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!

!
(Seasonal Influenza or Seasonal Flu)

H1N1H3N2 A / B
!

)Influenza Season or Flu Season*


7 1 6 30

)Influenza Epidemic Period*


11 3
6 9

(Complicated Influenza )

(
)()

(Pneumonia and Influenza, P&I)


1975
(ICD-9) 480-487
(480)(481)(482)
(485*(486)(487)

(Cluster)
!!!
!

33

2008

2009 H1N1
2011 9
2012
2011 12

H5N1

IC
Rapiacta

2013

44


(Description of Disease)

(Influenza )
(Influenza virus) 1 4 (1,2)

( 1-3) 10

3~7 2
(2,3)

common cold

(Influenza)
(Common Cold)

200

3-4

1-2
2-5

()

()

55

(Reye syndrome)(1-5)


1933 Wilson Smith(1-3)

A (H1N1)
ABC A B
A
B
1977 A (H1N1 H3N2)
B

)Orthomyxoviridae)
ABC Segmented RNA
80-120nmenvelope 2
- Hemagglutinin HA, H
NeuraminidaseNA, N (1-5) M1
M2A B 8
(-)RNA (-)RNA NucleoproteinNP
TranscriptaseRNA PB1PB2PA C
7 (1,3,4)
(HA NA) A
(H ) (N )

66

H 17 H1-H17N 10
N1-N10(1-4,7) HA NA H1N1 A
HA-1 NA-1 H3N2 A
HA-3 NA-2 B C (1-4)
A B

A H1N1H2N2H3N2
H1N1
H7N7 H3N8 B
B C
(1,3,4)

A
B

8
8

11
11

(Antigenic

(Antigenic drift)
drift)
(Antigenic shift)

C
7
9

(Antigenic drift)

1957 1968
( A/H2N2 A/H3N2)

77

2009 H1N1

1. A B 8 RNA C
7
2.

3.

4.

5. Zanamivir(Relenza ) Oseltamivir(Tamiflu )

6.
Reassortment

ABC 1980 (WHO)


H N
(1,6)A
(HnNn)
A/swine/Lowa/15/33 (H1N1)
A 1933 Lowa H1N1
15
A/Beijing/262/95 (H1N1) A 1995 262
Beijing H1HA N1NA(1-4)
B ///
HA NA B/Beijing/243/97 C
(1-4)

88

A
-

(1,3,4)
RNA RNA
"""#
HA NA
"Antigenic drift"""
A B

"Antigenic shift""
"
20-30 (1,3,4) A

48 60
5 10
(1,3,6)

(NA)
Shedding

Viremia
(1-4)

99


Mode of transmission!

Incubation period
1~4 2

12 3

Period of communicability
3~5 7-10 (1,2)
24-48
24-72 5 (4,6)
(1,2,5)
21

Susceptibility and resistance

Antigenic drift(2,5)

(2,5)

A B
A C

10
10

Pandemics
(Seasonal epidemics)(Sporadic cases*
(WHO) 3 5
25 50 (1)
(Centers for Disease Control and Prevention,
CDC) 10 3
10 4
2~4 5
100 (100/100,000)
500/100,000(2,6,8)2012-2013
11 1
A/H3N2 (9)
11 3 4 2012-2013 12 2
(EuroFlu) 2012-2013
94,000 63% A 2009 A/H1N1
( 66%) (9)!

Centre for

Health Protection
12
7 8 2012-2013 A/H3N2
2009 A/H1N1 (10) 2012-2013
11 2013 1 2
A/H3N2(9,11)
(National Notifiable
Diseases Surveillance System(NNDSS)) 5-6
8-9 2013 (1 1 8 4 )
1,164 A/H3N2
(12)

11
11

7 1 6 30
11 12 3 11%
(13)

B A/H3N2 2009 H1N1


2009 7 H1N1
2010-2011 A/H3N2 12
2009 A/H1N1 2011-2012 B
A/H3N2 2012-2013 2010-2011

2009-2013

2008-2009 28 2009-2010 1,312


2010-2011 1,785 2011-2012 1,704 2012-2013 968
7 49 140 155 68 2009

12
12

H1N1
2008-2009 65
1-6 2009-2010 H1N1 18
0-6 65 (13)2010-2011 65 0-3
2012-2013 65 50-64 2009
A/H1N1
0-6 65

(http://www.cdc.gov.tw)//

7 10

10-20
40-50

13
13

B A
B A Antigenic
drift Antigenic shift

1-2

6 65 65
1996 2011 7.7% 10.9%

14
14


(Surveillance System)

1.

