KHOA HOC KY THUAT THO Y TAP XXVI SO 6 - 2019
XAC DINH flOC t a c CUA VIRUS GAY BENH TIEU CHAY CAP TREN LdN CHUNG KTY-PEDOI DUCfC PHAN LAP TAI MIEN BAC VIET NAM
Truang Quang Lam, Nguyen Thi Lan, Nguyen Huu Nam, Nguyen Thi Ngpc, Nguyin Thi Hoa, Nguyin Thi Yin, Ngo Thi Hanh Khoa Thuy, Hoc Vien Nong nghiep VietNam
TOM
T A TTrong nghien ciiu nay, chiing t6i da tien hanh nghien ciru dgc luc cua chung vims KTY-PEDO 1 tren lgn 5 ngay tuoi. Lgn thi nghiem am tinh vai khang thi khang virus PED dugc gay nhilm chiing virus KTY- PEDOI vai heu 2 ml/con qua duong uong vai hieu gia 2xlO^TCID-Q/ml. Sau gay nhilm, lgn thi nghiem dugc theo doi ve trieu chimg lam sang va cac ton thuong dai the va vi the a m6t s6 co quan trong 15 ngay ke tir ngay gay nhiem. Ket qua nghien ciru trieu chiing lam sang da chi ra rang lgn sau gay nhiem co trieu chiing s6t, non mira, mat nuoc, gay com va dac biet la tieu chay phan long, mau vang. Ket qua xet nghiem mau phan lgn bang ky thuat RT-PCR cho thay su hien dien ciia chiing vims KTY-PEDO 1 trong phan sau 1 ngay gay nhiem. Ton thucrng dai the chu yen a lgn dugc gay nhiem voi chiing vmis KTY-PEDOI la mgt non cang ph6ng, thanh rugt mong, xuat huyet, ben trong rugt chiia djch long mau vang. Da day cang phong, chira thirc an kliong tieu. Cae hach lympho eo bieu hien sung, xuat huyet. Benh tieh vi the cho thay sir bong troe, diit nat cac ]6ng nhung mgt, sir xuat huyet, tham nhiem te bao viem, hach lympho xuat huyet.
Ket qua nghien ciiu cho thay chiing vims KTY-PEDOI co kha nang gay benh cho lgn thi nghiem voi bieu hien ve trieu chiing lam sang, bien doi benh tich dai the va vi the giong voi lgn mac chiing vmis PED dgc luc cao gay benh tu nhien dugc ghi nhan tai mien Bac Viet Nam
Tir khoa: Chiing virus KTY-PEDOI, dgc luc.
Determination of virulence of KTY-PED01 virus strain isolated in the North, Viet Nam
Truong Ouang Lam, Nguyen Thi Lan, Nguyen Huu Nam, Nguyen Thi Ngoc, Nguyen Thi Hoa, Nguyen Thi Yen, Ngo Thi Hanh
SUMMARY
In this study, we investigated the virulence of KTY-PEDOI virus strain in tlie piglet at 5-days old The experimental pigs (negative with PEDV antibody) were infected with PED virus strain.
Each experimental pig was orally inoculated with a dose: 2 ml of 2x103TCID5/ml (2ml/pig). After inoculation, the experimental pigs were recorded for the clinical signs, gross and histological lesions of some tissues in 15 days post inoculation. The studied results demonstrated that the infected pigs with PED virus strain presented the clinical signs and symptoms of fever, vomiting, dehydration, rough hair, thin flanks and diarrhea. The result of testing fecal samples through RT-PCR technique indicated that the KTY-PED01 virus strain was presented in the feces of the infected pigs at 1 day post inoculation. The gross lesions were consistently observed, including intestine inflammation, hemorrhage, thin wall and bloating stomach, containing undigested food, and kidneys with spotting hemorrhage, lung inflammation. The lymph nodes were characterized by swelling and hemorrhage of the pigs infected with KTY-PEDOI virus strain. The histological lesions included villous atrophy and lymphocytic inflammation in the intestine, hemorrhage, inflammatory cell infitration, haemorrhagic iymph nodes. The results of our study showed that the KTY-PED01 virus strain is capable in causing severe disease in pigs with clinical signs, symptoms, gross lesions and histological lesions were similar to those of highly pathogenic PED virus strain identified previously in the North, Viet Nam.
Keywords: KTY-PEDOI virus strain, virulence.
