 16 64 A/Finland/9/08 16.02.2009 1498555 8 ZH/2 128 128 64 128 A/Finland/9/08 23.02.2009 1498559 128 ZH/2 32 64 64 64 64 A/Finland/9/08 23.02.2009 1498565 128 ZH/2 64 128 64 128 128 A/Finland/9/08 23.02.2009 1498571 128 ZH/2 128 512 256 512 512 A/Bribane/10/2007

38/41

Antiserum 2009 A/Brib07 A/Tries07 A/Wisc05 A/Finl08 A/Nep06 A/Bribane/10/2007 256 512 512 2048 1024 A/Trieste/25c/07 512 1024 512 512 512 A/Wisconsin/67/05 1024 20248 4096 1024 512 A/Finland/9/08 64 128 32 256 64 A/Nepal/921/06 128 512 256 256 2048

23.02.2009 1498578 128 ZH/2 128 256 128 256 256 A/Finland/9/08 04.02.2009 1498583 16 ZH/1 256 256 64 128 64 A/Finland/9/08 02.03.2009 1498609 32 ZH/3 1024 4096 512 1024 1024 A/Bribane/10/2007 02.03.2009 1498619 32 ZH/3 1024 4096 1024 2048 2048 A/Bribane/10/2007 20.03.2009 1498644 128 ZH/3 256 512 128 256 256 A/Bribane/10/2007 02.03.2009 1498708 64 ZH/1 128 128 64 128 A/Finland/9/08 02.03.2009 1498716 32 ZH/2 1024 2048 1024 2048 8192 A/Bribane/10/2007 20.03.2009 1504733 16 ZH/2 128 512 64 128 128 A/Finland/9/08 23.02.2009 1504777 8 ZH/1 128 256 128 128 128 A/Finland/9/08 11.02.2009 1504804 128 ZH/1 64 32 16 32 A/Bribane/10/2007 10.02.2009 1504863 16 ZH/1 256 512 128 512 A/Bribane/10/2007 20.03.2009 1504900 32 ZH/2 256 512 128 256 256 A/Bribane/10/2007 20.03.2009 1505267 16 ZH/2 256 256 64 256 128 A/Bribane/10/2007 11.02.2009 1505296 64 ZH/1 256 256 128 256 A/Bribane/10/2007 10.02.2009 1506729 16 ZH/1 256 256 64 128 A/Finland/9/08 10.03.2009 1506733 16 ZH/4 128 512 128 256 128 A/Finland/9/08 11.02.2009 1506734 16 ZH/1 128 128 64 128 A/Finland/9/08 20.03.2009 1506736 128 ZH/3 256 512 64 128 512 A/Finland/9/08 20.03.2009 1506736 256 ZH/3 256 512 64 128 512 A/Finland/9/08 09.04.2009 1506738 16 ZH/2 256 1024 512 512 1024 A/Bribane/10/2007 23.02.2009 1514435 32 ZH/2 32 32 16 32 32 reduced 02.03.2009 1514440 16 ZH/2 512 1024 256 1024 256 A/Bribane/10/2007 17.02.2009 1514457 64 ZH/1 256 256 128 128 A/Finland/9/08 01.04.2009 1514461 16 ZH/1 256 512 256 512 512 A/Bribane/10/2007 16.02.2009 1514466 64 ZH/1 64 32 16 32 A/Bribane/10/2007 23.02.2009 1514470 128 ZH/2 128 256 256 256 256 A/Finland/9/08 13.03.2009 1514486 32 ZH/2 128 512 64 128 128 A/Finland/9/08 01.04.2009 1514524 32 ZH/1 512 2048 512 1024 512 A/Bribane/10/2007 01.04.2009 1514551 16 ZH/1 256 512 256 256 256 A/Bribane/10/2007 16.02.2009 1514568 64 ZH/1 256 32 32 64 A/Finland/9/08 02.03.2009 1514576 256 ZH/2 256 256 64 256 256 A/Bribane/10/2007 23.02.2009 1514848 128 ZH/2 64 128 32 128 64 A/Finland/9/08 23.02.2009 1514856 128 ZH/2 64 128 16 128 32 A/Finland/9/08 23.02.2009 1514865 128 ZH/2 32 64 64 128 64 A/Finland/9/08 19.03.2009 1520849 256 ZH/3 256 512 512 256 256 A/Bribane/10/2007 19.03.2009 1520864 128 ZH/3 64 512 256 256 256 A/Finland/9/08 19.03.2009 1520871 32 ZH/3 128 512 256 512 512 A/Bribane/10/2007 19.03.2009 1520871 32 ZH/3 128 512 256 512 512 A/Bribane/10/2007 19.03.2009 1520888 32 ZH/3 128 1024 128 256 256 A/Finland/9/08 19.03.2009 1520930 16 ZH/3 256 256 128 512 1024 A/Bribane/10/2007

