TỶ LỆ MANG KHÁNG NGUYÊN BỀ MẶT VI RÚT VIÊM GAN B HBSAg TRÊN CÁC ĐỐI TƯỢNG KHÁC NHAU TẠI TỈNH LÂM ĐỒNG
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BỘ GIÁO DỤC VÀ ĐÀO TẠO
TRƯỜNG ĐẠI HỌC ĐÀ LẠT
HOÀNG CÔNG LONG
TỶ LỆ MANG KHÁNG NGUYÊN BỀ MẶT
VIRÚT VIÊM GAN B (HBSAg)
TRÊN NHỮNG ĐỐI TƯNG KHÁC NHAU
TẠI TỈNH LÂM ĐỒNG
LUẬN VĂN THẠC SĨ
NGÀNH: SINH HỌC
ĐÀ LẠT - 1995
80 GlAD
Due vA flAo T.o.O
TRVONG E)~I HOC DALAT
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HoANG CONG LONG
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MANG KHANG NGUYEN BE MAT
VIRUT VIEM GAN B (HB sAg)
TREN NHUNG DOl nJc;5NG KHAc NHAU
TAl TINH LAM DONG
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CHUYEN NGANH: VI SINH vAT Y HOC
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GSTS. HoANG THUY NGUYEN
PTS. NGUYEN THU VAN
([)alal, 1995
El§ holm thanh Luiln an nay, toi xin chan thanh cam dn :
• GS. TS HoANG THUY NGUYEN - Vi~n trVClng Vi~n V~
sinh Djch Ie hoc Trung Vdng - Ha NQi.
• P.T.S. NGUYEN THU VAN - Trung tam Ki§m dinh Quoc
Gia - Ha NQi.
• GS. NGUYEN THI KE. Vi~n trvClng Vi~n Vaccin va cac
che pham sinh hoc.
• SS. PHAN THANH HAl, Giam doc Trung Tam Chan
doan Y Khoa - TP. HCM.
• TS. LE sA DONG - TrvClng Khoa Elao taO sau Elai hoc TrVClng Elai hoc DALAT.
• SS. El6 HOU CHiNH - Giam doc Trung Tam Y hoc Dl,!
phbng - Tlnh Lam Dong.
• PTS. LE VAN Hli;p - Vi~n ph6 Vi~n Vaccin.
• OS. NGUYEN THANH TONG - TrvClng Khoa xet nghi~m
- TTCElYK - TP/ HCM.
• P.T.S ElAO XUAN VINH. Phong vi sinh vi~n v~ sinh
Dich I§ Tay Nguyen.
• TRUdNG ElAI HOC ElA LAT.
• vA
cAc
ElONG NGHli;p.
Ela het long giup do va dQng vien.
HBsAg
Hepatitis - B Surface Antigen
Kheng nguyen be mi;it vinH viem gan B
HBV
Hepatitis B Virus
Virut viem gan B
CV
Cut off value: gia tri ngtlang.
NCx
Negative control Mean
Gia tri trung binh cac chung Am.
P
Positive: DtlClng tinh
N
Negative: Am tinh
AFP
Alpha Fetoprotein
HIV
Human Immuno Deficiency Virus.
RPHA
Reverse Passive Haemagglutination
Ngtlng ket hong cau thv d¢ng ngtl<;1c.
RIA
Radio Immuno Assay
Mien dich ph6ng
ELISA
XI;!.
Enzym linked immuno Sorbent Assay.
Mien dich Men.
CIEP
Counter immuno Electro Phoresis.
Elien di mien dich doi Itlu.
PLC
Primary Liver Cancer.
Ung thtl gan Nguyen phat
HCC
Hepatocellular carcinoma
Ung thtl bieu ma te bao gan
NIHE
National Institude of Hygiene ADN Epidemiology.
Vien V e sinh Dich Ie hoc Trung UCing.
WHO
World health organization
Hi chuc y te the gioi.
q.Jhdn LIlli Ilht1l
DAT VAN DE
2
Vinit Viem gan B (HBV) Iii lac nhan gay benh Viem gan B ",t nguy hiem.
Nglloi mang Khang nguyen be m?1 virut Viem gan B Iii mol lrang nhung
nguyen nhan lam lay Iruyen HBV, gay benh Viem gan sieu vi cap hoac
man, sau d6 c6 the chuyen Ihilnh Xo gan va cuoi cung la Ung Ihll gan
(carcinoma).
Hien nay Viem gan virut Iii mol van de dang dllc;Jc quan lam ti;li nhieu nlloc
Iren the giOi. Ty Ie nglloi mang virut nay
Bac My va Chau Au Iliong doi
thap: 0,1 den 0,2% khong Vll(jl qua 2%. Song li;li nhieu nlloc Chau A, Chau
Phi thi Iy Ie nay ral cao : 15 den 20%. Ve phllong dien dich Ie hoc clla HBV
ral giong voi HIV.
a
Thea 16 chuc y Ie the gioi (WHO) moi nam c6 kholmg 2 trieu nglloi tren
loan cau chel do virul Viem gan B, hon 285 trieu ngllai mang mam benh
trang d6 nhieu nhat la ngllai Chau A - chiem Iy Ie 75%.
