KHOA HOC CONG NCHE
B U i r C D A L I X A C O j I V H IVGUYEIXI IVHAIM G A Y BEIVH SOC DEIV THAIV TREIV C A Y H O A CIJC [Chrysanthemum
s p p . ) O LAIVI D O M G B A I V G K Y T H U A T PCR
Bui Cdch Tuyen\ Pham Due Todn^, Nguyen Thi Thu^, Pham Thi Hong Phi^ Le Cao Lugng^, H u ^ Van Bi^t^
T6MTAT
Hoa ciic {Cbrysantbemum spp.) thupc ho Asteraceae la cay than thao hang nien va la cay hoa co gia tn kinh le cao t^ii Da Lal - Lam Ddng. Muc dich cua nghien cuu nay la xac dinh nguyen nhan gay ra tneu chiing soc den than tren hoa cue, de tir do dua ra cac giai phap each ly, ngan ngira. phong tranh b^nh hieu qua. Tong sd 20 mSu dutjrc phan lich de nhan dang su hien dien cua Phytoplasma bdng ky thual nested PCR. Ket qua the hten tat ca 20 mau d^u am tinh voi Phytoplasma khi su dung hai cap pnmer Pl/Tmt va R16mF2/RI6mRl, Vi vay, ket qua da chi ra khong co hi^n dien cua Phytoplasma Uen cac miu hoa ciic dupe phan lich. Tuong lu, cac miu hoa ciic ciing duoc phan hch chan doan su hien dien ciia TSWV (Tomalo spotted will virus) bang ky Ihual PCR. Ket qua chdn doan voi cap primer dac hieu PTSW302 va fTSWr>75 da phdt hien 15/18 miu duong tinh voi TSWV. Cac mau ducmg tinh hien dien tr&n cac giong hoa ciic AT. Kim Cuong, Kim Cuong trang, day la cac gidng dupc Uong pho bien tai Lam Dong. Day li\ moi trong nhirng k^t qua buoc diu ket luan TSWV la doi tuong gay ra benh soc den than tren cay hoa ciic tai Lam Dong. Va kel qua nay cung la mpt trong nhung thong tin huu ich, giup cho cong tac quan ly. phong Irir loai benh nay co hi?u quii trong san xuat cay hoa cue tai Lam D6ng.
Tir khod: Cay boa ciic, l<ini Cuong. AT, Phytoplasma. TSW T.
I.DATVANDE
Hoa ciic {Chiysanlheniuni spp) thuoc ho Asteraceae la cdy than thao hang nien va la cay hoa CO gia tri kinh te cao dupc tr6ng tai Da Lal - Lam Dong. Dien tich cay hoa ciic chieni 37% trong long so gan 8.900 ha cay hoa tai Lam Ddng, chiem ty le cao nhat h'ong co' cdu cay hoa ciia tinh. Tuy nhien, nhung nam gan ddy cay hoa cue hi thiel hai nang do sau benh. Trong do, benh soc den than Id mot trong nhung ddi hrpng gay hai nghiem trpng nhat. benh nay xuat hien hi nam 2018 vd gay ra thiet hai nghiem trpng ve chat luong hoa thuoTig phdm ciing nhu thiet hai cao ve kinh te cua iigu'oi trong hoa.
