Tap chi Cong nghe Smh hoc 12(2): 221-227, 2014
H O A T T I N H K H A N G K H U A N V A K H A N G U N G THU" C U A C H U N G X ^ K H U A N B I E N Streptomyces variabilis H P 4 1 1
Pham Thanh Huy§n, Bach Thi Mai Hoa, Phi QuySt Tien, Nguyin Phirong Nhue, L6 Gia Hy Vien Cong nghe sinh hoc. Vien Han ldm Khoa hoc vd Cong nghe Viet Nam
Ngay nhan bai: 02.5.2014 Ngiy nhan dang: 12.6.2014 TOM T A T
Chiing xa khuan bien Streptomyces variabilis HP411 dirpc phan lap tir cac mau biin dat ven bd thupc vung bien Dong Bac Viet Nam. Ket qua nghien cuoi dac diem sinh hpc cho thiy chiing xa khuan HP411 thupc nhom xam, khong cd kha nang sinh sac td melanin, sinh sac to mau nau nhat tren moi truong ISP2, Cau tnic chuoi bao tir dai hoi liron song 6 dau, be mat bao tir xu xi. Chiing cd kha nang sinh tnrdng va phdt trien tot d nhiet dp 25- 37''C, pH 7-8, chm dupc ndng do NaCI d^n 10% (w/v) ChSt khang khu§n thu dupc tu: chiing HP411 co kha nang ire ch^ vi khuin Escherichia coli (ATCC 25922), Staphylococcus epidemidis (ATCC 12228) va aim gay bfnh Candida ablicans (ATCC 10231); dac biet la kha nang lire che loai vi khuin khang thuoc Staphylococcus aureus khang methicillin (MRSA) va Staphylococcus epidermidis khang methicillin (MRSE). Ben canh do, chit niy khong c6 kha nang gay dpc \ m dong te bao lanh - dong tS bao thSn Hek293- ma co kha nang tieu diet hen 50% tg bao ung thu bieu mo vii MCF7 o nong dp 70,2 ng/ml, giatn ICsola61,34±l,l (pg/ml). Cac ket qua nhan dupc mo ra triln vpng nghien cuu v^ ti^m nang tim ki6m va san xuit cac chat mdi cd hoat tinh khing khuan vi khang te bao ung thu tir xa khuan bien Viet Nam trong tuong lai.
Tit khod: Khang khuan, khdng ung thie. Streptomyces, vi sinh vgt bien, xg khuan bien, Streptomyces variabilis
MO DAU
N g i y nay, vdi sir phat trien khdng ngirng cua cdng nghe sinh hoc vi sinh vat da tao ra hang nghin loai thudc mdi, tuy nhien van khdng dap ling dil cho nhu c i u di6u tri nhftng can benh mdi nhu ung thu (Berdy, 2005; Lam, 2006), Vdi thuc t6 ngudn thudc chua benh cho con ngudi tir vi sinh vat trong dat hen khdng cdn mdi nira nen cac nha khoa hpc phai tim kiem mot ngudn nguyen lieu mdi cho nganh cdng nghiep s i n xuat thudc ciia the gidi. Cac nghien ciru sdm nhat ve cac chat cd hoat tinh ling dung trong y duoc tir vi sinh vat bien bat d i u v i o nam 1960 (Alphonse, 2002) Tir day da md ra mot hudng mdi de phat trien c i e ngudn chat trign vong tir sinh khOi ciia dai duong. Sau han nua thd ky, n g i y nay tiem nang san xuit thudc tir sinh vat bien van dugc coi l i hudng nghien cuu ldn day hira hen (Berdy, 2005).