Ung Ihll gan va Xo gan la nhung benh hiem nghE!O gay tU vong nhanh
ch6ng sau khi c6 bieu hien lam sang, song neu phal hien sam a giai doan
dau, kel qua dieu Iri S8 101 hem rat nhieu. £lay la nhung can benh tien Irien
ch"m, nhllng chac chan trong nhieu nam sau S8 c6 khong il ngllai bj Xo
gan dan cten Ung Ihll gan.
Thang 3.1993, lrang Hoi IhllO Khoa hoc ve chuyen de Ph6ng chong Ung
thll gan nguye~ phal ti;li Ha Noi, mot trong nhung muc tieu de ra la: lam Ihe
nao de pilat hien th"t sam Irong can bo, nhan dan nhung ngllai c6 cac yeu
to nguy co cao.
Hien nay, Viel Nam dang n1lm trong vung dich tEl virul Viem gan B Iliu hanh
muc do cao. Lam £long la mol Tlnh mien nui phia nam Tay nguyen, c6
Thanh pho 8a Li;lt la noi nghl mal Iy tllang cho du khach. Neu chung la
khong c6 bien phap ng{m ch?n virul HBV S8 bien noi nay thanh 1 vung
muc do ngay cang cao. £lieu nay S8 anh hllang ral Ion ve
nhiem virut
nhieu m?l: i
a
a
a
3
Vi vf:!y, de tai se tien hanh dieu tra Huyet thanh va giam sat tinh hinh mang
HBsAg l<;li Tlnh Lam EJong. Viee ThlJ nghiem mau xae dinh virut HBV tren
moi ngLloi binh thLlong va tn§n nhung doi lLlong c6 nguy co eao bi nhiem
HBV la dieu rat can thiel va d6 ehinh la mue lieu ella de ta;,
Xae dinh dLloe ty Ie cae nh6m ngLloi cae nh6m doi tLlong khae nhau e6
mang HBsAg DLlong tinh de sau d6 ve mf:!t Dieh te hoc se e6 nhung bien
phap h<;ln ehe eOng nhLl ngan chan st,llan truyen ella virut HBV trong dan
eLlt<;li cae Tlnh va Thanh pho trong ea nLlOe.
Ngoai ra mue lieu ella de tai la ung dung cae tien b(> khoa hoc ky thuat
trong chan doan de phat hien ngLloi e6 mang virut va dt,l ph6ng nhiem HBV
doi vol ngLloi am tinh.
4
rph.iiJL title ltai
;J
..,.,
TONG QUAN TAl L1I;U
5
Viem gan B la mot benh do Hepatitis B virus (HBV) ral phb bien va t",a
thanh mot van de nghiem trong ve y Ie cong cong <'J nhieu nLioc tren the
gioi ngay nay. NhLlng benh nay con la mol moi de doa nghiem trong han:
VinH gay ra Vi em gan B chinh la tae nhan gay Ung thLi thu ba dLlc;Je biet sau
thuoe la.
BLioe di dau tien Irang viee xae dinh da dlic;Je thl/e hien vaa nam 1983 b<'Ji
Baruehs. Blumberg t",i Vien nghien eLlu Ung thLi <'J Philadehphia trang khi
nghien cuu mot so Protein trang Huyet thanh ella mot benh ph€lm lay W
mot benh nhan bi benh L1a chay mau, ong da lam bi~u hien ro mot Khang
th~ phan ung voi mot Khang nguyen e6 trang mau mot thb dan Australia bi
Viem gan.
EJen nam 1968, Blumberg xae dinh d6 Iii Khang nguyen be m.;;t ella virut
Viem gan B.
1. Virut Viem gan B:
Thuoc hO Hepadnaviridae. NgLioi ta da xac dinh dLlc;Jc virut Viem gan B c6 5
dau an mien dich :
- Khang nguyen be m';;t virut Viem gan B (HBsAg).
- Khang lh~ chong Khang nguyen be m';;t virul Viem gan B (Anti-HBs).
- Khang th~ chong Khang nguyen 16i virut Viem gan B (Anti - HBc).
- Khang nguyen e Virut Viem gan B (HBeAg).
- Khang th~ chong Khang nguyen e virut Viem gan B (Anti - HBe).
Trang 5 dau an mien dieh Iren ngLioi la thay dau an Khang nguyen be mat
virul Viem gan B Iii quan Irong nhat (Hinh a).
HBsAg e6 th~ ton t",i trang mau bat eu lue nila <'J giai da",n nhiem cap, man,
vii c6 th~ ton I",i suot dOi. Nang do clla HBsAg DLiong tinh (+) giam dan
thea lubi nhLlng tae h",i ella Xo gan vii ung thu gan thi tang dan thea tubi <'J
nglioi e6 mang HBsAg (+) trang mau, thong thLiong Ung thLi gan chi xuat
hien sau mot thai gian tiem tilng W 30 den 50 nam, nhLlng cOng nh';;n thay
mot so ca carcinoma te baa gan <'J Ire em .
6
CAU TRUC VIRUT VIEM GAN B
HEPAnTISB:
THE VIRUS
-'
3
1. Va boc
2. Khang nguyen tien S
3. Khang nguyen S
4. HBeAg
5. HBcAg
6. M(>I Nucleocapsid mang nhung kh ang nguyen
7. Enzym ADN - Polymeraza
8. B(> gen ADN clla HBV
7
d~c
bi$t
HBV c6 mot eElU true hai Ian va protein dong tam. Va boe,
nghTa la va ngoai baa gam 3 protein dlioe gOi la "Ion hem",
"trung blnh" va "Ion". HBsAg, Khang nguyen be mat, e6 mat
tren ea 3 protein d6. Capxit hay va trong eau t",o blil mot
protein Ii trong vua baa boe vua tLidng tae voi ADN eua virut.
