Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi

PHN I. M U

1.1. t vn
Nc v thc phm l ngun nui sng con ngi nhng cng c th gy ng c hoc gy bnh cho con ngi do chng c th cha cc c t vi sinh vt, c t ha hc hoc vi sinh vt gy bnh. Trong nhng nm gn y, ng c thc phm c ghi nhn kh thng xuyn v tr thnh mi quan tm ca ton x hi. C nhiu nguyn nhn khc nhau c th gy ra cc v ng c thc phm nhng phn ln cc trng hp l do vi sinh vt gy ra. Nguyn nhn cc v ng c thc phm l do c s hin din mt lng ln cc vi sinh vt gy bnh hay c t tit ra bi cc vi sinh vt. Theo di v tng kt trong nhiu nm cc nh khoa hc trn th gii u xc nh nguyn nhn ch yu gy ng c thc phm trong n ung l do nhim vi khun gy bnh trong thc phm, nh vi khun gy bnh lao, st thng hn v dch t Canada hng nm c khong trn 2 triu ngi b ng c do thc n, nu tnh trung bnh theo dn s th c 11 ngi c 1 ngi mc bnh. M c khong 13 triu ngi ng c thc n trong nm v c 18 ngi c 1 ngi mc bnh do thc phm, trong 85% s ca ng c thc n l do nguyn nhn vi sinh. An ton thc phm hin l mi quan tm ln ca x hi. Ngi tiu dng ngy nay hiu bit v cc nguyn nhn chnh gy nhng cn i dch do thc phm v i hi nhng ngun cung cp thc phm an ton. Cc nh vi sinh vt thc phm c trch nhim phi nghin cu nh lng, phn lp, pht hin, miu t, ngn nga v kim sot cc vi sinh vt gy bnh thc phm, trong nc v trong c mi trng. Vic kim tra cc ch tiu vi sinh trong thc phm cn c quan tm nhiu hn khi sn phm nc ta ang ha nhp vo th trng th gii. Cng trong mi quan tm n Cht lng, v sinh an ton thc phm hin nay Vit Nam.

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 1

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi Xut pht t nhng l do trn chng ti tin hnh thc hin bi kha lun: Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi.

1.2. Mc ch
Nhm kho st t l nhim S. aureus nhim trong cc nhm thc phm nh ch c chin n ngay, ch la, ... Nhm ngn chn ng c thc phm t h gia nh hay cc bp n tp th.

1.3. Ni dung
Kho st t l nhim S. aureus trong cc loi thc phm ang lu hnh trn th trng. ti c ngha ng gp mt phn vo vic gip cho c quan qun l thc phm nh gi v vn cht lng thc phm trn a bn th x ng Xoi, tnh Bnh Phc v nhn nhn khch quan v mi nguy him tht s ca S. aureus. Do thi gian c hn nn y ch l nhng nh gi khi u t l nhim S. aureus gip ngn nga mt cch hiu qu ng c thc phm. Cc kt qu thu c cng l thnh qu bc u tip cn vi cng tc nghin cu trong phng th nghim nn ni dung chc chn cn rt nhiu thiu st do kha lun khng th trnh khi nhng khim khuyt. Rt mong nhn c s nhn xt, nh gi qu bu v s ng gp chn thnh ca qu thy c, cc anh ch v cc bn kha lun c xc tch hn, hon thin hn.

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 2

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi

PHN II: TNG QUAN TI LIU A. V TRUNG TM Y T D PHONG TNH BINH PHC I. LICH S PHAT TRIN
1.1.S thanh lp
Trc khi thanh lp, Trung Tm Y T D Phong tinh Binh Phc c mang tn la Tram V Sinh Phong Dich (VSPD), sau o i tn thanh Trung tm VSPD. n thang 4 nm 1997 theo quyt inh s 239/TTg cua thu tng chinh phu, a i tn Trung Tm VSPD thanh Trung tm Y T D Phong (TT YTDP).

1.2.Vi tri, chc nng

TT YTDP tinh Binh Phc la n vi s nghip thuc S Y t, co chc nng tham mu va t chc trin khai thc hin cac nhim vu chuyn mn, ky thut v y t d phong trn ia ban tinh.

1.3.Nhim vu
- Xy dng cac k hoach trin khai thc hin cac nhim vu chuyn mn ky thut v y t d phong trn c s inh hng chin lc cua B Y t va tinh hinh thc t cua tinh trinh giam c S Y t ph duyt. - Trin khai thc hin cac hoat ng chuyn mn, ky thut v: Phong chng dich bnh, dinh dng cng ng, kim dich y t, sc khoe mi trng, sc khoe trng hoc, sc khoe ngh nghip, phong chng tai nan thng tich va xy dng cng ng an toan. - Thc hin cng tac kim nghim cht lng, v sinh an toan thc phm. - Chi ao, hng dn va giam sat chuyn mn, ky thut v cac hoat ng thuc linh vc phu trach vi TT YTDP huyn, cac c s y t va cac tram y t trn ia ban.

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 3

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi - Phi hp vi trung tm truyn thng giao duc sc khoe va cac c quan thng tin ai chung trn ia ban tinh t chc trin khai thc hin cng tac thng tin, truyn thng giao duc sc khoe v linh vc y t d phong. - Tham gia ao tao va ao tao lai chuyn mn, ky thut v linh vc y t d phong theo k hoach cua tinh va trung ng cho can b chuyn khoa va cac can b khac. - Nghin cu va tham gia nghin cu khoa hoc,ng dung cac tin b khoa hoc, ky thut v linh vc y t d phong. - Quan ly va t chc trin khai thc hin cac d an thuc chng trinh muc tiu y t quc gia va cac d an khac c giam c S Y t phn cng. - Trin khai t chc thc hin cac dich vu v y t d phong theo s phn cng, y quyn cua giam c S Y t va theo quy inh cua phap lut. - Phi hp vi cac c quan lin quan thc hin cng tac thanh tra, kim tra, anh gia cac hoat ng thuc linh vc phu trach. - Thc hin cac ch thng k, bao cao theo quy inh cua phap lut. - Thc hin cac nhim vu, quyn han khac do giam c S Y t giao.

II.T CHC B MAY 2.1. T chc

2.1.1. Lnh o trung tm: Gm c 01 gim c v 01 ph gim c - Gim c : BSCKI T c Sinh - Ph gim c : Ths.BS Phm Hong Xun u c trnh t bc s chuyn khoa cp I hoc thc s tr ln. 2.1.2. Cc phng chc nng : SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 4

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi - Phng T Chc Hnh Chnh - Phng K Ton Ti V 2.1.3. Cc khoa chuyn mn : - Khoa Dch T - Khoa V Sinh Mi Trng Nha Hc ng - Khoa Y T Lao ng - Khoa Kim Dch Y T Bin Gii - Khoa Xt Nghim

S t chc ca TTYT D phng tnh Bnh Phc

GIM C

PH GIM C

P.HNH CHNH

P.K TON

KDYT BG

YTL

VSMTNH

XT NGHIM

DCH T

2.2. Bin ch v nh mc :
Bin ch nh mc lao ng ca TT YTDP tnh c thc hin theo quy nh ca php lut. SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 5

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi

2.3. Kinh ph hot ng :

- Ngn sch s nghip y t - Ngn sch chng trnh mc tiu quc gia y t - Thu ph l ph theo quy nh ca php lut - Vin tr v cc ngun kinh ph hp php khc.

III.GII THIU V KHOA XET NGHIM CUA TRUNG TM

V cng tac xet nghim cua trung tm: - Chi tiu cac loai mu trong nm 2012 la 650 mu. Nhn mu va tra mu ung thi gian quy inh, nu mu nhiu thi t chc lam ngoai gi ap ng mu cho khach hang. - Thc hin cac xet nghim theo yu cu cua cng tac y t d phong. Thanh lp cac labo ky thut ring bit theo tng linh vc. - Nghin cu tip nhn cac ky thut xet nghim mi phuc vu cho cng tac chn oan bnh. - Tham gia cng tac ao tao cho can b chuyn khoa va cac i tng khac. - am bao san xut, pha ch mi trng nui cy, hoa cht xet nghim cung cp theo yu cu cho tuyn huyn phuc vu cho cng tac phong chng dich bnhh. - Thc hin dich vu xet nghim trong linh vc y t d phong theo quy inh hin hanh. - Thng nht ap dung thng quy ky thut xet nghim a c B Y t ban hanh, ph bin ky thut cho cac tuyn. Kim tra anh gia vic thc hin theo thng quy ky thut a c thng nht. - Chiu trach nhim v kt qua xet nghim i vi cp trn va phap lut.

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 6

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi - Thc hin cng tac thng k bao cao theo quy inh. Khoa xet nghim gm co 07 phong: - Phng Lu Nhn mu - Phong Vi sinh Nc Thc phm - Phng Vi sinh Bnh phm - Phong Ly hoa Nc Thc Phm - Phng sinh ha huyt hc - Phng k sinh trng - Phng Hnh Chnh Khoa xet nghim a ky hp ng kim tra nc sach nh k 06 thng/ ln i vi cc nh my cung cp nc trn a bn tnh Bnh Phc nh : nh my nc B Gia Mp, nh my nc Ph Ring, nh my nc cao su ng Ph, ng Xoi, Bnh Long, trm cp nc Tn Tin, trm cp nc H ng Thoi, nh my nc Phc Long.....

IV. S TRUNG TM Y T D PHNG TNH BNH PHC

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 7

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 8

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi

Chinh Din
Ca chnh

Ca Phu

Ln Xung
Cu Thang

Ln Xung
Cu Thang

Phong Lu Nhn mu

Phong trng
Ging Tri

Tng Trt
Phong cha ha cht

Phng ph gim c

Li i

Labo l ha Nc Thc Phm

Phong Tim Nga

Sanh

Labo XN Vi Sinh Nc Thc phm

Phong Hanh Chanh

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn

Phong Trng khoa Xt Nghim

Khu V Sinh

Labo sinh ha huyt hc

Li i

Phong Nha Hoc ng

Phong Nui Cy Vi Trung

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi

WC N

WC Nam

Khoa v sinh mi trng - Nha hc ng

Khoa Sc Kho Ngh Nghip

Khoa Kim Dch Y T Bin gii

Hi trng

Li i

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn

Khoa dch t Li i

Ging Tri

Phong K Toan Tai Vu

TNG MT

Kho Dich T
Phong Gim c

Phong Hnh chnh khoa Xet Nghim

Phong Trng VSMT NH

Phong Trng Khoa Dich T

Ln Xung Cu Thang

Ln Xung Cu Thang

10

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi

V. AN TON TRONG PHNG TH NGHIM

m bo hiu qu v an ton trong phng th nghim, th nghim vin phi nghim chnh tun theo nhng quy nh v an ton trong phng th nghim. 5.1. Ni quy trong phng th nghim. - Th nghim vin cn phi nm vng v duy tr quy tc an ton trong phng th nghim ra v nghim chnh chp hnh. - Th nghim vin lm vic phi c qun o trang b chuyn mn ng tiu chun quy nh. - Cm lm vic bng nhng dng c cha c kim tra nh: bnh, cc, v cc ng ong dung dch... - Khng s dngdng c b h hng. - Khi lm vic phi gi trt t, cm an ung, ht thuc trong phng th nghim. - Khi s dng ho cht d chy, d n, dng c d v, t tin phi tuyt i tun theo ch dn. - Cn c thc tit kim ho cht, trnh gy v dng c. - Khng di chuyn ho cht khi ch quy nh. Trc khi m ho cht phi lau sch np v c chai. - Dng c dng ly ho cht phi tht sch v dng xong phi ra ngay. Khng dng ln cc dng c ly ho cht cho cc loi ho cht khc nhau. - G sch s ni lm vic, ra dng c v lau chi ngn np ni lm vic. - Phi thc hin quy nh v phng chy cha chy. - Khi ra v phi tt n, ngt cu dao in v kim tra cc vi nc

5.2. Quy tc lm vic vi ho cht c, d chy, d n.

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 11

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi a s cc cht hu c s dng trong phng th nghm u c hi, do cn phi nm vng qui tc chng c, chng chy n khi lm vic vi cc cht hu c. - Ho cht phi cha trong chai. L c np y, dn nhn. Khi cm chai ho cht khng c xch c chai m phi b y chai. - S dng cc cht NaCN, HCN,(CH3)2SO4,,CH3NH2, Cl2, NO2....Phi eo mt n, knh bo him v phi lm trong t ht v khng tt my khi cn cht c. - S dng Na, K.... phi dng kp st, lau kh bng giy lc v dng ru butylic hay amylic hu Na v K d. - Brm c cha trong bnh y, mu ti c nt nhm, rt brm phi tin hnh trong t ht, eo knh bo him v gng tay. Mi ln ly brm khng qu 10ml, khi cho vo bnh phn ng phi dng phu nh git th kn. - Khi lm vic vi H2SO4 c phi rt cn thn qua phu trong t ht. Pha long acid H2SO4 phi trong bnh chu nhit v rt t t acid vo nc khuy u. - Bao gi cng acid (baz) vo nc khi pha long ( tuyt i khng lm ngc li). - Khng dng ming ht acid(baz). Khng ht bng pipet khi cn t ha cht trong l. -S dng ho cht d chy nh cn,benzene,acetone,etylacetat,. . . phi xa ngn la, khng un nng trc tip trn ngn la m phi dng bp cch thu.

5.3. S cu trong phng th nghim.

5.3.1. i vi ha cht - Bng acid phi ra ngay vt bng bng vi nc mnh t 3-5 pht dng bng tm KMnSO4 3% bi nh ln vt bng, hoc dng natribicacbonat long (1%) ra vt bng.

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 12

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi - Bng kim c th tin hnh nh trn nhng thy nc bng dung dch acid axetic 1%. - Khi b ho cht bn vo mt phi ra ngay mt bng dng nc sch hoc NaCl 1% chy lin tc v a ngay n bnh vin. - Bng bi vt nng ( thu tinh, kim loi) th phi bi dung dch KMnSO4 3% ri bi m chng bng. - Bng bi P bi ch bng bng dung dch CuSO4 2%. - Trng hp ung phi acid th phi sc ming v ung nc lnh c MgO. - Trng hp ung phi baz th phi sc ming v ung nc lnh c cha acid acetic 1%. - Ng c kh Clo, brm th a ngay ra ch thong c khng kh trong lnh. - Ng c bi asen, thu tinh, mui xianua... phi nhanh chng a n bnh vin. - B t tay phi lau sch mu , st trng bng cn hay dung dch KMnSO4 3% ri cm mu bng dung dch FeCl3 v bng li. - Khi b chy qun o trn ngi vi din tch ln th tuyt i khng chy ra ch gi phi nm xung v ln dp tt la, nu din tch chy b th dng nc, gi lau dp tt. 5.3.2. i vi mu thc phm bnh phm (c VSV gy bnh) - Vt sc nhn m vo tay, chn trong khi lm vic vi tc nhn gy bnh phi bc l vt thng, x nc vo vt thng trong 15 pht. Dung bng, gc che vt thng sau n phng s cu ca c quan hay c s iu tr gn nht. - Nut phi cht c nguy c ly nhim phi sc ming ngay bng nc sch ri nh vo bnh cha dung dch dit khun. n phng s cu ca c quan hay c s iu tr gn nht. SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 13

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi - S c c kh dung: tt c mi ngi phi lp tc ri khi vng b nhim v ng ca phng khong 1 gi kh dung lng xung. Ty trng qun o bo h. ngi ht phi kh dung phi c bc s khm v tham vn. - Lm mu: phi dung khn giy thm kh mu b , p khn giy khc ln cht kh nhim ln khn, khong 30 pht thi gian cho cht kh nhim dit khun. Thu dn sch vng b mu.
B. TNG QUAN V STAPHYLOCOCCUS AUREUS

I. Gii thiu v Staphylococcus:

Staphylococcus c ngun gc t ting Latinh, staphylo (chm nho) v coccus (ht). Phn loi ca vi khun Staphylococcus nh sau: Gii: Prokaryote Phn loi: Firmicute Lp: Firmibacteria H: Micrococceae Ging: Staphylococcus

1.1. Hnh thi

Staphylococcus l vi khun gram dng, hnh cu ng knh 0,5 - 1,5 m, c th ng ring l, tng i, tng chui ngn, hoc tng chm khng u ging nh chm nho. y l loi vi khun khng di ng v khng sinh bo t, thng c tr trn da v mng nhy ca ngi v ng vt mu nng. Nm 1871, Recklinghausen thu c cu khun trong thn ca bnh nhn cht do bnh nhim khun huyt. Nm 1880, Alexander Ogston chng minh c p-xe sinh m l do cu khun dng chm v Ogston c cng nhn l ngi khm ph v t tn cho t cu Staphylococcus vo SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 14

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi nm 1882. Nm 1884, Rosenbach nghin cu v t tn cho cu khun to khun lc mu vng l Staphylococcus pyrogen aureus (Scott E Martin v John J Iandolo, 2000).

Hnh 1.1. Hnh thi Staphylococcus aureus ( Ngun: Theo <www.biotox.cz/.../staphylococcus_aureus_1s.jpg>)

1.2. Tnh cht

Staphylococcus l nhng vi khun hiu kh hoc k kh ty nghi, c c s trao i cht, h hp v ln men. Chng cho phn ng catalase dng tnh v c th s dng nhiu loi carbonhidrat khc nhau to acid lactic nhng khng sinh hi. Khun lc trn mi trng khng chn lc nh Tryptic soy agar thng t mu kem n mu cam. Thnh t bo cha peptidoglican hnh thnh mt hng ro cng vng chc xung quanh t bo v acid teichoic gip duy tr mi trng ion thch hp cho mng cytoplasma, ng thi gp phn bo v b mt t bo t cu. Staphylococcus c th mc nhiu iu kin, mi trng khc nhau, nhng tt nht nhit t 30-37oC v pH gn trung tnh. Chng khng c vi cc cht dit trng, kh nng v c kh nng tng trng trong mi trng cha n 15% NaCl (Scott E Martin v John J Iandolo, 2000).

1.3. Phn loi

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 15

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi Hin nay ngi ta bit c 33 loi Staphylococcus (Mary K. Sandel v John L. McKillip, 2002) nhng c mt s loi t c quan tm. C th chia Staphylococcus thnh 3 nhm: - Nhm cho phn ng coagulase dng tnh. - Nhm cho phn ng coagulase m tnh v nhy vi Nobovicine. - Nhm cho phn ng coagulase m tnh v khng vi Nobovicine. Trong s cc loi Staphylococcus th Staphylococcus aureus l loi thng gp nht, chng thuc nhm cho phn ng coagulase dng tnh.

1.4. c im sinh ha
S.aureus l nhng vi khun hiu kh hay k kh ty nghi, c enzyme catalase phn gii oxy gi gii phng oxy v nc:

catalase H2O2 H2O + O2

S. aureus cho phn ng ng huyt tng dng tnh do chng tit ra enzyme coagulase. y c xem l tnh cht c trng ca S. aureus, l tiu chun phn bit S. aureus vi cc t cu khc. C hai dng coagulase: coagulase c nh (bound coagulase) gn vo thnh t bo v coagulase t do (free coagulase) c phng thch khi thnh t bo. C hai phng php thc hin th nghim coagulase l thc hin trn lam knh v trong ng nghim. Phng php lam knh gip pht hin nhng coagulase c nh bng cch phn ng trc tip vi fibrinogen, phng php ng nghim pht hin nhng coagulase t do bng phn ng gin tip vi fibrinogen qua cng hp vi nhng yu t khc trong huyt tng (C H Collin v ctv, 1995).

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 16

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi Ngoi ra, chng cn cho phn ng DNAse, phosphatase dng tnh, c kh nng ln men v sinh acid t manitol, trehalose, sucrose. Tt c cc dng S. aureus u nhy vi Novobicine, c kh nng tng trng trong mi trng cha n 15% mui NaCl (Trn Linh Thc, 2002). Mt s dng S. aureus c kh nng gy tan mu trn mi trng thch mu, vng tan mu ph thuc vo tng chng nhng chng u c vng tan mu hp hn so vi ng knh khun lc. Hu ht cc dng S. aureus u to sc t vng, nhng cc sc t ny t thy khi qu trnh nui cy cn non m thng thy r sau 1-2 ngy nui cy nhit phng. Sc t c to ra nhiu hn trong mi trng c hin din lactose hay cc ngun hidrocacbon khc m vi sinh vt ny c th b gy v s dng (C H Collin v ctv, 1995).

II. Ng c thc phm do Staphylococcus aureus 2.1. Nhng triu chng thng gp
Staphylococcus aureus c xem l mt trong ba tc nhn chnh ca cc v ng c thc phm nhiu nc ch sau Salmonella v Clostridium perfringens (J.P.Rosec v ctv, 1996; J.M. Fueyo v ctv, 2000; Viktoria Atanassova v ctv, 2001; P J Bremer v ctv, 2004; G.Normanno v ctv, 2005). Triu chng thng gp cc v ng c do t cu l bun nn, nn ma, au bng, c hay khng c tiu chy, ngoi ra cn c th b au u, chut rt, thay i huyt p. Triu chng ng c xy ra nhanh, t 3-6 gi sau khi n thc phm b nhim, ty vo lng thc phm dng, lng c t c trong thc phm v nhy vi c t cng nh sc khe ca tng ngi (P J Bremer v ctv, 2004). Thng th cc triu chng ch ko di trong mt thi gian ngn, khong 6-8 gi (Trn Linh Thc, 2002) v ht bnh sau 1-2 ngy (P J Bremer v ctv, 2004). Tuy nhin khong 10% trng hp ngi bnh b mt nhiu nc cn phi nhp vin truyn dch (G.Normanno v ctv, 2004).

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 17

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi Nhng thc phm b nhim t cu v gy ng c thng gp l tht, c, g v sn phm t chng, rau ci, trng, nm, sa v sn phm t sa, kem, phomai, thc phm ln men(G. Normanno v ctv, 2004). Hu ht cc v ng c do t cu l do qu trnh ch bin hoc bo qun thc phm khng tt. T cu thng nhim trc tip vo thc phm do tay ngi ch bin b try xc hay do ho, ht hi. Vic bo qun thc phm nhit khng ph hp cng rt quan trng, mt khi thc phm nhim t cu, chng s tng nhanh s lng do t cu pht trin c trong khong nhit rt ln, t 7-48oC. iu ng lo ngi c t c to ra trong sut qu trnh pht trin ca t cu nhng li khng gy nh hng n cm quan ca thc phm, do t c ch (Mary K. Sandel v John L. McKillip, 2002).

2.2. Tnh hnh ng c do Staphylococcus aureus

Tuy thi gian gy bnh ngn nhng nhng v ng c thc phm do t cu li hu qu khng nh. Theo nh gi ca t chc Y t th gii, hng nm Vit nam c khong trn 3 triu ca ng c thc phm, gy tn hi trn 200 triu USD. Trong nhng nm gn y s v ng c thc phm nc ta ngy cng gia tng. Nm 2000 c 213 v ng c thc phm vi 4233 ngi mc 59 ngi t vong (Bi Th Hin, T Th Thu v cng s, 2003). Theo s liu t cc Qun l cht lng v sinh an ton thc phm, trong nhng v dch c tng kt t nm 1997 n 2002 th ng c thc phm do tc nhn vi sinh vt chim t l cao t 40-45% trong cc loi gy ng c, trong s c nhiu v c xc nh tc nhn l Staphylococcus aureus (Nguyn Phc v ctv, 2003). T nm 2002 n 2004 theo s liu ca trung tm y t d phng c 77 v ng c thc phm m phn ln nguyn nhn l do thc n b nhim khun, chim 66% (Nguyn L Hng v ctv, 2005). T d liu ca Cc v sinh an ton thc phm th nm 2005 s v ng c thc phm nc ta l 141 v, lm 4291 ngi b ng c, trong c 73 v (51,8%) l do SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 18

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi vi sinh vt gy ra. T thng 1/2006 n thng 3/2006, c 18 v ng c thc phm, trong vi sinh vt gy ra 5 v (27,8%). Qua cc cuc kho st tnh hnh v sinh thc n ng ph v cc thc phm ch bin sn ti cc ch cho thy mc nhim Staphylococcus aureus l rt cao, ph hp vi cc kt qu xt nghim trong cc v ng c ti thnh ph H Ch Minh trong nhng nm qua. ng ch hn c l cc mu bnh m tht ngui l 16/30 mu (53%), cc mu tht quay l 18/20 mu (90%) (Nguyn Phc v ctv, 2003; Nguyn L Hng v ctv, 2005). ng ch hn c l v ng c thc phm ca cn b, sinh vin khoa a cht v Sinh hc trng i hc Khoa hc t nhin TP.HCM vo ngy 27/7/2006 trong chuyn i thc t v n tra ti Nha Trang (Khnh Ha). Ba gi chiu cng ngy, nhiu cn b sinh vin au u, tiu chy, i ma, tt c 105 ngi phi vo bnh vin cp cu. Nguyn nhn c xc nh l do thc n ch bin khng hp v sinh, li lu nn nhim t cu trng vng v sinh c t enterotoxin. Enterotoxin do t cu to ra l loi c t cc mnh gy ng c cp tnh. My nm gn y, nhng v ng thc n do t cu c pht hin v ni n nhiu hn ( Th Ha, 2006). Ng c thc phm khng ch xy ra nc ta m cn xy ra nhiu nc trn th gii k c nhng nc pht trin. Theo WHO/FAO (thng 5/2005), ng c thc phm nh hng rt ln n sc khe con ngi v kinh t, lm 1,5 t lt ngi bnh (Nguyn Vn Hi v L Trung Hi, 2005). M, t cu gy ra khong 14% trong cc vu ng c thc phm; v hng nm, M mt khong 1,5 t la cho nhng v ng c do t cu (G.Normanno v ctv, 2005). Ngoi vn vin ph cn phi mt nhiu thi gian lm vic cng nh sn phm lao ng ca ngi bnh, v c vic phi loi b nhng sn phm b nhim V ng c ln u tin c lin quan n t cu xy ra vo nm 1884 Michigan do phomai. Tip n l Php vo nm 1894 do tht t b b bnh. Kh b b nhim t cu cng tng gy ng c Kalamazoo, Michigan vo nm 1907. Nm 1914, SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 19

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi Philippines, Barbert xc nh rng sa ly t b b vim v gy ra ng c ngi. Nm 1930, Dark li xc nh c v ng c do S. aureus t bnh ging sinh.(Sita R Tatini v Reginald Bennett, 2000). Cch y vi thp nin, cc v ng c do t cu chim 25% cc v ng c thc phm M (G. Normanno v ctv, 2004). T nm 1988 n 1992, S. aureus gy ra 5,1% trong s cc v ng c Chu u (G.Normanno v ctv, 2005). T nm 1988 n 1996, c xy ra nhiu v ng c thc phm do t cu gy ra v vo nm 2000 li xy ra mt v dch lm 297 ngi b ng c cng do tc nhn l t cu (Viktoria Atanassova v ctv, 2001). i Loan, S. aureus chim 30% trong s cc v dch t nm 1986 n nm 1995, Vo thng 6 nm 2000, v ng c thc phm do t cu ti mt trng trung hc Taichung County lm 10 trong s 356 hc sinh c biu hin ng c 2-3 gi sau khi n sng (H.-L. Wei v C.-S. Chiou, 2002) . Ti Brazil, vo thng 2 v thng 5 nm 1999 xy ra hai v dch lm 378 ngi b ng c do dng phomai v sa ti c nhim t cu (L. Simeo do Carmo v ctv, 2002). Ti Php, nm 1997 ngi ta tm thy S. aureus l tc nhn gy ra 569 trong tng s 1142 v ng c thc phm (J.P. Rosec v O. Gigaud, 2002). Nht, t nm 1994 n nm 1998 s trng hp ng c do t cu chim 3,111,9% tng s cc v ng c thc phm do vi khun. Ngy 17/6/1999, 21 trong s 53 cng nhn sau khi n tra ti cn tin cng ty Shizuoka Prefecter th c biu hin bnh, trong c 8 trng hp phi nhp vin (Norinaga Miwa v ctv, 2000). 2.3. Cc yu t c lc ca Staphylococcus aureus Staphylococcus aureus gy ra nhiu bnh nhim trng, to m v gy c ngi. Thng xy ra nhng ch xy xc trn b mt da nh nht, gy ra nhiu bnh nhim nghim trng nh vim phi, vim v, vim tnh mch, vim mng no, nhim trng tiu v nhng bnh nguy him khc nh vim xng ty, vim mng trong tim. S. aureus cng l nguyn nhn ch yu ca vic nhim trng vt m v nhng v nhim SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 20

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi trng do dng c y khoa. S. aureus cn gy ng c thc phm do to c t rut enterotoxin trong thc phm, v gy hi chng shock c t do chng to ra siu khng nguyn trong mu (Kenneth Todar, 2005).

Hnh 2.1. Cc yu t c lc ca Staphylococcus aureus (Ngun: Theo Kenneth Todar, 2005) S. aureus to nhiu yu t c lc (Kenneth Todar, 2005; Mary K.Sandel v John L.McKillip, 2002): Protein b mt: thc y vic bm dnh vo t bo ch. Ngoi ra, hu ht cc dng u to protein gn kt fibronogen v fibronetin lm kch thch s kt dnh cc khi mu v m b chn thng. Cc protein gn kt cht to keo cng thng gp nhng dng gy bnh vim xng ty v vim khp. Yu t xm ln (hemolysins, leukocidin, kinase, hyaluronidase): gip vi khun lan ra trn m, phn hy mng t bo eukaryote.

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 21

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi + Hemolysin: - toxin ( hemolysin): y l c t kh mng mnh nht ca S. aureus. N dng mt monomer gn kt vi mng t bo mn cm. ngi, tiu cu v bch cu c bit nhy vi toxin do chng c th th chuyn bit nhn din v cho php c t gn kt hnh thnh l nh m cation ha tr mt c th qua c. - toxin: y l mt mch enzyme phn hy mng giu lipid. Th nghim i vi toxin l phn ng phn hy hng cu cu. - toxin: l mt c t c peptide nh. toxin c th phn hy mt s dng t bo khc nhau. + Leukocidin: l protein a thnh phn, do nhiu thnh phn ring r hp li phn hy mng. Leukocidin cng phn hy mu nhng yu hn hemolysin. Ch 2% trong tt c cc dng S. aureus c th to leukocidin, nhng n gn 90% cc dng phn lp t vt xc trn da c to c t ny. + Hyaluronidase: lm gim cht gian bo ca t bo ch v c th gip t cu lan rng sang cc vng xung quanh. + Catalase: c chc nng bt hot hydrogen peroxide v cc gc t do hnh thnh do h thng myeloperoxidase trong t bo ch. + Coagulase v yu t gy ng: coagulase l mt enzyme ngoi bo s gn vi prothrombin trong t bo ch hnh thnh phc hp staphylothrombin. Coagulase l mt ch th thng dng pht hin S. aureus cc phng th nghim. Tuy nhin, a s bng chng cho thy rng y khng phi l yu t gy c, mc d chng c th t bo v khi s thc bo v p ng min dch bng cch gy ng. C mt s nhm ln v mi lin quan gia coagulase v yu t gy ng u l yu t quyt nh s gn kt fibrinogen trn b mt t bo S. aureus. Mt vi nghin cu cho thy tht s ch c mt lng nh coagulase trn b mt t bo vi khun v chng phn ng vi prothrombin lm ng si fibrin. Nhng nhng nghin cu di truyn ch ra rng khng th gii thch r l coagulase v yu t gy ng c tn ti ring bit hay khng. Bi v nhng t SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 22

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi bin thiu coagulase vn duy tr hot tnh yu t gy ng v nhng t bin thiu yu t gy ng vn biu hin hot tnh coagulase bnh thng (Kenneth Todar, 2005). +Staphylokinase: y l yu t phn gii fibrin. Mt phc hp s c hnh thnh gia staphylokinase v plasminogen kch hot hot tnh phn gii protein gip phn hy fibrin. Cng nh coagulase, khng c bng chng cho thy staphylokinase l yu t gy c, mc d vic phn gii fibrin gip cho s lan rng ca t cu. + Cc enzyme ngoi bo khc - TNase: l enzyme khng nhit, c kh nng hidro ha DNA v RNA ca t bo ch. - DNase, protease, lipase: cung cp cht dinh dng cho vi khun, c tc ng gy bnh thp. - FAME (fatty acid modifying enzyme): l enzyme rt quan trng nhng ch b p-xe, l ni chng c th bin i nhng lipid khng khun v ko di s sng ca vi khun.

2.4. Cc yu t chng li s t v ca t bo ch
Capsule polysaccharide: cn gi l microcapsule do ta ch c th nhn thy chng di knh hin vi in t. Trong cc mu bnh phm, S. aureus c th to ra mt lng ln polysaccharide nhng kh nng ny s gim nhanh khi a chng vo nui cy trong phng th nghim. Chc nng gy c ca v capsule khng r lm mc d chng c th ngn chn s thc bo. Protein A: l protein b mt c th gn phn t IgG nh vng Fc. Trong huyt thanh, vi khun s gn cc phn t IgG sai hng lm ph hy s opsonin ha v s thc bo. Cc chng S. aureus t bin thiu protein A cho s thc bo trong ng nghim hiu qu hn, v nhng t bin trn mu nhim s lm gim c tnh.

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 23

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi Leukocidin: S. aureus to c t r nht trn cc bch cu a nhn. S thc bo l mt c ch quan trng chng li s nhim t cu, do leukocidin l mt yu t gy c. Exfoliative exotoxin: gm hai loi ETA v ETB. Chng gy hi chng phng da tr s sinh (Scalded Skin Syndrome) v gy chc l (Bullous Impetigo) c tr em v ngi ln. Cc siu khng nguyn (supperantigen): S. aureus tit ra hai dng c t c hot tnh siu khng nguyn: - TSST-1( Toxic shock syndrom toxin) c tit ra gy hi chng sc nhim c t cu (TSS). - Enterotoxin gy nn ma v tiu chy, thng xy ra cc v ng c thc phm. 75% hi chng sc nhim c t cu l do TSST-1 gy ra v thng gp trn ph n giai on hnh kinh. Ngoi ra, enterotoxin B v C cng gy ra 50% hi chng ny nhng khng lin quan n kinh nguyt. TSS c th xy ra khi nhim t cu nu nh enterotoxin hoc TSST-1 c tit ra c h thng v t bo ch thiu nhng khng th trung ha thch hp. Nhng siu khng nguyn ny s kch hot nhng t bo T khng chuyn bit vn khng nhn c nhng khng nguyn thng thng. Hn 1 trong s 5 t bo T b kch hot, trong khi ch c 1 trong s 10000 b kch hot khi dng khng nguyn thng thng v cytokine c tit ra vi lng ln gy ra hi chng ny (Kenneth Todar, 2005).

2.5. c t rut enterotoxin ca Staphylococcus aureus

2.5.1. Cu trc Cc c t t cu SE thuc h c t gy st, lm biu hin min dch v tng nhanh s lng t bo T khng chuyn bit. Trong s cc siu khng nguyn th ch c SE l c hot tnh gy nn. Hot tnh siu khng nguyn v hot tnh gy nn l hai SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 24

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi chc nng ca hai protein ring bit nhng s lin h gia hai hot tnh ny vn cha c lm r. Tuy nhin trong hu ht trng hp, hai hot tnh ny tn ti song song nhau, nhng t bin lm mt hot tnh siu khng nguyn cng lm mt hot tnh gy nn (Yves Le Loir v ctv, 2003). 2.5.2. Nhng hot tnh ca c t SE Cc c t t cu SE thuc h c t gy st, lm biu hin min dch v tng nhanh s lng t bo T khng chuyn bit. Trong s cc siu khng nguyn th ch c SE l c hot tnh gy nn. Hot tnh siu khng nguyn v hot tnh gy nn l hai chc nng ca hai protein ring bit nhng s lin h gia hai hot tnh ny vn cha c lm r. Tuy nhin trong hu ht trng hp, hai hot tnh ny tn ti song song nhau, nhng t bin lm mt hot tnh siu khng nguyn cng lm mt hot tnh gy nn (Yves Le Loir v ctv, 2003). 2.5.2.1. Hot tnh siu khng nguyn Hot tnh siu khng nguyn l do tc ng trc tip ca SE vi th th khng nguyn t bo T v phc hp ha mng ca t bo nhn din khng nguyn (Yves Le Loir v ctv, 2003). S nhn din ca khng nguyn l bc u tin trong p ng min dch t bo v cng l vn then cht quyt nh mc chuyn bit ca p ng min dch. Mt khng nguyn thng thng nhn din c th th t bo T bng cch hnh thnh nhng peptide gn kt vi phc hp ha mng MHC lp I hoc II. Ch mt vi t bo T c th nhn din c mt khng nguyn chuyn bit trn phc hp ha mng ca t bo nhn din khng nguyn (Yves Le Loir v ctv, 2003). Trong khi , cc c t siu khng nguyn tc ng trc tip ln nhiu t bo T bng cch nhn din cc chui V chuyn bit ca th th khng nguyn t bo T. Cc c t ny c th lin kt cho vi th th khng nguyn t bo T v phc hp tng ng lp 2 ca t bo nhn din khng nguyn. Chnh s lin kt cho ny dn n vic hot ha khng chuyn bit lm tng nhanh lng t bo T v lng interleukin khng SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 25

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi l l nhng yu t c th lin quan n c ch gy c ca SE (hnh 2.5). Do cc SE c th hot ha 10% t bo T ca chut, trong khi nhng khng nguyn thng thng kch hot t hn 1% t bo T (Merline S Bergdoll, 2000).

Hnh 2.2. Hot tnh siu khng nguyn ( Ngun: Theo Kenneth Todar, 2005) 2.5.2.2. Hot tnh gy nn Hot tnh gy nn khng c m t r nh l hot tnh siu khng nguyn. Ch SE c th gy nn ma khi a vo c th kh bng ng ming trong khi cc siu khng nguyn khc th khng gy nn (Yves Le Loir v ctv, 2003). SE tc ng trc tip ln biu m rut v kch thch trung khu gy nn dn n nhng triu chng ca ng c thc phm. Liu gy ng c do t cu c khong 0,1 g, liu ny c th thay i nhng ngi nhy cm. c im chung nht gia cc SE l vng cystine v y c cho l yu t quan trong nht nh hng n hot tnh gy nn (Yves Le Loir v ctv, 2003). Tuy nhin, hai c t SEB v SEC1 c th b chn ngang vng cystein m vn khng trung ha phn ng gy nn (Merlin S Bergdoll, 2000), hay SEI thiu vng cystine nhng c c hai hot tnh khng nguyn v gy nn, d tnh gy nn yu hn cc SE khc (Yves Le Loir v ctv, 2003). SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 26

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi

2.6. C ch gy bnh
Tm hiu cc c ch m vi khun s dng xm nhp v gy bnh c ngha quan trng trong qua trnh phng chng bnh. Bc quan trng u tin trong qu trnh tng tc gia tc nhn gy bnh v vt ch l s bm dnh (adherence) ca chng vo cc b mt ca vt ch. Cc b mt ny bao gm da, nim mc(khoang ming, mi hu, ng tit niu) v cc t chc su hn (t chc lympho, biu m d dy rut, b mt ph nang, t chc ni m). Ging nh cc vi khun gram (+) khc l Streptococcus v cc Mycobacteria , S.aureus bm dnh vo b mt t bo vt ch nh cc adhensin c bn cht polypeptide. Mt khi bm dnh vo b mt t bo vt ch, tc nhn gy bnh c kh nng khi ng cc qu trnh ha sinh c hiu gy bnh nh tng sinh, bi tit c t, xm nhp v hot ha cc chui tn hiu ca t bo vt ch. S.aureus s tip tc tin su vo trong c th vt ch tip tc chu trnh xm nhp. S.aureus xm nhp ngoi bo bng cch tit mt s enzyme nh: hyaluronidase, hemolysine, leukocidin, exfoliatine, ph hy cc thnh phn cu to t bo vt ch. 2.6.1. Bnh v cc triu chng bnh: S.aureus c mt khp ni trong thin nhin nh: khng kh, nc, da, hng, mi,tc hay lng ca cc loi ng vt mu nngv ch gy c khi hnh thnh c t rut (Enterotoxin). Nhng trng hp nhim c u tin do n bnh ko gy ra bi t cuvng c ni n t nhng nm 1901-1914. Vit Nam cng nh trn khp th gii,ng c thc phm xy ra thng xuyn. Kh nng gy c ch xy ra khi n thc n cng c t ca vi khun. Cn nu ch nvi khun th khng gy ra ng c. iu chng t, ng c l do c t ca vi khunc sn xut ra trong mi trng thc n vi s hot ng ca vi khun, c t rut l mt loi c t mnh. 2.6.1.1. Vim phi: S.aureus c th xm nhp vo nhu m phi qua hai ng: ht vo ng h hp trn hoc ly lan qua ng mu. Thng sau khi mc bnh cm hoc ngi SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 27

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi suy gim min dch, t cu vng theo dch tit ng h hp trn b ht vo phi. Tuy y l bnh t gp nhng li l bnh nhim khun nghim trng. Vi khun to v bc d xm nhp vo mu gy nhim khun huyt. Vim phi thng gm nhiu , tm vim l ph qun hoc tiu ph qun vim hoi t, xut huyt. Cc vim ny v ra to ra cc p-xe. nhng ca nng, thnh ph nang thng b ph hy, khng kh vo ph nang b ph hynhng khng thot ra c, to ra cc ti kh thnh mng. Cc yu t gy nguy c mc bnh l iu kin sng km, s dng khng sinh ba bi, bnh nhn nm vin lu ngylm gim sc khng ca c th to iu kin thun li cho vi khun d dng xm nhp vo ng h hp, ng thi lm gim chc nng min dch ca c th. Triu chng: ph bin l st cao, mch nhanh, th nhanh, ho, t gp ho ra mu. C th suy h hp, sc nhim khun, nhim c h thng, kh th, hoi t v hnh thnh p-xe. Hai bin chng hay gp nht l trn dch mng phi v m mng phi. 2.6.1.2. Hi chng shock do c t: - L cn bnh gy ra bi c t TSST-1 c tit ra bi vi khun S.aureus pht trintrong iu kin c t hoc khng c oxy. Bnh thng gp nhng ph n trong k kinh nguyt c dng bng thm ht mnh, cc bng ny c kh nng gn cc magnesium do lm gim lng ion ny trong m o. Khong 20% ph n c mang t cu ng m o. Do lng ion magnesium gim xung, vi khun ny tng cng sn xut cc ngoi c t gy sc. - Hi chng Thukydides l th c bit ca hi chng sc nhim c. Hi chng ny c th gp thanh thiu nin c hai gii, thng gp do bi nhim t cu sau khi b cm.T l t vong ca hi chng ny kh cao trn 50%. Biu hin lm sng v mt h hp v tiu ha rt ging vi bnh dch hch do Thurykodides m t Athen v nm 430 trc cng nguyn. SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 28

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi Triu chng: t ngt st cao, cm gic mt mi, tiu chy ton nc, nhc u, au c, ni bang ngoi da, c mt vi du hiu ca sc, nn ma, h huyt p. C th c pht ban nh b chy nng, lt da, vt c hiu ngoi tng m hay lan ton thn nhanh, cth c biu hin xung huyt, kt mc, hng , ph ngoi bin. Thay i cc ch s cn lm sng nh: ur mu tng, albumin mu gim, calcim mu h, h phosphate mu, tng creatin phosphakinase, gim tiu cu. Gy ra cc ri lon chc nng thn v c tim, qu ti dch, suy h hp ngi ln. Sau khong 1 tun bnh nhn b trc da vng thn mnh v t chi. Di chng mun gm hoi th ngoi bin, mt mng c th phc hi yu c suy nhc, ko di v ri lon tm thn kinh. Nu bnh nng c th t vong t l kh cao. 2.6.1.3. Nhim trng da v m t bo, p-xe, vim no, vim ty xng: - S.aureus t lu c cng nhn l mt trong nhng vi khun quan trng nht gy bnh ngi. N l nguyn nhn hng u ca nhim trng da v m mm, n cng c th gy nhim trng nghim trng nh nhim trng xng khp, vim thn kinh Nhim trng da v m t bo, p-xe: nhng bnh nhim trng da v cc phn ph ch yu l cc chn lng v tuyn m hi to thnh bnh cnh p-xe kinh in ca t cu vng. N to fibrin bao bc p xe. Cc nhim trng ny c th ch nh nh u inhnu vim nang lng v kch thc nh qu to nh p-xe c. M ca p-xe do t cu vng thng c mu vng, c v khng hi. Triu chng: trn da hnh thnh nhng khu vc thng l mu , au v xng ln, cha y m, xung quanh khu vc b p xe c th cm thy m khi chm vo. + i vi m t bo th s nhim trng l cc lp c bn di da, cng b sng v au, ni mn nht v ly nhim. + Vi khun c th xm nhp vo t nhng vt ct, vt co hoc nhng thng tch nh, c th xy ra bt c ni no trn c th nhng thng xy ra trn hai tay v hai chn. SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 29

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi - Vim no, vim ty xng: + Nhim trng cc c quan bn trong c th c th do ng ni sinh: t mt nhim ngoi vi, vi khun theo ng mu v bch huyt n cc c quan khc. Chng ivo c th qua cc vt rch trn da sau chn thng hoc trong qu trnh phu thut. Triu chng: no sau vim ni tm mc, chng mt, vim mng no m, trn m di mng cng, vim tc tnh mch, vim xoang, vim xoang chm, yu tay, chn, thay i th gic. Gy st t vim mn, vim khp nhim trng, thng gp khp gi, hng v khp cng chu, gy vim mi c. 2.7. Cc bin php phng v x l bnh 2.7.1. Cc bin php phng bnh - nhng vng c iu kin v sinh c nhn km, ni ng dn c, c bit l thiu nc sch trong sinh hot, thi tit nng m l iu kin thun li cho bnh pht sinh mnh. - Phng bnh t cu khun cng nh cc bnh nhim khun khc th yu t v sinh c nhn l iu quan trng, ra tay sch trc khi n v sau khi i v sinh l yu cu c bn phng bnh, n ung hp v sinh, nu k trc khi dng, gi gn sch s nhng vt try xc trn da. Nhng bnh nhn tai bin mch mu no nm lit ging c th dn n xut hin nhng vt lot cn phi chm sc h tr vn ng nh xoa bp mch mu lu thng, cn i vi bnh nhn i tho ng cn gi gn bn chn trch b phng lot. - Cc bin php nh mang m hoc khn trm u, mang khu t r a n g v r a t a y thng xuyn l bin php phng chng ly truyn hu hiu trong bnh vin. - Nhng ngi lm vic trong cc bp n tp th hoc trong cc xng ch bin thc phm cng cn thc hin cc bin php phng h trn. SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 30

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi - i vi thc phm th nn bo qun cn thn, khng ln n sng ln vi n chn. Nu nghi ng thc phm b nhim khun th nn loi b. - Khm sc khe nh k cho cng nhn lm vic trc tip vi thc phm, ch khmtai, mi, hng. - Khng nhng ngi b vim xoang, vim mi hng, c mn m tay tham gia ch bin thc phm. - Dng kp khi phc v thc n. - Tng cng ung vitamin, nng cao sc khe, ra tay bng nc nng v x phng,h pH thc phm c ch vi khun pht trin. 2.7.2. X l bnh - i vi nhng tn thng khu tr trn da th khng cn phi u tr khng sinh tr khi nhim khun lan rng hoc c bin chng. Ti ch mng m cn lm sch da ti ch bng dung dch st khun, dng khng sinh trc tip bi ngoi da. Cc loi p-xe cn rch m v kt hp dng khng sinh ng ung hoc tim. Nhng loi nhim khun xng,no th cn dng cc loi khng sinh c hiu v khi iu tr th phi theo di cht ch nht l i vi nhng ngi cao tui ngi b x va ng mch hay y tho ng. - Cn s dng nhng loi khng sinh ph hp cho tng bnh nhn Thng dng gamma globuline iu tr bnh. Mc d S.aureus c th sn xut c men penicillinase ph hy vng betalactamase lm bt hot cc loi khng sinh nh peni cilline G, Ampicilline... Nhng hin nay trong iu tr nhim trng S.aureus ngi ta vn s dng peniclline v cephalosporine khng vi beta-lactamase . - Nafcilline v Oxacilline l hai loi peniclline c dng bng ng tim. Nu nhim trng nng th nn s dng cephalosporine th h th nht nh cephazoline.

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 31

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi - Nu bnh nhn khng dung np vi khng sinh nhm beta-lactamase th nn thay bng vancomycine v clidamycine bng ng tim. Dicloxaciline v cephalexine l khng sinh dng ung. III. PHNG PHP PHT HIN STAPHYLOCOCCUS AUREUS 3.1. Phng php truyn thng 3.1.1. Phng php nh tnh Cho dung dch mu vo mi trng TSB, 37 oC trong 24 gi, sau cy ria ln mi trng thch BP (Baird Paker), 37 OC trong 48 gi. Tm khun lc c trng ca S. aureus trn mi trng BP c mu en nhnh, sng, trn, li, ng knh 1-1,5 mm, c vng sng chung quanh khun lc cy sang mi trng BHI (Brain Heart Infusion), 37 OC trong 24 gi. Sau cy sang ng nghim nh c cha 0,3 ml huyt tng th phn ng ng kt (phn ng coagulase), thc hin song song mt ng i chng khng c cy dch vi sinh vt, theo di kt qu ng huyt tng sau cc khong thi gian 2, 4, 6, 8 v 24 gi. Mu c kt lun l c S. aureus khi th nghim coagulase dng tnh. 3.1.2. Phng php nh lng 3.1.2.1. Phng php m khun lc (phng php hp tri) p dng theo AOAC 2007 (975.55) Nguyn tc: + Hnh thi khun lc in hnh trn mi trng Baird-Parkersau khi 35oC/48h. + Th nghim Coagulase: dng tnh Chun b mu -Mu thc phm c ct nh hoc xay nhuyn bng my trong iu kin v trngcho ti khi c th ng . -Nu mu th ng phi lm tan bng trc khi lm xt nghim. SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 32

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi Chun b dung dch mu th 10-1 Cn chnh xc 45g mu thc phm c chun b hoc ht chnh xc 45 ml mu thc phm lng cho vo ti nilon v trng (hay cn 10g mu thc phm/10ml thc phm lng).Thm 450 ml nc m phosphate (hay 90ml nc m phosphate) v ng nht mu trong vng 2 pht,thu c dung dch mu th 10-1 Chun b dung dch mu th 10-2, 10-3, 10-4 -Ht chnh xc 10 ml (hoc 1ml)dung dch mu th 10-1 cho sang chai dural cha sn 90 ml nc m phosphate (hoc ng nghim c chha sn 9 ml nc m phosphate), lc u,thu c dung dch 10-2. - Tip tc lm tng t nh vy, ta thu c cc dung dch mu th 10-2, 10-3 Thc hin: Bc 1: cy mu - T nng pha long ( 10-1, 10-2, 10-3,), ly 1ml dung dch mu th cho sang 3 a thch Baird-Parker (v d: th tch mu th cho vo 3 alln lt l: 0,4, 0,3, 0,3 ml) + Ring vi mu thc phm lng: c th thc hin nh sau: ht trc tip 1ml dung dch mu th nguyn thy hoc 1 ml t cc nng pha long 10 -1, 10-2, 10-3,cho sang 3 a thch Baird-Parker (v d: th tch mu th cho vo 3 a ln lt l: 0,4, 0,3, 0,3 ml) - Dng que cy v trng, dn mu th cn thn ln mt a thch, c gng khng chm vo mp a. cc a c np y cho n kh trong khong 10 pht nhit phng. Nu cc a ny cha kh th c th t cc a (khng lt ngc a) vo t 1h - Lt ngc cc a, trong t m 35 0C trong 24-48 h. Bc 2: m v nhn nh khun lc in hnh ca S.aureus trn mi trng thch Baird-Parker SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 33

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi - m v nh du v tr cc khun lc in hnh ca S.aureus trn mi trng thch Baird-Parker : Khun lc trn, nhn, li, t, ng knh 2-3 mm mu xm n en nhnh; v thng c bao quanh bi mt vng sang hoc vng sang c. - Chn cc a thch Baird-Parker c s khun lc in hnh t 20 200 ( lu : vn chp nhn chn cc a thch ca nng pha long thp nht c sd khun lc < 20). - Nu trn mi a thch Baird-Parker c xut hin mt s dng khun lc nghi ng l S.aureus th m ring r s khun lc ca mi dng. - Nu trn cc a c xut hin > 200 khun lc in hnh ca S.aureus v cc khun lc in hnh khng c nhng nng pha long cao nht th s dng cc a ny nh lng S.aureus m khng cn m cc khun lc khng in hnh. Bc 3: Th nghim coagulase - Chn 5 khun lc ca mi dng khun lc chn m bc 2 v tin hnh th nghim coagulase nh sau: - Dng u que cy thng v trng, cy chuyn mi khun lc nghi ng chn vo mi ng nghim c cha sn 0,2 ml mi trng canh thang BHI v ng thi cy ria ln mi ng thch nghing ca mi trng bo qun chng nh TSA hoc Nutrient agar m 35oC/ 18-24h. + Thm 0,5 ml huyt tng th hon nguyn vo mi ng dch nui cy BHI, lc u. m 35o v kim tra s ng t ca huyt tng sau mi gi, 6 gi. S ng t huyt tng bt c mc no cng c xem l phn ng dng tnh. Nhng cc ng t huyt tng nh hoc yu c th quan st bng cch nghing nh ng nghim cho phn dung dch ca hn hp phn ng n gn pha ming ng nghim, cnnhng cc ng t huyt tng s nh ra trn b mt dung dch. + Kim tra li cc kt qu th nghim coagulase ca dch nui cy BHI trong khong thi gian > 18h nhng < 48h 350C. SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 34

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi + Cn thc hin ng ti cc th nghim chng m v chng dng cho th nghim coagulase. + Cc ng nui cy trn thch nghing c gi li nhit phng s dng trong trng hp th nghim coagulase c vn nghi ng cn th nghim li. - Nhng ng dch nui cy BHI c th nghim coagulase dng tnh c coi l S.aureus. Bc 4: Tnh ton v bo co kt qu + Tnh ton s khun lc A trn mi a thch Baird-Parker cho keets qu th nghim coagulase dng tnh theo cng thc sau: A= BCCC / Ac +BncCnc /Anc Trong : Ac l s khun lc in hnh em th phn ng coagulase. BC l s khun lc in hnh em th phn ng coagulase dng tnh CC l tng s khun lc in hnh thy trn a Anc s khun lc khng in hnh em th phn ng coagulase. Bnc s khun lc khng in hnh em th phn ng coagulase dng tnh Cnc l tng s khun lc khng in hnh thy trn a Ly kt qu trn s + Tnh s lng vi khun S.aureus (N) c mt trong 1g hoc 1 ml mu th theo cng thc N= Trong : trn 3 a chn SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 35 l tng s khun lc cho kt qu th nghim coagulase dng tnh

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi d: pha long ca 3 a chn Lm trn kt qu tnh c v biu th kt qu theo biu thc: nx10x CFU/g hoc ml thc phm. Trong : n s thp phn tng ng t 1,0 n 9,9. x: s m ph hp ca 10. + Nu c 3 a thch ca nng pha long u tin khng cha bt k khun lc no cho kt qu th nghim coagulase dng tnh th bo co kt qu nh sau: < 1 CFU/ml (thc phm dng lng) < 10 CFU/g (thc phm dng c, st)

3.1.2.2 . K thut MPN p dng theo AOAC 2007 (987.09) Nguyn tc : - Tiu chun xc nh Staphylococcus aureus l da vo cc c im sau: + S.aureus mc c trn mi trng canh thang TSB c b sung thm 10% NaCl v 1% sodium pyruvate. + Hnh thi khun lc in hnh trn mi trng thch Baird-Parker sau khi 350C/48h + Th nghim Coagulase: dng tnh Chun b mu - Mu thc phm c ct nh hoc xay nhuyn bng my trong iu kin v trng cho ti khi c th ng nht. - Nu mu th ng phi lm tan bng trc khi lm xt nghim. Chun b dung dch mu th 10-1 SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 36

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi Cn chnh xc 45g mu thc phm c chun b hoc ht chnh xc 45 ml mu thc phm lng cho vo ti nilon v trng (hay cn 10g mu thc phm/10ml thc phm lng).Thm 450 ml nc m phosphate (hay 90ml nc m phosphate) v ng nht mu trong vng 2 pht,thu c dung dch mu th 10-1 Chun b dung dch mu th 10-2, 10-3, 10-4 Ht chnh xc 10 ml (hoc 1ml)dung dch mu th 10-1 cho sang chai dural cha sn 90 ml nc m phosphate (hoc ng nghim c chha sn 9 ml nc m phosphate), lc u,thu c dung dch 10-2. - Tip tc lm tng t nh vy, ta thu c cc dung dch mu th 10-2, 10-3 Thc hin: Bc 1: Cy mu - Ly 1ml dung dch mu th 10-1, 10-2, 10-3cho sang ng nghim cha sn 10ml canh thang TSB c 10% NaCl v 1% Sodium pyruvate. Mi m cy 3 ng. + Ring vi mu thc phm lng c th thc hin xt nghim nh sau: ht trc tip 1ml dung dch mu th nguyn thu (100) v nng pha long 10-1, 10-2, 10-3 cho sang ng nghim cha sn 10ml canh thang TSB c 10% NaCl v 1% Sodium pyruvate. Mi m cy 3 ng. - m 350C/48h. Bc 2: Cy chuyn - Staphylococus aureus pht trin trong mi trng canh thang TSB vi 10% NaCl v 1% Sodium pyruvate lm c mi trng. - Dng que cy vng cy ria mt vng dch nui cy ln b mt a thch BairdParker sao cho thu c cc khun lc tch bit r rng, mi ng dch nui cy cy ria ln mt a thch. cc a thch ny 350C/48h

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 37

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi Bc 3: Quan st v nhn nh khun lc in hnh ca Staphylococus aureus trn mi trng thch Baird-Parker - Trn mi trng thch Baird-Parker: Khun lc trn, nhn, li, t, ng knh 2-3mm mu xm n en nhnh; v thng c bao quanh bi mt vng sang hoc vng trng c. - Thnh thong cc chng khng phn gii lipit c th c hnh dng khun lc tng t nhng chng c vng trng c v vng sng xung quanh khun lc. - Nhng khun lc c phn lp t m ng hoc sy kh m c bo qun trong thi gian di th khun lc thng t en hn v khun lc c th c dng nhm v kh. Bc 4: th nghim khng nh - T mi a thc Baird-Parker, chn ly 1 khun lc nghi ng l S.aureus thc hin th nghim Coaglulase. - Dng u que cy thng v trng, cy chuyn mi khun lc nghi ng chn vo mi ng nghim c cha sn 0,2 ml mi trng canh thang BHI v ng thi cy ria ln mi ng thch nghing ca mi trng bo qun chng nh TSA hoc Nutrient agar m 35oC/ 18-24h. + Thm 0,5 ml huyt tng th hon nguyn vo mi ng dch nui cy BHI, lc u. m 35o v kim tra s ng t ca huyt tng sau mi gi, 6 gi. S ng t huyt tng bt c mc no cng c xem l phn ng dng tnh. Nhng cc ng t huyt tng nh hoc yu c th quan st bng cch nghing nh ng nghim cho phn dung dch ca hn hp phn ng n gn pha ming ng nghim, cnnhng cc ng t huyt tng s nh ra trn b mt dung dch. + Kim tra li cc kt qu th nghim coagulase ca dch nui cy BHI trong khong thi gian > 18h nhng < 48h 350C.

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 38

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi + Cn thc hin ng ti cc th nghim chng m v chng dng cho th nghim coagulase. + Cc ng nui cy trn thch nghing c gi li nhit phng s dng trong trng hp th nghim coagulase c vn nghi ng cn th nghim li. - Nhng ng dch nui cy BHI c th nghim coagulase dng tnh c coi l S.aureus. Bc 5. Tnh kt qu -i vi mi nng pha long, ghi li s ng dng tnh (+) da vo kt qu th nghim coagulase. - Tra bng MPN nng tng ng, tnh s MPN ca S.aureus trong 1 g thc phm hoc 1 ml thc phm. V d tnh Nng 10-2: c 1 ng (+): 1 ng cho th nghim coagulase dng tnh Nng 10-3: c 0 ng (+) Tra bng MPN, xc nh trong 1 g mu th c 43 MPN. 3.2. Phng php hin i Phng php truyn thng tuy c s dng t rt lu, c nhiu quc gia cng nhn nhng cng c nhng nhc im nht nh nh mt nhiu thi gian, cng sc, tn km, thng phi mt khong 4-5 ngy mi xc nh c mu c nhim S. aureus khng. khc phc nhng nhc im ny, gn y vic ng dng k thut sinh hc phn t hin i pht hin v phn bit cc tc nhn gy bnh ngy cng tr nn quan trng hn trong v sinh thc phm. 3.2.1 Phng php RPLA (Reversed Passive Latex Aggulutination) Nng 10-1: c 3 ng (+): 3 ng cho th nghim coagulase dng

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 39

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi K thut RPLA c ng dng pht hin c t trong thc phm cng nh pht hin nhng dng Staphylococcus c c t dng tnh. Trong k thut ny, ht nha c ph khng th ca c t trc khi cho vo ging. Cho mu vo to phn ng. Nu c c t trong mu th phn ng gia c t v khng th s to ra s ngng kt, ty mc ngng kt m xc nh c lng c t. Phng php ny nhy pht hin c t trong hu ht thc phm ca cc v ng c, cng nh trong vic pht hin nhng dng Staphylococcus to ra lng c t thp m phng php gel -diffusion khng pht hin c, ch vi khong 1020 ng/ ml. B kit RPLA c gii thiu bi cng ty Denka Seiken, Niigata, Nht Bn v c bn ti M (Merlin S Bergdoll, 2000). 3.2.2. Phng php PCR (polymerase chain reaction) Gn y phng php PCR c s dng ph bin pht hin c t Staphylococcus da vo vic tng hp mi chuyn bit cho on gen m ha c t. Cc phn ng PCR thng dng l uniplex v multiplex PCR, v gn y l real-time PCR (Beatriz Pinto v ctv, 2005). Phng php ny cng c ng dng pht hin cc gen to c t mi nh gen m ha cho c t SEU trn vng egc ca S. aureus (C. Letertre, 2003). Cc phng php da trn PCR nhy v chuyn bit, cho php pht hin Staphylococcus to c t trong thi gian tng i ngn vi lng mu nh. u im ca phng php ny l nhng t bo sinh c t cht vn c th pht hin, iu ny rt quan trng trong vic phn tch nhng thc phm x l nhit lin quan n nhng v ng c thc phm (Merlin S Bergdoll, 2000). Tuy phng php PCR c nhiu u im l nhanh, nhy, chuyn bit nhng ch c th xc nh c s hin din ca cc gen m ha c t m khng th xc nh c c to c t hay khng cng nh vic nh lng c t (J.A. Boerema, 2005). 3.2.3. Phng php hp ph min dch dng enzyme - ELISA (Enzyme Linked Immunosorbent Assay) SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 40

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi Phng php ELISA c ng dng pht hin c t S. aureus trong thc phm kh ph bin (Nighat P.Kokan v Merline S Bergdoll, 1987; J.P.Rosec v ctv, 1997; Susana M. Portopcarrero v ctv, 2002; Hiroshi Fujikawa v Satoshi Morozumi, 2005). y l phng php n gin, nhy, nhanh, c th pht hin cng nh nh lng c t v c nhiu b kit pht hin c t ang c bn trn th trng. Kiu ELISA thng gp nht l phng php sandwich. Trong phng php ny, khng th c gn vi mu cha bit, sau phc hp khng th - c t c gn vi cng hp enzyme khng th. Qui trnh ny c s dng nhiu v lng enzyme v mu c to ra t phn ng enzyme c cht t l thun vi lng c t c trong mu cha bit. Hai enzyme alkaline photphatase v horseradish peroxidase u c dng trong phng php ny, nhng alkaline photphatase d gn vi khng th hn. Tuy nhin horseradish peroxidase cho phn ng nhy hn, khi kt hp vi c cht cho mu xanh l, xanh dng hay cam, cn mu c to ra vi c cht khi s dng vi alkaline photphatase, t/ml (Merlin S Bergdoll, 2000). Nhiu nh khoa hc s dng phng php ELISA pht hin c t S.aureus trong phng th nghim vi nhiu qui trnh khc nhau. Phng php ELISA ch yu s dng a ging (microtitre plate hay strip) khng th gn vo. Mu c cho vo ging, nu nh trong mu c khng nguyn mc tiu, khng nguyn s gn vi khng th. Sau , ly mu ra v ra a. Cng hp enzyme khng th (khng th c gn vi enzyme horseradish peroxidase hoc alkaline photphatase) c thm vo v cho php phn ng vi phc hp c t - khng th. Ra a, thm c cht c hiu ca enzyme vo. Enzyme xc tc phn ng thy phn c cht to ra sn phm c mu. i chng dng v i chng m cng c a vo th nghim. Phn ng dng tnh c ghi nhn nu nh mu c c m hn mu i chng m. Bng cch theo di s i mu, c th pht hin s hin din v nh lng lng khng nguyn (c t) (Merlin S Bergdoll, 2000; Trn Linh Thc, 2002). SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 41 -nitrophenyl photphatase (pNPP) l mu vng. Hu ht cc phng php ELISA nhy t nht l 0,5 ng c

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi

PHN III. VT LIU V PHNG PHP TIN HNH

3.1. Thi gian v a im thc hin
3.1.1. Thi gian thc hin Thi gian thc hin ti t 12/03/2005 n 08/04/2012. 3.1.2. a im thc hin Ni ly mu: Mu c ly ti cc ch ng Xoi Ni tin hnh phn tch th nghim: Phng vi sinh nc thc phm, khoa Xt Nghim, Trung Tm Y T d phng tnh Bnh Phc.

