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1
- -
,
- (RSV),
(MPV)
2004-2009 .
„ ”
01.06.13
:
. - , . .
2015
2
167 , 23 , 31
338 (12 326 ).
„ ”, ,
( )
-
, 06/11/2014 .
.....................2015 .
............ . ,
№26, ,
.
, .
„
”, , - .
3
I. ............................................................................................................................... 1
II. ......................................................................................................................... 4
III. ............................................................................................................... 5
1. RSV..................................................................................................... 5
2. , ................................... 5
3. , RSV……………………….. 6
4. .................................................................................................................................... 6
4.1. RSV .. ............................................. 6
4.1.1. ............................................................. 6
4.2. ( ) RSV..
7
4.3. RSV ....... 7
4.4. .................................................................................. 8
4.4.1. Trizol LS (Invitrogen, USA)............................................ 8
4.4.2. RIBOSORB (SACACE, Italy)........................................... 8
4.4.3. QIAAMP® VIRAL RNA MINI KIT (QIAGEN)............. 8
4.4.4. IPREP PURELINK TM
VIRUS KIT ............................... 9
4.5. RT-PCR RSV………………………… 10
4.5.1. RT-PCR RSV.............. 10
4.6. RT-PCR MPV……………….……….. 11
4.6.1. RT-PCR MPV................... 12
4.7. PCR ………………………………................................................ 13
4.8. Real Time RT- PCR ............................................................. 13
4.9 ................................................................. 13
IV. ……………………………………………………………………... 15
1. ...................................... 15
1.1. RSV MPV ...........................................................................................
16
1.2. ( 1N1)2009 ........................................................................
17
2 . RSV ............................................................. 18
3. RSV -
–Directigen RSV......................................................
23
4. RSV MPV............................................................... 26
4.1. RT-PCR 2006/ 2007................................................................ 27
4.2. RT-PCR 2007/ 2008............................................................... 30
4.3. RT-PCR 2008/ 2009............................................................... 34
4.4. ................................................................................................................................ 38
5. ( 1N1)2009....................................................................... 44
V. ......................................................................................................................... 58
VI. . ........................................................................... 65
VII. . ............................................................................... 67
VIII. ……………………………………... 68
4
:
- OP - RSV- - MPV-
PCR-
SARS-CoV, NL63, HKU1- . Coronaviridae
hBoV- KI, WU- . Polyomaviridae
ECDC-
CDC –Centers for Diseases Control and Prevention
AMPV- -
-
-
m -
ORF-
-2-
MRC-5-
BHK-
-
LLC-MK2-Rhesus Monkey Kidney Epithelial Cells -
ELISA ( - ) EIA-
- Nested PCR- -
Real Time PCR - -
PBS -Phosphate buffer solution / / RT-PCR- -
DEPC -diethylpyrocarbonate
FITC -fluorescin isothiocyanate
c.DNA - dNTP s –
-
5
I.
( )
.
,
. ,
,
.
World Lung Fondation
4,25
.
, - (RSV) -
3
. , 50%
15-35%
.
,
( ,
.).
(
)
. , ,
, ,
- (RSV), , ,
, .
,
. ,
.
, .
2001 . van den Hoogen
,
(MPV). ,
7-10% .
6
, MPV ,
RSV, .
- , , ,
. ,
RSV MPV ,
.
,
RSV,
, ,
RSV MPV . in
vitro,
, RSV,
,
.
.
,
- -
(PCR).
,
-2001 ., SARS-CoV -2003 , NL63 -2003 . HKU1 (2005 .),
(С V) -2005 . KI 2007 . АU-2007 .
( 5N1/) (1997 .)
, MERS-CoV2012 .
,
RT-PCR MPV .
RSV
(ECDC),
RSV
.
7
( 5N1/) (1997 .)
.
20- , 1999 .
,
.
17 2009 . CDC
A(H1N1)2009,
, .
24 2009 . ,
, 10 2009 .,
,
6- . 10
2010 .
„ ”, 2002 .
( 1N1)2009, ,
RSV
,
, ,
.
.
8
II.
,
- ,
,
( 1N1)2009 .
:
1.
.
2. RSV
- .
3. RT-PCR
RSV MPV Real Time RT-PCR
.
4. RSV MPV
5
.
5. Real Time RT-PCR
2009 .
.
6.
(H1N1)2009.
9
III.
1. RSV
RSV- Randall
-2, .
2. ,
RSV MPV
5 . 2004-2009 . RSV
846 , 452
MPV, 371 .
04.05.2009 . - 08.01.2010
( ), ,
. 2429
( 9 ).