KHOA HOC KY THUAT THU Y TAP XXVI SO 6 - 2019
I. DAT VAN DE
Benh tieu chay cap tren lgn do vuus PED (Porcine Epidemic Diarrhea Virus) thuoc giong Coronavirus, hg Coronaviridae biing phat dau tien tai Bi va Anh nam 1971 (Oldham J, 1972), sau do nhanh chong lay lan va xay ra ph6 bien 6 cac quoc gia chau Au khac nhu Dire, Phap, Ha Lan... va a chau A nhu Trung Qu6c, Han Quoc, Nhat Ban, Thai Lan vai ty le chet len dgn 50 - 90% 6 Ign con nho hon 2- 3 tuan tuoi (Chen et ai, 2014; Song va Park, 2012). Nam 2013, dich PED bung phat tai My (Stevenson et ai, 2013), gay thiet hai h-en 7 trieu lgn trong nam d§u tien c6 djch (Cima, 2014). Day la benh truyen nhiem, lay lan nhanh, gay thiet hai ldn voi dac diem xay ra tren lgn 6 tat ca cac liia tuoi vai trieu chiing lam sang chii yeu gom; chan an, n6n va tieu chay. Ty le lay lan va tu vong O' lgn con duoi 7 ngay tuoi rat cao, co the len toi 100%
bai tieu chay va mat nuac nang.
Ci Viet Nam, benh tieu chay cap a lgn lin dau tien dugc phat hien vao nam 2008 tai tinh D6ng Nai (Toan effl/.. 2011) va bao cao tai mien B5c lan dau tien vao nam 2012 (Nguyen Van Diep, 2012). Theo cac nghien ciiu da c6ng bo, benh tieu chay cap xay ra tren lgn mgi Ilia tuoi.
Benh nhanh chong Ian rgng, gay anh huong Ion den kinh te chan nuoi lgn do lam tang ty le benh, ty le chet cao, dac biet tren lgn con theo me tir 50 - 100%. Hien nay, viec dieu tri cho dan lgn hi mac PED gap rat nhieu kho khan do chua co che pham dieu tri dac hieu; de ngan chan va tien toi kh6ng che dich benh thi viec dimg vacxin tir chinh chiing virus phan lap tai Viet Nam cho dan lgn dugc xem la giai phap toi uu. Cac nghien ciru truoc do tai Vien Thii y, Trung tam chan doan Thii y Trung uong, va phong thi nghiem trgng diem CNSH thii y - Hgc vien Nong nghiep Viet Nam da xac dinh dugc cac ca nhiem PED tren lgn a phia Bac Viet Nam, tuy nhien cho toi nay chua co bat ky c6ng bo rong rai nao ve kha nang gay benh ciia chitng virus PED phan lap, De phuc vu cho cong tac san xuat vacxin thi viec xac dinh kha nang gay benh cua chimg virus PED phan lap la viec lam rat can thiet. Do
do, chiing toi tiSn hanh nghien cmi de tai ''Xdc dinh doc luc cua virus gdy benh tieu chdy cap tren lan chiing virus KTY-PEDOI duac phdn lap tai mien Bdc Viet Nanf\
II. NOI DUNG, VAT LIEU VA PHlTONG PHAP NGHIEN ClTU 2.1. Noi dung nghien cihi
Xac dinh dugc kha nang gay benh cua chiing virus KTY-PEDOI tren lgn con theo me.
Xac dinh trieu chung lara sang, benh tich dai thg, benh tich vi the cua lgn con theo me dugc gay nhigm thuc nghiem bang chiing virus KTY- PEDOI .
2.2. Nguyen vSt lieu
Dgng vat: Nghien ciiu dugc tien hanh trgn 2 16 lgn: 16 thi nghiem (5 lgn: TNI, TN2, TN3, TN4, TN5) va 16 doi chiing (5 lgn: DCI, DC2, DC3, DC4, DC5), tit ca la lgn con theo me (4 - 5 ngay tuoi) thugc gi6ng Yorkshire. Truoc khi tien hanh gay nhiem, toan bg lgn thi nghiem dugc lay mau kigm tra khang the khang PEDV bang phuong phap ELISA va kiem tra virus PED bing phucmg phap RT-PCR. Lgn dugc chgn lam thi nghiem phai co kgt qua am tinh vai khang the khang virus PED, am tinh vai virus PED va mgt so virus khac nhu: TGE, Rotavirus, Circo virus type 2 (PCV2), hgi chimg roi loan h6 hap va smh san o lgn (PRRS), dich ta lgn (CSF) va Porcine Deltacoronavinis (PDCoV).
