DAC DIEM CHUNG NOROVIRUS VA SAPOVIRUS 6 TRE VIEM DA DAY RUOT CAP TAI BENH VIEN NHI TRUNG UONG NAM 2007-2008

Nguyen Van Trang, Le Thi Luan*. Le Thi Kim Anh. Vu Thi Bich Hau. Le Thi Hong Nhung. Hoang Thj Bich Ngoc**. Pimmnapar Phasuk***. Orntipa Setrabutr***, Jan Vinje****, Carl J. Mason*** . Dang Due Anh

Vien Ve Sinh Dich Te Trung Vang, Hd ndi "Trung lam nghien citu san xuat vdc xin vd sinh pham y te, Hd nai **Benh vien IS'hi Trung Vcmg, Hd noi ***Vien nghien cuv y hoe quan su Hoa ky, Bangkok, Thai lan ****Trung lam kiem sadi vd phdng ngira hinh tat (CDC) Atlanta, Hoa ky

TOM TAT Chung toi thu thap 501 mau phan tir tre em bj viem da day ruot cap tinh tu" thang 11/2006 den het 10/2007 tai benh vien Nhi Trung Udng d Ha Noi. Norovirus (NoV) and Sapovirus (SaV) du'dc phat hien bang ky thuat real time RT-PCR su" dung doan probe Taqman dac hieu va dinh genotyp bang giai trinh tii gen. Ty le nhiem NoV genogroup (G)ll va SaV tren cac mau nay tuong jog la 36,3% va 1,4%, trong khi do khong phat hien du'dc cac trUdng hdp nhiem vdi NoV genogroup I trong thdi gian nay. Tinh trang nhiem NoV xay ra quanh nam tai benh vien Nhi, cao nhat trong mua khi Rotavirus du'dc phat hien thap nhat. Trong 182 mau diidng tinh vdi NoV GII, 62 mau du'dc chon ngau nhien de xac dmh genotype bang giai trinh tu" gen. NoV GII.4 la genotype pho bien nhat (60 mlu) va co 2 tru*dng hop NoV-G13. Chung NoV-GII thuoc cluster Minerva GII.42006b, tuy nhien khac vdi chung GII.4 du'dc phan lap trong nhurig nam 1998-1999 tai benh vien nay. Tre d cac do tudi (tir 6 thang den 5 nam) deu bj nhiem NoV va khong co su" khac biet ve ty le nhiem giCfa cac nhom tudi.Chung toi xac dinh genotype cua bon trong so bay mau dUdng tinh vdi SaV, trong do co genotype GI.2 va GILL Trong nghien CLAJ nay, chung toi nhan manh vai trd cua NoV (chi sau virus rota) trong benh viem da day ruot cap d tre em tai mien Bac Viet Nam I. DAT VAN DE

Tren the gidi, Noroviruses (NoV) va Sapoviruses (SaV) lii nguyen nhan quan trong ciia cac vu dich tieu chay d ngUdi Idn va tre em. Dong thdi. NoV da du'pc cdng nhan la nguyen nhan quan trong gay viem da day rudt cap d tre em. trong mdt sd trUdng hdp. chi ddng thu hai sau virus Rota [1]. Ty le nhiem vdi NoV d tre em cac nddc giao ddng tii khoang tii 5% -48%, Ty le tre nhiem Sapoviruses (SaV). thap hon nhieu. thUdng la 1-5% d

tre em bi tieu chay. NoV va SaV. thudc ho Caliciviridae. la virus khdng vd. sdi ARN ddn. dudng. Virus thudc ho nay cd ty le dot bien cao. tao nen nhieu chung khac nhau. do vay viec giam sat lien tuc de xac dinh cac bien the mdi ciia virus nay la rat can thiet.