2.
3.
4.
5.
6.

IC

1999
2000
2004
2007
2011 9

15
15

3 39

(
)

+
(
)()
*
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O
( )
1.

()
2.-
(https://ida4.cdc.gov.tw/hospital/)

3.
-(https://ida4.cdc.gov.tw/phb/)

4.

16
16

21382

1.:https://ida4.cdc.gov.tw/hospital
2.
()

(
)

1.:https://ida4.cdc.gov.tw/phb

()

2.(
)

17
17

(
)

()()

1.Possible case
2.Probable case

3.Confirmed case
(1)
(2) 14 ()

(3)

1.()
2.
3.
4.4

( )

(http://nidss.cdc.gov.tw) /

18
18

1. ()
2.

1.
2.

1. (
)

2. (
)

21

14
()

**

* ;
!

14 )*
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19
19

()
1999 3

255

1.

2.

3.
()
1.

(1)38
(2)

2. 255 (
)
3.

2 2012 8

20
20

4.
(1)

()

(
)

(A)

Transtube

(B)

21
21

(2)
a. 4
b. 24
c.
(
*

(3) (real-time PCR*

()()
1.
(1)
(2)
a. 4
b. 24

(3)(real-time RTPCR*
2.

Real-time PCR()

Virus isolation ()
Real-time PCR()

Virus isolation ()
1. Virus isolation Real-time PCR()

Virus isolation ()
2. Real-time
Real-time PCR()

Virus isolation ()
PCR

Real-time PCR()

Virus isolation ()
Real-time PCR(+)rapid antigen testvirus isolation(+) 14

21 21
21

H1H3 INF untype,

!Real-time PCR

22
22

(
)


(http://cdc.gov.tw)//
//

( )

IC
Real-time Outbreak and Disease
SurveillanceRODS1999 9

2004 RODS
2006 11
ICD-9-CM

1701080%
ICD-9-CM

RODS
2008
IC
IC

RODS

23
23

RODS
(http://nidss.cdc.gov.tw)//
!

2009H1N1
H1N1

( )

(1) 38

24
24

(2)
(3)
(
)
1.

https://ida4.cdc.gov.tw/phb/
2.

( )
1.
2.

3.
bar-code (
)

()

(http://www.cdc.gov.tw)//

25
25


!!

!!https://203.65.72.161/ida2/
(https://ida4.cdc.gov.tw/phb/)!

2/,!

3/A+H1H3

26
26

2002
665

( )


1. 38C
2.

()
1.

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27
27

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28
28

!!

24

24

24

..

1:
&

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&

29
29

Long term care facilityThe elderly


apartmentInstitution for disability
ProtectoryVeterans homePrison
Nursing homeDay-care center for mental recovery

( )

1.

(1) 38
(2)

2.

!
1.
38
37.5
2. 3

( )
1.
(http://iss.cdc.gov.tw)

30
30

2. 24

3.
10

4.

( )

( )

( )

31
31

;
--
)*

--<-
/

(http://www.cdc.gov.tw)//

32
32

()
1.Nasal swab
2.Nasopharyngeal swab
3.Nasal wash fluid
4.Throat swab
5.
Transtracheal aspirateBronchoalveolar lavage fluidSputumLung
biopsy Post-mortem lung or Tracheal tissue
6.
Acute serum
Convalescent serum

()

3 (Nasopharyngeal
aspirate)Throat swab

Immunofluorescence staining
assay, IFA 4
4
48-72
-70

33
33

()
1. 10g veal infusion broth 2g bovine albumin fraction V
400 ml
2. 0.8 ml gentamicin sulfate solution (50mg/ml) 3.2 ml amphotericin
B (250ug/ml)
3.
() 0.8NaCl