Tren thuc t^, khoang 60% cac loai thudc co ngudn gdc tu nhien thi chiem tdi 50% chat khang sinh tu nhien do cd ngudn gdc tit xa khuin (Ravikumar el al, 2010). Da phin cac chat thu dugc hi xa khuin bien cd cac hoat tinh mdi nhu: khang sinh, khing nam, gay ddc te bao, gay ddc thin kinh.
chdng vims, chdng ung thu (Pereira, 2004; Molinski el al., 2009). Trong cac tai lieu da cdng bd, cac chimg xa khuan bien tiem nSng thudc chi Streptomyces phan lap tir vimg bien cd kha nang ire ch€ 2 ddng t6 b i o ung thu vu la MCF7 va MDA- MB-23I (Ravikumar et al, 2012). Chung xa khuin bien S. tropica cd k h i nang sinh ra chat uc che m^nh vdi proteasome salinosporamide A-chat niy hien dang dugc thu nghiem dieu tri ung thu tren ngudi (Fleming et al, 2003); va nhdm sporolide A va B- l i cac chat mdi cila macrolide cd gan them nhdm halogen (Bunchanan el al., 2005).
Trong han mgt thap ky trd lai diy, cac nha khoa hgc d Viet Nam da cd cac cdng bd ve nghien cuu hoat tinh smh hoc ciia cac chat thu dugc tir xa khuan bien nhung chua dugc nghien ciru sau. Do dd, dinh hudng nghien ciru sang lgc c i e chit khang khuin v i chdng ung thu mdi tir xa khuan bien nham tao nguOn cung cap cie chat thay the thudc chua benh hien nay li cin thiet.
Trong bai b i o nay, chiing tdi trinh bay ket qua nghien ciiu hoat tinh khing khuan va khang ung thu ciia chiing xa khuan S. variabilis HP411 phan lap ttr viing bien phia b i c Viet Nam.
221
Ph?im Thanh Huyen el al VAT LIEU VA PHU'ONG PHAP
Vit li|u
Chiing xa khuan bien HP411 cd hoat tinh ddi khing vdi mpt sd loai vi sinh vat ki6m dinh (vi khuin Gram am, vi khuan Gram duong v i nam) dugc phan lap tir viing bien ven bd phia Bac Viet Nam (vT do B i c 19° d^n 21" va Ddng 105" dan 107").
C i e chung vi sinh vat kiem dinh (5 aureus khang methicillin (MRSA), 5. epidermidis khing methicillin (MRSE). Esclienchia colt (ATCC 25922), S epidemidis (ATCC 12228), S typhimurium (ATCC 14028) v i nam Candida albicans (ATCC 10231) nhan tir bd suu tap gidng cua phdng Cdng nghe len men, Vien Cdng nghe sinh hgc.
Moi trtrtmg
Mdi trudng SCA (g/l): Tinh bgt 10; casein 10;
KH2PO4 0,5; MgSOj 0,5 NaCl 30; nudc cit 500 ml;
nudc biln 500 ml; thach 18,0; pH 7,0. Mdi trudng NA (g/l): Cao thit 5; pepton 10; NaCl 5; thach 20;
nudc cat Hit, pH 7. Mdi trudng khoai t i y (g/l): nudc chiet khoai tiy 200 ml; dextrose 20; thach 20, nudc cat Hit, pH 7. Cac moi trudng phan loai xa khuin I S P l , ISP2. . ISP9 theo Shirhng, 1966. Moi trudng M I A S W (g/l): Tinh bdt tan 10; cao men 4, pepton 2;
th^ch 18; nudc biln 1 lit.
Nghieii clhi dac diem sinh hoc cua xa khuin Nghien ciiu dac diem hinh thai hgc ctia chiing x^
khuin HP411 dugc quan sit bing mit thudng va kinh hien vi theo phucmg p h i p nudi tren mat thach ciia Kawato va Shinobu, 1959. Mau sic khuin ty va sac td cua chung dugc quan sat tren cac moi trudng khac nhau theo Khda phan loai ISP (Shirling SB v i Gottlieb D, 1966; Tresner HD va Backus J, 1963).