39/41

Antiserum 2009 A/Brib07 A/Tries07 A/Wisc05 A/Finl08 A/Nep06 A/Bribane/10/2007 256 512 512 2048 1024 A/Trieste/25c/07 512 1024 512 512 512 A/Wisconsin/67/05 1024 20248 4096 1024 512 A/Finland/9/08 64 128 32 256 64 A/Nepal/921/06 128 512 256 256 2048

20.03.2009 1520940 32 ZH/3 64 256 32 128 128 A/Finland/9/08 20.03.2009 1520953 128 ZH/3 64 64 32 128 32 A/Finland/9/08 19.03.2009 1520961 64 ZH/3 128 128 64 128 128 A/Finland/9/08 20.03.2009 1520988 32 ZH/3 256 512 64 256 256 A/Bribane/10/2007 20.03.2009 1520995 64 ZH/3 256 512 128 512 256 A/Bribane/10/2007 18.03.2009 1522877 8 ZH/1 64 64 16 64 16 A/Finland/9/08 18.03.2009 1522896 64 ZH/1 64 64 16 128 32 A/Finland/9/08 01.04.2009 1528756 128 ZH/2 256 1024 512 512 512 A/Bribane/10/2007 20.03.2009 1528968 16 ZH/1 128 256 64 128 64 A/Finland/9/08 18.03.2009 1528997 32 ZH/3 256 1024 128 512 256 A/Bribane/10/2007 20.03.2009 1529031 32 ZH/1 64 64 16 64 32 A/Finland/9/08 20.03.2009 1530769 16 ZH/1 64 128 128 64 128 A/Finland/9/08 19.03.2009 1530805 32 ZH/1 256 512 128 256 256 A/Bribane/10/2007 12.03.2009 1539334 64 ZH/2 64 128 64 256 32 A/Finland/9/08 10.03.2009 1539347 16 ZH/2 32 64 64 64 64 A/Finland/9/08 12.03.2009 1545836 128 ZH/2 512 2048 512 512 512 A/Bribane/10/2007 19.03.2009 1545896 64 ZH/2 64 128 64 128 128 A/Finland/9/08 12.03.2009 1546163 256 ZH/2 256 1024 1024 1024 1024 A/Bribane/10/2007 10.03.2009 1553909 8 ZH/2 256 1024 64 512 128 A/Bribane/10/2007 10.03.2009 1561644 32 ZH/2 256 1024 256 512 512 A/Bribane/10/2007 12.03.2009 1562431 64 ZH/2 256 512 256 256 256 A/Bribane/10/2007 12.03.2009 1568535 128 ZH/2 256 512 256 256 256 A/Bribane/10/2007 18.03.2009 1570509 128 ZH/1 256 512 256 256 256 A/Bribane/10/2007 13.03.2009 1570597 128 ZH/2 128 256 128 128 256 A/Finland/9/08 09.04.2009 1580835 32 ZH/2 16 32 <16 32 16 reduced 13.03.2009 1588808 64 ZH/2 512 2048 512 1024 512 A/Bribane/10/2007 13.03.2009 1596358 32 ZH/2 64 128 16 64 32 A/Finland/9/08 10.03.2009 1611155 512 ZH/1 512 A/Bribane/10/2007 10.03.2009 1611171 512 ZH/1 512 A/Bribane/10/2007 13.03.2009 1612685 16 ZH/1 128 64 64 128 64 A/Finland/9/08 13.03.2009 1615266 128 ZH/1 256 512 512 512 A/Bribane/10/2007

40/41

ANNEX 3: INFLUENZA B HEMAGGLUTINATION Antisera 2008 B/HK05 B/Vict06 B/Malay04 B/Barce08 B/Flori06 B/Brisb07 B/Bengl.07 B/Hong Kong/45/05 32 128 512 <8 <8 <8 <8 B/Victoria/304/06 16 256 256 <8 <8 <8 <8 B/Malaysia/2506/04 32 256 512 <8 <8 <8 <8 B/Barcelona/143/08 <8 <8 <8 64 32 32 64 B/Florida/04/06 <8 <8 64 64 512 1024 512 B/Brisbane/3/07 <8 <8 64 32 1024 2048 512 B/Bengladesh/3333/07 <8 <8 32 32 128 256 256