Bo gen eua HBV Ian dau lien dLioe ph an lap nam 1974 blii William
S.Roblnson Ii TrLiong Y thuoe 8",i Hoc Tong HOp Stanford, Iii mot philn tU
ADN vong baa gam khoang 3200 Nuelectit. 86 la bo gen nha nhat virut
dong vat dLioe biet; bo gen (jenom) eua HBV eau t",o blii hai soi nhLing n6
I",i e6 1 dae tinh bat thLiong : Ha; sol e6 ehieu da; khong biing nhau. Chieu
dal eua sOi ngiin con gOi la sOi "dLidng" thi thay doi va chi biing 50 den 80
ehieu dai eua soi "am". Cau true dae biet nay la ket qua eua cd ehe nhan
len doc nhat dLioe vi rut su dung.
Cau true vong eua virut dLioe baa dam blii SLi gMp do! giua cae baz Ii m(Jt
dau eua cae soi. Vlrut HBV e6 eau true dae biet xit chat. N6 baa gam e6 4
gen dLioe gOi la gen S, gen C, gen P va gen X. Chum phu len nhau rat
rQng. NhQng dO<;ln dieu hoa kiem soat sL,i san xuat CaG protein eua vi rut va
ehu ky nhan fen eOng baa gam trong cae do",n ghi rna.
Gen S ghi rna hay chua dl,lng thong tin eho protein Ion hdn eOng nhLi eho
HBsAg. Mot doan khoang 500 Nueleotlt Ii phia trLioe g"n S eOng e6 the
dLioe phien rna eung lue (Wolfram H. Gerlich) ta e6 the ph an blat trong
do",n nay 2 vDng Ii phia trLioe, tien S, va tien S2 tham gia vao Sl,l tong hop
cae protein kMe eua va boe : Protein trung binh dLioe ghi rna blil tien S2 va
gen S, Protein Ion dLioe ghl rna blii tien S" tien S2 va gen S. VDng lien S,
e6 val tro quan trong de vi rut xam nhap vao te baa gan (A. Robert Neurath.
eua Trung tam mau Ii New York da chung minh) (40).
Gen C ghi rna eho protein eua Capxit, eOng nhLi gen S, gen C eOng e6 1
do",n tien C ngiin Ii trLioe gh; rna eho 1 pept;t ky nLioe tham gla vao viee liip
ghep h",t virut. Gen P e6 ehieu dai tOi mue n6 eiit cae phan eua tat ea cae
gen, ghi rna eho cae Enzym eeJ ban eua ehu ky nhan len eua HBV. Vi tri
eua gen X niim tran cae dau nol eua cae soi ADN eua Virut. San pham
Protein eua n6 kieh thieh sl,l the hlen eua tat ea cae gen khae biing tLieJng
tae va; m(Jt do",n ADN dae biat e6 mat trong b(J gen HBV.
8
Cac Hepadnavirul nhan len Iheo mot ceJ che bal Ihuang. Ela phan vinH
chua AON lai lao Iruc liep b(> gen cGa chung nho nhung Enzym gol la
Polymeraza, cac polymeraza xli dung cac sol AON mol sinh lam khuon
mau Iflp ghep cac soi mol bo sung. Tuy nhien cac hepadnavirul sli dung 1
phueJng phap glan tlep c6 st,l Iham gla cGa ARN lam Irung gian ceJ che nay
da duoc mo la Ian dau (;en nam 1982 blii Jesse W. Summers
Williams.
Mason cGa Trung lam Ung Ihu Fox Chase (Philadelphie).
CeJ che nay nhu sau: Khi m(>1 vlrut cGa hepadnavirut xam nhaP vao te bao
n6 dl loi nhan, Ii do nhung polymeraza te bao phlen rna Ihanh m(>t phan tti
ARN da;. Philn Iii ARN nay co 3500 Nucleolil dU(jc goi la lien jel10m. Tien
Jenom duoc boc b~ng m(>t AON polymeraza cGa virul lrong m61 capxit moi
sau do Ira I"i nguyen sinh chal Ie bao. 0 day, polymeraza cGa
1"0 Ihanh
vlrut lam cong viec phlen rna nguoc cGa tien jenom Ihanh m6t sol AON am
moi. Elen khl soi am da hlnh Ihanh, tien jenom bj pM hGy bai cac Enzym,
luc do cac AON - Pclymeraza co Ihe xay dt,lng s(jl dueJng sli dung soi am
lam khuon mau. Capxil
AON cGa virul cuoi cung duoc dal Irong mol
boc ngoai moi khi virus roi Ie bao gan. Khl vlrut duoc hinh thanh sol dueJng
ngung dai ra laP ltJc. Elieu nay giai Ihich t<;ll sao chieu dai cGa s(jl dueJng
thay do;'
va
va
va
va
Tal ca cac virul deu la nhung vi sinh val ky sinh Ie bao, cau 1<;10 blJl 1 Ihe
nhlem sflc la ARN hay AON (b6 gen) chua dt,lng trong 1 va boc protein nhu
da mo ta Ii phan tren. Muon nhiln len vlrul phal dl vao Irong Ie bilO va xli
dung b6 may cGa Ie bao de tong hOp cac protein cGa
boC
b6 gen cGa
virul klli b6 gen da dU(jc bao Irong
boC thl cac hal virul moi roi Ie bao
c6 Ihe gay nhiem cac Ie bao Ian Can
va
va
va
va
CeJ che hap dan cGa st,l nhiln len cGa hepanavirul lam oM l<;Ii CeJ che sli
dung bal cac Relrovirul, nhu virul SIOA co b6 gen la ARN
sli dung m61
philn tli AON lam Irung gian. cac hepadnavirut con co nhung diem chung
kMc nua vol Retrovirut. ea 2 deu gay nhiem te bao m61 cach Ilem lang va
khong pha hGy Ie bilO do d6 Iht,lc Ie ta co the t11ay r6 hi en tuong nhlem vi
rlll nhung khong xual hien Irieu chung, ceJ Ihe
hoan loan khae m<;lnh,
sinh hO<;l1 van binh Ihuong, Iinh Ir<;lng nay co the keo dai tU 8 - 10 nam. Sau
do moi chuyen qua gial dO<;ln benh lam sang.