23.09.2009 .-12.10.2009 . 152
.
RSV
, RSV.
-
. ,
+4 .
.
. RSV MPV,
Directigen RSV,
RT–PCR. BSL3
( )
,
rRT PCR.
10
9 ( ).
,
. ,
2ml ,
,
.
3. ,
RSV
RSV -2,
. -
MRC-5,
.
4.
4.1. RSV
-2 MRC-5 24
37 5% 2
. 0,2ml/ ( -
).
10 . 50%
, - ,
.
+4 .
4.1.1.
RSV
,
.
,
, .
11
- 24-36
. -
. ,
.
.
4.2. ( )
RSV
(chamber-slides)
,
, 16 , 0,25ml/ .
IMAGENTM
RSV Dako Cytomation.
, . ,
, 25 μl
.
16 0,4.
, .
-
, 3
.
4.3.
DIRECTIGEN RSV (BECTON DICKINSON, USA)
RSV
-
,
, 48 10 . -
RSV -
, ,
.
12
4.4.
- ,
.
4.4.1. TRIzol LS (Invitrogen, USA)
TRIzol LS e
. .
,
,
.
, .
. ,
.
.
. 250µl
, 25µl dd 20
(RNAse free), –70º
RT-PCR.
4.4.2. RIBOSORB (SACACE, ITALY)
Ribo-Sorb , -
.
;
100µl, -50µl.
4.4.3. QIAamp® Viral RNA Mini Kit
(QIAGEN)
- ( , )
.
„ ” ,
Carrier RNA, AVL
( ). ,
13
. ,
- , -
. ,
.
, .
140µl,
60/80l .
4.4.4. IPREP PURELINK TM
VIRUS
KIT
,
iPrep Purification
instrument, 40 .
Magtration® ( ) PSS (Precis ion System
Science),
, .
,
. ,
.
ChargeSwitch®
Technology (CST),
, .
. . CST®
.
.
8,5, ,
50 100µl. 200µl
400 µl.
14
4.5. RT-PCR
-
RSV in-h use
RT-PCR. RSV
N ( 858
1135– ).
:
N1 5’GGAACAAGTTGTTGAGGTTTATGAATATGC3'
N2 5'CTTCTGCTGTCAAGTCTAGTACACTGTAGT3'
RT-PCR Mastercycler
(Eppendorf) :
READY-TO- GO RT-PCR BEADS (AMERSHAM BIOSCIENCES, UK)
ABGENE REVERSE IT-ONE STEP RT-PCR (INVITROGEN, USA)
4.5.3. ABGENE ABSOLUTE MAX QRT PCR
e - ,
.
-Randall, - DEPC-treated H2O.
4.5.1. я RT-PCR RSV
RSV-Randall, -2,
RNA Real Time PCR (Respiratory Syncytial Virus
Real Time RT-PCR Kit, Liferiver, Shanghai Bio-Tech)-
RSV Chromo 4 (BioRad).
FA BHQ1
HEX/VIC/JOE .
1 107cop/ml RNA, .
,
15
RSV
30 380cop/ml . 3 104cop/ml.
( 10-1
10-6) 3 104
cop/ml
( 3 000 0,03 cop/ml).
RT-PCR. RT-
PCR
(H3N2) (H1N1). . 1
2%
RT-PCR. -
, RSV , 10-3
( 1:1 000 30 cop/ml).
.
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15
. 1 RT-PCR
RSV-Randall
- ; 1- ; 2 3 RSV – ; 4 5
; RSV 10-1 (6) 10-6
(11)
4.6. RT-PCR
MPV
M ,
16
.
:
M1 5`CCC TTT GTT TCA GGC CAA C 3`
M2 5`GCA GCT TCA ACA GTA GCT G 3`
RT-PCR Mastercycler
(Eppendorf) : Qiagen® One Step RT-PCR Kit (Qiagen) kit
e - ,
.
MPV Catacusino Institiut, Romania,
- DEPC-treated H2O.
4.6.1. я RT-PCR MPV
Cantacuzino Institute, Romania (Tecu C, et al. 2007)
MPV, LLC-MK2 1/10 1/10000
( 2) - , , 10-4
. (Tecu
C, et al. 2007)
MPV,
MPV Qiagen,
,
.
.2 RT-PCR
MPV. (Tecu C, et al. 2007)
17
4.7. PCR
PCR ( ),
з280bp RSV ~416bp MPV,
2% 0,5g/ ml.