Theo thong hn hi Chi cue Trong trpt va Bao ve thuc vat tinh Ldm Dong thi dien tich hoa ciic bi nhi^m benh spc den Ihan khoang hon 900 ha vd benh dang phdt tri^n rdt phiic tap, chua co ddu hieu dimg lai. Hien nay, d6i voi cac benh do ndm, vi
' Khoa Moi uuang v:i Tai Nguyen. Tnrcmg Dai hoc Nong Lam TP. H6 Chi Minh
" Vien Nghien cuu Cong nghc Sinh hpc va Moi iruong - Khoa N'dng hoc. Truong Dai hpc Nong Lam TP. Ho Chi Minh
Email: [email protected]
khuan gay ra thi nguoi h-ong khong lo ngai, vi hau nhu nguoi dan da co day dii kien thiic va ky nang trong phong hi nam va vi khuan gay benh tr^n cdy hoa ciic. Tuy nhien, doi voi cac dang benh do viroid, phytoplasma, virus thi nguoi h-ong chua co thong tin day dii v& cdch nhdn dang, vi khong co phuong U?n chdn doan, cung nhu chua co ddy dii thong hn ve cdc bien phap phong hir va each ly nguon benh va con triing moi gioi. Tren the gioi, trong chuong trinh kiem dinh gicing ciia EU, doi voi hoa cue can phai kiem dinh cac doi tupng nhu Chiysantiieinum B carla virus {CBN); Tomato Aspermy Cucumovirus CTAV): Tomato Spotted Wilt Virus CTSWV);
Chiysantiienmni Stimt Viroid (ChSVd) (Krczal 1998). Trong do, dang luu y la Tomato Spotted Wilt Virus CTSWV), day la mpt Tospovlrus va la chi duy nhat trong hp Bunyaviridae gay benh cho thuc vat.
TSWV CO pho ky chti rat rpng, khoang 800 loai thuc vat va hem 80 hp gom ca hai va mpl la mam (Krczal 1998). TS\W cung da duuc ghi nhan gdy ra trieu chung spc den than tren cay hoa ciic b cac nuoc tren th^ gioi nhu Nhat Ban, An Dp, Argenhna (Bo va cs, 1995: Matsuura va cs, 2007; Renukadevi va cs 2015;
Senthilraja va cs, 2018). Hien nay co khodng 14 lodi Tospovirus gay benh cay (Brian va cs, 2010). Trong
NONG NGHIEP VA PHAT TRIEN NONG THON - KY 2 - THANG 10/2019 29
s o d o . ngoai T S W ^ dd d u o c c h i n h thtic phdn Ioai vao l(i</unini>. c o n CO m p t virus dupc cho Id t h u o c nhom ndy {Chnsanthemum Stem .\ccrosis Virus - C S N \ ' ) T n ? u c h i i n g s o c t h a n cdy h o a cue h-en d 6 n g r u p n g tgi Ldm Diioj^ voi cdc vet s*.c chet hoai c h o thdy k h d n a n g h o a c u e bi virus T S W \ ' hoac CSNA g a y hai. h o a c c i i n g c o k h d n a n g la do Ph\loplasma.
T u y n h i ^ n , tai Lam D o n g viec ki^m dinh cdc b e n h t r e n cdy con ciia h o a ciic c o n h a n ch& nen viec phdi h i e n b e n h h-en r u p n g s a n xudt t h u d n g gap nhieu kho k h a n U o n g k h a u p h o n g t r u va cdch ly b e n h lay Ian.
Do vdy, m u c dich c u a n g h i e n cuu nay Id xac dmh n g u y 6 n nhdn gdy ra tr\^u c h i i n g b e n h sue den than tri'ii cdy h o a c u e . d e l u d o dira r a cdc giai phdp each
ly, n g a n n g u a , p h o n g t r a n h b e n h i i u i ^'i- •'-''^"' ^' d i n h d i e n h c h s a n x u a t cdy h o a c u e lai L ' i P ^ " ^ '
2. VAT UBJ VA PHUDNG PHAP 2 . 1 . V d t l i § u
M a u cay h o a ciic d u o c t h u t h a p tai cac n h a v u o n U-ong h o a ciic tai t h a n h p h d D a U t , h u y e n Lac D u o n g , tinh Lam D P n g . T o n g s o m d n thu thdP d u n g cho p h a n tich Id 20, b a o g d m cdc m d u chira b i ^ u h i e n t n e u chiing va c a c m d u voi c d c trieu c h i m g s o c d e n than, g a y r a di d a n g h o a , h e o c h e t cay ( b a n g 1, h i n h 1). Mau thu thdp d u p c dira v e p h o n g thi n g h i e m va ti^n h a n h li h-ich D N A , RNA, va Iuu m d u u tii dm 40'C d^ p h u c vu c h o cdc n g h i e n ciru c h u y e n sdu khdc.
, S'lT 1 3 4 5 6 7 8 9 10 U 12 K!