O Viet Nam, trong thdi gian gan day, tam quan trong ciia calicivirus da bat dau dupe nhan ra. Mdt sd nghien cdu d mien Nam ciia Viet Nam da xiic dinh NoV trong - 5,5-6,4% tre em nhap vien vdi benh tieu chay va SaV trong 0.8-1.2% ciia

Tac gia chinh: Nguyen Van Trang Dia chi: Vien Ve Sinh Dich Te Trung L'dng. sd 1 Yersin, Ha ndi Dien thoai: 04-39718491 Email: nvtrang@gmail.com

Tap chi Y hoc dir phdng. Tap XX. so 3(113) 79

nhung tre em nay [2], Mot nghien ciiu khac d Xha Trang, Khanh Hoa cung cho ty le nhiem trong 6 thang d tre cm la 7,1% [3], Trong khi ty le phat hien SaV trong nhdng nghien cdu la tudng tu vdi ty le nhiem tre the gidi, ty le nhiem NoV d tre thap hdn so vdi cac nghien cdu d cac qudc gia khac. Trong nghien ciiu nay. chung tdi danh gia tinh trang nhiem NoV va SaV d tre em dudi 5 tudi nhap vien do viem da day rudt cap. Ddng thdi, dac diem cac chung gay nhiem cung nhd hien tddng da nhiem vdi virus khac nhau d tre em nay cung dUdc xem xet. Muc dich cudi ciing ciia chung tdi la danh gia dung tam quan trong ciia 2 virus nay trong tieu chay cap d tre. I I . PHUONG PHAP NGHIEN CUU

2.1 Ddi tUdng va thdi gian nghien ciiu

Nam tram va mdt mau phan ciia chUdng trinh giam sat rotavirus thu thap td 11-2007 ddn het 10 -2008 da dddc sii dung de sang lpc NoV va SaV. Cac mau tii tre em dudi d tudi. nhan vao benh vien Nhi trung ddng Ha Noi do viem da day rudt cap tinh. Bon mUdi mau phiin dat du'dc thu thap nam 1998-1999 tai cac benh vien tai Ilk Ndi xet nghiem dddng tinh vdi RV cung da dUdc thd nghiem NoV. 2.2. PhUdng phap nghien ciiu

Mau phan dii du'dc hoa trong nddc cat thanh hdn dich 20% va ARN ciia virus chide tach chiet sii dung Qiagen viral RNA mini kit (Qiagen. GmbH, Diic), theo hudng dan cua nhii san xuat. ARN tach chiet dupe, chia thanh cac tuyp nho va lu'u tru d -80"C. Ba phan dng real-time RT-PCR ndng biet dUdc dung de phat hien NoV-Gl. NoV-GII, SaV (chi tiet quy trinh d phan sau). Chdng am va chung dddng cho qua trinh chiet tach ARV. doi

chiing khdng cd khuon du'dc su' dung de kiem tra mdi lan chay.

Phat hien NoV phan nhom I va II (GI va GII) bang real-time RT-PCR

Qua trinh real-time RT-PCR du'dc tien hanh mdt bu'dc trong 25ul cua hdn hdp phan Ung cd chu'a oul ARN mau. dem lxTaqMan EZ (Applied Biosystems, Foster. CA). 300uM cua dATP. dGTP. dCTP va dUTP, 150uM TTP. 400uM mdi xudi va mdi ngudc. lOOnM cua dau do huynh quang (Taqman). 0.2U uracyl gly-cosylase-N (UNG). enzyme chiu nhiet polymerase 2U rTth. va manganese acetate 3mM de phat hien NoV GI hoar manganese acetate 3.5 mM de phat hien NoV GIL Trinh tii cac mdi va dau dd dddc lay tii bai bao da du'dc cdng bd [4]. Viec khuech dai dUp'c thiic hien tren may ABI 7.700 va Q-Rotorgene (Qiagen. GmbH. Diic) theo cac dieu kien sau day: 50oC ban dau trong 2 phut de kich hoat UNG. tiep theo la 60°C trong 30 phut cho cac phan ling phien ma ngUdc, va bien tinh d 95"C trong 5 phut va 40 chu ky td 94°C trong 20 giay va 60"C trong 45 giay.

Phat hien SaV bdng ky thuat real-time RT-PCR

Quy trinh phat hien SaV bao gdm 2 budc[5], cADN du'dc tdng hdp d 42oC trong 30 phut trong mdt hdn hdp phan dng chda: 12ul ARN mau. enzyme Multiscnbe. hexamers ngau nhien. dT ohgo, dNTPs, MgC12. chat dc che ARNase. Khuech dai cua cADN da dUdc thiic hien theo rac dieu kien sau day: Platinium PCR supermix, mdi xudi va mdi ngUdc (hdn hdp mdi ciia GI, GII, GIV va GV). dau dd (mot dau dd de phat hien GI. GII. va GIV va mdt dau do khac cho GV). Dieu kien khuech dai nhu sau: 50"C trong 2 phut. 95"C trong 5 phut. 50 chu ky d 95°C trong 10 giay va 60"C trong 40giay. Ddi chung du'dng. ddi chdng am

80 Tap chi Y hoc dir phdng. Tap XX. sd 5(113)

cho qua trinh tach chiet ARN, do'i chdng am rieng cho ca bddc phien ma ngUdc va cho khuech dai deu dddc bao gom d moi lan chay.