Chiing giong virus KTY-PEDOI dugc phan lap tir rugt non ciia lgn con theo me c6 trieu chimg lam sang va benh tich dien hinh ciia lgn mac PED a tinh Hung Yen nam 2017. Hieu gia cua chung virus KTY- PEDOl dat 10^ TCID^y ml. Chiing virus dang dugc luu giii va bao quan tai Phong thi nghiera trgng digra Cong nghe sinh hgc Thil y, Khoa Thii y. Hoc vien N6ng nghiep Viet Nara.
2.3. PhiroTig phap nghien ciru 2.3.1. Phuorng phdp RT-PCR
Bao g6m cac buoc tach chigt R -\ cua virus
KHOA HOC Kt THUAT THU Y TAP XXVI SO 6 - 2019
va cac buac thirc hien ky thuat RT-PCR. RNA ciia virus dugc tach chigt bang kit QIAamp cua hang Qiagen (Diic) theo huong dan cua nha san xuat. Phan img RT-PCR dugc thuc hien theo quy trinh ciia hg kit.
Cap ra6i dugc sir dung la: Cap m6i PMl (moi xu6i: 5' CCC CAG TAC TGT TAT TGA CGT ATA AAC 3', m6i ngugc: 5' GTT TAG ACT AAA TGA AGC ACT TTC 3') (Song et al., 2006), cap m6i nay cho phep xac dinh ragt phan thugc doan gen M cua virus PED co kich thuoc 751 bp. Tien hanh khugch dai san phSra trong may PCR vai chu ky nhiet: 50''C: 30 phiit, 9500: 2 phut; 35 chu ky (950C: 30 giay, SZ^C:
60 giay, 72''C: I phut); 72"C: 10 phiit, 4"C: kgt thiic. Dien di kigm tra ket qua RT-PCR a hieu dien the 100 V trong 30 phiit. Quan sat va chup anh kgt qua dien di san pham RT-PCR trgn may chup anh gel.
2.3.2. Gay benh thuc nghiem
Bo tri thi nghiem: Lgn thi nghiem dugc chia thanh 2 16: Lo thi nghiem (L6 TN) g6m 5 lgn (ky hieu tir TNI den TN5) va 16 d6i chiing 5 Ign (kyhieuDCI dgnDC5),
Thi nghiem: Khu thi nghiem dugc dam bao CO do an toan sinh hgc cap II: Ve sinh sach se, thuong xuyen phun thuoc sat trimg va v6 tning, cac dung cu sii dung dimg quy dinh, c6 mang loc khong khi ngan chan mam benh tii bgn ngoai vao va tir phong gay nhigm sang phong d6i chiing. Nhiet do phong duy tri o 30 - 32''C, do am kh6ng qua 90%i, lgn dugc cho an 3 biia/
ngay (sang, trua, t6i) theo khau phan cua lgn con theo me.
Gay nhiem cho lgn: Nhom lgn thi nghiem dugc gay nhigra chung virus KTY-PEDOI qua duang uong bang syringe voi lieu 2 ml/con, hieu gia 2xl0^TCID5yml.
Theo doi lgn sau khi gay nhiera: the trang, tinh hirih siic klioe, tinh trang an uong bang quan sat va ghi chep hang ngay. Theo doi than nhiet cua lgn thi nghiera bang each do bang nhiet kg a hau m6n ciia lgn 2 lan/ngay vao 9 gio va 17
gio. Tien hanh ISy mhx phan hang ngay (ngay 1 d6n ngay 15 sau gay nhilm) d6 kigm tra su c6 mat cua vims trong ca the lgn thi nghiem. Mlu phan dugc bao quan lanh trong thiing da va van chuyen vg phong thi nghiem trong ngay.
2.3.3. Xdc dinh benh tich dai thi vd vi thi Quy trinh m6 kham: Lgn dugc m6 kham theo tigu chuin TCVN 8402-2010 (Bg Klioa hgc va cong nghe, 2010).