va
van
Ta co Ihe Iham khao sl/ xam nhap
(hlnI12)
9
va
nhiln len cGa Relrovlrul duoi day
Hinh 2:
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1. VirLiI HIV
2. ThV IhEi CD,
3. PhOng ARN vil Enzym pllien
ma ngl1<;1c vila ben Irong Ie bilo
4. ARN cua virLiI
(,)
t.l 0
.)
~O
9. Te bilo san xuat ra nhieu ARN va
prolein virLiI
10. Prolein virLiI
11. ARN virLiI
12. NhGng virLiI moi hinh thitnh IV I$p
a mimg Ie bilo
13. NhGng mam virLiI moi da ra
ngoai Ie bilO chuan bi gay nhi§m
Ie bilo Ian c$n
5. Enzym phien ma ngl1<;1c
6. ADN cua virLiI
7. Nilan Ie bilo Bach cau
8. ADN cua Ie bilo Bach cau
10
VinH HIV e6 eau 1<;10 don gilm, chI can 1 ph an tU ARN va mang Polypeplit.
Chung ky sinh tren cae te baa B<;Ieh eau, te baa than kinh va 1 so te baa
kMe eua co thEi ngliai. HIV xam nhap vao B<;Ieh eau bang each bam vilO
mang B<;Ieh eau, ph6ng ARN vao ben Irong, can mang Polypeptid thi nam I!
ngoai. Nha e6 Enzym phien ma nglioe ma ARN d6 tli nhan len thanh ADN,
ADN nay gan voi ADN eua b<;leh eau trl! IMnh vat lieu di truyi'm moi, tU d6
san xuat ra cae ARN eva HIV ma t<;lO tMnh 2 - 4 - 8 mam HIV kMe trong
b<;leh cau. Chung pM va vaeh B<;Ich eau roi I<;Ii bam vao cae b<;leh eau khae
dEi tiep tue nay nl! va phat triEin. Bang each d6 HIV pM hvy dan he thong
mien dieh eva co IhEi ngliai.
Thli tli va ehlie nang cae gen eva Retrovirut : gag, pol va env deu e6 thEi so
sanh trlie tiep voi cae gen eva hepadnavirut : C, P va S (gag ghi ma eho
protein eva capxit retrovirut, pol ghi ma eho Enzym phien ma va EnV ghi
ma eho cae prolein eva v6 boe). Ca 2 ho virut nay deu e6 anh hlil!ng doi
voi ung thli.
2. Soj lien quan giG'a HBV va PLC :
Cae nghien cliu Dich t8 hoe da ehling minh r6 rang la nhiem HBV man tinh
dv dEi gay Ung thli gan. Cae dong vat c6 vu bi nhiE'im man tinh hepadnavirut
e6 Iy Ie mac Ung thli gan eao : tren 80% cae s6e maemot bi nhiE'im phat
triEin mot Ung thli gan tien pMt (HCC) trang vong 2 nam. Hans Popper
thuoe El<;li hoc T6ng hop Cornell va John L. Gerin thuoe El<;li hoc T6ng hOp
Georgelown da ehling minll rang e6 thEi gay ra Ung thli gan thlie nghiem
tren s6e Maemot bang each tiem virut Viem gan rieng eva giong nay
(30,40).