Xac dinh genotype ciia GII-NoV vd SaV bdng gidi trinh tie gen

Chi GlI-Nov dddc phat hien trong cac man phan, giai trinh tii dddc thdc hien tren 5 mau dudng tfnh thu dddc til 1998-1 999 va 62 mau dddng tinh dddc Ilia chon ngau nhien tu 182 mau dddng tinh NoV-(111 cua nam 2007-2008. De giai trinh tu. san pham 468 bp ctia No-GIl dddc khuech dai bang each sii dung bp mdi G2FB-a. G2FB-b, G2FB-C va G2SKR bao gdm td nucleotide 4922 den 5389 [4]. Nhiing mdi nay khuyech dai phan nhd ciia gene poly­merase va doan giao nhau gnia gene poly­merase va capsid. Dd khang dinh cac genotyp cua NoV-GII khi sii dung bp mdi tren. chung tdi khuech dai 14 mau vdi bp mdi khac d vung D ciia gen capsid (nham den vung 1' cria protein VP!) sii dung mdi dii dUdc rdng bd |6]. Sau dd. cac san pham PCR du'dc dda vao vector -T pGKM (Promega. Madison, Wl. USA) va chuyen thanh cac chung E.coli TOPO10 de tach ddng va giai trinh tit vdi mdi T7 va SP6.

Ddi vdi SaV. cac cADN cua mau du'dng tinh vdi SaV bdi real lime RT-PCR dddc khuech dai bang each sii dung cap mdi SV5317 va SV5749 theo cac dieu kien sau day: 94"C trong 3phut. 45 chu ky: 30 giay d 94°C, 55°C trong 30 giay va 72"C trong 1 phiit. va 72"C trong 7 phut [7]. San pham PCR (433bp) sau dd dddc dda vao pGFM -T vector (Promega. Madison,

Wl. USA). Plasmid tinh khict da dddc chen san

pham PCR cua NoV va SaV dUdc gin idi Cdng ty Macrogen, Han Qudc de giai trinh tii sii dung cap mdi T7 va SP6. Qua trinh giai trinh tii thiic bien trdn may Sequencer 3730x1 (Applied Biosystem, Foster City. CA, USA). Cac phan tich dddc thuc hien Iron cac chudi nucleotide. Du lieu chudi gdc dau tien du'dc phan l ich boi phan mem CHROMAS va dii lieu trinh tii xudi. ngUdc cua mdi mau du'dc sap xep bang each sii dung chUdng trinh MegAlign va q Edit Seq (phan mem Mega 4,1 [23]) de cd chide nhdng chudi cudi cung. Sap xep hoan chinh du'dc thu'c bien vdi ('luster W (Mega 1.1). Thuat toan Maximum Parsimony dddc sii dung de xay diing cay phat smh loai vdi 1.000 lan lap lai. I I I . KET QUA

Ty le nhiem NoV va SaV d tre em nhap vien do viem da day rudt cap tai Benh vien Nhi TW

Ty le nhiem NoV d Ire em bi tieu chay cap tai benh vien Nhi Trung Udng rait cao.

Trong so 501 mau kiem tra. cdi 82 mau dddng tinh vdi Gil NoV va khdng cd mau GI NoV nao.ty le nhiem NoV-GII chung la 36,3% (Bang 1). Cac man dUdng tinh vdi NoV-GII dUdc tim thay d tat ca cac thang (11-2007 dim 10-2008). Trong cac mau am tinh vdi RV. ty le nhiem NoV thap nhat trong thang td (9,5%) va cao nha't trong thang bay (87%) (Hinh 1).

Tap chi Y hoc du phdng. Tap XX. so 5 (113) 81

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1. Phan bd cac trirdng hdp nhiem NoV theo thang tai Benh vien Nhi Trung Udng trong nam 2007-2008, tudng quan vdi tinh trang nhiem virus Rota d tre.