Lgn thi nghiem se duoc mo kliam khi chgt trong qua trinh theo doi hoac khi ket thiic theo doi (15 ngay), ghi chep, chup anh de thu thap th6ng tin vg benh tich dai thg. Cac mau co quan tir nhiing lgn m6 kham nhu da day, rugt non, rugt gia, hach lympho, gan, phoi, than... se co dinh trong formol 10% de lam tigu ban xac dinh bien doi vi the.
Lam tigu ban vi thg: Benh pham dugc c6 dinh bing fonnol 10%, diic parafin bing he th6ng chuygn due mo Ur dgng STP 120. Tigu ban dugc cat a do day 2 - 7 \im, sau do nhugra bang phuong phap HE (Haematoxyhn Eosin).
2.3.4. Phuffng phdp nuoi cay te bdo Vero M6i truang diy du DMEM c6 10% FBS dugc lam im trong tu am 37"C trong 30 phiit truac khi lay te bao ra nuoi cay. Giai dong tg bao a nhiet do phong. Ly tam 3000 v6ng/5 phiit dg loai bo dung dich bao quan va giii lai can te bao. Hoa tan can te bao trong 1ml moi truong da chuan bi a tren. Chuygn te bao vao binh nuoi cay chua 5ml raoi truong day dii, Giii' tg bao 6 37''C voi 5% CO,, hang ngay theo doi su phat trign cua tg bao. Cay chuyen tg bao khi te bao rage mgt lap a binh nu6i cay
2.2.5. Phuffng phdp phan lap virus Buac mgt, chuan bi tg bao phan lap: Te bao Vero dugc dua vao nuoi cay trong moi truang va dieu kien thich hgp, raoi truong DMEM (Dulbecco's Modified Eagle Medium) co bo sung 1 D%o FBS (Fetal bovine serum) nuoi te bao a 37°C voi 5%) CO,, khi tg bao mgc vira kin day binh thi tien hanh gay nhilm virus.
KHOA HOC KY THUAT THU Y TAP XX\^I SO 6 - 2019
Budc hai, chuan hi raau phan lap: Mau benh pham la rugt non cita Ign dugc gay nhiem PED dugc xir ly theo quy trinh sau: rugt nghign bang chay coi vo tning pha loang trong DMEM theo ty le 1:5, pha vo' te bao bang may phat song sieu am, ly tam 10.000 vong/lO phut/4"C loai bo can, xu ly khang sinh r6i Igc qua mang loc 0,22 pm, sau do gay nhigra len m6i truang te bao Vero.
Buo'c ba, gay nhiera virus va quan sat kgt qua: Tir cac tg bao da chuan bi a buoc mgt, hiit bo m6i truong nu6i cay va bo sung lOO/il mau phan lap da chuan bj a buoc hai, u trong thoi gian 60 phut a 37''C, sau do bo sung 5 ml ra6i truang phan lap gora (DMEM co chiia tryptose phosphate Broth (0,3%>), dich chigt nam men (0,02%), va trypsin dg o 37''C voi 5% CO,.
Hang ngay theo doi su pha buy te bao bang kinh hien vi soi ngugc. Ngu xuit hien benh tich tg bao thi thu lai virus khi te bao hi pha hiiy dat 80% - 90%) dien tich day ciia binh nuoi. Neu 5 ngay khong thay xuat hien benh tich thi thu lai te bao va m6i truang nuoi cay, d6ng tan 3 lan va tiep tue ciy chuygn cho den khi xuat hien benh tich te bao. Ngu cay chuygn 9 lan lien tiep ma kh6ng thay xuat hien benh tich te bao thi coi nhu am tinh phan lap.
2.3.6. Xu ly so lieu
So lieu dugc xu ly bang phan mgm GraphPad Prism version 7.03.
HI. KET QUA NGHIEN CU U 3.1. Kit qua trieu chung lam sang chu y§u ciia Iffn dirtrc gay nhiem
3.1.1. Than nhiet cua Iffn duffc gay binh thuc nghiem v&i chiing KTY-PEDOI
Sau khi gay benh, ehiing toi tien hanh do than nhiet hang ngay cua Ign vao thoi diem co dinh la 9 gio va 17 gio. Sau do ghi chep lai, xii ly so lieu qua phan mem va dugc thg hien a hinh 1. Lgn s6t ngu than nhiet ldn hon 39,5''C.