Cae Thl! nghiem nay da khi!lng dinh l<;Ii vai tro sinh Ung thu eva HBV va 10<;li
tru Sli can thiet cva cae dong yeu to eva sli phal sinh Ung thli C6 nhung
hien tliong kMc e6 thEi tham gia vao benh I! ngliai: Thai ky tiem lang di
trlioc Sli pMt triEin eva Ung thli gan I! ngliai dai hall rat nhieu so voi s6e
maemot va cae Ung thli thliang xual hien tren gan bi xa. Cae chat gay Ung
thlilrol1g moi trliang va slilieu thu qua nhieu rliou eung luon luon e6 thEi
neu len Irong sli pMt sinh Ung thli gan I! ngliai.(31)
Van de co ehe gay Ung thli cva HBV van chlia dlioe giai quyet. HBV e6 thEi
trlie tiep gay ra sli phat triEin cae u. Milt kMc, u eva gan e6 thEi xay den
mot each gian tiep do Viem gan m<;ln tfnh, Xa gan va slitai t<;lO Ie baa xay
ra trong cae mo bi benh. Nhieu virul ehiu traeh nhiem gay ra Ung thli mang
cae oneogen, la nhung gen e6 thEi lam bien d6i trlie tiep cae Ie baa bi
nhiE'im eva co thEi. (Nhung oneogen tliang \lJ eung dlioe tim thay trong cae
Ie baa binh thliang trong d6 chung kiEim soal sli sinh trlil!ng va pMt triEin)
tuy nhien bo gen eva HBV hinh nhli khong mang oncogen. Them vao d6,
11
thai gian tiem tang dai giua lue nhiem HBV tOi lue xuat hi$n u khong phu
hOp voi
y kien
eha fang mot oncogen eua virut ch(u trach nhi$m gay Ung
thLi gan. Tuy nhien, ngLiai ta e6 the quan niem riing Protein hoat h6a cae
gen san xuat bai gen X eOa HBV e6 Ihe can thiep VilO mot giai doan sam
eOa qua trlnh phat sinh u.
o nLiae ta PLG la Ung thJ dung himg thu 3 sau Ung thLi pheii va Ung thLi da
day (pheii : 22.5%; Da day 15%; Gan 12%). Gae eong trinh nghien euu tren
the giai da chung minh moi lien quan chat ehe giua nhiem HBV va PLG.
Nhiem HBV man linh dLioc xem la nguyen nhan quan trong nhat gay HGG.
Elae biet (j Ghau A va Ghau Phi (S.Z.Yu. T.LSung. Ma Kew, T.G. Tan
1991) chi diem de tiep can nhat eOa nhiem HBV la sl,! e6 m;;;t eOa HBsAg
trong Huyet thanh.(7,12,13,26).
Mot so cae eong trinh eOng da chung minh: Huyet thanh ngLiai e6 mang
HBsAg (+) nguy e(J tLiang doi de dang m;:'c HGG eao gap 223 Ian so vai
ngLiai Am linh (Beasley-1981).
3. Cae vung d!eh Ii! Iren Ihe giai
Thea tei ehue y 1e the giai (WHO) thi hien nay tren the giai e6 3 mue do
nhiem HBV khac nhau:
(1) Vung dieh tEi nhiem HBV thap: (0 cae nLiae Tay Au, B;:'e My): Ty Ie
mang HBsAg (+) VaG khoang 0,2 - 0,5%.
(2) Vung dieh te nl1iem HBV trung gian: 0 cae nLiae Elong Nam Ghau Au,
Trung Elong: Ty Ie mang HBsAg (+) tU 2 - 7%.
(3) Vung dieh te nhiem HBV eao: 0 cae nLiae nhLi Trung Quae, £long Nam
A, GhilU Phi ... Ty 1$ mang HBsAg (+) tU 8 - 20%.
Ngoai ra con e6 mot so quae gia niim trong nhung vung dieh te nhiem HBV
rat eao, ty Ie mang HBsAg (+) tren 20%.
WHO eOng eho riing 3 10ai ty Ie dau eOng dO tap hOp dai da so cae nLiae.
Nhung euoe dieu tra ve ty Ie nhiem HBsAg dLioe tien hanh trong khu vl,!e
cae dao (j Nam Thai Binh DLiang eho thay nhling bien deii nhieu khi rat Ian
ve ty Ie mang HBsAg tU quan dao nay sang quan dao kia ho;;;e tU dao nay
sang dao khae va ngay ea trong eung mot dao(3). Tham khao bang sau
day ta se thay r6 nhung bien dong nay.
12
Muc dO d[ch Viem gan B ILlu hill1h dia phLiong IJ Nam Thai Binh DLiong.
HBsAg (+)
Mue dQ virut
lLiu hanh
Niim
< den 7%
Trung gian
1989
Australes
8-20%
Cao
1989
Marquises
...............................
Loyoute
>20%
Rat cao
1989
8-12%
Cao
1981
Tim Caledoni (Grand Terre)
0,6%
Trung gian
1981
Tan CalMoni (Grand Noumea)
4,2%
Trung gian
1987
Vanuatu
17,8%
Cao
1986
29%
Rat cao
NLide - Dao hay quan dao
Societe, Tuamotu
Tonga
Trong mot nghien cuu Djch t~ h<;>c ve HBsAg va Anti-H8s da dLic;JC thvc
hi$n t<;ii mot trLiang EJ<;ii H<;>c V khoa Hanover (Germany) tU nam 1974 den
1975 doi tLic;Jng la nhan vien b$nh vi$n da cho thay huyet thanh cua nhung
ngLiai chuyen lau chui san nha c6 HBsAg (+) la cao nhat (26,3%), ke d6 la
nh6m nu y ta (20,5%) roi den V-Bac Sl (18,2%), sinh vi en y khoa (11%),
thap nhat la cac h<;>c vien nu y ta khoang 5% mang HBsAg (+), voi n = 1825.