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Hinh

Ddng nhiem gida NoV-GII va RV xay ra d 27 trUdng hop (chiem 5,4% cua tat ca car trudng hpp). cao nhat trong 10-2008 (10 trudng hpp. 32,3%). Ty le nhiem Sapovirus la 1,4%, thudng xay ra trong

Bang 1. Ty If tre dUdng tinh vdi NoV va SaV va dac diem cac chung phan lap dude trong nam 2007-2008

nhiing thang lanh tii thang If) - thang 2 (6 trong tdng sd 7 trUdng hop) va chi cd mdt trUdng hdp xay ra thang 5 (ket qua khdng dUdc trinh hay).

Tac nhan So mau

duong tinh (%)

Tong so mau

56 mau giai trinh tir Genogroup/genotype

NoV 182 (36,3) 501 62 GIIM Minerva, Gl 1/13

SaV 7 (1.4) 501 4 GI/2, GII/1*

Tdng .so tre phan tich ditdv dudi 501 do thii u thong tin ve tudi/ngay sinh ciia 2 tre.

Phan hd cac trudng hap nhiem NoV vd SaV theo nhdm ludi ciia tre

Phan bd car trudng hop nhiem NoV-(111 theo nhdm tudi ciia tre dddc torn tat trong Bang 2. Tre em ciia tat ca cac nhdm tudi (tu 6 thang ddn 5 nam) deu bi nhidm NoV, cao nhat 0 tre em 7-12 thang tudi

(41.2";,) va thap nhat d tre em tren 2 tuoi (20%). luy nhien sU khac biet khdng cd y nghia thdng ke. Mot dieu dang chu y la mac du so'tre o nhdm tudi Idn dddng tinh vdi NoV thap (chi chiem 3.4",, trong long so mau dddng tinh cua cac Ida tudi). sd tre d Ilia tudi Idn nay nhap vien vi benh

82 'lap chi Y hoc dir phdng. Tap XX. sd 5(113)

Bang 2. Phan bo theo lua tuoi cac trudng hdp nhiem NoV va SaV

NoV SaV

Nhom tuoi So tre Sd duong tinh

Ty le dudng tinh trong ciing nhom

tudi (%)

No of positives

Ty le duong tinh trong ciing nhom

tudi (%) < 6 thang 121 41 33.9 2 1,7 7-12 thang 170 70 41,2 3 1,8 13-24 thang 178 62 34.8 1 0,56 >=25 thang 30 6 20.0 1 3,3 Tdng 499* 179 7

Tong so tre phan lich ditclc dtfiri 501 do thieu thdng tin vi tuoi/ngay sinh cua 2 tre.

Da nhidm vdi cac loai virus khac nhau

Da nhiem phd bien nhat d tre em la gida rotavirus (RV) va NoV (27 trudng

hdp). trong do 1 trddng hdp nhidm ca 3 virus RV. NoV va SaV. Nhiem ddi cua SaV vdi Nov hoac RV da xay ra vdi 1 trddng hdp.

Tap chi Y hoc dir phdng. Tap XX. so 5(113) 83

Phan tich ti inh tii cac chiing NoV va SaV

100 | I ...

HUN/02 Lordsdale/97 US/MD145-12/87 US/Miami Beach/326/95 Grimsby US/Burwash Landing/331/95 UK/Oxford/B5S22/03 USA/Farmmgton Hills/02 DE/Langen1061/02 AU/Hunter 284E/04O JP/Sakai/04-179/05 USA/Minerva/OSD-CS/06 VN/NTD55D/07 VN/NTD164D/08 VN/NTD49D/07 VN/NTD110D/08 VN/NTD95D/07 VN/NTD74D/08 VN/NTD166D/08 VN/NTD11D/08 VN/NTD277D/08 VN/NTD357D/08 VN/NTD346D/08 VN/NTD87D/07 VN/NTD427D/08 VN/NIHE245 4D/98 Kashiwa47 GII.13 VN/NTD115D/08 JP/SaitamaU1/97 JP/SaitamaU3/97 JP/SaitamaU201/98 VN/NIHE33 3D/98

Gill

GII.4

GII 13

GII.3

Hinh 2. Cay phan chung loai dua vao trinh tir doan ndi giura gen ma hoa polymerase va gene ma hoa capsid cua NoV genogroup II tU 11/2007- het 10/2008, so vdi chung 0 Viet Nam trong nhung nam 1998-1999 (do phdng thi nghiem phan lap dUdc) va cac

chiing tren the gidi. Chi cd 14 chiing trong so 62 chung (2007-2008) va 5 chiing (1998-1999) dUdc the hien trong cay phan chiing loai nay. Chiing EU 360814HUN2002 (Gill) duoc sU dung lam chung ngoai nhdm (out-group). Cac chiing phan lap trong nghien

ciru nay dude ddng khung.

84 Tap chi Y hoc du phdng. Tap XX. so 5(113)

tieu chay thap (n = 30), do do ty le nhiem d Ilia tudi Idn nay cd the khdng dUdc danh gia dung. Cd 5 tre (71,4%) dUOng tinh vdi SaV dudi 1 tudi.

Trinh tu ctia san pham 468bp tren doan giao giiia gen polymerase va capsid cho thay trong sd 62 mau phan tich. 60 mau thudc genotyp GII.4 . gidng vdi chung Minerva (GII.42006b) (Hinh 2). cho thay sd Uu the ciia genotyp NoV-GII-1. Co 2 chiing NoV Gil dUdc xac dinh la Gil. 13. Khi phan tich cac mau thu thap dUdc nam 1998-1999 dc so sanh. chung tdi thay sii cd mat ciia chiing GI1-4 (tUdng ddng vdi chiing Minerva), chiing tddng ddng vdi US/MiamiBeach/95 va chung GH-3 (cum Saitama/U201). Trinh tii vung D ciia gen capsid d 14 chiing (2007-2008) va 2 chung cua nam 1998-1999 deu khang dinh ket qua dinh genotyp khi su dung doan gen gida gen polymerase-cap-sid. Doan mdi cho vung I) cd the sii dung de nhan len phan Idn cac chiing NoV-

GII.4 phan lap trong nam 2007-2008. tuy nhien lai khdng sii dung dUdc de nhan cac chiing GII.4 phan lap trong nhiing nam 1998-1 999. dieu nay cho thay tuy ciing la chung GII.4, cac chiing nam 1998-1999 da thudc cluster khiic vdi cac chiing thudc 2007-2008. Cd 2 chiing phan lap vdi geno­type GII.13, va deu dupe khang dinh bdi trinh tit ciia ca 2 doan (doan ndi giiia gen polymerase va capsid va doan gen capsid, Dieu nay cung cho thay rang doan mdi nham vao viing danh gidi giiia gen poly­merase va capsid khdng dii do dac hieu dc phan bien cac cluster khac nhau ciia Gil.4. nhat la cac chiing tii nhiing nam trUdc.

Trong sd 3 chiing SaV cd the giai trinh tu dddc. cd mdt chiing xac dinh la GI (nhiing khdng xac dinh dUdc geno­type), mot chiing GI.2 gidng vdi chung Parkville, va hai chiing GII.] gid'ng vdi Bnstol/98 (Hinh 3).

l ap chi Y hpc dir phdng. Tap XX. sd 5 (113) 85

i L J ,1 LI L .

L_.

VN-NTD137/08 UK/Bnstol/98 F/Lyon598/97 VN/VN387/03 Mc2 VN/NTD280/08 London Ehime2K-1948/00 Mc10

- JP/Chiba990763/99 Jena HUN/PEC swine-ld3/05 Ehime01-1669/01 Arg39

- VN/VN337/03 JP/Chiba000764/00 Houston/86 Manchester VN/VN779/03 JP/Chiba010250/01 HCU65427 JP/Chiba000496/00 JP/Chiba010658/01 SE/Stockholm318/97 VN/NTD430/08 DEU/Potsdam/00 Parkville Houston/90 JP/Chiba000671/99 Hou7-1181

Hinh 3. Cay phan chung loai dua vao trinh tir nucleotide doan gen capsid cua cac chung SaV. Co 3 chung phan lap tir 11/2007 den het 10/2008 dude bieu hien trong hinh

nay. Chung SaV-GIII PEC dUdc siirdung la chiing ngoai nhom. Cac chung phan lap trong nghien curu nay d trong khung.

IV. BAN LUAN Trong nghien ciiu nay. lan dau tien

chung toi danh gia tinh hinh nhiem NoV va SaV ri tre em tai mien Bac nddc ta. So vdi cac nghien cdu khiic d Nha Trang hoac ri thanh phd HCM, ty le tre nhiem

NoV trong nghien cdu nay cao hon nhieu. Cu the. cac nghien cdu ri thanh phd I [CM cho ty le nhiem tii 5-6%, con nghien cdu tai Nha trang. Khanh Hoa cho ty le 7,1% [8], Cd the cho rang cd sU khac nhau gifla ty le nhiem virus ri tirng vung mien khac nhau. Tuy nhien do cac phUdng phap sii

86 Tap chi Y hpc du phdng. Tap XX. so 5 (113)

(lung kha khac nhau giiia cac nhom nghien cu'u d 2 vung Iron (phUdng phap RT-PCR truyen thdng hoac NASBA) vdi phUdng phap ciia chiing tdi (real time RT-PCR. tinh trang nhiem NoV cd the chda dUdc danh gia dung mdc d 2 dja diem tren. Nghien cdu d Thai Lan cho ty Id nhiem tii 8,1-14% khi sii dung phUdg phap RT-PCR truyen thdng. Cung trong thdi gian nay (2006-2007). ty le nhidm NoV tang hdn dd'n 45,7% khi sii dung nested RT-PCR [9, 10]. Ty le nhiem NoV d cac ndiioc khac cung cao do viec sii dung real time RT-PCR nhu 1 5% 6 Han Qudc. 17.4% d Tunizia, 15.1% d Velore. an dp, 48.4% d Italy va 3:1,3%. d Hang Sao Paulo, Brazil |1 1| [12, 13] [14. 15],

Trong khi ty le nhiem NoV dao ddng kha Idn d cac nddc, cac khu vUc, ty Id nhiem SaV thUdng thap (<3%), va ky thuat real-tiem RT-PCR nhay hdn |2. 16. 1 7]. Trong nghien cdu nay 2 genotyp phat hien dupe bao gdm genotype (11/2 va (ill/I. Mdt so lddng mau nhd d benh vien tinh Bac Ninh cho thay mot chiing virus SaV-GIV. NhU vay ca 3 genotyp phd bien gay benh cho ngddi la (II. (Ill va GIV deu dupe tim thay trong nghien cdu cua chung tdi. Gidng nhU d thanh phd HCM. ciic chiing SaV-GI.2 gan gid'ng vdi chung Postdam cdn chung SaV-Gll.l gan gidng vdi chung Lyon/97 va UK/Bristol/98). d thanh phd HCM. car chung khac nhd 01.1 va GIL 1 ciing dtidc tim thay.

Mac dii mdi chi cd 62 mauNoV-GIl (trong tdng so' 182 man dUdng tinh) dddc giai trinh tii trong nghien ciiu nay . cd the thfiy rang genotype GII.4, thudc cluster Minerva GII-4 2006b chiem so ddng. gidng vdi cac chung phan lap d Thai lan. Mini Quoc cho thay tinh trang phd bien cua chung nay. Tuy nhien nghien cdu cua Nguyen Tuan Anh d thanh phd HCM

trong nhiing nam 2005-2006 cho thay tdn tai song song Gil.3 va GII.4 vdi ty le tUdng dUdng nhau (tddng ling 40.6% va 50% respectively) |2], Ddng thdi d thanh phd HCM, cac genotype khiic nhd GII.12 va GII.6 cung hay gap. cho thay sii da dang ciia virus nay d mien Nam nude ta. Ngay ca ddi vdi genotype GII.4. trong khi mien nam cd 2 variants cua GII.4 2006. d mien Bac chiing GII.4 ciing thudc Gil.4 2006b Minerva strain. Do trinh tii cac chiing phan lap d TP HCM ngan, do vay cac chiing nay khdng dtidc phan tich ciing vdi cac chimg ciia chung tdi. Tuy nhien khi so sanh (alignment), chung GII.4 phan lap d TP 11 CMC gid'ng vdi chimg Sakai hdn cac chiing d mien Bac.

0 mien bac ciing nhd d mien Nam nddc ta, NoV lay nhiem quanh nam vdi ty le nhiem cao trong nhiing thiing tii thang 5 den thang 10. Thdi diem cd ty le tre nhiem NoV cao ngUdc lai vdi thdi diem cd ty le nhiem RV cao: cu the la trong nhiing thang 6 va 7. khi ty Id nhiem RV thap, ty le nhiem NoV tang len 84%. Mdt diem khdng thuan Idi ciia nghien cu'u nay la so lu'dng mau khdng hoan thien tii thang 4-thang 8/2008. trong dd cac mau phan dUdng tinh vdi RV khdng cdn cho phiin tich NoV va SaV ciia chung tor Do vay, chdng tdi khdng the xac dinh ty le nhidm theo thang cua virus nay. Tuy nhien chung tdi van cd the tide tinh ty le nhiem trong nam. ddng thdi ty le ddng nhiem giua NoV va RV (ty le chung 5.4%. 0-37.5% tiing thang) cho thay tam quan trong cua NoV. chi thd 2 sau RV giiy viem da day rudt cap d tre.