()Rapid antigen detection


Enzyme immunoassay
Nucleoprotien, NP A B
15-30

()Virus culture
7-14 MDCK (MadinDarby Canine Kidney cell)
Cytopathic Effect, CPE

Hemagglutination- inhibition, HI

() RT-PCR Real-time RT-PCR


7-14
Reverse
transcriptase-Polymerase chain reaction (RT-PCR) Realtime RT-PCR

34
34

RT-PCR Realtime RT-PCR


(Primer)
(RT-PCR )(Realtime RT-PCR )

(
)

Paired sera
10
4

Neutralization test, NT
Hemagglutination inhibition test, HINTHI
SubtypeStrainHI

Immunofluorescence staining assay, IFA


IFA
A B
A H1 H3 H5
Anti-mouse IgG FITC conjugate

Virus Culture

MDCK cell
(HI)
()

35
35

RT-PCR Real-time RT-PCR


RT-PCR
RNAPCR
Reverse transcriptase, RTRNAcDNAPCR
HANAM

WHO
RT-PCR
RT-PCR

RT-PCR
Internal control

36
36

27

10
(2,3,4)
A
B
A B B
A

(Viral pneumonia)
(Bacterial pneumonia)(Encephalitis)(Encephalophy)
(Myocarditis)(Pericarditis)(Tansverse myelitis)
.(Guillain-Barr syndrome)Reye syndrome(2-5)

65
()
72

37
37

48
48
65


M2 protein
(Neuraminidase)M2 protein A
neuraminidase AB
() M2 protein
Amantadine Rimantadine M2 protein
(ion channel) M2 protein
Amantadine Rimantadine
M2 protein M2 protein

Amantadine
Rimantadine Amantadine

Amantadine Catecholamime Rimantadine


Amantadine 10%
2%
Rimantadine

A (
*

() (Neuraminidase)

38
38

Zanamivir (Relenza TM)Oseltamivir (Tamiflu)


Peramivir (Rapiacta)A (Hemagglutinin)
(Neuraminidase)N-acetylneuraminic acid ( Sialic acid)
hemagglutinin
neuraminidase
Sialic acid
neuraminidase
neuraminidase

A B
Zanamivir 78%
15% 5-15%
2%
Oseltamivir
oseltamivir carboxylate 75%) 6-10
99%Rapiacta

ZanamivirOseltamivir Peramivir
36-48 1-2
0.4% neuraminidase
4%

)*!M2 protein
A Amantadine Rimantadine

() Neuraminidase
Oseltamivir Zanamivir
1-1.5

39
39


() M2 protein
M2 protein
Amantadine Rimantadine
H3N2 H1N1 100%

() Neuraminidase
1.5% Oseltamivir

M2 protein Neuraminidase M2
protein A B

Neuraminidase Oseltamivir
Zanamivir A B
M2 protein Neuraminidase
M2 protein
Amantadine
Rimantadine

Symmetrel,
Flumadine

Symadine
A
A

1
1

(10%)
(2%)

(90-100%) (90-100%)

Neuraminidase
Oseltamivir
Zanamivir
Peramivir
Tamiflu
Relenza
Rapiacta
A B

1-1.5
(2%)
(1.5%)

A B

1-1.5
(1.5%)

A B

() Oseltamivir (Tamiflu)
13() 40
1()

40
40

1.(10 ) 2 5
2.(10 ) 1 10
() Zanamivir (Relenza)
5()

1. 2 (25 ) 2 5
2. 2 (25 ) 1 10

41
41

-ACIP
6185 50

body-mass indext35)
<5t506
65
6

60-64

Inactivated trivalent influenza vaccines, TIV (Liveattenuated influenza vaccine, LAIV)(2,6)

Whole virus
vaccine
Split-virus vaccine
Subunit vaccine
Purified surface-antigen vaccine
HANA
(cell based)
influenza virus-like particle, VLP
WHO
83110
2

42
42


A(A/H1N1A/H3N2)B

3(2A1B)
4(2A2B)
1B
2013-2014

11
T
2

6570-90%(3-5)
30 70% ( Pneumonia and
Influenza, P&I)3040%
50~60%80%(2,5,6)

7612
8(15)

universal influenza vaccine

43
43

12 3 9
11

65 66

1-6

6 3 0.25 ml3
0.5 ml8
1
1

1 2

44
44


Guillain-Barr
1976
2009 H1N1
Guillain-Barr

1.
2.
3.