–φ -174 (4l) DNA Ladder 100-1500bp
(Invitrtogen)
4.8. REAL TIME RT- PCR
Real Time RT-PCR (rRT-PCR) Super Script One-Step
RT-PCR with Platium Taq (Invitrogen, USA) iScript One Step RT-PCR Kit for
Probes (BioRad, USA) Real Time PCR Chromo 4 (BioRad),
(Taqman®)
( /H1, A/H3 B), CDC Atlanta, .
FAM/BHQ
( /H1, A/H3 B)
.
: InfA -
; swInfA
; sw 1
; InfB ; Inf 1 Inf 3
.
,
.
rRT-PCR:
A/Brisbane/59/2007(H1N1),
A/Brisbane/10/2007(H3N2) /Brisbane/60/2008,
(WHO-CCs)- ,
.
A/California/04/2009
CDC Atlanta, USA ,
0.01M PBS.
.
18
я :
CDC,
,
, “ ” (swInfA / swH1),
(threshold line)
40- ,
.
,
. “Case definition” ,
, ( - )
PCR A(H1N1)2009
.
.
.
4.9.
:
4.9.1.
4.9.1.1.
4.9.1.2. : ,
4.9.2. -
. α=0,05
.
4.9.2.1. -
(PОКrsШЧ χ2 tОst, CСТ sqЮКrО tОst).
4.9.3. (MS E cel
2007)
19
IV.
1.
2004/2005 .,
RSV 2 ,
.
.
,
.
5 ,
.
-
- , .
RSV
CDC
, - .
RSV
. -
.
MPV
5 , ,
.
17 2009 ., CDC
,
.
A(H1N1)2009
. .
„ ” ,
20
.
.
,
,
, .
1.1. RSV
MPV
№1 №2
846 RSV, 452
MPV 371 ,
.
MPV 2008 .
.
. №1 RSV
MPV 0 5 . /
2004/2005 . 2 .
** MPV 81 .
/
2004/2005* 2005/2006 2006/2007 2007/2008** 2008/2009
- - 33 - - -
II - - 141 101 130 241
13 - - - -
- - 18 - 32
- - 29 - -
- 10 - - - -
- - - - 98
23 174 148 130 371
21
. №2
1.2. ( 1N1)2009
№ 3 4
2429
( 1N1) , 04.05.2009 .-08.01.2010 .
. №3
/
. №4
(H1N1)2009
2004/2005 2005/2006 2006/2007 2007/2008 2008/2009
13 36 54 51 137
- 65 66 72 98
10 73 28 7 136
23 174 148 130 371
1174
596
659
2429
396
446
1 518
# 31
* 38
** 2 429
22
* – , , , , ,
, , , , ,
, , , , , ,
# - , , , .
**
.
2 . RSV
RSV ,
,
.
- , -
, . . « » .
, -
. ,
- MPV
2001 . RT-PCR
RSV,
.
,
.
2004-2009 . RSV
-2, „ ”,
. -
, -2
, ,
95%,
23
-2 24-29% .
2006/2007 2007/2008 .
MRC-5 ( ), „
”, . ,
. № 5
№5 RSV
-2
MRC-5
MRC-5
-2
2004/2005 23 2 (8,7%) - -
2005/2006 174 0 - -
2006/2007 148 2 (1,4%) 4 (2,7%) 3 (2%)
2007/2008 130 2 (1,5%) 0 0
2008/2009 371 2 (0,5%) - -
. 24 .
, -
.
( 48 10
).
, –
, , ,
.
, 48 .
-
.
, . ( .3 , 3 3 , .4 , 4 , .5)
24
3 . -2 , - ,
10 0,25
3 . -2 RSV-
, - , 10 0,25
25
3 -2 RSV-
, - , 10 0,25
4 . MRC-5 , - ,
10 0,25
26
4 . MRC-5 RSV –
, - , 10 0,25
5. -2 RSV -
, , 20 0,5,
IMAGEN RSV (Dako Cytomation)
27
№5 ,
( 846) RSV
15 .
-2 , MRC-5 -
- .
2004/2005 2006/2007
, (2007/2008
2008/2009) .
, . -
(9 ) 2006/2007,
RT-PCR.
. ,
.
, ,
.
.
- (4 ) MRC-
5 -2 2006/2007 ., ,
RSV
,
.
.
3.
RSV -
–DIRECTIGEN
RSV
( )
, PCR .
( 85%)
28
RSV, .
, .
, RSV (
15-20 .)
.
2003/2004 ,
48 ,
.
RSV,
– ,
. RT-PCR
( 2006/2007),
RSV
. №6
.
. №6 Directigen RSV
-
2004/2005 2005/2006 2006/2007
13 16 24
Directigen RSV 11* 4** 16
IF RSV 12* 10*** -
* .