Date HA Prel. TITRE

HA Cell./Passage B/HK05 B/Vict06 B/Malay04 B/Barce08 B/Flori06 B/Brisb07 B/Bengl.07 Result IHA 09.12.2008 1334249 16 ZH/1 <16 <16 <16 128 64 128 128 B/Barcelona/143/08 22.01.2009 1429326 1024 ZH/1 16 <16 16 64 32 64 64 B/Barcelona/143/08 02..03.2009 1553456 1024 ZH/1 16 64 64 <16 <16 <16 <16 B/Hong Kong/45/05 02.03.2009 1576803 256 ZH/1 16 32 64 <16 <16 <16 <16 B/Hong Kong/45/05 02.03.2009 1576819 256 ZH/1 16 32 64 <16 <16 <16 <16 B/Hong Kong/45/05 02.03.2009 1588667 256 ZH/1 16 32 32 <16 <16 <16 <16 B/Hong Kong/45/05 01.04.2009 1593374 1024 ZH/1 16 32 64 <16 <16 <16 <16 B/Hong Kong/45/05 18.03.2009 1599302 1024 ZH/1 16 32 64 <16 <16 <16 <16 B/Hong Kong/45/05 12.03.2009 1611379 512 ZH/1 16 16 32 <16 <16 <16 <16 B/Hong Kong/45/05 12.03.2009 1611431 512 ZH/1 16 16 64 <16 <16 <16 <16 B/Hong Kong/45/05 12.03.2009 1611458 1024 ZH/1 16 16 16 <16 <16 <16 <16 B/Hong Kong/45/05 13.03.2009 1612706 512 ZH/1 16 32 32 <16 <16 <16 <16 B/Hong Kong/45/05 26.03.2009 1621443 512 ZH/1 16 32 64 <16 <16 <16 <16 B/Hong Kong/45/05 19.03.2009 1621593 512 ZH/1 16 16 32 <16 <16 <16 <16 B/Hong Kong/45/05 19.03.2009 1631388 512 ZH/1 16 32 64 <16 <16 <16 <16 B/Hong Kong/45/05 18.03.2009 1631402 512 ZH/1 16 32 64 <16 <16 <16 <16 B/Hong Kong/45/05 18.03.2009 1638463 512 ZH/1 16 16 32 <16 <16 <16 <16 B/Hong Kong/45/05 18.03.2009 1638465 512 ZH/1 16 32 32 <16 <16 <16 <16 B/Hong Kong/45/05 26.03.2009 1656833 512 ZH/1 16 32 32 <16 <16 <16 <16 B/Hong Kong/45/05 10.03.2009 1538680 16 ZH/2 32 <16 <16 <16 <16 <16 <16 B/Hong Kong/45/05 12.03.2009 1538680 16 ZH/1 32 <16 <16 B/Hong Kong/45/05 12.03.2009 1561775 16 ZH/2 32 <16 <16 B/Hong Kong/45/05 13.03.2009 1573595 128 ZH/2 32 <16 <16 <16 <16 <16 <16 B/Hong Kong/45/05 19.03.2009 1621469 512 ZH/2 32 32 32 <16 <16 <16 <16 B/Hong Kong/45/05 19.03.2009 1621652 2048 ZH/1 32 64 64 <16 <16 <16 <16 B/Hong Kong/45/05 26.03.2009 1623234 512 ZH/1 32 32 64 <16 <16 <16 <16 B/Hong Kong/45/05 18.03.2009 1623276 512 ZH/2 32 32 32 <16 <16 <16 <16 B/Hong Kong/45/05 17.03.2009 1639141 1024 ZH/1 32 64 64 <16 <16 <16 <16 B/Hong Kong/45/05 12.03.2009 1647791 1024 ZH/1 32 64 64 <16 <16 <16 <16 B/Hong Kong/45/05 01.04.2009 1514818 16 ZH/1 64 <16 <16 <16 <16 <16 <16 B/Hong Kong/45/05 12.03.2009 1561775 32 ZH/2 64 <16 <16 <16 <16 <16 <16 B/Hong Kong/45/05 10.03.2009 1562056 32 ZH/2 64 <16 <16 <16 <16 <16 <16 B/Hong Kong/45/05 26.03.2009 1652056 16 ZH/2 64 <16 <16 B/Hong Kong/45/05 23.02.2009 1504699 16 ZH/1 128 <16 <16 <16 <16 <16 <16 B/Hong Kong/45/05