Qua hinh 1 the hien nhiet do trung binh cua lgn 0 15 thi nghiem va 16 d6i chiing, co the thay lgn thi nghiem deu co bieu hien sot trgn 40*'C sau 1 ngay gay nhiem va than nhiet van giii 6 mire s6t tren 39,5"C nhung co xu huang giam din trong 3 ngay dau sau gay nhigm. Tir ngay thir 4 dgn ngay 15 sau gay nhiera, than nhiet ciia tat ca cac lgn thi nghiem deu dao dgng a rairc sinh ly binh thuong. Ngay thii 5 sau gay nhilm, CO 1 lgn (TN2) a 16 thi nghiera bi chet va ngay thii 6 CO 1 lgn thi nghiem (TN4) bi chgt, cac lgn bi chgt khi than nhiet 6 miic nhiet sinh ly binh thuong khoang 39,2'*C. Lgn co hien tugng s6t la do virus tan cong vao co thg, pha huy tg bao va san sinh cac chat dgc, kich thich vao trung khu digu hoa nhiet, lara r6i loan trung khu dieu hoa nhiet, gay hien tugng s6t cao, kem theo do la hien tugng met moi, giam an hoac bo an. Lgn d6i chihig c6 than nhiet dao dgng trong pham vi sinh ly binh thuong.
41.0- 40 5- 40,0- 39 5 39 0' 38.5- 38.0- 37 5 37.0- 36 5' 3B 0- 35 5- 35,0-
Hinh 1. Than nhiet trung binh cua Ion sau khi gay benh thwc nghiem v&i chunq vims
KHOA HOC KY THUAT THU Y TAP XXVI SO 6 - 2019
3.1.2. Trieu chung ldm sdng chu yeu cua Iffn duffc gdy nhiem v&i chung virus KTY-PEDOI Lgn thi nghiem va lgn doi chiing dgu dugc theo doi trieu chiing lam sang trong vong 15 ngay, ket qua theo doi dugc ghi chep lai theo fling ngay. Kgt qua cho thiy cac lgn d 16 thi nghiem deu co bieu hien trieu chirng ii rii, n6n, tieu chay, mat nude ciia lgn mic virus PED. Ci 16 thi nghiera trong thdi gian nghign ciru CO 2 lgn chet vao ngay thii 5 va ngay thii 6 sau gay nhilm, chiem ty le 40%) tdng s6 dgng vat d 16 thi nghiem. Trieu chiing lam sang chii ygu ciia lgn dugc gay nhiem vdi chimg virus KTY-PEDO 1 la: Sdt trong 3 ngay dau tien sau gay nhiem, ii rii, met moi, bd an, non, tigu chay phan long vang, da khd, long xo xac, phan vang dinh bet vao hau mon, gay com. Mlic do nang nhe ciia cac tneu chirng gan nhu gi6ng nhau d cac lgn thi nghiem.
Rieng trieu chiing da kho, long xo xac, mat trung sau, bung hop, gay com dugc quan sat thay sau 5 ngay gay nhiem, do lgn tigu chay lign tue dan den hien tugng mat nude va chat dien giai ngn co thg bi suy kiet. Trong khi do lgn ddi chiing van an u6ng va khoe manh binh thudng.
Trong nghien ciiu gay benh thuc nghiera 4 chiing IN 19338/2013, NC35140/2013, NC49469/2013 va
IL2097/2014 cho lgn con theo me 5 ngay tudi, tac gia Chen et al. (2016) da m6 ta mdt s6 trieu chiing cua lgn dugc gay nhigm cac chimg virus tren nhu sau:
0 cac 16 lgn dugc gay nhiem chiing INI9338/20I3, NC35140/2013 va NC49469/2013 diu co hien tu'gng u ru, tigu chay, bd an, mat nude, cac trieu chiing nay nghigm d-gng nhat khoang 4 ngay sau gay nhigm va keo dai din hit thdi gian theo doi. K6t qua nghign ciiu trieu chiing lam sang ciia chung tdi tu'ong hr vdi kgt qua nghien ciiu trieu chiing lam sang ciia tac gia Chen et al. (2016) khi tien hanh gay nhiem 3 chung virus trgn, nhung khac vdi ket qua lam sang ciia lgn thi nghiem dugc gay nhigm chiing IL2097/2014. Chimg virus nay chi gay ra nhimg tneu chimg c6 bieu hien nhe trong khoang 1-2 ngay sau gay nhigm trgn lgn thi nghiem: chi cd 1 lgn thi nghiem bi tigu chay nhe d ngay dau tien sau gay nhiem, Ign kh6ng mat tinh them an, kh6ng hi mat nude va lgn phuc hoi sau 4 ngay gay nhilm IL2097/2014.