14 15 Ki 17 IS 19 20
Binst 1. Danh sdch cac miu c4y hoa ciic co c ^ tri§u chung soc den than dupc ti Ma so
01 02 03 3A 04 05 5A DK 07 08 09 lOA lOB 11 12 13 15 16 17 L J . S
f)iii chi Thi tran Lac Dinmi;
Phmmg7,TP. DalM Phimng7.TP.DaLil Phuong 7. TP. Da Lat Phuimg 7, TP. Da Lal PhmVng7.TP. Dabil Phuong 7. TP. Da Lat i'liutmg 7. TP. Da Lit Phirung 7. TP. Da Lal Phuong 7. TP. Da Lat Phuong 7, TP. Da Lat Phmmg 7, TP. Da LiU Phuong 7. TP. Da Lat Phucmg 7. TP. Da Lat Phuong 7, TP. Da Lal Phu(ms;7.TP. DaLat Phuong l l . T P . Da Lat Phuong 11. TP. Da U t Phirong l.TP. Da U t Phirtmg l.TP. Da U t
Trieu chung S<,K than, la vang Soc den than Soi- den than Mau canh hoa Soc den than Soc den than Mau canh hoa Soc den than Sgc den than Spc den than Soc den than Mau khoe Cay nho. vang la Thap cay. vang la Soc den than Soc den than Spc den than Soc den than Soc den than Soc den tll;in
Ghi chii gi6ng AT
AT AT AT
Kim Cuong vang Kim Cuong vang Kim Cuong vang Kim Cuong vang Kim Cuong trang Kim Cuong vang Kim Cuong vang Kim Cuong vang Kim Cuong vang Kim Cuong vang Kim Cuong vang Kim Cuong vang Kim Cuong vang Kim Cuimg vang Kim Cuimg vang i^ini Cu'mg vang
u thap ChibSu
V / P V / P V / P P V / P V / P P V / P V / P v / p V / P v / p v / p v / p v / p v/p v/p v/p
~v7f' Ghichu: V: nnis: P Pbi-taplasnia:mau 03 va 03A tren cimg 1 cay lav iiuhi 2 hi, phan ' W ^ dd mau 03 la mau la. 03.A la mau canh hoa: miu 05 va iC.-t cung Hr hai ho phan khac nhm, h li T '™"-*'
-. - ^3 hoa tren cung mot cay
M) N O N G NGHIEP VA PHAT TRIEN NONG THON - KY 2 Tl-;,
•10/2019
KHOA HOC CONG NGHE
Hinh 1. Tri^u chiing b§nh spc den thdn tr6n cdy hoa ctic du(jc thu thdp t?ii Ldm Ddng 2.2. Phiremg phdp nghifin ciiu
2.2.1. Li tiich DNA vaphan ung nested PCR phit hien Phytoplasma
DNA tong so cua cac mdu cdy hoa ciic duoc Iy trich bdng kit ly trich DNA long sd cua hang Promega, My. Quy trinh Iy trich duuc tien hdnh theo nhu huong ddn cua nha san xuat. DNA Iy trich tir cac mdu hoa ciic nghi ngu nhifem Phytoplasma dugc sii dung lam khuon DNA de thiet lap quy trinh nested PCR. O PCR buffc 1 dupc tien hdnh theo tham khao tu nghien ciiu ciia Smart va cs (1996) voi cap primer Pi/Tint- San pham khuech dai cua PCR buoc 1 cb kich thuoc khoang 1,6 kb. Ti^p theo, san pham PCR buoc 1 dupc pha loang 10 ldn de su dung ldm khuon DNA cho PCR buoc 2. Quy h-inh PCR buoc 2 dupc dua tren nghi&n cuu ctia tdc gia Gundersen va Lee (1996), voi cap primer R16mF2/R16mRl. Kich thuoc DNA dupc khudch dai boi PCR buoc 2 khoang 1,4 kb. Sau khi thuc hien nested PCR (PCR buoc 2) san phdm PCR dupc dien di tren gel agarose 1%, hieu dien the 80 V. cuong dp dong dien 400 mA, ti-ong 30 phiit. Gel dupc nhupm va quan sdt ducri den tia UV.