Chudi b i o tir va be mat bao ttr dugc quan sat dudi kinh hien vi dien tii sau thdi gian nudi 14 ngiy theo phuong phap ciia Stanley (Stanley T et al, 1989) Dac diem sinh ly, sinh hda nhu: Nghien ciiu kha ning sii dung c i e ngudn dudng v i nito tren mdi trudng ISP 9 v i 8; nghien ciiu kha nang chiu nhiet do (10-45''C). kha nang phat trien d gia tri pH thay ddi (4-11) va ndng do NaCl (1-10%) ciia chiing tren moi trudng ISP 2 (Shirling SB, Gottlieb D, 1966;
Stanley T e / i / / . , 1989),
Phan loai dya vao phan tich trinh tir 16S rDNA Tich DNA tdng sd: chiing H P 4 i l sau khi nudi tren moi trudng SCA d 28°C trong 2 ngay, xir ly \k bao bing sdc nhiet 3 7 ^ trong 5 phut, lim lanh
nhanh tai -70"C trong 10 phut. Dich te bao sau sdc nhiet duac ly tam 7.500 vdng/ 3 phiit, dmh ndi dugc b i o quin d 4''C d l sir dung (Sambrook et al., 1989).
Su dung c i p mdi FC27 (5'- AGAGTTTGATCCTGGCTCAG-3') va RC1492 (5'- TACGGCTACCTTGTTACGACTT-3') (Genset) d l nhan gen ma hda 16S rDNA. Phan iing dugc thuc hien theo chu trinh nhiet nhu sau: 94°C: 5 phiit; 30 chu ky ( 9 4 T : 1,5 phiit; S T C : 1,5 phiit; 7 2 " ^ 2 phiit); 72"C: 10 phut. Sin p h i m ciia phin img PCR dugc kiem tra bang dien di tren gel agarose 1%. Kich thudc ctia cac doan DNA thu dugc sau phan irng PCR dugc so sinh vdi thang DNA chuin (Fermentas). Sin p h i m PCR dugc tinh sach bang bg kit PureLinkTM-DNA Purification (Invitrogen) va giii trinh tu trSn m i y dgc trinh tu tu ddng ABI PRISM®3100-Avant Genetic Analyzer (Applied Biosystems, Foster City, CA, USA) tai Vien Cdng nghe sinh hgc, Vien Han lam Khoa hgc v i Cdng nghe Viet Nam. So sanh trinh tu gen tucmg ung tren CO so dif lieu GenBank (www,ncbi.iilm.nih.gov), T i c h chiet cac chat cd heat tfnh khing khuSn tir xa khuan
Dich len men dugc ly t i m 8000 vdng/phiit d nhiet do 4°C trong 10 phiit, tach thu djch va sinh khdi. Dich sau iy tam dugc bO sung dung mdi thudng dimg trong tach chiet chit khang sinh theo ty le sau: Ethyl acetate . dich len men (2:1) (v/v), lie 200 vdng/phiit d 4°C trong 2 h. Dich sau lie d l tmh 6-8 gid d nhift dd^4°C, sau dd dimg phlu tich chiit phan dung mdi. Hdn hgp n i y dugc loai bd dtmg mdi bang phuong p h i p cd d nhiet dd 60°C (Liu el al, 1986) Chat khd thu dugc sau khi loai bd dung mdi dugc sir dung cho cac thi nghiem tiip theo.
Thu hoat tinh khang khu^n cua xa khuan Phuong phap xie djnh hoat tinh khing khuin dugc thuc hien theo phuong phap khuich tin tren thach cua B a n y (Barry AL. Thomsberry C, 1985), moi trudng su dyng cho vi khuin la mdi trudng NA, cho nim men la mdi trudng thach khoai tiy. Cich lim nhu sau:
Lay 1 mg chit chiit thd hda vao 1 ml nudc cit vd tning, sau dd nhd v i o mdi lo thach 50 pi dich da hda tan, nrang duong 50 pg chit thd. Sau khi nuoi ttong tii am, do vdng vd khuin xung quanh Id thach.