Viec NoV-GII.4 ndi len va chiem Uu the cung dddc ghi nhan d cac nghicn cu'u d cac nddc khac tren the gidi (I loa Ky, cac nu'dc khac). Ngudi ta van chUa ro tai sao car chiing NoV lai trd nen ddng nhat

Tap chi Y hpc du phdng. 'lap XX. so 5(113) 87

trong vong 10 nam trd lai day. Mot gia thuyet duoc dUa ra la chiing virus nay da dat dddc mot Idi the la cd the gay nhiem ngddi vdi cac nhdm mau khac nhau. khdng chi ngddi cd cd nhdm man O [18].

IV. KET LUAN Vai trd cua NoV trong viem da day

rudt 0 tre la kha Idn (36,3% tr e nhiem), chi sau virus rota. Chiing NoV-GII,4 dang chiem du the d nddc ta tddng tu nhd tren the gidi. Nhiem NoV d moi hi'a tudi. trong khi nhiem SaV thddng tap trung d tre dddi 1 tudi va vdi ty Id thap. Can cd nhiing giam sat dich te hoc phan tii' ve linh trang nhiem NoV d tre em tren loan qudc va theo ddi sii lu'u hanh chung, de danh gia diing vai trd cua virus nay d nUdc ta.

TAI LIEU THAM KH \() 1. Kstes MK, Prasad BV, Atmar HL.

Noroviruses everywhere; has some­thing changed'.' Curr Opin Infect Dis 2006;19(5):467-74.

2. Nguyen TA. I loang L, Pham le D. Hoang KT, Okitsu S. Mizuguchi M. et al. Norovirus and sapovirus infections among children with acute gastroen­teritis m Ho Chi Mmh City during 2005-2006. .) Trop Pediatr 2008;54(2);102-13.

,'i. Nguyen Thi I lien Anh LPM, Nguyen Cong khanh. Nguyen Dmh hien Cat, Bien Thi Nga. Hiroshi Suzuki and Dang Due Anh. Rotavirus and Norovirus infection in hospitalized children less than 5 years old in Nha Trang. Journal of Practical Medicine 2007:57 1(71:20-22.

4. Kageyama T. Kojima S, Shmohara M, LJchida K. Fukushi S. Hoshino FB, el al. Broadly reactive and highly sensi­

tive assay for Norwalk-like viruses based on real-time quantitative reverse transcript ion-PC R. 3 ( Im Microbiol 2003;41(4):1548-57.

5. Oka T, Katayama K, llansman GS. Kageyama T. Ogawa S. Wu FT, et al. Detection of human sapovirus by real­time reverse transcription-poly-merase chain reaction. 3 Mod Virol 2006;78(10):1347-53.

6. Vinje .J. Hamidjaja RA. Sobsey MD. Development and application of a capsid VP! (region D) based reverse transcription PCR assay for genotyp-ing of genogroup 1 and II noroviruses. J Virol Methods 2001:1 1 6(2): 109-1 7.

7. Okada M. Yamashila Y. Oseto M. Shinozaki K. The detection of human sapoviruses with universal and genogroup-specific primers. Arch Virol 2006;151(12):2503-9.

8. Nguyen TA, Yagyu F. Okame M. Phan TG. Trinh QD, Yan II, et al. Diversity of viruses associated with acute gas­troenteritis in children hospitalized with diarrhea in I lo Chi Minh City, Vietnam. .J Med Virol 2007;79(5):582-90.

9. Kittigul L, Pombubpa K, Taweekate Y. Yeephoo T. Khamrin P, Ushijima H. Molecular characterization of rotaviruses, noroviruses, sapovirus. and adenoviruses in patients with acute gastroenteritis in Thailand. 3 Med Virol 2009;81(2):345-53.

10. Kittigul L, Pombubpa K. Taweekate Y. Diraphat P. Sujirarat D, Khamrin P, et al. Norovirus Gil 4 2006b vari­ant circulating in patients with acute gastroenteritis in Thailand during a 2006-2007 study. J Med Virol;82(5):854-60.