3.2. Vt liu
3.2.1. Dng c v thit b 3.2.1.1. Dng c Bnh tam gic, Cc 50, 100, 500 ml

ng ong hnh tr 50, 100, 500 ml Pipetman, u cn SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 42

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi ng nghim Que cy vng n cn Ko ct Giy th Ph bng gn. 3.2.1.2. Thit b Cn phn tch knh hin vi T sy T m 37C My m khun lc T lnh. 3.2.2. Mi trng v thuc th - Nc em phosphate (Merck-c) - Canh thang BHI (Merck-c) - Moi trng thach Baird-Parker (Merck-c) - Moi trng thach TSA (Merck-c) - Huyet tng th (Merck-c) - Nc muoi sinh ly (Merck-c) - Dung dch egg yolk (Merck-c) 3.2.3. Vt liu th nghim my o pH Ni hp autoclave T cy an ton sinh hc cp II L viba My ra dng c bng sng siu m, a petri Que cy thng Ti nylon v trng Pince kim loi gng tay y t

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 43

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi Mu thc phm s dng trong th nghim bao gm cc loi thc phm n nhanh c thu t ch ng Xoi. Cc mu c thu i din cho 2 nhm thc phm: ch la v ch c chin n ngay. Trnh by trong bng 3.1. Bng 3.1 Cc nhm thc phm c s dng trong th nghim STT 1 2 Nhm thc phm Ch la Ch c chin n ngay S lng 9 9 Khi lng/mu 300 g 300 g

3.3. Phng php

B tr th nghim Mu ch la, ch c chin n ngay

Thu mu ti cc a im kho st

nh gi cc ch tiu

Staphylococcus (lp li 3 ln) Cm quan aureus

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Nhn xt, Kt lun Vn 44

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi

3.3.1. Phng php ly mu v bo qun mu Mu c thu ti ch v c cha trong bao nylon v trng, phi bo qun mu bng nc cho ti khi vn chuyn n phng th nghim phn tch. Nu cha phn tch, mu c bo qun trong t lnh 2 - 8C trong 24 - 36 gi. 3.3.2. Phng php bo qun chng vi sinh vt Cc S. aureus phn lp c t thc phm c bo qun trong cc ng nghim thch nghing TSA dng trong cc bc th nghim sinh ha v phn ng ng huyt tng. T cc ng thch nghing TSA giai on phc hi, ly mt t sinh khi bng que cy vng cy chuyn sang cc ng nghim thch nghing TSA mi nhn ging v bo qun ging, 37C qua m. Bo qun cc ng vi khun ging ny trong t lnh 2 8C n khi s dng tip vo cc th nghim. Thi gian bo qun t 1 2 thng. Ht hn bo qun, cc chng c hot ha v cy chuyn nh trn. Mi ng chng phi c ghi cc thng tin nh: tn, k hiu mu, k hiu chng, ngy, s ln cy chuyn. 3.3.3. nh lng S. aureus bng phng php m khun lc p dng theo AOAC 2007 (975.55) Nguyn tc: + Hnh thi khun lc in hnh trn mi trng Baird-Parker sau khi 35oC/48h. + Th nghim Coagulase: dng tnh Chun b mu -Mu thc phm c ct nh hoc xay nhuyn bng my trong iu kin v trng cho ti khi c th ng . SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 45

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi -Nu mu th ng phi lm tan bng trc khi lm xt nghim. Chun b dung dch mu th 10-1 Cn chnh xc 10g mu thc phm c chun b cho vo ti nilon v trng Thm 90ml nc m phosphate v ng nht mu trong vng 2 pht,thu c dung dch mu th 10-1. Chun b dung dch mu th 10-2 , 10-3 -Ht chnh xc 1ml dung dch mu th 10-1 cho sang ng nghim cha sn 9 ml nc m phosphate, lc u,thu c dung dch 10-2. - Tip tc lm tng t nh vy, ta thu c cc dung dch mu th 10-3 Thc hin: Bc 1: cy mu - T nng pha long 10-1, 10-2, 10-3, ly 1ml dung dch mu th cho sang 3 a thch Baird-Parker (th tch mu th cho vo 3 al ln lt l: 0,4, 0,3, 0,3 ml) - Dng que trang v trng, dn mu th cn thn ln mt a thch, c gng khng chm vo mp a. cc a c np y cho n kh trong khong 10 pht nhit phng. Nu cc a ny cha kh th c th t cc a (khng lt ngc a) vo t 1h. - Lt ngc cc a, trong t m 35 0C trong 24-48 h. Bc 2: c kt qu Sau 24 gi trn mi trng Baird Paker agar, khun lc S. aureus c ng knh 0,5 1 mm, li, en bng c vng sng rng khang 1 2 mm bao quanh. nh du trn mt y ca a cc khun lc c c im nh trn v tip tc n 48 gi. Sau 48 gi khun lc S. aureus c ng knh khong 1 1,5 mm, mu en bng, li, c vng trng c hp v c vng sng rng khong 2 4 mm bao quanh. C mt

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 46

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi s dng S. aureus cho khun lc khng c trng. Cn m v nh du hai dng khun lc.

Hnh 3.1. Khun lc S. aureus trn mi tr ng Baird Paker Bc 3: Khng nh - Chn 5 khun lc ca mi dng khun lc chn m bc 2 v tin hnh th nghim coagulase nh sau: - Dng u que cy thng v trng, cy chuyn mi khun lc nghi ng chn vo mi ng nghim c cha sn 0,2 ml mi trng canh thang BHI v ng thi cy ria ln mi ng thch nghing ca mi trng bo qun chng nh TSA. m 35oC/ 1824h. + Thm 0,3 ml huyt tng th hon nguyn vo mi ng dch nui cy BHI, lc u. m 35o v kim tra s ng t ca huyt tng sau , 2, 4, 6, 8 v 24 gi. S ng t huyt tng bt c mc no cng c xem l phn ng dng tnh. Nhng cc ng t huyt tng nh hoc yu c th quan st bng cch nghing nh ng nghim cho phn dung dch ca hn hp phn ng n gn pha ming ng nghim, cnnhng cc ng t huyt tng s nh ra trn b mt dung dch.

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 47

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi + Cc ng nui cy trn thch nghing c gi li nhit phng s dng trong trng hp th nghim coagulase c vn nghi ng cn th nghim li. - Nhng ng dch nui cy BHI c th nghim coagulase dng tnh c coi l S.aureus.

nh 3.2. Kt qu phn ng coagulase Bc 4: Tnh ton v bo co kt qu + Tnh ton s khun lc A trn mi a thch Baird-Parker cho kt qu th nghim coagulase dng tnh theo cng thc sau: A= BCCC / Ac +BncCnc /Anc Trong : Ac l s khun lc in hnh em th phn ng coagulase. BC l s khun lc in hnh em th phn ng coagulase dng tnh CC l tng s khun lc in hnh thy trn a Anc s khun lc khng in hnh em th phn ng coagulase. Bnc s khun lc khng in hnh em th phn ng coagulase dng tnh Cnc l tng s khun lc khng in hnh thy trn a Ly kt qu trn s SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 48

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi + Tnh s lng vi khun S.aureus (N) c mt trong 1g mu th theo cng thc N= Trong : trn 3 a chn. d: pha long ca 3 a chn Lm trn kt qu tnh c v biu th kt qu theo biu thc: nx10x CFU/g hoc ml thc phm. Trong : n s thp phn tng ng t 1,0 n 9,9. x: s m ph hp ca 10. + Nu c 3 a thch ca nng pha long u tin khng cha bt k khun lc no cho kt qu th nghim coagulase dng tnh th bo co kt qu nh sau: ng nht mu v pha < 10 CFU/g (thc phm dng c, st) long mu thnh cc dy nng thp phn 10-1, 10-2, 10-3 l tng s khun lc cho kt qu th nghim coagulase dng tnh

Quy trnh nh lng S. aureus Tri ln a thch BP 35oC/24-48h.

m s khun lc in hnh

m s khun lc khng in hnh Ly 5 khun lc khng in hnh cy vo TSA . 35oC/24h

Ly 5 khun lc in hnh cy vo TSA . 35oC/24h

Th nghim ngng kt coagulase

T l khun lc in hnh c T l khun lc khng in hnh coagulase Anh; Nguyn Th Dim coagulase (+) inh Th c Hng; SVTH: Nguyn N Lan (+) Vn 49 Kt qu

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi

3.3.4. Phng php x l s liu S liu c x l bng phn mm Excel.

PHN IV. KT QU V THO LUN

4.1. Kt qu cm quan

Hnh 4.1. Mu c mua ch Ch la V mu sc Mu trng c trng V mi Thm c trng ca tht, V tnh cht B mt ca ch mn,

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 50

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi ca tht Ch c chin mu Ch c vng u, khng b chy. khng c mi l Mi thm c trng ca tht c c chin chin do, dai c trng Kt cu do, dai

V cm quan v mu nhn chung l tt, tnh cht, mu sc v mi v ca mu khng c g l. Nhng nh gi cm quan khng xem xt mc sch ca mu.

4.2. Kho st t l nhim S. aureus trong cc nhm thc phm ang lu hnh trn th trng
Theo quyt nh s 46/2007/Q-BYT ngy 19 thng 12 nm ca B trng B Y t. Quy nh gi hn ti a nhim trong thc phm s dng trc tip ( qua x l nhit): Sn phm Ch la Ch c chin n ngay Loi vi sinh vt S. aureus S. aureus Gii hn vi sinh vt cho php ( trong 1g/1ml thc phm) 102 10

Mc ch ca kho st ny l nhm nh gi t l nhim nhim S.aureus trong cc loi thc phm ang lu hnh trn cc ch trn khu vc th x ng Xoi. Kho st c tin hnh trn 18 mu thuc 2 nhm thc phm khc nhau: 9 mu thuc nhm ch c chn n ngay, 9 mu thuc nhm ch la. Tt c cc mu thc phm c thu mua ti ch ng Xoi, tnh Bnh Phc. Cc mu c phn tch bng phng php nui cy truyn thng. Kt qu kho st c th hin trn bng 4.1v trn biu 4.2 v 4.3. Bng 4.1: Kt qu kho st t l nhim S. aureus trong cc nhm thc phm STT 1 Nhm mu Ch la Tng s mu 9 T l mu khng t (%) 77,78 22,22 T l mu t (%)

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 51

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi 2 Ch c chn n ngay
22,22 % 44,44% 77,78% 55,56%

55,56

44,44

a.Ch la Biu 4.1: T l nhim S.aureus trn mu th

b. Ch c

Cn c theo quyt nh s 46/2007/Q-BYT ngy 19 thng 12 nm ca B trng B Y t. Quy nh gi hn sinh hc ti a nhim trong thc phm s dng trc tip ( qua x l nhit): Nhng mu b nhim S.aureus vt qu 102 vi khun / 1g thc phm i vi mu ch la v 10 vi khun/1g thc phm i vi mu ch c chn n ngay c xem l khng t v vy khng c s dng nhng mu thc phm ny. i vi mu ch la: khi phn tch 9 mu th c n 7 mu c S.aureus vt qu gii hn cho php, chim 77,78%. i vi mu ch c chin n ngay: khi phn tch 9 mu c 5 mu nhim S.aureus vt qu gii hn cho php, chim 55,56 %. T kt qu trn cho thy, trong tng s 18 mu thc phm c phn tch bng phng php nui cy truyn thng c 12 mu c khng nh l nhim S. aureus vt qu tiu chun, chim 66,67% (12/18) , nhng mu ny phi c loi b v c 6 mu nhim S. aureus nm trong gii hn cho php, chim 33.33% (6/18), nhng mu thc phm ny vn c php s dng. SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 52

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi T l mu ch la khng t chim t l cao hn so vi mu ch c. Kt qu ny cho thy cc loi thc phm c by bn ti cc ch trong khu vc ng Xoi c t l nhim S. aureus kh cao. iu ny khuyn co cc nh qun l thc phm cn c nhng bin php tng cng gim st v sinh an ton thc phm ang lu hnh trn th trng, nhm m bo an ton cho sc khe cng ng. Vi kt qu phn tch nhn c nu trn, c th kt lun rng: tnh hnh nhim S. aureus trong cc thc phm ti ch ng Xoi, tnh Bnh Phc l rt ng quan tm. T l nhim S. aureus c kh nng gy bnh vn chim mt t l kh cao trong tng s mu phn tch.