Thu- hoait tfnh gSy dOc te bao cua ch^t tho dirffc chiSt tir dich Iln men xa khuan
Ddc tinh te bao ciia chit dugc chiit tir chiing HP4I1 tren cac ddng t l bao dugc xac dinh bing phuong p h i p so mau MTT (MTT assay). Phuang
Tgp chi Cong nghe Sinh hgc 12(2): 221-227, 2014
phap MTT dugc tiSn hanh dua tren phin img khir mau cua MTT- 3-(4,5- Dimethylthiazol-2-yl)- 2,5- diphenyltetrazolium bromide, 1-tetrazol) cd mau vang thanh formazan cd mau tim trong ty the cua te b i o sdng. Do vay, khi so mau se cho biet so lugng te bao sdng sdt trong cie mau thi nghiem (Van et al, 1994). Cac ddng te bao dugc nuOi cay trong dia 96 gieng (lO* te bao/ gieng) trong 24 h d 3 7 ^ va 5%
CO2. C i e dia nudi te b i o dugc thii dgc tinh bang cich bd sung chat chiet tir chiing xa khuan bien HP411 d cie ndng dd khac nhau: 4,68; 23,4; 46,8;
70,2 v i 93,6 pg/ml, mau doi chirng khong bo sung chit rai su dung DMSO de xir ly te b i o . Sau dd, bo sung 10 pi ciia dung dich thuOc thii MTT (5mg/ml trong dem muOi phopphat -PBS) vao trong 90 pi dung dich ciia cac gieng, c i e dTa dugc u d 37''C trong 4 h va sau dd cac giSng dugc do mat do quang (OD) d budc sdng 540nm bang may ELISA (TECAN, Thyy Si). Phan tram sdng sdt cua te b i o dugc xac dinh bing each nhu sau. [ I - (OD te bio/OD mau ddi chung)] xlOO.
Gia tri IC;o (ndng do ire che 50%): diing g i i tn CS (ty le te bao sdng sdt) cua 10 thang ndng do, dua vao chuong trinh Table curve theo thang gia tri
logarit cua dudng cong phat trien te bao va ndng do chit thir d l tinh gia tn IC50. COng thiic: l/y=a+b!nX, trong dd y: nong dp chat thir; X. g i i tri CS (%)
K.ET QUA VA THAO LUAN
Chimg xa khuin HP411 cd m i u sac da dang tir mau tring (a, b) d i n mau xam (d, 3fe, 5fe) va m i u xam nhat tren cac mOi trudng ISP (Hinh I A). Ddi chilu vdi khda phan loai ISP cho thay, chimg dugc xep vao nhdm xam (Gray), tren mdi trudng ISPl, 2, 4 v i 7 cd m i u x i m xanh, chiing cd mau trang den tring xam nhat tren mOi trudng ISP 3, 5 va 6. Chiing nay khdng cd kha nang smh sac td melanin, chi sinh sac td mau nau nhat tren mdi trudng ISP 2, mep khuan 'ac cd mau trang den trang xam va x i m dam (Tresner HD, Backus J, 1963).
Quan sit hinh anh hien vi dien tir (Hinh IB) cho thay, chudi bao tu ciia chimg HP411 cd do dai trung binh khoang 10-30 b i o tir tren mgt chudi, cau tnic phin d i u cua chudi b i o tir xoan, hai lugn sdng (RA), be mat bao tti xii xi (wa). Cudng sinh b i o tir cd hinh truy khi mdi nay mam (Stanley et al, 1989).
Hinh 1. Hinh thai khuin lac (A) va hinh thai bSo tiJ (B) cua chung xa khuan HP411.
Dac di£m sinh ly, sinh hoa cua chung xa khuan HP411 Ket qua nghien ciiu cac d i e diem sinh ly, sinh hoa ciia chiing HP411 duoc trinh bay tren b i n g I cho thiy, chiing HP411 thudc nhdm ua i m (mesophilic), cin nhiet dd trong khoang 25- 37''C de sinh trudng, pH tOi uu l i 7-8, chiing cd the sinh trudng d dii pH kha rgng 5-10 va cd Idii nang phat trien tot d ndng do NaCl tdi 7%>, phit triln kem d ndng do NaCl 10%
(Shirling SB, Gottlieb D, 1966, Stanley e/A/., 1989).