41/41

Antisera 2008 B/HK05 B/Vict06 B/Malay04 B/Barce08 B/Flori06 B/Brisb07 B/Bengl.07 B/Hong Kong/45/05 32 128 512 <8 <8 <8 <8 B/Victoria/304/06 16 256 256 <8 <8 <8 <8 B/Malaysia/2506/04 32 256 512 <8 <8 <8 <8 B/Barcelona/143/08 <8 <8 <8 64 32 32 64 B/Florida/04/06 <8 <8 64 64 512 1024 512 B/Brisbane/3/07 <8 <8 64 32 1024 2048 512 B/Bengladesh/3333/07 <8 <8 32 32 128 256 256 12.03.2009 1561747 16 ZH/1 128 <16 <16 <16 <16 <16 <16 B/Hong Kong/45/05 26.03.2009 1663390 1024 ZH/1 32 32 32 <16 <16 <16 <16 B/Hong Kong/45/05 26.03.2009 1663403 1024 ZH/1 32 32 32 <16 <16 <16 <16 B/Hong Kong/45/05 26.03.2009 1665530 1024 ZH/1 32 64 32 <16 <16 <16 <16 B/Hong Kong/45/05 26.03.2009 1665566 1024 ZH/1 32 64 64 <16 <16 <16 <16 B/Hong Kong/45/05 26.03.2009 1665574 1024 ZH/1 16 32 32 <16 <16 <16 <16 B/Hong Kong/45/05 09.04.2009 1665582 1024 ZH/1 16 64 64 <16 <16 <16 <16 B/Hong Kong/45/05 02.04.2009 1673467 512 ZH/1 32 16 32 <16 <16 <16 <16 B/Hong Kong/45/05 02.04.2009 1673538 1024 ZH/1 16 16 16 <16 <16 <16 <16 B/Hong Kong/45/05 09.04.2009 1673909 1024 ZH/1 32 32 32 <16 <16 <16 <16 B/Hong Kong/45/05 02.04.2009 1673921 512 ZH/1 32 32 32 <16 <16 <16 <16 B/Hong Kong/45/05 02.04.2009 1673941 1024 ZH/1 32 32 64 <16 <16 <16 <16 B/Hong Kong/45/05 02.04.2009 1690435 1024 ZH/1 32 32 64 <16 <16 <16 <16 B/Hong Kong/45/05 02.04.2009 1690466 1024 ZH/1 16 32 32 <16 <16 <16 <16 B/Hong Kong/45/05 02.04.2009 1690482 1024 ZH/1 16 16 16 <16 <16 <16 <16 B/Hong Kong/45/05 09.04.2009 1698777 512 ZH/1 16 16 16 <16 <16 <16 <16 B/Hong Kong/45/05 09.04.2009 1699078 512 ZH/1 16 16 32 <16 <16 <16 <16 B/Hong Kong/45/05 09.04.2009 1716891 256 ZH/1 16 16 32 <16 <16 <16 <16 B/Hong Kong/45/05 16.04.2009 1716945 256 ZH/1 16 16 32 <16 <16 <16 <16 B/Hong Kong/45/05 26.01.2009 1451508 512 ZH/1 <16 <16 64 <16 <16 <16 <16 reduced 02.03.2009 1573575 512 ZH/1 <16 32 64 <16 <16 <16 <16 reduced

02.03.2009 1576753 256 ZH/1 <16 16 32 <16 <16 <16 <16 reduced

B/England/393/08 12.03.2009 1611155 - 12.03.2009 1611171 - 13.03.2009 1612759 1024 ZH/1 <16 16 32 <16 <16 <16 <16 reduced 19.03.2009 1612804 1024 ZH/1 <16 16 32 <16 <16 <16 <16 reduced 19.03.2009 1638475 512 ZH/1 <16 16 32 <16 <16 <16 <16 reduced 26.03.2009 1639437 512 ZH/1 <16 32 32 <16 <16 <16 <16 reduced 02.04.2009 1673493 512 ZH/1 <16 32 32 <16 <16 <16 <16 reduced 16.04.2009 1690839 512 ZH/1 <16 32 32 <16 <16 <16 <16 reduced 16.04.2009 1724714 512 ZH/1 <16 16 32 <16 <16 <16 <16 reduced

: