ONG DUNG KY THUAT PCR DA MOI PHAT HIEN CAC TAC NHAN GAY BENH VA SANG LOC CAC KIEU GEN MA HOA ENZYME SINH ESBL VA CARBAPENEMASES TRONG BENH PHAM NHIEM KHUAN VET M6 0 VIET NAM Ngé Tt Trung’, Tran Thj Thu Hién', Tran Thj Thanh Huyén', Bao Thanh Quyén', Mai Thanh Binh’, Phan Quéc Hoan', Christian G Meyer', Thirumalaisamy P Velavan', L@ Hiru Song'. Téng quan: Nhiém khudn vét m6 (Surgical site infection) ld bénh thudng gép 6 céc BN sau phiu thugt tai Vigt Nam. Bénh nay lam ting nguy co phoi nhiém cho BN, kéo dai thoi gian nim vign lam tang mitc chi phi va nguy c@ tir vong cao déi voi truéng hop phau thugt phitc tap nhu dat stent hog mé tim... Cho t6i hign nay, phuong phép chdn doan bang nudi cdy van duzge coi li tidu chudn ving é xéc dinh téc nhan gy bénh va tinh Khéng thuéc trong nhiém khudn vét mé. Tuy nhién, véi tinh chat da dang va phic tap cia logi bénh pham thi vin c6 khé nang tn tai hién turgng 4m tinh gia va thoi gian nudi cy kéo dai, Muc tiéu: Ung dung phurong phap PCR da méi phét hign ede téc nhin vi khuan thumg gép nhat gay bénh va céc kiéu gen mii hda cho enzyme betalactamases khang ESBL phé rong hoc carbapenemases tir bénh nhan nhiém khudn sau md @ bénh vién. Déi twong va phuong phép: Nghién ctu duge thyc hign trén 91 mau bénh phim ahiém khudn sau m6 tir BN tai Bénh vién Trung wong Quin d6i 108. Nghién ciru dnh gié dc Jap va so sinh két qud gta hai phuong phap nudi ca cy, lam khang sinh 4d va PCR da mi. Két qué vat két Iudin: Sit dung phuong phap PCR da mdi da t6i wu lam t8ng kha nang phét hign céc te nhin gay bénh tit céc bénh phim nhiém khudn vét mé so voi hytong phap nudi edy truyén théng. PCR da mdi co thé nhanh chéng dua ra két qua vé céc kiéu gen ma ha enzyme betalac- tamase lién quan dén khang betalactam cia vi khudn. Nghién eitu lin dau tién phat hién duge kiéu gen Turkey - specific ESBL (PER - 1) bag hai hg gen (Oxa23 va Oxa58) tai Viét Nam. Tir khéa: PCR da méi, nhiém khudn sau mé (SSI), ESBL, carbapenem. PAT VAN DE Nhi&m Khun bénh vign fa van G8 thang g§p & hing BN sau phu thugt, lim ting nguy od mac bénh, kéo dil thai gian ndm vién, chi phi ting va tng nguy oo tir vongtl, Nhiing BN bj nhigm Khun sau mé sé duc chim séc oBin thin, dude du tr céc liu php khang Khun hodc khéng nim, Tuy nhign, higu qua diéu ti cia (ign Lam sang ¥ hoc nbigt déi, Béah vién Trung wong Quan doi 108, ‘Nehy i bi: 1572015. Nedy phn blfn song: /0/122015 Ney day ding: 2601/2076 [Ng chie eric nhifm oft dung koa ge: Td Trg, Mn Lm sng Y he nid. Bénin Trane ome Qin di 108. Dif thoi: 0919119618. Ema murargi@email.com 30 - TRUYEWNHIEM VIET NAM & SO 1(19) - 2016 khang sinh chi yéu phy thuéc vao chinh cdc téc nhin vi sinh gy bénh. Cho d&n nay, nudi cy vi khudn vn dude coi [8 tiéu chu8n vang 68 xAc dinh tée nhdn gy bénh trong nhiém khudn sau md, Cac tac nhin gay bénh nay thuting gp nhdt nhw’ Staphylococcus aureus, Escherichia coli, Enterococcus spp., Pseudomonas aeruginosa, Enter obacter spp, Candida albicans, Klebsiella pneumonize, ‘nhém vi Khun Gram dung ky Khi va Proteus mirabilis Tuy nhién, phn tgp nudi cBy gp phai hai van 08 tn tai: {) Nubi cy chi cd thé phat hién céc vi sinh vat cb kha ming sinh truding trong diéu kién nudi cy nh&t dinh va chua xi ly khang sinh; ii) thdi gian cy khudn dai (ti 24 - 48h). ‘Trong khoang thdi gian nay tinh trang cla bénh nhan obthé thay di, thm chi nguy hiém hon), Mac di cdc khang sinh phd rng 48 dugc sir dung dé kiém soat SSI, nhung Sif gia ting cdc vi khudin Khang thudc Ia van 48 dang bso dng. S. aureus khéng Methiciline (ARMS) hoc ho Enter- obacteriace khang betalactamases (ESBL)/carbapene- mases#H1, Cho dén nay, hon mét ngan gen mi héa cho betalactamase (ESBL) hoc carbapenemase 43 dug thita, nh§ni®, Cc cBng bé cing chi ra ring, biéu hign lm sng inde nhau theo vi tri dia ly va dia hinh cia dia phung, Néu nudi ey vi khu8n that bai thi vide xc dinh khang sinh 46 s khéng thé thy hién due. ‘Vidt Narn 18 63 eutic 0b khi hu nhidt dt gid mira den, noi cb nguy od cao ala céc bénh truyén nhiém. Ty 6 mac SSI dupe xéc dinh bing nudi cSy 18 33967”. O nude ta, ob mt vai bao cdo vé SSI én quan tdi mim bénh nhung khéng 06 dif liéu vé kiéu hinh khang thude va chua 06 Inghién city hay bo co vé ki6u gen betalactamase. Nghién ‘ctu ca chiing tai cho thay, Staphylococaus epidermidts, E. coli, Pseudomonas auriginosa, Streptococcus sp, Klebsiella preumoniae, Enterobacter spp, Staphylococcus aureus va ‘Candida spp la nhiing téc nn nubi cy thuting gp nhSt gay nhiém Khugn vét mé (SSI) va khd ning ching mang cdc gen mé héa VEB, CTX - M cia nhém ESBL hoc NDM ~ cla carbapenemase. Nghién cit nly véi muc tiéy nm. tng dung PCR da mdi dé phat hién nhanh téc nhén gay bénh 6 mat trong mau nhiém khudn vét mé va khdo sét 138 cc ki@u hinh khéng ESBL hoc carbapenem 6 bénh nhdn nhiém khuén vét m6 (SSI), NGUYEN LIEU VA PHUONG PHAP Su bénh phdm: 91 mau bénh phdm dugc lay tir BN nhdp vién thang 2/2012 dén thing 12/2012 tai Bénh vién Trung udng Quan d6i 108 vdi mét trong cdc tiéu chudn sau: (1)C6 mid tai céc vét mé kin su 6 phn md mem. trong ving 30 ngay ké tir khi hoat déng hoc trong ving m@t nim d6i vai viéc cay ghép nhiing b} phan gid; (2)Tai Vi tri vat m8 cé triGu chimg dau, sung dé, phir né véi cde vét mS md va c6 su giém dinh cia bac sf. Bude sur dong {ca BN va cho phép cla Hi dng Y dite BEnh vién ‘Trung vong Quan 44i 108. Bénh ph3m tif vét mé sé dude chia lam hal va tén hanh song song hai phuding php nudi cy va PCR, Phan Ip vi khudin: Dé thiét lp dugc quy trinh POR da méi, ching t6i phn Igp cdc lodi vi Khudin dé lam ching duung Candida albicans, Acinetobacter baumanni, coccus epidermidis va E. coll tai Khoa Vi sinh vat, BEnh vign Trung wong Quan 46i 108. Nuéi cy vi khudn: Viéc nudi cy vi khudn duge ten hanh theo hung dan), Cu thé nhur sau: SOOp! dich ti vvét thutong dugc tra du trong 800 dung dich PBS. Dich. 1n6i dug nudi cay 6 bén loai mai truding khéc nhaur nhut ‘thach mau city, thach MacConkey, thach sécola va méi ‘truding thach 6 than hoat va cao men (BCYE) ¢ diéu kién 37°C trong ving 7 ngay. Khuan lac cla cdc vi khudin duc inh danh tai Khoa Vi sinh vat ‘Thiét k& mai cho cdc téc nhn cy thé trong phan ting PCR da mai: BE phat hién 8 téc nhan gay bénh ‘wén, chiing tdi chia ra lam hai b6 moi MicroSHPT @ SleX va MicroSHPT @ 3IeX, cdc bing dién di cb khodng cach tir 50 - 100bp dm bao phn bit 16 rang trén ge! dién di agarose (bang 1). Bang 1: Trinh ty mBi va gen dich dugc sis dung 48 sang loc tac nin vi sinh vat gy béah vamozem ———_ ASSersi™ rormurartevere Prine #08 39 socom Fig conensertion Pemacmann — avysen Smeets o =| “emmmoma Resin Tren » 2 Seotrmnte aroma atmcrnvncwcnnermonarris a sp « Sescanenan nasi om 2 tot cucrinernmaeae : TRUYEN NEEM VIET NAM x SO 1(13) - 2016 - 31‘Thiét ké mdi cho phan ing PCR da mdi sing loc cc. ho gen md héa cho ESBL hoc carbapenemase. ‘BE sng foc cic gen ma héa cho céc enzyme beta- tactamsase (ESBL) va carbapenemases, chiing tai sf dyng indim bd mi nhu/ sau: SHPT@ESBL - 1 (SHV, TEM, CTX - M), SHPT@ESBL - 2 (VEB, GES, PER), SHPT@Carba - 1 (NOM, SPM, VIM), SHPT@Carba - 2 (IMP, AIM, KPC/BIC, DIM), SHPT@Carba - 3 (0xa23 nhu nhém, OXA48 nhut nhém, Oxa58 nhu nhém) dB duigc thiét ké. Trong cic b6 ‘mdi trén, thi bd mdi SHPT@Carba - 3 {OXA nhémm) ching $61 thiét kE nh’ hudng tO bat tt cd cdc myc tiéu dua trén khu vc bdo tin cita ho gen ESBL (SHV, TEM, CTX - M, VEB, GES, PER) hoc ho gen carbapenemase (NDM, SPM, VIM, IMP, AIM, KPC/BIC, DIM). Céc b6 moi sif dung 48 sang loc cdc gen ma héa ESBL hoc carbapenemase ‘Gutde liét k@ trang bang 2. Céc chiing vi khuan mang gen. ‘OXA48, VIM, SIM, SPM, AIM, IMP, KPC, BIC, DIM dugc Laurent Poirel cung cfip!), Bang 2. Trinh by mai sif dung 46 sang I9c céc gen md héa cho ESBL hog carbapenemase [Ning 65] svwrecn- 9 Tm COM) = (omy sn senecns[eoeneuwonwce | an aenteecualomuenacacconoccee | ea cal sermons feemamocwrcoewee | om enieeTonin [ocomwenecaceciwe | om Tal senor femnneuneerucawene| arcecreTeoe senaensne ra SHPTOESBL - 2(VER, GES, PER) zl rneomner [oseermeneamne [ow eisoremerlorearerecrmancrm | oo a a oerinensont [oomaraemacc | 0 a ral POH cerremenoxen o ‘SHeTi ne | CTGAGCA ACC TGC GOA ATR ATA GCT T 008 Sree ‘SHPT@Carba - (MDM, SPH, VIM) Tal ewioemin Javonarenecmmene | on Ss ee ail sions lormeuarreemcecnes | artinonnn [erenrenmemnmnecne ost 1a SOO Lcocneemae | ow ‘SHPTIOGONOM-ER [eaccecocaGATOCTOMCTE 008 = 32 - TRUYENNEREM VIET NAM & SO 1(13) - 2016 bog i ‘SHPTOESIL - (SH, TEM, CTX) = (om) ‘SHPTocarta - 2CMP, ADM, KPC/BC, ODM) 710] surricw@ronee [aT TCAGCTTOTCTICOCTOCTAACG 8 seriveer-piw [craccarTocccTecarTearTTG cc} a|memcece — [crrermancractseactnoeracr | 02 TROGIR —_[AGCOMTOMCINCNNGIETECTECINGGE | 02 [2s] SeTIoOMMF | COCICANGCTETACGENMMCAC 2 sewrimoamte | GasrrescccaccTocaaTts 008 oa leer Hence Tener TIEAGGIOTIMNEROMACCASSTINT] 0. ‘SerTocarta -3(0na23 le grou, OXA48 the grup, Ora He rom) 585] irigeeca- 504 | CCOCTCTGCGCTCIACATACANCATC co ‘SHPTIOBGO@ SER | ANGTATTGGGCTTTECTGAGCATAG 8 [AGAATATETGIC)OCENTSCICATTCCVACA | 8.2 aeironcon je moaazn — |econamecemenemmccncnne | 2 aoe) cea cetF2 | CACCANGTCTTIANGTEGGATEGACA 08 Toa Get-r2 —_[COGATACGTGTANCTIATIGTGATACAGCTT | 008 ‘Tach chiét ADN va thyc hién phan ting PCR Miu bénh phém duige ly tim thu cin, sau 46 thém v0 300y dung dich universat lysis solution (200m NaOH, 1% SDS) dé U 6 nhiét 46 95°C trong 5 phit Sau 46 trung haa bing 250pl trisHCL 1M sao cho pH dat mite 7,5. Thém 400i dung dich phenal/chlorofornyisoamys at cohol vao hn hgp trén sau 6 ly tém 13,000g trong $ phiot™!, LSy khodng 600p! dich ndi chuyén sang mét Gng ‘eppendorf méi va thém vao dung dich thu dug nay iso- propanol theo ty [@ 1:1. Trén déu dung dich va ly tam 16,000g trong 30 phit thu duc cin ADN, tiép tye riva hal fan bing cBn 70% va kam khé ADN. Cuéi cing thém 150g ‘TE (251M Tris - base pHi 8.0, ImM EDTA) va si dyng Spl cho phan ting PCR thé tich 2Sul bao gém: 10mm Tis - HO pH 8.3/50mM KCI/1.5mM M2, 250uM dNTP vi idng moi theo bang 1 va 2. Chu trinh nhigt 46 nhu sau: 95°C trong 4 philt, (95°C trong 25 gidy, 58°C trong 45 gidy, 72°C trong 1 phiit) x 35 chu ky; 72°C trong § phit. Tong thi aghiém nay, cdc sin phim khuéch dai duigc tinh sach lam khugn cho phan ding PCR giai trinh ty gen. Két qué gidi trinh ty dudc doc va so sdnh 661 chigu vet cdc erinh ‘Y’ gen 3 6uge céng b6 trén ngén hang gen qué tf NCBI (hitp://blast.ncbi.nim.nih.gov/Blast.cgi).KET QUA Kt qué PCR da mdi phat hign tac nhSn nang cao ty 1@ dufang tinh cia SST ‘Sau khi ti uti hda digu kién phan ng cho hai b moi MicroSHPT@SIeX vi MicroSHPT@3IeX (Hinh 1, Bang 1), chiing t6i tién hanh sang lgc trén 91 mu bénh phdm. K&t qua cy thé 3 bang 3, LHEEled phe 1100 " so 0 00 20 rr 200 100, 0 Hinh 1: C4c phan ting Multiplex PCR assays sang loc vi sinh vat gay nhiém khugin viet mo Panel tréi phan Ging MicroSHPT@SleX dc higu cho Candida albicans, Acinetobacter baurmanni, Pseudomonas aeruginosa, Staphylococcus aureus, Escherichia coli, trong hi 46 panel phdi MicroSHPT@3Iex Khuéch dat cic gene ‘3c higu cho Klebsiella pneumonia, Streptococcus pneu- monia, Staphylococcus epidermidis. Control - 1, control - 2 1B DNA nguts. Bing 3: Két qua PCR da mdi va nudi cy phat hign téc nhn gay bénh trong mu bénh phim sshiém khudn vet mé Mam bénh Sa an, Candida abicons 0 0 Acinetobacter baumanni 4 n Pseudomonas aeruginosa 9 8 Kebsiela pneumonia 5 4 Staphylococcus aureus 2 B | Steetococeus pneumania 0 0 ‘Staphylococcus epidermidis. 3 3 Escherichia coli 6 10 Citrobacter sp. 1 Q Téng 55 mBu (derong tinh) 40 0 Téng s6 mau a1 9 Két qua cho thay, 40 trong s6 91 bénh pham SSIs cho ké&t qua dufdng tinh bang nudt cy Khuda (moc khudn lac cca it nht 01 fodi vi sinh vat) trong khi dé 60 bénh phém cho két qud dung tinh bing PCR da moi (Mi- ‘croSHPT@SleX or MicroSHPT@3leX reactions) vot it nat 01 loai vi sinh vat (bang 3). Ngoai tri’ Citrobacter spp, tat cd cdc khudn lac déu dugc khéng dinh igi bing két qua dufdng tinh boi MicroSHPT@SleX hoc MicroSHPT@3IeX. C6 bon mau dutie xac dinh [a 4m tinh véi nudi cy nhumng dufong tinh vai PCR. Két qua cing chi ra rng, nudi cy vi kkhudin 6 ty 1é ducing tinh thap hon so vél PCR da moi. (BiBu nay chimg to rang this nghiém PCR da mdi ma ching 46i thidt ké 06 4 nhay cao v8 chinh xéc trong chén doan ‘SSI hon phung phap nudi cay. Két qua sing loc kiéu gen m3 héa enzyme betalac- tamase tiy cc miu bénh phdm SSI Két qua PCR da mi sir dyng céc b6 moi SHPT@ ESBL +1, SHPT@ ESBL - 2, SHPT@Carba - 1, SHPT@Carba - 2, SHPT@Carba - 3 cy thé d hinh 2. N&m phan ting PCR da mdi dugc tién hanh aé sing loc céc gen ma héa enzyme ‘sinh ESBL hoc carbapenemase tir 60 mu dud xde dinh 18 dong tinh bing PCR (Bang 3). Két qua sang loc cho thay ty I6 duong tinh vai ho gen SHV cao nhdt 13 10/60 ca (chiém 16,6%). Ngoai ra cdc ho gen sinh ESBL khac nh’ CTX - M_ hoe TEM cling dugc phat hign trén cdc mau SSI. Bc biét 06 nm mBu PCR dung tinh ntut (NKSMS, NKSM11, NKSM28, NKSM33, NKSM40) cho két qué dung ‘tinh voi NDM - 1 mai héa enzyme carbapenemase va cling duong tinh vdi SHV, CTX - M hodc TEM. Mu NKSM 28 dung tinh vdi kiéu gen Turkey - specific ESBL (PER - 1) va cic gen thudc ho gen oxaciline nhu (Oxa23 v3 Oxa58) ‘a bang 4 duéi day: SLOPE PP a sa ==~ SHPT@ESDL2 | SHPT@Cube? ioe bry ‘SHPT@Carbe| Hinh 2: Phan ifng Multiplex PCR sang loc gene ESBL va carbapenemase ‘SHPTI@Cabe> ‘TRUYEN NHRERA VIET NAM & SO 1(13) - 2016 - 33Panel trén cing SHPT@ESBL - 1 sin tim cdc gene ‘ESB thuc ho SHY, TEM, CTX M; pane! gia b&n trai phan (ing SHPT@ESBL - 2 phét hign céc gene ESBL thudc tie VEB, GES, PER genes; trong kh d6 panel gta bn phai 43 phan ing SHPT@Carba - 2 phat higa IMP, AIM, Bang 4: Két qua sang loc kiéu gen ma héa cho enzyme KPC/BIC, DIM; panel dudi cling bén trdi 13 phan dng ‘SHPT@Carba - 1 phat hién NDM, SPM, VIM; panel dudi cing bén phai la phan ding SHPT@Carba - 3 phat hin 0xa23 like group, Oxad8 like group va OxaS8 like group; Mu ching ld genomics DNA ngutt. e betalactamases detected ti’ mu bénh phim Sst aR qua xbe Gah mlm bent 1 gud PR 0s ng ie cic Rd gen kha ony bfalaomaser sr | PER de mel wodicky | wom | soy | cmem] tem | os | Peri | omea | ves [ous asi | £5 poagmonee, © cas [vena | Ouong tn] Oven Oven nh sau Fo Bet | dustnoemg tin oven tt dg sn woes | 4 taunant rte ou et oso ts |Duagt g i {xt be | ug 190 ERS en earn [oes on orn oye somo | preumeiee | pramense [ovo tng i] Deng seh erg son ste | Ranma Raines uma sang |? acon Same | ay oven | ‘cu in wore | A naman ‘nt oxen | vwsiss | panini Ravi Due | mcs | A bamond Atamesi vg [ang sah (nas eas ne) BAN LUAN 63 iu tr khéng sinh thi vigc nudi cay s& gp kis Kin, (Céc téc nhén gay nhiém triing bénh vién sau phu thuit va tinh Khang khang sinh [& mdi de doa tdi stfc khde ong dng. & nghién city ndy, chiing tid t6i vu héa hai this nghiém PCR da mii: i)Phat hién tam téc nn gSy bénh thuting g&p nhat nhu Candida albicans, Acinetobac- ter baumanni, Pseudomonas aeruginosa, Klebsiella pneu- moniae, Staphylococcus aureus, Streptococcus ‘pneumoniae, Staphylococcus epidermidis va Escherichia ‘coli 06 kha ning phét hign tir 20 - 50 CFU/m! mu nguts (s6 figu chua cing bd). 861 chiéu vii két qua nudi cy, ket qua PCR da mdi 6 56 truding hop duting tinh cao hon & mu nhiém khudn vét m6. Bc biét Id PCR nhay hon so vi phuttng phép cay méu, (Mt phn khdc khGng thé khdng nhc tdi trong nhiém tring bénh vién Ia hién tung Khang khang sinh ng’y ng gia ting va vide di hdi phat trign cing cy giém sat ‘va theo dai la cn thiét. Tuy nhién, voi nhing truting hop. 34 - TRUYENDISEM VIET NAM x SO 1(13) - 2016 thim chi viée nuBi cSy $8 thSt bai thi vide xSc din tinh khang cia mm bénh [a di@u khéng thé, Do vay trong thi nghiém nay ching t8i 68 xudt phuiong phap PCR da mii ‘sang loc cc gen ms héa cho enzyme betakactamase vit kd niing thiy phn duc chat betalactam 6 cic mit 65 hdc nhau va hogt tinh carbapenemase. Ching t6i 65 tii ‘uu hdéa phan ing PCR da mdi huting dich phat hign ofc kiéu gen Vietnam - specific ESBL nh VES, ho gen DlaCTX - M = 15, BlaCTX - M - 228! hoc ho gen E58. khée nhu NDM - 1, OXA48, GIM - 1. Két qua cla ching ti cho thy rang, mac di Khéng co két qua nudi cay phan ip cla cic mu NKSM28, NKSM54 (bang 4) nhung bing PCR 6a mdi chiing t6i vn 6 thé xc dinh 6ugc mmm bah ob trong cdc méu nay va sing loc duc cdc gen ma héa cho cc enzyme betalactamase nhu (NDM - 1, SHY, CTX-M, GIM - 1, VEB, OXA - 58 trong mau bénh NKSM28, hose SHV trong NKSMS4, bang 4).Mic di Ky thut PCR 0b 48 nhay cao va 06 thé xd ink tc nhén vi sinh vat g8y nhiém khudn vét m6, ching ta van chua thé bd qua phudng phdp cdy khugn truy8n théng, Thém vao 46 phan img PCR khéng thé cho phd toan dién céc vi sinh vat g8y bénh cling nhu tinh chit khang khéng sinh cla vi Khun. Vi thé cd hai phuting php nn dude six dung theo cach bé trg cho nhau. kéT LUAN ~ Sif dung phuong phdp PCR da mdi 68 téi uu lam ‘ting kh ning chin doan cde téc nhan gy bénh trong cdc truting hgp nhiém Khun vét md SSI so véi phudng phap nudi cy truyn thing, ~ Sif dung PCR da moi o6 thé nhanh chéng dua ra két ‘qua vé cdc kiéu gen ma héa enzyme betalactamase sinh tinh khéng betalactam cia vi khuén, ~ Nghién citu an dau tién phat hién dugc kiéu gen ‘Turkey - specific ESBL (PER - 1) bag hai ho gen (xa23 va Oxa58) tai Vit Nam. TAL LIU THAM KHAO 4. Owens, C.D. and K, Stoessel, Surgical site infec- tions: epidemiology, microbiology and prevention. Hosp Infect, 2008, 70 Suppl 2: p. 3 - 10. 2. Qadan, M. and W.G. Cheadle, Common microbial pathogens in surgical practice. Surg Giin North Am, 2009. 89(2): p. 295 - 310, vil 3. Mancini N., et aly The era of molecular and other ‘non - culture - based methods in diagnosis of sepsis, Cin Microbiol Rev, 2010. 23(1): p. 235 - Si. 4, Cao, \), et al, Distribution of extended - spectrum beta - lactamases in clinical isolates of Enterobacteriaceae in Vietnam, Antimicrob Agents Chemother, 2002. 46(12): 1. 3739-43, 5, Nordmana, P, T. Naas, and L. Poirel, Global spread of Carbapenemase - producing Enterobacteriaceae. Emerg Infect Dis, 2011. 17(10): p. 1791 - 8. 6. Jacoby, G.A. and LS, Munoz - Price, The new beta ~factamases. N Engl J Med, 2005. 352(4): p. 380 - 91. 7. Nguyen, D, et al, Incidence and predictors of sur- gical - site infections in Vietnam. Infect Control Hosp Epi- demiol, 2001. 22(8): p. 485 - 92. 8. Patrick R. Murray, E..B,, Michael A. Pfaller, Fred C. ‘Tenover, Robert H. Yolken, Manual of Clinical Microbiology. Washington. D.C. ASMPress, 1999. 9. Poirel, L., et al,, Multiplex PCR for detection of ac- quired carbapenemase genes, Diagn Microbiol Infect Dis, 2011. 70(1): p. 119 - 23. 10, Ausubel, FB, R.; Kingston, R.; Moore, D.; Seid- man, 1.G.; Smith, J.;Struhl, K. , Short Protocols in Molec- Ular Biology 3rd ed. 1995( ): p. page 2- 3. 11. Naas, T, et al,, Integron - located VEB - 1 ex- tended - spectrum beta - lactamase gene in a Proteus mirabilis clinical isolate from Vietnam. J Antimicrob Chemother, 2000. 46(5): p. 703 - 11. 12. Trang, NL, et al, The characterization of ESBL genes in Escherichia coli and Klebsiella pneumoniae caus- jing nosocomial infections in Vietnam. J Infect Dev Cries, 2013. 7(12): p. 922 - 8. ‘TRUYENNHIEM VIETNAM & SO 1(13) - 2016 - 35‘SIMPLE MULTIPLEX PCR ASSAYS TO DETEC COMMON PATHOGENS AND ASSOCIATED GENES ENCODING FOR ACQUIRED EXTENDED SPECTRUM BETALACTAMASES (ESBL) OR CARBAPENEMASES FROM SURGICAL SITE SPECIMENS IN VIETNAM ‘Summary Surgical site infection (SSI) is common in Vietnamese Post - operative patients, it contributes to increased mor- bidity, mortality, hospitalization time and health care ex- penditure. Bacterial culture is considered the gold standard procedure to identify SSI pathogens and antibi- otic resistant properties; However, it can detect microbes, that can readily grow and is time - consuming. We pro- pose optimized multiplex PCR assays to detect the mast relevant microbes and associated genes encoding for ac- quired extended spectrum betalactamases (ESBL) or car- bapenemases from Vietnamese patients with SSI in a hospital setting in Hanoi, Methods: Ninety one samples from patients (n = 91) were collected in order to identify microbial pathogens and associated genes encoding for acquired extended spectrum betalactamases (ESBL) or carbapenemases by both conventional bacterial culture and in-house multiplex PCR assays. Resuits and Condu- sions; The novel in - house multiplex PCR assays were ‘comparable to the bacterial culture approach in screening for common pathogens causing SSI and for relevant geno- types conferring betalactam/carbapenem resistance for bacteria. This is the first report of Turkey - specific ESBL gene {PER - 1) and two Oxacilinase families (Oxa23 and ‘Oxa 58) in Vietnam. Key words: Multiplex PCR, surgical site infection (SSI}, ESBL, carbapenem. 36 . TRUYENINHIEM VIETNAM % SO 1(13) - 2016.