Vi$c danh gid mdu cdy hoa ciic thu thap co su hien di6n ctia Phytoplasma dua vao viec xuat hien bang dich voi kich thuoc khoang 1,4 kb cua san pham PCR buoc 2.
2.2.2 Ly tiich RNA vi phan ung PCR phit hien TS^^
RNA tong sd cua cac mdu la cay ciic dupc ly tiich bang kit Iy h'ich RNA (Gene Jet Plant RNA
Purification Mini kit) cua Hang Thermo Fisher Scientific, My. Quy trinh ly trich dupc ti^n hdnh theo nhu huong ddn cua nhd sdn xuat. RNA ly trich tir cdc mdu cay hoa cue dupc su dung lam khuon cho quy trinh cDNA- Quy trinh cDNA dupc thuc hidn theo bp kit CTetro cDNA Synthesis Kit) cua Hang Thermo Fisher Scientific, My. San pham cDNA dupc Idm khuon mdu cho PCR nhdn dipn virus TSWV. Primer diing de chdn doan TSWV dupc thuc hi§n theo nghien cuu ciia Vaira va cs, 1995 (13). Trong do primer duoc str dung Id PTSW302 (5- GGGTCAGGCTTGTTGAGGA-3') va PTSW575 (5TTCCCTAAGGCTTCCCTGGTG-3'). Sau khi thuc hien phan ling PCR, san pham PCR dupc dien di tr6n gel agarose 1%, hieu dien the 80 V, cuong dp ddng dien 400 mA, trong 30 phiit. Gel dupc nhupm va quan sat duoi den tia UV. Vi6c danh gia mau thu thap cd su hien dien cua TSWV dua vao viec xuat hien bang dich vdi kich thudc doan khu&h dai khoang 273 bp cua san pham PCR.
3 . KET QUA VA n u o LUAN
3.1. K^t qua nested PCR chdn d o ^ Phytoplasma txba. cay hoa ciic
Nested-PCR dupc ddnh gid la mot trong nhiing phuong phap gia tang dp nhay, d6 ddc hieu va dupc xem la phuong phdp hang dau de nhan dang Phytoplasma khi chiing hien di&n d' mtic do thdp trong mach ddn cua cay chu hay cd chat tic che trong mdu ly trich (Gundersen va cs, 1996). Trong nghien ciiu ndy, long sd 20 mdu hoa cue duoc ly trich DNA
NONG NGHIEP VA PHAT TRIEN NONG THON - KY 2 - THANG 10/2019
'jJl®/ l©it^ll^-i'!^i ',V. IMA!:;:. 1)L;<II V-J
d r p h a n Uch c h a n d o a n s u h i e n dien P h j l o p l a s m a ( b a n g 1, h i n h 1). Cac m a u DNA lung s o s a u k h i d u g c ly Uich d a d u o c d i i n g lam k h u o n d e c h a y n e s t e d PCR vol hai c a p p n m e r P l / T i n l va R 1 6 m F 2 / R 1 6 m R l . Ket q u d phdn Uch Uen g e l a g a r o s e s a n p h a m PCR buoc 2
B a n g 2. K^t q u d chin dodn s u h i e n d i ^ T _ c u a j n i v t Q p l ^ m a va T S W V h-^n c a y ho,
cho thdy h a u n h u c a c m d u k h o n g ili.^>' ^"'' ^^ ' , hicn d o a n k h u e c h d g i 1.4 k b wvfi ^^^ ' P h \ i o p l a s m a t r e n c a y h o a c u e (ban.c -• 'i''^'^-; , vav ket q u a d a c h i r a r d n g k h o n g CD s u h i c " du
phan hi--h.