Tu van de tren ngLiai ta khuyen cao: Tat ca nhan vien b$nh vien
nhung
ngLiai tNc tiep hoac khong trvc tiep lam ve V deu c6 nguy co cao bj lay nhiem
vi rut Viem gan B. Do d6 tat ca can phai dLic;JC tiem chUng (18,25,29).
ca
Viet Nam nam trong vung c6 muc dO ILiu hilnh virut HBV cao, theo nhung
bao cao bLicJc dau muc dO nhiem virut tU 15 den 20% tuy Iheo tUng d[a
phLiong. T<;ii Ha Noi theo tac gia Hoang Thuy Nguyen va Nguyen Thu Van
(Vi$n V$ sinh Djch t~ h<;>c Ha Noi)(5,6,9):
18,02%
- NgLiai cho mau c6 HBsAg (+)
24,74%
- NgLiai khoe m<;inh c6 HBsAg (+)
43,50%
- B$nh nhan Viem gan c6 HBsAg (+)
25,24%
- B$nh nhiin khong phai Viem gan c6 HBsAg (+)
T<;ii thanh pho He, Chi Minh thea lac gia Tran Van Be (Trung tam truyen
mau va huyet thanh):
- NgVai cho va hien mau c6 HBsAg (+)
9,5%
11,4%
- Nhfll1 vien y te c6 HBsAg (+)
10.0%
- Phu nu c6 thai c6 HBsAg (+)
61,8%
- Benh nhan Ung thV gan c6 HBsAg (+)
Theo tac gia Ngo Quang Lvc (Ha Noi) Ty 1$ ngVai cho mau c6 HBsAg (+) la
9 - 10%. Mot nghien cuu khac tren nh6m doi tVc;Jng la nhiin vien y te IJ Ha
Noi la: 9,83%.
13
Va theo tac gia VO Tri$u An, VO Thuy Hien (16), thLJ nghi$m tren 43 nguoi
trang d6 Nhan vien Y te, Nhan vien Philng xet nghi$m sinh hoa, B$nh vi$n
Nhi va m¢t so can b¢ thu¢c ph6ng mien djch b¢ men sinh Iy b$nh de. c6
8/43 truong h<;lp HBsAg (+), chiem ty 1$ 18,6%.
Tren nh6m phu nv c6 thai ty
Ie mang HBsAg (+):
• Tai Singapore
4%
• Y (17)
2,4
• Trung Quoc
15,7%
• Vi$t Nam (Hai ph6ng)(11, 15)
12,59% (32 HBsAg (+) /254 ba me)
T?i Portugal (23): Ty 1$ nguoi kh6e m?nh nhiem virut Viem gan B la 24,3%
(1984)
Ta c6 the tham khao ban do Dich te hoc virut Viem gan B dudi day
Vung Djch te nhiem HBV thap
•
Vung Djch te nhiem HBV trung gian
•
Vung dich te nhiem HBV cao
14
4. Nguy eel nhiem HVB :
Tal ca nhung nguoi chua m<'lc b~nh Irong cong dong deu co Ihe nhiem vinH
(24,32). Trang so do cac doi luc;1ng sau day co nguy cd cao hdn:
• Tre sinh ra tU nguoi me co HBsAg Dudng Hnh. Tren 70% cac Ire sd sinh
nay se Ira Ihanh nguoi mang mam b~nh. Khi Ion len chling co nguy cd bj
Ung Ihu gan gap 200 Ian so voi Ire khong mang virli!.
• Nhung nguoi Irong gia dinh. Nhung nguoi liep xlic Ihan mal voi nguoi co
HBsAg Dudng Hnh.
• NMn vien y Ie, cac ky Ihual vien phong xel nghi~m va nhung nguoi lam
cong lac nghien cuu thuong xuyen liep xlic voi mau va cac che philm tU
mau.
•
B~nh nhan bi cac b~nh ve mau, Ihuong xuyen phlli Iruyen mau va cac
philm val tU mau.
•
Nguoi co quan h~ linh duc blia bai, nguoi co nhieu b,m linh, nguoi dong
linh luyen ai, gai mGli dam.
• Nguoi chich xi ke, ma lliy.
• Nguoi xam minh, xo
10 lai
hay b~nh nMn duc;1c dieu Iri biing cham cuu.
• Khach du Ijch Ihuong xuyen den vung dich.
5. 51! lay Iruy;;n vii lien trien eua nguoi mang virlil Viem gan B.
• 51! fay 'ruyen:
Eluong lay Iruyen clla vi rlit HBV la qua cac D[ch sinh Iy cua Cd the nhu:
mau, nuoc bol, linh dich, d[ch am dGlO va qua cac vel rach clla da. Gho nen
st/ liep xlic chal cile hay giao hc;1p co Ihe Iruyen HBV. Elieu nay gilli thich
duc;1c st/ lay Ian virlil Vi em gan B cao lrang n¢i b¢ m¢1 gia dinh, hay m¢t
c¢ng dong nho (34,35).
15
• ruin triifn ella nglf6i mang virtU Viem gan B.
NHIEM VI RUT VIEM GAN B
VIEM GAN CAP
VIEM GANMAN
~\
1/
VIEM CAN DQT PI-JAr
CVCNI-IANI-I
(Tlf yang 80 - 90%
XOGAN
InJb'ng hqp)
UNGTIIVGAN
NGUYEN PHAT
(Tl( vong 100%.
IrtJong hQpl
16
MANG 1-I~,Ag KIIONG
XUAT HIEN TRIEU
OILING
rpJullL LJul ba
PHudNG PHAp NGHIEN CUU
- --- ----;.:.- -_.
1 ~' • • I ' - . \ " t"" \ . ,..
I 1
!f
'I.'
':
~:'
I
I
\
,
u.,,'"
n.,I
4.
fI:' /.. ...