PHN V. KT LUN V NGH

5.1. Kt lun
T cc kt qu kho st trn cho php rt ra cc kt lun nh sau: - Trong tng s 18 mu c kho st, s mu pht hin t l nhim S. aureus chim 12/18 tng ng 66,67%. - Trong s cc nhm thc phm c kho st th t l nhim S. aureus trong nhm ch la chim t l (77,78%) cao hn trong nhm ch c n ngay (55,56%) - Qua qu trnh lm bi kha lun ny chng ti nhn ra rng Staphylococcus aureus l 1 loi vi khun nguy hin cho c con ngi v ng vt. - Vi khun ny rt nguy him c bit l cc c t ng rut ca n. N gy ra cc v nhim khun ng rut cn c coi l 1 siu khng nguyn rt nguy him v l s bt u ca cc triu chng sc c hi. Ngoi ra vi khun ny cn ly nhim trong cc bnh vin, y l 1 iu ng bo ng v hu ht nhng v nhim trng ny nh hng n cc bnh nhn mi m xong v c bit l tr s sinh. SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 53

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi - S.aureus cn gy cc triu chng sc c hi rt nguy him i km vi nhim trng huyt. Ngi khe mnh hu ht u c S.aureus nhng ch khi c iu kin thch hp th vi khun ny mi sn sinh ra cc c t. Tm li, trn th gii hin nay c rt nhiu loi vi khun nguy him khng ch l S.aureus m cn l Clostridium botulinum, E.coli, Salmonella. y u l nhng vi khun nguy him n sc khe ca con ngi. Vic tm hiu v nghin cu v chng l rt cn thit ra cc bin php phng nga thch hp nhng c l thc ca mi con ngi l bin php phng nga hiu qu nht. 5.2. ngh Do gii hn v thi gian v iu kin th nghim nn c nhiu vn cha th thc hin c. Nu ti c tip tc, chng ti c cc xut sau: - Kho st tnh trng vi sinh ca khu bun bn. - Ly mu pht tay ca nhng ngi ch bin v bun bn thc phm. - Nn lp cc a im bn thc phm tp trung d dng kim tra v sinh khu bun bn. Thng xuyn t chc cc t kim tra m bo v sinh an ton thc phm - Tin hnh xt nghim vi s lng mu ln v trn nhiu i tng mu trong phm vi a bn rng c s nh gi tng qut v t l S. aureus trong thc phm. - Nghin cu s tc ng n cc vi sinh vt gy bnh khc nh Salmonella, Listeria monocytogenes. - M nhng lp tp hun v v sinh an ton thc phm cho ngi sn xut v kinh doanh thc phm.

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 54

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi

TI LIU THAM KHO

Ti liu ting Vit 1. Nguyn Vn Hi v L Trung Hi. Nhn xt 173 trng hp ng c thc phm ti Khnh Ha 2001-2004, Trung tm y t d phng tnh Khnh Ha, Thng tin khoa hc, Cc an ton v sinh thc phm, 2005. 2. Bi Th Hin, T Th Thu v Cng s. Tnh hnh nhim thc phm do vi sinh vt ti hai x huyn Kin Xng tnh Thi Bnh nm 2001, Trung tm y t d phng Thi Bnh, Thng tin khoa hc, Cc an ton v sinh thc phm, 2005. 3. Nguyn L Hng, Nguyn th Phn v Bi Th Kim Dung. Kho st tnh hnh nhim vi sinh vt trn mt s mt hng thc phm n lin bn ti cc ch Tp.H Ch Minh trong 3 nm 2002-2004, Trung tm y t d phng Tp.H Ch Minh, Thng tin khoa hc, Cc an ton v sinh thc phm, 2005. 4. Vng Th Vit Hoa, 2002. Gio trnh thc tp Vi sinh thc phm. Trng i hoc Nng Lm TP.HCM. 74 trang. 5. Th Ha, 2006. Phng chng t cu trng vng. Khoa hc ph thng, s 30/06.

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 55

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi 6. Cc an ton v sinh thc phm, 2006. C s d liu ng c thc phm. 7. Nguyn Phc, Hong Hoi Phng v Bi Kiu Nng. nh gi mc nhim vi sinh vt thc n ng ph ti thnh ph H Ch Minh nm 2002, Vin V Sinh Y t Cng Cng Tp HCM, Thng tin khoa hc, Cc an ton v sinh thc phm, 2003. 8. B mn vi sinh khoa Y, 1996. Thc tp vi sinh hc v min dch hc. Trng i hc Y dc TP.HCM. 134 trang. 9. Trn Linh Thc, 2002. Phng php phn tch vi sinh vt trong nc, thc phm v m phm. Nxb Gio dc. 230 trang.

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 56

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi Ti liu ting nc ngoi 10. Beatriz Pinto, Empar Chenoll v Rosa Azna, 2004. Identification and typing of foodborne Staphylococcus aureus by PCR-based techniques. Systematic and Applied Microbiology 28: 340-350. Elsivier Science. 11. Capucine Letetre, Sylvie Perelle, Francoise Dilasser v Patrick Fach, 2003. Detectio and genotyping by real-time PCR of the Staphylococcal enterotoxin genes sea to sej. Molecular and Cellular Probes 17: 139-147. Elsivier Science. 12. C H Collins, Patricia M Lyne v J M Grange, 1995. Staphylococcus and Micococcus. Collines and Lynes Microbiological Methods, (C H Collins, Patricia M Lyne v J M Grange). Butterworth-Heinemann Ltd. p.353-359. 13. C. Letertre, S. Perelle, F. Dilasser v P. Fach, 2003. Identification of a new putative enterotoxin SEU encoded by the egc cluster of Staphylococcus aureus. Journal of Food Microbiology 95: 38-43. The Society for Applied Microbiology. 14. C. Vernozy-Rozand, C. Mazuy-Cruchaudet, C. Bavai v Y. Richard, 2004. Comparison of three immunological methods for detecting staphylococcal enterotoxin from food. Letter in Applied Microbiology 39: 1390-394. The Society for Applied Microbiology. 15. D.L.K.Ng v L.Tay, 1992. Enterotoxigenic strains of coagulase-positive Staphylococcus aureus in drinks and ready-to-eat foods. Food Microbiology 10: 317320. Academic Press Limited. 16. Ernest Jawetz, Joseph L. Melnick, Edward A. Adelberg, George F. Brooks, Janet S. Butel v L. Nicholas Ornston, 1989. The Staphylococci, Medical Microbiology (Ernest Jawetz, Joseph L. Melnick, Edward A. Adelberg, George F. Brooks, Janet S. Butel v L. Nicholas Ornston). Pretice-Hall International Inc, USA. p.187-192. 17. G. Normanno, A. firinu, S. Virgilio, G. Mula, A. Dambrosio, A. Poggiu, L. Decastelli, R. Mioni, S. scuota, G. Bolzoni, E. Di Giannatale, A.P. Salinetti, G.La Salandra, M. bartoli, F. Zuccon, T. Pirino, S.Sias, A. Parisi, N.C. Quaglia v G.V. Celano, 2004. Coagulase-positive Staphylococci and Staphylococcus aureus in food products marketed in Italy. International Journal of Food Microbiology 98: 73-79. Elsivier Science. 18. Hiroshi Fujikawa v Satoshi Morozumi, 2005. Modeling Staphylococcus aureus growth and enterotoxin production in milk. Food Microbiology 23: 260-267. Elsivier Science.

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 57

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi 19. H.J.Jogensen, T. Mork, H.R.Hogasen v L.M.Rorvik, 2004. Enterotoxigenic Staphylococcus aureus in bulk in Norway. Journal of Applied Microbiology 99: 158166. The Society for Applied Microbiology. 20. Kenneth Todar. Todars Online Textbook of Bacteriology University of WisconsinMadison Department of Bacteriology (Staphylococcus). Kenneth Todar University of Wisconsin-Madison Department of Bacteriology, 2005. 21. Scott E Martin, John J Iandolo, J Harvey, A Gilmour, Sita R Tatini, Reginald Bennett v Merlin S Bergdoll, 2000. Staphylococcus. Encyclopedia of Food Microbiology, (Richard K.Robinson, Carl A.Batt v Pradip D.Patel). Academic Press, San Diego - San Francisco - New Yolk Boston London Sydney - Tokyo. p.20622083. TI LIU WEBSITE:
22. http://vi.wikipedia.org/wiki/T_cu_khun 23. http://vi.wikipedia.org/wiki/C_ch_c_lc_ca_vi_khun 24. www.bacsidakhoa.com 25. http://www.ncbi.nlm.nih.gov/pubmed/1820332 26. http://www.ncbi.nlm.nih.gov/sites/pubmed 27. http://www.benhhoc.com/index.php?do=viewarticle&artid=1427&title=dap-ung-

mien-dich-chong-nhiem-trung
28. http://tusach.thuvienkhoahoc.com/wiki/Min_dch_bnh_sinh_ca_nhim_trng

_huyt

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 58

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi

PH LC
S liu S.aureus trn mi trng BP trong mu ch la (vi khun/1g) TN 1 TN 2 TN 3 Trung bnh Mu 1 Mu 2 Mu 3 260 300 50 250 290 60 240 280 50 250 290 53,33 Mu 4 400 410 395 401,66 Mu 5 310 300 315 308.33 Mu 6 Mu 7 Mu 8 70 170 150 80 165 145 75 160 155 75 165 150 Mu 9 360 355 365 360

S liu S.aureus trn mi trng BP trong mu ch c (vi khun/1g) TN 1 TN 2 TN 3 Trung bnh Mu 1 Mu 2 Mu 3 160 10 170 150 20 150 140 10 165 150 13,33 161,67 Mu 4 195 185 175 185 Mu 5 Mu 6 Mu 7 30 50 150 40 55 145 45 40 160 38.33 48,33 151,67 Mu 8 15 14 20 16.33 Mu 9 260 255 265 260

PHA CH MI TRNG V THUC TH 1.Dung dch nc mui sinh l SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 59

Kho st t l nhim Staphylococcus aureus trong thc n nhanh ch ng Xoi Cn 8,5 g NaCl, ha tan trong 1000 ml nc ct. Hp tit trng 121oC/ 15 pht. Bo qun 2-8oC. 2.Thch Baird-Parker (mi trng ng kh) Chuan b: Cn 58g, ha tan trong 950ml nc ct bng cch un nng cho tan hon ton. Chnh pH 7.0 0.2 250C. Hp tit trng 1210C/15pht. ngui n 45 50 oC cho them 5ml dung dch egg yolk, lc nh cho tan u, trnh to bt. rt khong 12-15 ml mi trng vo a petri v trng. Bo qun mi trng 1 thng 2- 80C. 3.Canh thang BHI Chun b: Cn 37 g/l. Ho tan trong nc bng cch un nng nu cn. Chnh pH = 7.4 0.2. Phn phi 0.2ml vo cc ng nghim. Hp tit trng 1210C/15pht. Bo qun mi trng 1 thng 2- 80C 4. m phosphate - Dung dch m: Ho tan 34g KH2PO4 trong 500ml nc ct, chnh pH ti 7.2 bng cch thm 175ml dung dch NaOH 1M, v thm nc ct cho va 1000ml. Bo qun mi trng 1 thng 2- 80C - Dung dch pha long: Pha long 1.25ml dung dch m vi 1000ml nc ct. Phn phi vo chai Dural hoc ng nghim np vn c dung tch thch hp. Hp tit trng 1210C/15pht. Bo qun mi trng 1 thng 2- 80C 5. Dung dch huyt tng th hon nguyn Hon nguyn huyt tng th theo hng dn ca nh sn xut. 6. Mi trng thchTSA Chun b: ha tan trong 950ml nc ct bng cch un nng cho tan hon ton. Chnh pH 7.0 0.2 25oC. Hp tit trng 1210C/15pht. Phn phi mi trng vo ng nghim to thch nghing Bo qun mi trng 1 thng 2- 80C

SVTH: Nguyn N Lan Anh; Nguyn Th Dim Hng; inh Th Vn 60