P h W o p l a s m a h'en c a c m d u h o a ciic diroc (
S T T 1
-
4 5 6 7 8 _ 9 10 11 12 13 M 15 16 17 18 19 20
M a so (11 ('2 0 3 _ 3A _04_ _ 1 05
SA 06 07 08 09 lOA I OH 11 12 13 15 16 17 18
T r i ^ u chirng Soc than, la vaiit;
Soc d e n thdn S()c d e n than M a u c a n h h o a SDC d e n ihan Soc dijn tlian M a u c a n h h o a Soc d e n thdn Soc d e n than Soc d e n thdn Sue d e n than M a u k h d e Cav nhd. vdng la
l l i a p c a v . vang ia SDC den than Soc d e n than Sue den than Soc den than Soc den than Soc den than
_ Ghi c h u giong __ __
A T ^ _ _ _ _ _ _ _ _ .•\T
.AT
Kim Cuonj,Maiig Kim C u o n g vang Kim C u o n g v a n g Kim C u o n g Uang Kim C u o n g v a n g Kim C u o n g vdng Kim C u o n g v a n g Kim C u o n g v a n g Kim Cirong v a n g Kim Cucmg v a n g Kim C u o n g v a n g Kim C u o n g v a n g Kim C u o n g v d n g Kim C u o n g v a n g Kim C u o n g v d n g
1 P h y t o p l a s m a Am t m h A m tinh A m linh A m t m h A m tinh A m tinh A m tinh A m tinh A m t i n h A m tinh A m tinh A m tinh A m t i n h A m tinh A m tinh A m tinh A m tinh A m tinh A m tinh A m tinh
T S \ \ ^ 1 D u u i v : > i i i l i _ _ i nmniL: t i n h _ _
Duow: tinh___
Khoii>: Dha:itich_
Uirong'inh _ Duong Unh Kliong phdn tich
Duong tinh Duong tinh Duoiig linh Duong lmh Am tinh Am tinh Am tinh Duong tinh Duong tinh Duong tinh Duong tinh Duong tinh Duong tinh
Hinh 2. K^t qua difen di san phdm PCR nhdn dang su bign dien Phytoplasma tren 20 mdu hoa c g dm; (+): ddi chiing duong; cdc s«
o r d^n "Ma so 18" cua bang 1
a cue, }j[. ^ chuan H\perLadder Ikb; (-): doi chung dm; (+): ddi chiing duong; cdc sd hi 1 d^n 18 hrong ting voi ten "Ms A
NONG NGHIEP VA PHAT TRIEN NONG THON - KY 2 - THANG l0/2n
KHOA HOC CONG NGHE
3.2. K^t qua PCR chdn dodn virus TSWV h-^n cdy hoa ciic
Ky thudt PCR duoc danh gia Id phuong phap hieu qua. nhay va cho kel qua chinh xac vi su dung primer dac hieu vd dupc xem la phuong phdp co uu the de nhdn dang virus khi chung hien dien d- muc do Ihap trong mo cua cay chti. Tdng sd 18 mau cay hoa ciic bao gdm nhiing cdy chua bieu hien trieu chiing va cd cdc trieu chiing spc den than, heo den than, hoa bien dang da dupc thu thap (bang 1, hinh 1). Cac mdu RNA long sd sau khi dupc ly trich da dupc su dung Iam mdu cho phan irng cDNA. Ket qua san pham cDNA dupc dung Iam khuon de chay PCR vdi primer dac hieu PTSW302 vd PTSW575. Kel qua dien di tren gel agai-ose san pham PCR cho thay cac
mdu xudt hien bang dich voi kich tliiioc khoang 273 bp khi ddi chieu vdi Uiang 100 bp. Kich tiiuoc ciia sdn pham khuech dai hoan loan tuong ung voi nghien ciiu ciia Vaira va cs (1995), khi sir dung primer PTSW302 va PTSW575 de nhan dang va phat hien TS^\'\' tren mdu la cay thuoc id Nicotiana benthamiana dupc chiing TSWV.
Ket qua phdt hien 15 h-en tdng sd 18 mdu cay hoa cue dupc thu thap duong tinh voi TSWV (bang 2). Cdc mau duong tinh cd chung trieu chiing spc den than, than cdy bien dang, cay bi cong veo. trang va cdnh hoa mdt can ddi. Voi cdc dieu chiing spc den than, lain cho cay kem phdt trien dan ddn chet thdn va chet cay.