** X, XI, XII 2005 .
*** I, II, III, IV 2006 .
29
2004/2005
RSV 23 . 11
Directigen RSV ( 12 )
.
10 , , 2005 .
RSV RT-
PCR RSV .
4 , ,
.
-2005/2006
,
( №6). 16 ( -2) 14
(87,5%) RSV,
.
,
,
( 177 ). ,
.
,
,
(RSV
). , Directigen RSV
RSV , ,
.
2006/2007,
RT-PCR. 7
24
.
30
. №7 24
, 2006/2007
Directigen
RSV
Directigen
RSV
RT-PCR
24
9
(37,5 %) 0
16
(66,6 %)
24
(100 %)
24 RT-PCR
, 9 , Directigen RSV
, 16
48 .
4. RSV MPV
RSV MPV RT-
PCR ( )
.
RT-PCR “ ”,
2005 ., 2006/2007 .
RSV 5 .
RT-PCR , RSV,
MPV.
5 ,
- .
MPV RT-PCR 2008 .
5 .
31
4.1. RT-PCR 2006/ 2007 .
2006/2007 . RT-PCR
RSV 148 ,
,
. RSV
.
RSV , 2 2007 . (
6.) RSV
,
2007 ., RSV
.
RSV 148
5 . , -
(631 ), ,
. ,
– RSV
( 3N2) .
RSV,
. 2006 . . .
.
№ 8 , 148
RT-PCR, 56 (37,8%) RSV,
.№9 , ,
RSV RT-PCR 84%. ,
,
. ( №10) -
(37 ) - ,
- . RSV
RT-PCR 24-48 .,
- .
32
RSV ( ) 27,7% 39,2%
, - ,
.
.7 RT-PCR
2% .
~280b ,
.
. №8 RSV RT-PCR, 2006/2007
(p<0.001, Pearson chi square=17.47).
. №9 RSV RT-
PCR
2006/2007
49 50-51 52-1 2 3
4 5 6
12 12 5 38 26 28 10 17
( )
2
(16,6%)
3
(25%)
4
(80%)
32
(84,2%)
7
(26,9%)
5
(17,9%)
1
(10,0%)
2
(11,8%)
RT-PCR
II - 101 49 (48,5%)
“ ” 29 2 (6,9%)
18 5 (27,7%)
148 56 (37,8%)
33
6. R -PCR RSV , 2006/2007 .
. №10. 56 RSV RT-PCR
, 2006/2007,
0- 11 12-35 36-59
RSV
/
RSV
/
RSV
/
54
20 /36
55,5%
7/18
38,8% 0/0
66 2/20
10%
3/22
13,6%
3/24
12,5%
28 15/17
88,2%
6/8
75% 0/3
37/73
50,6%
16/48
33,3%
3/27
11,1%
34
RSV 0-11
(p<0.001, Pearson chi square=15,37), 12-35
(p<0.001, Pearson chi square=10,35), 36 .
(p<0.001, Pearson chi square=13,69)
.7 RT-PCR 2% -
з280 . φX 174
4.2. RT-PCR RSV MPV 2007/ 2008
,
.
2007 . EISS (European Influenza Surveillance Scheme),
35
ECDC (European Centre for Disease
Prevention and Control),
, .
EISS ECDC,
RSV,
RSV. 2008 .
5 RSV MPV.
-
- 5 .
( 2007 .- 2008 .) RSV
130 5 ( .№1,
11, . 8), 2008 . 81
. RT-PCR RSV 11
(8,46%), 13 ( 81 ) (16.05%) MPV. (
15) . №12 24
.
MPV -
0 11 . (10 ), .
RSV - ,
( №12).
RSV,
,
2006 .
2007 ., RSV
.
RSV -2.
.8
,
, .
RSV MPV,
, ,
. MPV
36
(81 ),
.
. №11. RT-PCR RSV
MPV 2007/2008
. №12. RSV MPV 24
, 2007/2008
2007
2007
2007
2008
2008
2008
RSV
18 13 18
42 32 7
MPV
0 0 0
0 0 0
3 RSV
7%
2 MPV
4,7%
8 RSV
25%
8 MPV
25%
3 MPV
42%
0- 11 12-35 36-59
RSV MPV RSV MPV RSV MPV
/
/
/
51
7/33
21,2%
9/33
27,2%
2/17
11,7%
2/17
11,7% 0/1 0/1
72 2/39
5,1%
1/39
2,5% 0/27 0/27 0/6
1/6
16,6%
7 0/0 0/0 0/6 0/6 0/1 0/1
9/72
12,5%
10/72
13,8%
2/50
4%
2/50
4% 0/8
1/8
12,5%
37
RSV MPV 0-11 ,
( p<0.05, Pearson chi square=4,24 p<0.05,
Pearson chi square=9,16).