Nhdm nghien cim tap trung trinh bay mdt sd trieu chung chinh cua lgn dugc gay nhigm chiing KTY-PEDOI nhu: tieu chay, ndn bang phuong phap cham digm. Ket qua dugc thg hien dhinh 2.
0 5 O 0-
D i e m
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t
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f
—
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12 13 14 15 N g a y Hinh 2. Bieu do the hien gia tri diem trung binh trieu chu'ng tieu chay cua Imi
diPOfc gay nhiim v&i chung virus KTY-PEDOI
Ghi chu: 0: Khong tieu chdy: 1: Tieu chdy it (dang phdn mem, long); 2: Tieu chdy manh (phdn dang nu&c)
KHOA HOC KY THUAT THU Y TAP XXVI SO 6 - 2019
Qua ket qua RT-PCR, chiing toi thay 4/5 lgn d 16 thi nghiem dugc gay nhiera chung virus KTY-PEDOI phat hien dugc virus PED Urong mau phan vao ngay thii 1 sau gay nhigm (chigm 80%.). Kgt qua nay tuong tu vdi ket qua nghien ciin dgc luc cua 3 chitng virus PED nguygn goc US phan lap tai My cua Chen et al. (2016) va ket qua ciia Chang et al. (2017) khi nghign ciiu dgc luc ciia chiing PEDVPT-P5. Tir ngay thii 2 sau gay nhilm den bet thdi gian nghien cim ghi nhan thay tat ca lgn trong 16 thi nghiem dgu duong tinh vdi virus PED trong man phan tieu chay, dat 100%o. Do den ngay thit 5 sau gay nhilm cd 1 lgn (TN2) d Id thi nghiem bi chit va ngay thii 6 sau gay nhiem cd 1 lgn (TN4) chgt, nhiing lgn nay deu dugc rao kham kigm tra benh tich dai thg, kigm tra virus bang phuang phap RT-PCR. Kit qua phan lap va RT-PCR diu duong tinh vdi virus PED, va am tinh vdi cac vims gay tigu chay khac (Rotavirus, TGEV, CSFV, PDoV), chiing to lgn thi nghiera bi chit
diu do virus PED gay ra. Tir ngay thir 7 cho den hit thdi gian thi nghiera, khong ghi I'fhan thgm ca lgn thi nghiem nao bj chit, ket luan sd lgn chit chilm 40% tong s6 lgn thi nghiem. 016 dpi chiing, 5/5 Ign diu khde manh, khong cd bilu hien trieu chiing ciia lgn mac PED va ket qua RT-PCR diu am tmh vdi vuns PED, dieu nay chiing td nhdm nghign ciiu da gay nhiem thanh cdng virus PED cho lgn thi nghiem.
3.3. Nghien ciru bi§n d6i benh ly cua Iffn thirc nghiem
3.3.1. Kit qud benh tich dai the cua Iffn duffc gdy nhiim chung virus KTY-PEDOI
Sau khi gay nhiem virus KTY-PEDOI, cac Ign bi chgt se dugc m6 kham ngay dg quan sat benh tich va nhimg lgn cdn lai ciing dugc mo khara d ngay 15 (sau khi ket thuc thi nghiem) de quan sat, ghi chep lai henh tich dai the. Benh tich dai the tren cac ca quan ciia lgn thi nghiem dugc trinh bay trong bang 1.
Bang 1. Benh tich dai the cua Iori difcrc gay nhiem chung virus KTY-PED01 Benh tich tai mot so c c quan
Ruot Da day Hach lam ba Phoi Co' quan khac cang ph6ng, thanh mong, cang, chifa thifc sirng to, xuat huylt, viem Than sung, xuSt chira dich vang. Xult huyit an khong tieu bSm tim huylt; lach su'ng L6TN
(con) L6DC (con)
5/5 5/5 5/5
0/5
1/5 2/5
Kgt qua cho thay, lgn dugc gay nhiem chung vims KTY-PEDOI co cac benh tich chii yeu nhu: rugt non cang phdng, thanh mdt mdng, cd thg nhin thay dugc bgn trong. Chat chira trong mgt la dich long raau vang, lgn cgn. Ong duong chap cang, mau dd, ndi rat rd. Bgc 16 nigm mac phia trong mgt thay cd hien tugng xuat huyet lan tran. Da day cang phdng, chira thiic an kh6ng tigu, xuat huyet. Hau hgt cac hach diu sung, dac biet la hach mang treo mgt sung rat Idn, va CO hien tugng xuat huyet. Day la benh tich dien hinh nhat ciia lgn dugc gay nhiem
chiing vmis KTY-PEDOI. Ngoai ra con cd cac benh tich khac nhu phdi sung huyet nhe, than xuat huyet diem, lach sung, gan sung huyet, mat cang. Day la nhirng benh tich khdng dac trung vi khdng phai m6 khara lgn nao ciing thiy xuit hien. Khdng quan sat thiy benh tich dai thi bit thudng nao d 16 lgn d6i chung.