1.
2. 6
3. Guillain-Barr
4.

2-8 C

45
45

1.
2.
3.

1.

IC

2.
1
A.

B.

C.

D.

46
46

2
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3
LED
4 (http://www.cdc.gov.tw)

3.
12 3 9
11

23
10 1

4.
H5N1H5N1
H7N9

(BMI35)
/

5.
1 -

6.

47
47


1.

2.
3.
4.

1.

2.

3.
4.

1.

2.

1.

2.

3.

1.
2.

3.

4.

1.

2.
3.
4.

48
48


123

1.
2.
3.
4.

1.

2.
1
A.
B.
C.

D.

2
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B.
3
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49
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52
52

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(1)

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A.
B.

C.
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!!!!
5.
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)http://eir.cdc.gov.tw/ *
7.
(1)
7
(2)
)http://eir.cdc.gov.tw/ *!

53
53

!
!

1. World Health Organization.The world health report 2007 : a safer future : global
public health security in the 21st century. World Health Organization; 2007: 45-48.
2. Mandell, GL., Bennett, JE., and Dolin, R. Principles and Practice of Infectious
Diseases, 6th eds. Elsevier Inc. 2005: 2060-2078.
3. U.S. Centers for Disease Control and Prevention, National Center for Prevention
Services. Prevention and Control of Vaccine-Preventable Diseases: Influenza, 7th
eds. U.S. Dept. of Health and Human Services. 2003; 2006: 235-56.
4. Cox N.J. and Subbarao K. Influenza. Lancet; 1999;354: 1277-82.
5. Nicholson K.G., Wood J.M. and Zambon M. Influenza. Lancet; 2003; 362:1733-45.
6. U.S. Centers for Disease Control and Prevention. Prevention and control of
Influenza.Recommendations of the Advisory Committee on Immunization Practices
(ACIP). MMWR 2006;55(RR 10):1-42.
7. Centers for Disease Control and Prevention. Influenza Type A Viruses and
Subtypes, 2013. Available at: http://www.cdc.gov/flu/avianflu/influenza-a-virussubtypes.htm
8. World Health Organization. Influenza. Available at
http://www.who.int/csr/disease/influenza/en/
9. World Health Organization. Weekly Epidemiological Record. Review of the 20122013 Winter Influenza Season, Northern Hemisphere; 2013; 88:225-232
10. Sentinel Surveillance. Centre for Health Protection in Hong Kong. Monthly
summary tables of influenza virus isolation 2012, 2013. Available at:
http://www.chp.gov.hk/en/sentinel/26/44/292.html
11. National Institute of Infectious Disease. Isolation/detection of influenza virus in
Japan, week 36/2012-week 11/2013 (as of March 18, 2013). Available at:
http://www.nih.go.jp/niid/en/iasr-inf-e.html#graph
12. Australian Government Department of Health and Ageing. Australian Influenza
Surveillance Report; 2013 No.4
13. 2002
2004; 20:478-487
14. 2010
2012; 28:113-126
15. Masahiko Katayose et al. The effectiveness of trivalent inactivated influenza
vaccine in children over six consecutive influenza seasons. Vaccine 2011;29:184449.

54
54

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!
! !

!
55
55

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25/
15.https://ida4.cdc.gov.tw/hospital

()()
!
56
56

57

C Headache Myalgia Malaise Rhinorrhea Cough Sore throat Vomiting

!
!

! ! ! ! ! !



! ! ! ! ! ! ! !

Diarrhea Pharyngeal vesicles or ulcers Paralysis Skin rash Others! ! ! ! ! ! ! !

Fever!

VL-001

!!!!!!!!::/19/37 211/12/12 !

!!!

___________________

Bar-code

Throat swab
Nasopharyngeal secretion or swab
Others____________
Throat swab
Nasopharyngeal secretion or swab
Others____________

1382

!!
!!!!!!!!!!!!!!!!!!!!!!!!!

!
!

02-23123456*66908

02-87923311*13568

03-2118800 5497

04-23592525 5701

04-7238595 5930

06-2353535 2653

07-7317123*2578

03-85618255 2090

58
58

!!