, - .
.
. 8 RT-PCR RSV, MPV
, 2007/2008
.9 RT-PCR MPV 2%
– з 416bp . φX 17
38
4.3. RT-PCR RSV MPV 2008/ 2009
2008/2009 . 371
5 , Real
Time -PCR RT-PCR RSV MPV,
.
« », . ( . №13)
. №13 5 ,
2008/2009
/
-
Real Time RT-PCR :
RSV MPV
241
16
6,6%
47
19,5%
18
7,5%
2 RSV (0,8%)
3 AdV (1,2%)
5 / 3N2/ (2%)
98 2
2%
4
4%
1
1% 0
32 0 7
21,8%
6
18,75%
10 AdV
31,3%
371 18
4,8%
58
15,6%
25
6,7%
RSV
,
, . RSV
- (15,6% 4,8%).
-2 RSV.
, (58
) .
,
, RSV, MPV AdV.
39
№13 ,
5 , -
,
RSV -58 (15,6%), MPV 25
(6,7%) -18 (4,8%). . №14
83 RSV MPV
18 ( )
. RSV
- , MPV e
.
.№14. RSV, MPV
, 2008/2009
0-11 ,
, (p<0.05, Pearson chi square=7,06) 36-
59 , (p<0.05, Pearson chi square=7,02).
,
,
0- 11 12-35 36-59
RSV MPV
RSV MPV
RSV MPV
/
/
/
137
28/100
28%
5/100
5%
3/100
3%
2/31
6,5%
3/31
9,6%
1/31
3,2% 0/6
1/6
16,6% 0/6
98 9/39
23%
2/39
5,1%
5/39
12%
6/41
14,6%
4/41
9,7%
1/41
2,4% 0/18
3/18
16,6%
3/18
16,6%
136 7/31
22% 0/31 0/31
3/46
6,5%
5/46
10,8%
4/46
8,7%
3/59
5%
2/59
3,4%
1/59
1,7%
44/170
26%
7/170
4,1%
8/170
4,7%
11/118
9,3%
12/118
10,2%
6/118
5%
3/83
3,6%
5/83
6%
4/83
4,8%
40
RSV (p<0.001, Pearson
chi square=22,69).
. 10 , MPV 2008/2009
,
.
10 , RSV, MPV Ad, 2008/2009
-
, ,
( ).
,
, (
)
, , -
.
, 3
, 7
.
,
41
.
. ( .№15) AdV
,
-2 RSV MPV.
AdV , -
,
.
, -
RSV MPV.
– ,
.
RSV. ( . 10).
, , 22
. ,
, ,
- AdV MPV.
.
. №15
5 . .
PCR
„
”
,
38
20
7 RSV
2 MPV 0
„
”
,
15
7 7 AdV
3 MPV 10 AdV
5 3 AdV
1 MPV
42
4.4.
RSV MPV RT-PCR
( 24 48 .),
,
,
.
,
.
. . „ ”
,
.
2006/2007 RSV ,
RT-PCR 37,8% 6,1%
8,46% 1,5% 15,6% 0,54% . ( .№16)
-
RT-PCR .
RT-PCR ( 19,2% 2%
, )
,
, RSV,
,
.
.
43
. №16 RSV
RT-PCR
RSV
:
2006/2007
2007/2008
2008/2009
9
6,1%
2
1,5%
2
0,54%
13
2%
RT-PCR 56
37,8%
11
8,46%
58
15,6%
125
19,2%
148 130 371 649
%
(p<0.001, Pearson chi square=16,89),
% RT-PCR (p<0.001, Pearson chi square=45,71).
,
.
RSV
.11.
, RSV
2006/2007 , - (
№ 17),
.
9 RSV
2006/2007, .
RSV
RT-PCR - 84%,
.
, RSV
– ,
,
-
. 2008/2009
( № 17), MPV .
44
. №17 RSV MPV
* -
RSV
(p<0.001, Pearson Chi square=33.95).
%
2006/2007 .,
, .
2008/2009 . ((p<0.05, Pearson Chi square=4,42).
% MPV
2007/2008 2008/2009 . (p<0.05, Pearson Chi square=7,74).
% 2008/2009
.
%
2008/2009 .
(p<0.05, Pearson Chi square=4,79).