Nhu vay cho thay trgn lgn ma chiing toi nghign cuu deu cho kgt qua benh lich dai thi phii hgp vdi cac nghien ciiu eiia cac Xv': tJia Nguyen Van Diep va Nguyen Thi Lan (2; i 31. |gn mic PED c6 tdn thuong dai thg gi6nj: oi ]oii mfic
KHOA HOC KY THUAT THU Y TAP XXVI SO 6 - 2019
TGE: da day trdng rdng do lgn non, 6ng dudng chap kh6ng chifa nhieu dich duong do su kem hap thu d rugt. Cac doan rugt non chiia diy dich, cang ph6ng, thanh rugt mdng
tdi mire c6 the nhin thay ben trong do su teo lai cua tang niem mac, chat chiia ben trong rudt Ign cgn, hach mang treo rugt sung to thanh dai ro ret.
Hinh 7. Benh tich dai the cua lgn thi nghiem tfuw gay nhiem virus PED (A) Da ddy, ruot cdng, thdnh mong. (B) Hach mdng treo rugt simg.
(C) Thdnh ruot mong, xudt huyet. (D) Than xudt huyet diem
3.3.2. Kit qud benh tich vi thi cua iffn duffc gdy nhiim chung virus KTY-PEDOI
Chiing tdi nhan thay rang benh tich vi thg
chil yeu nhat cita ca 5 lgn la nhimg bien ddi d da day, rugt va hach lympho. Cu the dugc trinh bay d bang 2.
Bang 2. Benh tich vi the cua Io'n dwtyc gay nhiem chung virus KTY-PED01
L6TN (con) L6DC (t;on)
Ruot Niem mac bong troc.
Long nhung dOI nat.
Tham nhiem t l bao viem. Xuat huyet
5/5 0/5
Benh tich tai mpt s6 co- Da day
Bieu mo bi dui nat. Xuat
huylt
5/5 0/5
Hach lympho Cac nang lympho
thoai hoa. Hing cau lan tran. Tham nhiem t l bao viem
5/5 0/5
quan Phoi Tham nhilm cac
t l bao viem
2/5 0/5
Co* quan khac Than, lach tham nhilm t l bao viem, xult huylt
3/5 0/5
Kgt qua cho thay, benh tich vi the chu ygu ciia 5 lgn thi nghiem la: Hien tugng xuat huygt, bong troc, dut nat ldng nhung mot va nigm mac da day, tham nhigm te bao vigm a cac ca quan vdi ty le cao. Ngoai ra cdn thay hoai hi tg bao va thoai hoa te bao. Trong do cd 5/5 mau mdt deu CO cac tdn thuong nhu: Niem mac bong troc, long nhung dirt nat va tham nhiem cac tg bao
viem, sung huylt, xuat huygt. Da day d ca 5 mau nghien ciiu deu cd bigu hien bong trdc nigm mac, xuit huyet va sung huyet. Hach lympho cd bigu hien chii yen la xuat huyet va thoai hda cac nang lympho. Phdi trong cac mlu nghien ciiu chil ygu c6 bieu hien xuat huygt, tang sinh td chirc ke, tham nhilm t l bao viem. Cac benh tich nay dgu khdng xuat hien d 16 ddi chiing.
KHOA HOC KY THUAT THU Y TAP XX\'l SO 6 - 2019
Hinh 8. Benh tich vi the cua lgn thi nghiem dwryc gay nhiem virus PED (A) Bieu mo ruot dut ndt (HExlO). (B) Ruot non sung huyit, xudt huyet (HExlO).