"rr. S".O..3L fU:D.Ab.G
~----------
17
1. Elia dii!m :
Cac mau mau dllc;Jc lay tV cac diem dEW cll. Phan bo deu trong Tlnh Lam
£)ong, tV cac B$nh vi$n T]nh, cac Cc;J SO Y te trong Tinh va Thanh pho, cac
Benh vi$n Huy$n v.v..
2. Eloi tliong :
Gam cac Nh6m nglloi nhll sau :
a. Nh6m kh6e m;;lnh: (can bo cong nhan vien, sinh vien, nhEw dan ..).
b. Nh6m benh nhan Viem gan: (Sau khi d8 c6 chtin doan lam sang).
c. Nh6m benh nhan khac: (Gam cac b$nh nhan den ph6ng kham cua cac
Benh vien hoac dang dieu tri t",i cac khoa).
d. Nh6m benh nhan Xc;J gan: (sau khi c6 chtln doan Lam sang la xd gan).
e. Nh6m Ung thll gan: (Sau khi da dllc;Jc chtin doan Lam sang va xet
nghi$m Alpha - fetoprotein Dlldng tinh).
f. Nh6m cho mau: (T",i B$nh vi$n Tlnh Lam £)ong va Benh vi$n II BllO Lac).
3. Lay mau va xU' II' mau :
Lay mau tlnh m",ch 2ml, sau d6 tach chiet huyet thanh. Trllong hop Irong
huyet thanh c6 hong dIU bi va phai lam s",ch bAng iy tam. Tat ca cac mau
huyet thanh sau khi tach neu chlla dung ngay phai bao quan I",nh tV +2
den +8°C. Muon bao quan lau hdn phai lam dong I",nh (-20°C).
4. Ky thu~t phat hien Khang nguyen be m~t virut Viem gan B.
a. Khao sal lam sang:
Khong thay vang da, vang mal tru b$nh nhan Viem gan , Xd gan va Ung
thll gan.
b. Khao sat mau:
- Ky Ihu(it SERODIA - HBs.
· Kit SERODIA - HBs clla Hang Fujirebio inc (Nh(it).
- Ky lhu(it ELISA (Enzyme linked Immuno Sorbent Assay).
· Kit micro Elisa HBsAg clla Labo Viem gan - Vi$n ve sinh Dich te
hoc Ha Noi.
· Kit 1M x HBsAg cua hang ABBOTT Laboratories (USA).
18
1. Ky
lhu~l
SERODIA - HB s :
SERODIA - HBs Ii. Phliang phap Ngling ket hong cau thu dong Ngli<,1C
(Reverse Passive Hemagglutination test) Dung de xac dinh Khang nguyen
be mat virut Viem gan - B.
a. Nguyen Iy eua thii' nghiiim:
Bil Sinh ph am HBsAg - test gam cac thuoc thu cho Thu nghi$m ngling
ket hong cau thu dong ngli<,1c dung de xac dinh Kh8.ng nguyen be mi;lt
virut Viem gan - B (HBsAg) trong huyet thanh, huyet tliang nglioi, dl,la
tren II' thuyet Hong cau cliu ho",t h6a dli<,1C gan voi Globulin mien dich
Anti-HBs (lgG) tho da dli<,1C hap phu c6 do tinh s",ch cao va ngling ket
mot cach di;lc hieu khi c6 mi;lt HBsAg trong huyet thanh, huyet tliang
nglioi.
Thu nghiem nay c6 mot so liu diem sau :
- So voi cac Thu nghiem thuoC the hi) 1 va 2 nhli Khuyech tan tren gel.
8i$n di mien dich doi lliu thi Thli nghiem nay c6 do nh",y cao han.
- Quy trinh thl,lc hi$n don giim.
- Tiet kiem thai gian. 80C ket qua sau it nhat la 1 gio va doc biing mat
thliong.
b. 89 sinh phii'm:
Bil sinh ph am Serodia HBsAg bao gam cac thuoc thu:
- Control celis: Kh8.ng HBs - Hong cau cliu ho",t h6a - £long kho, Hoan
nguyen biing dung dich d$m trlioc khi dung.
- Antibody sensitized cells (D",ng dong kho).
- HBsAg chLlng dLiang (Positive control) : chlia bat ho"t.
- 8$m muoi Phosphat.
- Serodia - HA hap phu (D"ng dong kho).
e. Dl!ng el!:
Cac dung Cu sau day can chuan bi trlioc khi bat tay thl,lc hien voi bo Kit
serodia - HBs.
- Phien nhl,la day hinh chCi "U".
- Que pha loang 25;t1 (# 0,025 ml).
19
- 6ng nhl> gi<;>1 25~d 1 gi<;>t.
Khong dung cac ong nhl> giot khac.
- Micropipets (0 - 50~d); pipet 1cc
- Dung dich glutaraldehyde, hoilc sodium hypophosphate.
d. Thl/c hi~n thy nghi~m:
- Gi<;>t 2 gi<;>t (# 50~1 ) dung djch d$m
vao gieng II.
VilO
trong gieng I va 1 gi<;>t (# 25~1)
- Dung Micropipet cho 5~d huyet thanh vao gieng I.