2006/2007 2007/2008 2008/2009
RSV RSV MPV RSV MPV
/
/
/
51/130
39,2 %
11/130
8,5%
13/130
13%
47/241
19,5%
18/241
7,5%
16/241
6,6%
*
5/18
27,7% - -
11/130
8,4%
7/130
5,3%
2/130
1,5%
45
. 11 RT-PCR
RSV .
RSV MPV
. - , -
.
, 2008/2009 ( . №15)
RSV MPV 2 , -
, MPV .
,
.
12 MPV
2008 . ,
,
-13 25 ,
.
- ,
.
MPV
46
2008/2009.,
. (
17).
12 RT-PCR
MPV
RSV
- , .
RSV - ( 0 4 .)
- 40,8% RSV
, 11% .
5 ,
- 0 11 (n=315), 12 35
(n=216) 36 59 (n=118). 13 , -
RSV 90 (28,5%) 11
. 169 (53,6%) ,
47
55 (32,5%) RSV . -
- 29 (13,4%) RSV
, 66 (30,5%) -11 (16,6% RSV ),
6 (5%) RSV ,
. RSV
-
, 35
, .
MPV ,
-(7%,
8,3% 6,5%), , ,
,
5 .
13 RT-PCR
48
5. ( 1N1)2009
2009 . ( )
,
.
, ,
.
. . „ ”
,
. . 2008 .
« » rRT-PCR
/ 1, / 3
/ 5. CDC- Atlanta.
, CDC
, ,
CDC , .
,
, 100%
,
.
14.05.2009 .
rRT-PCR
,
BSL3, .
CDC ,
,
.
.
04.05.2009 .-08.01.2010 . rRT-PCR 2429
. 792 (32,6%)
. ( . №18)
49
. №18
A/H1N1/2009
%
1174 428 37
596 186 31
659 178 27
2429 792 32,6
01.07.2009 .,
22
, .
№20 , - /
(48,3%),
„ ” -565 .
, ,
(p<0.001, Pearson chi square=20,32), .
/
25 .
,
( ),
.
. . „ ” ,
.
,
, ,
.
.
50
,
,
.
-30
.
-38
, 32 .
( )
.№19
( 1N1)2009. e
, (ECDC working
group on influenza A(H1N1)v, 2009)
.
- 11 (7%) 155 . ( 14)
396 , 109
(27,50%) .
18% , -
, PCR,
17% .
. -
, , -
– .
40-50%
.
. №19 38
.
20 12 1 5
( 1N1)2009
3
(7,8%)
3
(7,8%) 0
1
(2,6%)
51
14 ,
2009 .-08.01.2010 .
, - - 94 (41,4%)
.
(p<0.001, Pearson chi square=74,41).
15 ,
/ . -
537 (35,4%),
, -1 518 (62,5%).
110 (24,6%),
-109 (27,5%).
,
.
- 14 ,
. 7
31
- , .
52
(p<0.001, Pearson chi square=17,16),
(p<0.05, Pearson chi square=9,86).
,
5 7 .
,
.
. 15 /
, , 2009 .-08.01.2010 .
* 54
/ ъ я
,
- .
15 29 . (36,7%) ( 16)
- 1918 .
-
. -
(5%) - 0 4 .,
.
53
4%
. (p<0.001, Pearson chi
square=161,88).
16 A(H1N1)2009
, 2009 .-08.01.2010 .
,
. , ,
. 1918 .
1957 . - 20
27%.
. 42
5 . 24 (51%)
, .
-2 .
,
.
,
,
54
. ,
,
- , ,
. .
,
.
9
3 .
,
rRT-PCR.
,
.
WHO CC, .
/
(
1-1/2) /California/4/2009, A/California/7/2009
A/England/195/2009/H1N1/ ( .№20 №21)
55
. №20
( 1N1)2009, ,
.