11. Colomba C, Saporito L, Giammanco

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GM, De Grazia S, Ramirez S, Arista S, et al. Norovirus and gastroenteritis in hospitalized children, Italy. Emerg Infect Dis 2007; 13(9): 1.389-91.

12. Monica B, Ramam S. Banerjee I, Primrose B, Iturriza-Gomara M. Gallimore CI, et al. Human cali-civiruses in symptomatic and asymp­tomatic infections in children in Vellore, South India. J Med Virol 2007;79(5):544-51.

13. Yoon JS, Lee SG, Hong SK. Lee SA. Jheong WH, Oh SS, et al. Molecular epidemiology of norovirus infections in children with acute gastroenteritis in South Korea in November 2005 through November 2006. J Clin Microbiol 2008:46(4): 1174-7.

14. Sdiri-Loulizi K, Gharbi-Khelifi II. de Rougemont A, Chouchane S, Sakly N, Ambert-Balay K. et al. Acute infantile gastroenteritis associated with human enteric viruses in Tunisia. J Clin Microbiol 2008.

15. Castilho JG, Munford V. Resque I1R. Fagundes-Neto U, Vmje J. Racz ML. Genetic diversity of norovirus among children with gastroenteritis in Sao Paulo State, Brazil. J Clm Microbiol 2006:44(1 1 ):3947-53.

16. Khamrin P, Maneekarn X. Peerakome S, Tonusin S, Malasao R, Mizuguchi M, et al. Genetic diversity of noroviruses and sapoviruses in children hospitalized with acute gas­troenteritis in Chiang Mai, Thailand. J Med Virol 2007;79(12): 1921-6.

17. Chan MC, Sung JJ, Lam RK. Chan PK. Lai RW, Leung WK. Sapovirus detection by quantitative real-time RT-PCR in clinical stool specimens. J Virol Methods 2006; 1 34(1-2): 146-53.

18. Lindesmith LC, Donaldson EF, Lobue AD, Cannon JL, Zheng DP. Vinje J, et al. Mechanisms of GII.4 norovirus persistence in human populations. PLoS Med 2008;5(2):e31.

Tap chi Y hpc dir phdng. Tap XX. so 5 (113) 89

DETECTION AND CHARACTERIZATION OF NOROVIRUSES AND SAPOVIRUSES IN CHILDREN WITH ACUTE GASTROENTERITIS IN NATIONAL PEDIATRIC HOSPITAL

Nguyen Van Trang, Le Thi Luan*. Le Thi Kim Anh. Vu Thi Bich Hau, Le Thi Hong Nhung. Hoang Thi Bich Ngoc**. Pimmnapar Phasuk***. Orntipa Setrabutr***, Jan Yinjc****. Carl J. Mason*** . Dang Due Anh

Department of Immunology and Molecular Biology. The National Institute of Hygiene and Epidemiology, Hanoi, Vietnam * Center for Research and Production of Vaccines and Biologicals, Hanoi, Vietnam ** National Pediatric Hospital, Hanoi, Vietnam *** Department of Enteric Diseases, Armed Force Research Institute for Medical Science,Bangkok, Thailand **** Center for Disease Control and Prevention, Atlanta, Georgia, USA

Fecal specimens collected from 501 children with acute gastroenteritis between November 2006 and October 2007 in the National Pediatric Hospital in Hanoi were screened for the presence of norovirus (NoV) and sapovirus (SaV) by real time RT-PCR. A subset of positive samples was sequenced for genotyping of the viruses. NoVs Gil and SaV were detected in 36.3% and 1.4% samples, respectively. NoV genotype I was not detected during this time. NoV could be detected year-round, highest in the sea­son when rotavirus detection is lowest.

NoV GII-4 was the dominant genotype, Minerva GII.42006b cluster which is dif­ferent from GII.4 strain isolated in 1998-1999 in the same region. A single case of GII-13 was also found. Among the 7 sam­ples positive for SaV, GI.2 and GJ.1.1 were identified in 4 samples. There is no differ­ence in NoV prevalence between age groups (from 6 months to 5 years). In this study, we confirmed for a very first time that NoV is the 2nd most important cause (after rotavirus) of acute diarrhea in children in Northern Vietnam.

90 Tap chi Y hoc du phdng, Tap XX. sd 5(113)