Hinh 3. K^t qua di^n di sdn phdm PCR nhdn dang su hi^n di^n TSWV tren 18 mdu hoa ciic dupc phdn tich. M:
thang chuin HyperLadder Ikb; cic sd tir 1 ddn 18 tuong ung vm tdn "Ma sd 01" ddn "Ma sd 18" cua bing 1 Nhin chung trong 18 mau cay hoa ciic bi benh
dupc tiiu tliap a cac dia diem kliac nhau, cd 15 mau duong tinh vdi TSWV. Tuy bieu hien benh ben ngodi da dang, khong gidng nhau nhimg kdt qua phan tich cho thay phan ldn nhiem TSWV. Jane va cs (2018) da ghi nhan cd nhieu loai vii-oid. viiiis, vd phytoplasma gay benh U-en cay hoa cue. Trong dd tac nhan gay hai ndng la vuus TSWV - Tomato Spotted Wilt virus gdy ra. Tuy vay, a Viet Nam chua tim thay bao cao nao da dupc cong bd ve benh virus tren hoa ciic (Vu Trieu Man va cs, 2018). Nhu vdy, da phat hien 15 mdu trong tdng so 18 mdu thu thdp cho ket qua duong tinh vdi TSWV. DiSu nay da khSng dinh sy hien dien cua TSAW tren cac mdu nghi ngd. Hon the nua, kdt qua cua nghien ciru nay la mot ti'ong nhiing bao cdo dau tien ve su hien dien cua TSWV tren cay hoa cue d Lam Ddng bdng ky thuat PCR.
4 . KETLUAN
Ke't qua nghien cuu khong phat hien su hien dien ciia Phytoplasma tren tat ca cdc mdu hoa cue dupc thu thdp chan doan. Tuy nhien. ddi vdi TSWV tili ket qua the hien tdng sd 15/18 mau cay hoa ciic thu thap a Lam Ddng duong tinh voi TS\W. Do do.
ket qua ndy buoc ddu kel ludn TSWV la tac nhan gay ra trieu chiing benh dom spc den than tren cay hoa ciic. Cac mau cdy hoa ciic duong tinh voi TSWV tren cac gidng dupc trong phd bien b Lam Dong nhu AT, Kun Cuong trdng, Kmi Cuong, Ke't qua nay Id Ihong Un buoc ddu rat htru ich cho cong tac quan Iy va phdng trir benh spc den than tren cay hoa ciic tai Lam Dong.
LOI CAM ON
Nhom tic gii gui Idi cim an chan thinh den tat ci Quy ddng nghiep & cic dia phuong noi da thu thap mau hoa cue tai Lam Ddng va quy ddng nghiep tai Tnrdng Dai hoc Nong Lam thinh phd Hd Chi Minh da tan tinh giiip da vi hd ti'a tivng sudt qua trinh thuc hien nghien ciru niy.
TAILEUTHAM KHAO
1. Bo, E. D., Ronco, L, Alippl, A. M. and Fernandez, R., 1995. Tomalo spotted wilt virus on Chiysanthemmn in Argentina. Plant Disease, 79(5), pp.538-538.
2. Brian W. J. Mahy, Marc H. V. Van Regenmortel (2010). Desk Encyclopedia of Plant and Fungal Virology. Academic Press.
NONG NGHIEP VA PHAT TRIEN NONG THON - KY 2 - THANG 10/2019 33
3. (mndersi-n D. E. Lee I. NL S^haff D. .A.
Hamson \ .A Chang C. J. Da\ns R. E. Kin^bun D T., UM(i Gt'ncnomic diver-^ity among plnioplasma strain-^ in 16s rRN.-\ group I i.-V-ier VeUow<; and related phyloplasma) and HI (X-Discasc and related phytoplasma). Intemahonal journal of systematic bactenology, -16, 64-75.
4. Gundersrn D. E va Lee I. M, 1996.
Ullrasensitive detection of phuoplasina by nested- PCR assays usmg two universal pairs. Phytopalh.
Mcdit. 35. 144-151.
5. Jane C. Trolinger. Robert J. McGovem, Wade H. Elmer. Nancy A Rcchcigl and Chnshne M Shoemaker (2018). l)lst•a^cs of Chrysan tliem um.
Trong: Handbook of Illinois" Crops Diseases.
Spnnger publisher.