Antigenic analyses of pandemic influenza A(H1N1) viruses (25/08/2009)
Haemagglutination inhibition titre1
Post infection ferret sera
Viruses Collection Passage A/NJ2 A/NJ A/Cal A/Cal A/Eng
date History 8/76 8/76 4/09 7/09 195/09
R15/79 F34/82 C4/F14/09 C4/16/09 NIBSC
F17/09
REFERENCE VIRUSES
A/New Jersey/8/76 Ex >5120 640 320 320 320
A/California/4/2009 C1,M1,E1 2560 160 2560 1280 2560
A/California/7/2009 E1+4 2560 160 2560 1280 1280
A/England/195/2009 MDCK3 2560 160 2560 1280 2560
TEST VIRUSES
A/Bulgaria/179/2009 26.6.2009 MDCK2 1280 80 1280 1280 1280
A/Bulgaria/340/2009 24.7.2009 MDCK2 1280 80 1280 1280 1280
A/Bulgaria/293/2009 15.7.2009 MDCK2 2560 160 2560 2560 1280
A/Bulgaria/330/2009 22.7.2009 MDCK2 2560 160 2560 1280 1280
A/Bulgaria/311/2009 20.7.2009 MDCK2 2560 160 2560 2560 1280
A/Bulgaria/324/2009 22.7.2009 MDCK2 2560 320 2560 2560 2560
A/Bulgaria/197/2009 08.6.2009 MDCK4 2560 160 1280 1280 1280
A/Bulgaria/325/2009 22.7.2009 MDCK3 2560 160 1280 1280 1280
A/Bulgaria/335/2009 23.7.2009 MDCK3 5120 160 1280 2560 2560
A/Bulgaria/363/2009 28.7.2009 MDCK3 5120 160 1280 2560 2560
A/Bulgaria/342/2009 24.7.2009 MDCK3 5120 160 1280 2560 1280
A/Bulgaria/354/2009 24.7.2009 MDCK3 5120 160 1280 2560 1280
1. < = 40; 2. hyperimmune
rabbit serum
56
. №21
( 1N1)2009, ,
.
Antigenic analyses of pandemic influenza A(H1N1) viruses (23/03/10)
Haemagglutination inhibition titre1
Post infection ferret sera
Viruses Collection Passage A/Cal A/Cal A/Eng A/Auck A/Bayern A/Lviv
date History 4/09 7/09 195/09 3/09 69/09 N6/2009
C4/F14/09 C4/31/09 NIBSC F17/09
C4/17/09 C4/33/09 C4/34/09
REFERENCE
VIRUSES
A/California/4/2009 C1,E2 2560 2560 1280 2560 1280 2560
A/California/7/2009 E6 2560 2560 1280 2560 1280 2560
A/England/195/2009 MDCK3/SIAT1 1280 2560 1280 2560 1280 2560
A/Auckland/3/2009 Ex+3 2560 2560 1280 5120 1280 2560
A/Bayern/69/2009 MDCK4/SIAT1 80 320 40 80 320 320
A/Lviv/N6/2009 MDCK4/SIAT1 640 1280 160 320 2560 2560
TEST VIRUSES
A/Bulgaria/1325/2009 03.11.2009 SIAT2 2560 5120 1280 5120 2560 2560
A/Bulgaria1369/2009 03.11.2009 SIAT3 2560 2560 1280 2560 1280 2560 1. < = 40
WHO CC
,
. ( NA),
- V106I N248D,
,
H275Y. ( .17, 18) 03.02.2010 . 225
H275Y,
, .
( 1N1) ,
57
2008 . .)
H275Y, ,
275
N1
.
(N2
N9), 274 .
. . N2 (H274Y), N1 -
(H275Y).
.
,
.
D222G
. 20
.
,
- α- 2-3
(
),
-
. ( 17, 18)
58
17
, N
59
18
,
60
( 2008 .)
, 25% ( 1N1)
, - -67%.
, ,
.
(H1N1),
(H1N1)2009.
.
40 (28.09.-04.10.2009 .)
, 45 (48%.),
. ( 19)
19. A(H1N1)2009
rRTPCR, 40-53 2009 .
61
20 ,
,
.
- ,
. ,
, .
, 99%
, .
20. -
2009 .
152 28 , 2009 .,
,
. rRT-PCR
5 (0,9%), - 7 (1,3%).
,
.
,
WHO/Europe.
62
V.
RSV, MPV
2004-2009 .
,
.
RSV .
RT-PCR 2005 .
2006/2007. 2008 .
RT-PCR
2001 . ,
2008/2009 5
RSV, MPV
. 2009 .
rRT-PCR ( 1N1)2009.
RSV 846
, 452 MPV, 371
. ,
04.05.2009 .- 08.01.2010 . 2429
157 ,
,
.
, RSV MPV
- , 0 5 .
0 11 ., 12 35 .
36 59 .
.
RSV
- , MPV
63
.
5 , ,
.
,
-
, , .
„ ”
RSV .
-
RT-PCR, . ,
RSV ,
, ,
,
- .
15 RSV,
- -2 MRC-5.
RSV MPV
, MPV
, ,
.
2009 . ,
- BSL3
,
.
Directigen RSV
RSV, ,
.
64
,
48 .,
.
,
,
,
.
RSV
,
, (2005 .)
,
.
RSV,
,
RT-PCR , .
MPV - ,
. ,
, RSV MPV.