Chiing HP411 sinh trudng tdt hen cac ngudn acid amin nhu: L-phenylalanine, L-leucine, L-asparagine monohydrat, L-tryplophan, L-isoleucine, L-lysine, L-
threonine va 2 amino 2 hydroxy-methyl 1,3 promo, sinh hudng kem tten mdi trudng ISPS; cd k h i nang ddng hda cie ngudn cacbon nhu D-glucose, mannitol, D-xylose, mannose.. khdng sinh trudng tren radi trudng ISP9 va saccharose. Chiing nay cd khi nang sinh mdt sd enzyme ngoai bao tMy phan tinh bgt, casein v i cellulose, dieu nay rat thuin lgi cho lira chgn nguon dmh dudng khi nudi cay chiing cho sinh tdng hgp chat khang khuan.
Chiing HP411 cd cie dac diem phan loai gin gidng vdi chiing chuin S. variabilis ISP 5179 hay A. variabilis ISP 5179, do Preobrahenskaya, R y ^ o v a va Blinow md t i nam 1958, thudc nhdm A-5 (Shirlmg SB, Gottlieb D, 1966).
223
Pham Thanh Huyln et ai
B i n g 1 . 0 | c iSem hinh thai, sinh ly, sinh hoa cCia chung x^ khuan HP411
Nhi^t dp phat then K i t q u i Cuong smh bdo ti>
Be mat bSo ti>
So bao tip tren chuoi Ho^t tinh Catalase Hoat tinh Oxidase Hoat tinh Urease Thuy phan casein Thuy phan tinh bpt Thuy phan cellulose Kha nang sinh sac td Sinh melanin Phat triin d ndng d^ NaCl
Xoan, X i i x i 10-30 +
, m6c hoi l u p n s6ng (RA) (wa)
4°G 15°C 25°C S C O 37°C 45°C pH ph^t trien 5 7 8 10
Ghi chu: (-). Khong sinh t r u i n g ; (±) Cd sinh trudng, y i u hon ddi chung d u o n g ; (+): smh I n r a n g binh t h u 6 n g ho$c duong linh; (++): sinh tmidrng tot
Phin loai theo phuong phap phSn tich trinh ti^
16S rDNA
Sau khi tich DNA tdng sd ciia chung HP4il kiem tra tren gel agarose 1%, sau dd thuc hidn phan ling PCR sit dung cap mOi dac hieu FC27 vi RC1492 khuich dai 16S rDNA ciia chting HP411 cho mdt bang DNA ro net duy nhat cd kich thudc khoing 1500 bp. Sau khi giai trinh tg dugc so sanh vdi cie trinh tu gen tuong irng tren ngin hang dS lieu co sd GenBank bang cdng cu
BLAST tr6n NCBI cho thay, 16S rDNA cua chiing HP4n cd dg tuong dong cao tdi 99% so vdi gen tuong dng ciia chiing S variabilis 65-AD 7' 4 UFPR va S. variabilis 19-AS G34 3B 18 UFPR (Bang 2).
Ket hgp cie ket qui nhan dugc tit phuong phap phan loai truyen thdng vi phuang phap giai trinh tg 16S rDNA, chiing HP411 la xa khuin, cd dac dilm phan loai gidng loii S. variabilis. Do do, chiing niy dugc dat ten la S. variabilis HP411.
Bang 2. Ket qua so sanh trinh t u gen ma hoa 16S rRNA cua chung HP411 vd'i gen tuo'ng Cmg cua cac chung xa khulin d u p c dang ky trSn GenBank.
Trinh t y gen ma hoa 16S rDNA ciia cac chung d u w : so sanh Ace. No. 0 6 tuo'ng dong (%) Streptomyces vanabilis 65-AD 7^ 41 UFPR
Streptomyces vanabilis 19-AS G34 3B 18 UFPR Sfreptomyces sp, A4-3
Sfrepfomyces sp. ZZY-2013 TRM46795-87 Streptomyces sp. IWBRC-82 Streptomyces sp 14CM012
JX997144.1 JX997143.1 JN627185 1 KC336333,1 KC179812 1 JX9123351
9 9 % 9 9 % 9 9 % 99%
99%
99%
Hoat tinb khang khuan cua chung xa khuan HP411 Chiing xa khuin S. variabilis HP4I1 duoc len men tren mdi trudng MIASW. Dich len men dugc chiet xuit bing dung moi ethyl acetate, sau khi loai bd dung moi dich duac sir dung dl xac djnh hoat tinh khing khuan vdi cac chiing vi sinh vat kilm dinh,
2 2 4
ket qui tren bang 3 vi hinh 2. Djch chiit cua chiing
xa khuin HP411 cd kha nang tie chi manh vdi
cac chiing vi smh vat kilm djnh nhu E. coli
(ATCC 25922), S. typhimurium (ATCC 14028)
va nim C. ablicans (ATCC 10231) dudng kinh
vdng khang khuan kha cao dat >40 mm. Die
biet, hoat chit niy cdn cd kha nang ire chi tOt doi
Tgp chi Cong ngh4 Sinh hgc 12(2): 221-227, 2014 vdi cie chiing vi khuan khang thudc methicillin nhu 5. aureus (MRSA) vi S. epidermidis (MRSE). Tuy nhien, nghien ciru niy thuc hien
tren dich chiet thd ctia chat khing khuan, can phai tien hinh nghien ciru tinh sach chat khing khuin de cho ket qui tdt han.