(C) Biiu mo da ddy dut ndt (HExlO). (D) Thdm nhiem te bdo viem a ruot (HEx40)
3.4. Ket qua phan lap lai chung virus KTY- PEDOI tren moi trudng te bao vero
Sau khi m6 khara tit ca lgn d Id thi nghiera.
chiing tdi tien hanh thu mau va phan lap lai chiing vims KTY-PEDO 1 firen mdi trudng te bao Vero. Ket qua thg hien d bang 3.
Bang 3. Ket q u a p h a n lap lai c h u n g v i r u s KTY-PED01 t r e n m o i t r y i n g t§ b a o Vero
STT LOT thi nghiem RT-PCR sau phan Igp (P3)
TNI TN2 TN3 TN4 TN5
Ghi chii: P^: D&i cdy chuyin virus tha n; 0: Khong xudt hien benh tich tebao; 1: Xudt hiin binh tich te bdo; + .• Kit qud ducmg tinh vai virus PED
Qua bang kgt qua c6 the thay chiing viras KTY-PEDOI khdng gay benh tich trgn tl bao Vero ngay trong ddi dau tign gay nhiem (PO).
Den ddi cay chuyen thir 1, c6 4/5 mlu gay benh tich trgn tg bao, chiem 80% t6ng s6 mau thi nghiera. Tir ddi cay chuyen thii 2, quan sat thiy tat ca cac mau gay nhigm dgu xuat hien benh tich trgn te bao. Vdi benh tich dac trung la eac tl bao
dung giai mang tao thanh te bao khdng 16 chiia nhieu nhan ben trong (thg hgp bao - syncytium).
Khi xuat hien benh tich te bao, dich nudi cay dugc thu de kigm tra su co mat cita vims PED bang phuang phap RT-PCR. Kit qua cho thiy 100% cac mau dgu duong tinh vdi virus PED, va benh tich xuat hien trgn te bao chinh la do chiing vims KTY-PEDOI gay ra.
Hinh 9. (A) Te bao Vero khd>>
(B) Benh tich cua PEC: I u i r v v »*•*y '
I B l^ pen
KHOA H O C KY T H U A T T H U Y TAP XXVI SO 6 - 2019
IV. KET LUAN
C h u n g virus K T Y - P E D O I cd k h a n a n g g a y benh tren lgn thi n g h i e m t u o n g d u a n g v d i chiing virus c u d n g d d c , v d i trieu chiing v a b e n h tich rat dien hinh c u a lgn m a c P E D . Trieu c h i m g lam sang chii y e u ciia lgn d u g c gay n h i l m la:
b o an, sdt, ndn miia, gay c d m v a dac biet la tigu chay p h a n m a u v a n g , dinh bet d h a u m d n .
B e n h tich dai thg chii y e u cua Ign d u g c g a y n h i l m chimg KTY-PEDOI la m g t n o n c a n g ph6ng, thanh m g t m d n g , xuat huygt, b e n trong mgt chiia dich ldng m a u vang. Da d a y c a n g phong, chiia thiic an k h o n g tieu, xuat huyet. Cac hach lympho, d a c biet la h a c h m a n g treo m g t cd bieu hien s u n g to, xuat huyet.
Benh tich vi thg cua lgn d u g c gay nhiem chimg KTY-PEDOI la: niera m a c m d t b o n g troc, ldng nhung diit nat va hoai tir, cd b i l u hien sung h u y l t , xuat huygt va sir tham nhiem cac te bao viem d h a niem mac, ngoai ra cdn hinh thanh cac khdng bao nam o dau 16ng nhung. Bieu ra6 da day bi b o n g trdc, diit nat, phan tuygn bi p h a buy, xuat huygt.
Cac hach lam ba cd bigu hien sung huyet, xuat huyet, cac nang lympho bi thoai hda. Ngoai ra cdn CO sir thoai hoa, hoai tir xuat huygt d mgt so c a quan khac nhu: gan, lach, than, phoi.
D u a vao cac ket q u a trgn, chiing v i m s K T Y - PEDO 1 CO the d u g c lua chgn d e tien h a n h nghign ciiu sau h o n p h u c vu cho cac nghign ciiu c h g n chiing v i m s tigm n a n g d e san xuat vacxin, thii hieu luc eiia vacxin hoac san xuat cac c h e p h a m sinh hgc phuc v u cho chan d o a n , p h d n g va tri benh P E D .
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