!
!
!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!
!!!!!!!!!!!!!!!!!!!!!!!!!! 1.34 !
!!!!!!!!!!!!!!!!!!!!!!!!! 1.34 !
!!!!!
!!!!!!!!!!!!!!!!!
!!!! 49!!!!!!!!

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59
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61
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62
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64
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!!!
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65
65

66

MDCK cell

*MK-2 cell
CPE!

(-)

(+)

Blind
passage

z MK-2 cell H292 cell


z 200 l 1 mL
0.45m MDCK 7-10 CPE 3,000rpm
15 1 mL PBS 21
Acetone Influenza A Influenza B monoclonal antibody
indirect immunofluorescence staining assayIFA
apple green

3. Influenza A virus HA typingby RT-PCR method

2. Respiratory virusesfollow DAKO system direct IFA


10 l wet chamber 3715 PBS PBS
5

1.

!!

67

PCR follow Qiagen PCR core kit procedure


a. PCR
5X Q solution
10 l
10X buffer
5 l
25mM MgCl2
4 l
10mM dNTP
4 l
RT template
2 l
A/H1 (H3)-110 M 1 l
A/H1 (H3)-210 M 1 l
Taq polymerase
0.5 l
H2O
22.5 l
b. PCR program
40cycles
94 2
94 1
52 2
72 3
72 7

RT follow Promega RT procedure


(a). 15 l RNA 70 5 RNA
(b). 40 l
RNA
15 l
5X buffer
5 l
100mM dNTP mix
0.5 l
RT
1 l
Random primer
1 l
H2O
17.5 l

Qiagen Viral RNA Mini Kit


140 l RNA

50 Pl AVE buffer RNA RT

RNA

68

Roche (LightCycler 480 RNA master hydrolysis probes reagents procedure)

RNase-free water
0.3 L
Real-time RT-PCR
FluA-F primer10 M 1.0 L
RT reaction63
3 !
FluA-R primer10 M 1.0 L
Taq activation95 30
FluB-F primer10 M 1.0 L
PCR reaction
FluB-R primer10 M 1.0 L
95 10
FluA probe5 M
0.5 L
58 30!
FluB probe5 M
0.5 L
72 3 45 cycles
Enzyme master mix
7.4 L
Enhancer
1.0 L
Activator
1.3 L
RNA sample
5.0 L

ABI (fiollow TaqMan one-step RT-PCR master mix reagents procedure)

Real-time RT-PCR
2X Master mix buffer
12.5 L
RT reaction48
30
FluA-F primer10 M 1.0 L
Taq activation95 10
FluA-R primer10 M 1.0 L
PCR reaction
FluB-F primer10 M 1.0 L
95 15
FluB-R primer10 M 1.0 L
60
145 cycles
FluA probe5 M
0.5 L
FluB probe5 M
0.5 L
RNA enzyme mix
0.67 L
RNase-free water
1.83 L
template
5.0 L

FLUA/B

69

Run gel and read data


1.5agarose gel100 40 Ethidium bromide 5 5

!
!
!
!
!
!
!
!

!!
Primer A/H1 H1-1: 5-GAT GCA GAC ACA ATA TGT AGA GG-3
Primer A/H1 H1-2: 5-CNC TAC AGA GAC ATA AGC ATT T-3
Primer A/H3 H3-1: 5-TCA GAT TGA AGT GAC TAA TGC T-3
Primer A/H3 H3-2: 5-AAT TTT GAT GCC TGA AAC CGT-3
FluA-F :5- AAG ACC AAT CCT GTC ACC TCT GA -3
FluA-R :5- CAA AGC GTC TAC GCT GCA GTC C -3
FluB-F: 5- GAG ACA CAA TTG CCT ACC TGC TT -3
FluB-R: 5 TTC TTT CCC ACC GAA CCA AC -3
FluA probe- FAM -5 TTT GTG TTC ACG CTC ACC GT 3-TAMRA
FluB probe -VIC-5AGA AGA TGG AGA AGG CAA AGC AGA ACT AGC 3-TAMRA
FluA probe: 5FAM - TTTGTGTTCACGCTCACCGT 3BBQ
FluB probe: 5HEX-AGAAGATGGAGAAGGCAAAGCAGAACTAGC

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(1)38
(2)
(3)

1.
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2
3 MIS

70
70

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()
MIS

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71
71

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