Bang 3. Hogt tinh khang khuan ciia chiing HP411 v d i mot sd vi smh vgt kiem dinh.
Vi sinh vat kiem dinh Ou-Qing kinh vong d6i khang (mm)
Staphylococcus aureus (MRSA) S aureus (MRSE) Eschenchia coli (ATCC 25922) Salmonella typhimurium (ATCC 14028) Candida albicans (ATCC 10231)
50,1 ± 0 , 1 49,5 ± 0 , 1 45,3 ± 0,1 40 ± 0,1 44 ± 0 , 1
Hinh 2. H o g l t i n h khang khuan cila djch chiet chung xa khuan HP411vgi m 6 ! s 6 vi sinh vat kiem dmh
Hoat tinh gay dgc te bao ciia chSt khing sinh thd thu nhan tir dich len men chung xa khuan HP411
Chat tho dugc tich chiet tir dich len men chiing HP411 dugc thir boat tinh gay ddc vdi 3 ddng te bao theo phuong phip MTT: ddng te bao lanh- te bao than Hek293, ddng te bio ung thu cd tii cung Hela vi ddng te bao ung thu bieu md vii MCF7 d cac ndng do khac nhau. Ket qui thu ddc tinh te bao (Bing 4) cho thay, chat thu dugc khdng gay dOc vdi ddng te bao than Hek293 va ddng te bio ung thu Hela d cac ndng do khic nhau, dich chiet chi cd kha nang lie che su phit trien ciia ddng te bio MCF7 d ndng do 70,2 va 93,6 pg/ml cho phan trim sOng sdt cua te bio- SC% li 43,64% va 32,58%, tuang iing vdi gia tri IC50 la 6l,47±l,I
(pg/ml) Vdi cac ket pua nay da dua ra tilm nang nghien ciiu ve cac chat cd kha nang ire chi vi tieu diet te bao ung thu MCF7 va khdng gay hai tdi cie te bio linh ciia chat thu dugc tir chung xa khuin bien 5 variabilis HP411. Vi ket qua niy cung tuang ling vdi cie nghien ciiu da cOng bd tren thi gidi ve chit khing ung thu ctia cac chiing xa khuan bien thudc chi Streptomyces. Theo nghien cu:u ctia Ravikumar, cie chung xa khuin biln Streptomyces sp. ACTOl va ACT02 cd kha ning diet ddng tl bao MCF7 vdi gia tri lCs„ tuong iing la 10,13±0,92 va 22,34±5,82 (pg/ml) (Ravikumar el al, 2012). Theo tic gii Sunanda nam 2013 thi chiing xa khuin bien Streptomyces sp. KS1908 cd khi nang diet cie ddng te bao ung thu nhu Hela, vi MCF7 vdi gii tri IC50 tuong iing la 1,13 va 4,44 (pg/ml) (Sunanda et al., 2013).
225
Ph^m Thanh Huyen el al Bang 4. Hoat tinh gay doc te bao cila cac hop ch^t phan lap d u p c .