RT-PCR RSV
.
„ ” 2006 . ,
INSTAND, RSV .
RT-PCR .
, -
RSV ,
.
RT-PCR
, - ,
, ,
65
. 2006 . RT-
PCR RSV .
2006/2007 . RSV -
,
,
.
MPV.
5 -
.
-
, , RSV MPV
.
RSV MPV
.
RSV
2006/2007, ,
RSV MPV,
.
,
2008/2009.
, RSV
(ECDC). 2007 .
ECDC
RSV.
2008 .
.
H5N1, 1997 .
.
66
, ,
2007 . ,
- External Quality Assessment (EQA).
„ ”
H1N1 2009 .
, 2009 .
. ,
,
.
. 2006 .
.
,
, ,
.
2429
15 . 29 .,
,
.
1918 . ,
, . . „ ”,
. ,
65 . . «
», .
. 34% 1950 .
.
, -
„ ” ,
67
.
.
, ,
90% 18% .
, A(H1N1) 2009
,
.
-
.
0.3-
1.5%. ,
. ,
. . „ „
.
.
ECDC,
.
„ ” Global
Influenza Surveillance Network (GISN)
WHO CCs
. ,
, ,
,
.
68
, RSV MPV,
.
( ) 80- 20–
RSV
(
.
.
,
.
( 1N1)2009
2009 .
e
Real Time RT-PCR.
69
VI. .
1.1.
RSV MPV,
-
.
1.2.
15 29 .,
.
2. (
) RSV,
- ,
,
.
3.1. in-house RT-PCR
RSV MPV
.
3.2. RT-PCR RSV
, .
3.3.
, 2009 .
Real Time RT-PCR,
.
4.1. RSV, MPV
5
.
4.2. RSV
( , ) -
(0-11 )
70
4.3. RT PCR
MPV 5
.
5.1.
( 1N1)2009 , ,
.
5.2.
.
6.
(
) ,
.
A/California/7/2009.
71
VII. .
1.
( )
RSV,
- ,
,
.
2. in-house RT-PCR
RSV MPV
.
3. RSV MPV
.
4. CDC Real Time RT-PCR,
.
5.
- Real Time RT-
PCR.
6.
,
.
72
VIII. :
:
1. ., ., ., ., ..
. , 3, 2004, . 28-32
2. Pavlova S., Hadziolova T, Kotzeva R. Diagnostic studies on the etiological role
of respiratory syncytial virus and influenza viruses in hospitalized children.
Problems of Infectious and Parasitic Diseases. 2006, vol 34/2, pp 29-31
3. ., ., . RT-PCR ,
. , 2007, LIV, Supplement I ,119-122.
4. ., ., . “ ”
, . , 2008, LV, ,
. 30-34
5. Pavlova S., Hadzjiolova T., Arabadjieva P., Kotseva R.. Application of PCR for
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Virology in Bulgaria), S 2008,151-159.
6. . , . , . . 2008/2009 .
, IV, . 1/2012, 17-22
7. . , . . (H1N1)2009 . 48/2012, №2, 17-
26
1. Hadzhiolova T., Pavlova S., Kotseva R. Possibilities for laboratory diagnosis of
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2. Pavlova S.,T.Hadzhiolova,P.Arabadjieva, R.Kotseva. Application of RT-PCR for
diagnosis of respiratory syncytial virus and human metapneumovirus infections
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3. Hadzhiolova T., Pavlova S., Kotseva R, Laboratory investigation of the first
suspected human cases of infection with avian influenza A(H5N1) virus in
Bulgaria, 2008, Eurosurveillance, vol 13, issue 30, 1-6.
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1. ., ., , . -
. . “ ”, Vol XL., 3, 2004, . 10-14
2. ., ., . -
. . . 2006; VIII; 2: 6-9
3. ., ., . . . 2007, 6, 5-8
4. ., ., . – . . 2008, 3, 16-18
, , :
1. ., ., . DТrОМtТРОЧ FХЮ – A+ DТrОМtТРОЧ RSV
RSV. II , , 30 -1 2003.II - , 30
-1 2003.
2. ., ., ., ., ..
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4. ., ., ., ., .
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; . 2006 . .
6. ., ., . -
. XI . . 05-07. 10.2006 .
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, BIO-RAD, , 12-14.11.2008 .
10. . , . . (H1N1)2009 . 8- , , 2010 .
:
1. . : . , . , .
, . , . . 2003 .
2. RT-PCR - . : . , . , . , . .
2004 .
3. PCR / .
: . , . , . , . , .
4. Real Time RT-PCR ( 1N1). . , . .
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