- Dung
se c6
gieng
gieng
que pha loang (Io"i 25~d) d~ tron deu, Huyet thanh trong gieng 1
dO pha loang la 1/10, cung dung que pha loang d6 chuy~n qua
II (Ela c6 siln 25~1 dung dich d$m). Tron deu, Huyet thanh trong
II luc nay c6 do pha loang la1/20.
- Nhl> mot gi<;>t (25J.l1) khang HBs hong cau cliu ho"t h6a (Tli d"ng dong
kho dil hoi chlnh trtidc d6 30' va bao quan CJ +2 oC den +1 O°C) vao
gieng I, va 25J.l1 Ie bao khang the nh"y cam vao gieng II. Nhti vaY nang
do cuoi cung cua Huyet thanh sa la:
1/20 trong gieng I
1/40 trong gieng II
- Song song tien Mnh mau Chung Am va mau ChL1ng Dtiong, dung d$m
muoi phosphate lam ChL1ng Am.
•
So d6 lhl,l'c hi~n:
Gieng so
(I)
·:~~;i;~::n~$m
_---_
_.....•....._......•...._
_
- Elo pha loang huyet thanh
....... _ _...................... ....................• .
::II~ ···-~~~r~;:
_ - _ _ -............ -.
1 : 10
...................................................................................................................................................................
- Anti - HBs Hong cau cliu ho"t h6a (Control cells) (J.ll)
- El o pha loang huyel thanh cuoi cung
20
1 : 20
25
(J.lI)
- Antibody sensitized cells
(II)
25
1 20
1 : 40
e. Nh~n dinh kif!
qua:
- Eloc kat qua sau it nhat la 1 gio, doc bang mat thllong.
- Tuy vao muc do Ngllng kat rna ph;m bi$t : Am tinh va Dliong tinh
80000
(- )
(+)
(++)
(+++)
(++++)
f. Khu- !rung:
- Ele tn,nh lay nhiem vinH HBV co the ap dung mot trong cac qui trinh
sau:
- Toan bo dvng cv va Nguyen vat li$u sau khi lam xong can phai ngam
vao dung dich Formaline it nhat la 72 gio /37 0 C.
- Hoi;ic ngam vao dung djch Glutaraldehyd 2% it nhat la 1 gio.
- Hoac dung djch Hypophosphate : 1 gio.
- Hap lIdt (Autoclave) : 121° C / gio.
2. Ky
lhu~l
ELISA:
a. Nguyen fy cua Thu- nghi~m :
Thu nghiem cua HBsAg Micro-Elisa la mol Thu nghl$m tn,/c tiap, khong
cl;lnh tranh de xac dinh Khang nguyen be mat vinH Viem gan - B, dung
khang the don dong khang HBs la khang the gan ban de phu cac 10 cua
phian nhlla va khang the W huyat lhanh de khang HB s lam cong hop vdl
Enzym Peroxidaza.
: HBsAg
C2 :
Khangthe
~E
+
~~O~--.
Mau
21
~E: Cong hqp
Mau din Thli nghiem dlic;1c u voi Khang the giln ban trong 10 nhl,la. Sau khi
rlia 10 nhl/a de 10<;1i b6 cac chat kh6ng giln, cho them c(>ng hop khang the
men va luc do S8 xay ra phan ung giua c(>ng hc;1p khang the men nay voi
phuc hc;1p Khang the· Khang nguyen da hinh thanh trong Ian u thu nhat,
sau khi ph an ung ket thuc va nia Ian thu 2, tien hilnh u tiep voi dung dich
co bim cua men va chat hien mau, luc do mau S8 xuat hien neu mau Thli
nghiem la Dliong tinh voi HBsAg.
Neu kh6ng thay mau xuat hien co nghia la mau Thli nghiem Am tinh.
b. Cac thuo'c thu kern theo: (cho 96 thu nghi~m)
· Phien dEW dung cho chuan d(> vi Ilic;1ng.
· M(>t phien dec (8 x 12 10) giln khang the don dong khang HBs.
• Dung d[ch c(>ng hc;1P (2 x 5 ml).
· Khang the Khang nguyen be mat virut Viem gan B (de) c(>ng hc;1P voi
Peroxidaza.
· £long kho.
· Chat bilO quan : Gentamycin Sulfat.
· Dung djch nia (1 x 50ml).
· £lem muoi Phosphate dac 10 lan, phai pha loang 10 Ian voi nlioc cat
trong khi dung.
· Chat bao quan: Merthiolate.
· Dung d[ch dem co chat (2 x 3ml).
Dung dich dem Phosphate • citrat dac 10 Ian co chua Hydrogen
peroxyde. Khi dung pha loang 10 Ian.
· Chat hien mau (8 x 5ml).
· 1,2 Phenylenediamine Dihydroclorid tinh the.
· Chung dliong ( 1x 1,5ml).
· HBsAg nglioi pha loang trong huyet thanh nglioi thliong.
· Chat bao quan: Natri azit.
· Chung am (1 x 1,5ml).
· Huyet thanh nglioi thliong pha loang khong chua HBsAg.
· Chat bao quan: Natri azit.
· Dung d[ch dem cacbonat (2 x 6,5ml).
· Dung d[ch dem carbonat dung de hoan nguyen khang the giln ban.
· Chat bao quan: Natri Azit.
c. Cac nguyen v~t Ii{iu can thilt:
· NlioC cat.
· H2S04 4N (dung trong pMn tich)
· Micropipet.
· Noi cach thuy 37°C ± 1°C hOac tu am co d(> am cao.
22