Nong do cua mau (^g/ml) Dg song sot cua te bao (CS) (%)
Hek293 MCF7 Hela
23.4 46.8 70,2 93,6
100,0 ± 0 , 4 93,38 ± 0,5 103,21 ± 0,6 102,0 + 0,4 103,58 ± 1,1 109,96 ± 0 , 4
100,0 ±0,6 102,04 ±0,3 106,93 + 0.7 84.76 ±1,5 43,64 ± 1,1 32,58 ±1,7
100,0 ± 0 , 5 115,39 ± 0 , 6 117.33 ± 0 . 9 103,41 ± 0 , 4 113,79 + 1,2 117,55 ± 0 , 6
KET LUAN
Chiing xa khuan bien Streptomyces variabilis HP411 phan lap dugc tir viing ven bien phia Bac Viet Nam cd cie hoat tinh khang khuin mdi nhu sau:
Cd k h i ning ire che cie chiing vi khuan gay benh khang thudc methicillin l i S. aureus (MRSA) v i S.
epidermidis (MRSE), n i m benh C. ablicans (ATCC 10231) va cac chung vi sinh vat kilm dinh khac. Dac biet, hoat chit thd dugc chiet tir dich len men chiing xa khuan bien HP411 cd k h i nang gay dgc vdi ddng t l b i o ung thu MCF7 vdi gia tri ICjo l i 61,47±1,1 (pg/ml), khong gay ddc vdi ddng t l bao lanh Hek293 De khang dinh chit n i y cd phii li chit cd hoat tfnh mdi can phii thuc hien them cie nghien ciru ve x i e djnh cau tnic hda hgc d l dinh hudng tim kiem v i phat hien ra cac ngudn nguyen lieu thudc mdi diing trong tri benh hien nay.
Ldi cam gn: Cong trinh dugc thuc hien thuoc di tai
"Nghien cicu tuyen chon cdc chdt co hoat linh khdng sinh m&i ditng cho muc dich y duoc tu vi .-iinh vgt bien Viet Nam " do Vien Han ldm Khoa hgc va Cong nghe Viet Nam cdp.
TAI LIEU THAM K H A O
Alphonse K (2002) Secondary metabolites from marine micro OTgmisms[}]. An Acad Bras Cienc74(\y 1-2], Barry AL, Thomsberry C (1985) Susceptibility tests:
diffusion test procedure. In Manual of Clinical Microbiology, 4th edn,, eds Ballows EA, Hawsler Jr WJ, Shadomy HI. Washington DC. Am Soc Microbiol ISBN 0-914826-65-4. pp 978-987,
Berdy J (2005) Bioactive microbial metabolites. ^ Anlibiotechnol (Tokyo) 58: 1-26,
FelingRH, Buchanan GO. Mincer TJ, Kauffman CA, Jensen PR, Fenical W (2003) Salinosporamide A: A highly
cytotoxic proteasome inhibitor from a novel microbial source, a marine bacterium ofthe new genus Salinospora.
Angew Chem Int Ed. 42: 355-357
Buchanan D, Fitzgerald L, Kelley D, Gollop R, Louise Jones J, Saint Lamoni S, Neath A, Whitby E (2005) Not going back: A review of the literature on sustaining organizational change. Inil J Manag Rev 7(3): 189-205.
Kawato N and Shinohu R (1959) On Streptomyces herbaricolor, nov SP supplement: a simple technique for the microscopic observation. Mem Saka Univ. Lib. Arts EdMC.JNatSci^. 114-119.
Lam KS (2006) Discovery of novel metabolites from marine actlnomycetes. Curr Opin Microbiol 9: 245-251.
Liu CM, JW Westley, TE Herman, BLT Prasser, N Palleroni, RH Evans and PA Miller (1986) Novel polyether amibioUcs, X- 14873 A, G and H produced by Streptomyces. Taxonomy of the producing culUire, fermentation, biological and lonophores properties of the antibiotics. y^«/;/iio; 39(12) 1712-1718,
Molinski TF, Dalisay DS, Elevens SL (2009) Drug development from marine natural products. Nat Rev Drug
£>fscov8(i): 69-85.