6. Krczal G. (1998). \irus Certificahon of Ornamental Plants - the Europian Strategy. Trong:
Planl Vuus Disease Control. APS publisher.
7. Matsuura, S.. Kubota, K and Okuda, M..
2007. First re[)ori of Chrysanthemum stem necrosis vims on ChrysanUicmums in Japan. Planl disease, 91(-l).pp.468-l(i8
8 Renukadevi, P., Nagendran, K., Nakkeeran.
S.. Karthikeyan. G., Jawaharlal, M.. Alice, D., Malalhi. V G. and Pappu. H. R, 2015. First report of
r.'ii!.!'" spoiled Clir>-;inilK'inum ii PP.IUK)-li;!0,
wilt India.
vmis Plant
of
\akkeei-;in S..
.nukadevi P., Spotted Wilt s Travel fi'om si,.,n Cuttings ppl 13.
, ^_ L •'' r-'iiiist and ri.isnia-^i'ccific PCR of tiie 16S-23S rRNA and Envuoninental isd
Scntliilraja C , Malathi V t.
Sivakumar U., Suganthy .\I, and (2018). Charactei-ization of Toni.n Mrus hifecting Chrysanthemum and Motiier Stock Plants to N^M Generation. J Mycol PI Patiiol
10. Smart C. D, B. Sch.-.
L J Guerra., 1996. Pli."
Pnmers Based on Sequence:
Spacer Region. Applied Microbiology, 62. 29SS-2993.
U. Vu Tieu Man. Nguyen Van Tudt, Biii Cach Tuye'r Pham Van Kim (2018). Benh hai cay trdng
\it't Nam. Nha xudl ban Hpc vien Nong nghiep, Ha X6i.
12. \'aira AM, Semeria L, Crespi S, Lisa V, Allavena A. Accotto GP (1995). Resistance to Tospoviruses in Nicotiana benthaniiana ti-ansfonned v«tii tiie N gene of Tomato Spotted Wilt Virus:
CoiTelalion between h-ansgene expression and protection in primai-y transformants. The American Phytopathological Society, 8 (1), 66-73.
INITIAL IDENTIFICATION OF PATHOGEN CAUSING BLACK-STREAKED STEM DISEASE OF Chrysanthemum spp. IN LAM DONG PROVINCE USING PCR TECHNIQUE
Bui Cach Tuyen, Pham Due Toan, Nguyen Thi Thu, Pham Thi Hong Phi, Le Cao Luong, Huynh Van Biet Summary
Chrysanthemum spp belong to the Asteraceae family, an annua! herbaceous planl/flower with high economic value grown m Da Lat city and vanous areas of Lam Dong province. Viel Nam. The purpose of this stud) was to identiij' pathogen(s) causing black-streaked slem disease on Chrysanthemum, that suggest solutions for effective disease isolation and prevention. A total of 20 samples were analyzed to Kk'iitily die presence of Phytoplasma by nested PCR Results showed that all 20 samples were neirafive to Phytoplasma using Pl/Tint and R16mF2/R16mRl primer pairs proving that Phyloplasma is nm \A esent in the analyzed Cho'santiieniuni samples. Similarly. Chrysanthemum samples were also diagnosed for the presence of i'S\V\' by PCR. 15/18 samples detected positive with TSWV using specific primers PTSW302 and ITSWSy.T 'rS\\'\' were present on Chrysanlbenmm vaneUes AT, Diamond and White Diamond, which .ire commonly grown in Umi Dong. This result is among the flrst ones proving that TSWV is the virus causing black-streaked slem disease on Chrysanthemum plants This result is a useful information for effective managemem program of this disease on Chrysanthemum in Lam Dong province.
Keywords: Chrysanlhenmm. Diamond. .A T. Pin-toplasma. TSWV.
Ngudi phan bi^n: GS.TS. Nguyin V3n Tudt Ngdy nhdn bdi: lO/S-^JoiO
Ngay thong qua phdn bien: l',V9/2019 Ngav duyet dint.; . ' -I'l'-'
N O N G NGHIEP V A PHAT TRIEN N O N G THON - KY 2 - THANG ]^i^