Pereira HS, Lea'o-Ferreira LR, Moussatche N, Teixeira VL, Cavalcanti DN, Costa LJ, Diaz R, Frugulhetti IC (2004) Antiviral activity of diterpenes isolated from the Brazilian marine alga Dictyota menstruahs against human immunodeficiency virus type 1 (HIV-1). Antiviral Res 64:
69-76.
Ravikumar S, Gnanadesigan M, Suganthi P, Ramalakashmi A (2010) Antibacterial potential of chosen mangrove plant against isolated urinary tract infectious bacterial pathogens. Int J Med Sci 2(3): 94-99.
Ravikumar S, Fredimoses M, Gnanadesigan M (2012) Anticancer property of sediment actinomycetes against MCF-7 and MDA-MB-231 cell lines. Asian Pac J Trop Biomed 92-96. Sambrook J, Fritsch E F, Maniatis T (1989) Molecular cloning, A laboratory manual, 2"'' ed..
Cold Spring Harbor Laboratory Press, New York
Tgp chi Cong nghe Sinh hgc 12(2): 221-227, 2014
Shirling EB, Gottlieb D (1966) International of Stanley T, Williams M, E Sharpe, and JG Holt (1989) systematic bacteriology: Method for characterization of Bergey's Mannual of Systematic bacteriology, Williams &
Streptomyces species. Derpartment of Botany and Wilkins Vol. 4:2452-2492.
Bacteriology Ohio Wesleyan University, Delaware, — TTT-. j r - i r . i .-.n^-nc . IT > L i
^L- J ^ .- . r n i . T, ,1. 1 IT -. r TresnerHD, and EJ Backus (1963) System of color wheels Ohio and Derpartment of Plant Pathology University of ^ ^ . . . , , ' / , , ,, . . - . - „ .... . . , u III- for streptomycete taxonomy, .^f/j/Mcraojo/. 11:335-338.
Van De Loosdrecht AA, Beelen RH, Ossenkoppele GJ, Sunanda Kadin, Nagendra Sastry Yarla, Siddaiah Broekhoven MG, Langenhuusen MM (1994) A Vidavalur (2013) Isolation and Identification of A Novel tetrazolium- based colonmetric MTT assay to quantitate Aporphine Alkaloid SSV, An Antitumor Antibiotic from human monocyte mediated cytotoxicity against leukemic Fermented Broth of Marine Associated Streptomyces sp. cells from cell lines and patients with acute myeloid K.S190S. J Marine Sci Res Dev 3(4): 1-5. leukemia. y/mrnHMo/MerAoiA 14: 311-320.
A N T I B A C T E R I A L A N D A N T I C A N C E R A C T I V I T I E S O F M A R I N E A C T I N O M Y C E T E S T R A I N Streptomycetes variabilis H P 4 1 1
Pham Thanh Huyen, Bach Thi Mai Hoa, Phi Quyet Tien, Nguyen Phuong Nhue, Le Gia Hy*
Institute of Biotechnology, Vietnam Academy ofScience and Technology
SUMMARY
Active substances from marine actinomycetes and mainly from the genus Streptomyces have wide application for medical as antibaterial and anticancer among others In this paper, we showed antibacterial and anticancer activity of marine actinomycete strain S. variabilis HP411, which isolated from sediments of the North coastal region of Vietnam. The strain HP411 was gray group, mature spore chains of HP411 showed an elongated and circular shape with 10 to 30 spores per chain, spore surface warty. This strain could grow rapidly on starch casem agar and other media with temperature at 25-35"C, pH optimum 7-8, and high salt containing 7-10% NaCl (w/v). Antimicrobial and anticancer activity were carried out against test microorganisms (resistant methicillin S. aureus and S. epidermidis - MRSA, MRSE- Escherichia coh (ATCC 25922), 5 epidemidis (ATCC 12228) and yeast sfrain C albicans (ATCC I023I) Further studies revealed that the crude extract of sfrain HP41I could obtain the anticancer activity for cancer cell line: MCF7 (breast adenocarcinoma cell line) and the IC50 value with 61.47 ± I.l (pg/ml). The anhbiotic from the strain S.
variabilis HP411 can hirther research as antimicrobial and anticancer agents in clinical Gials.
Keywords: Antibacterial, anticancer, marine microorganism, marine actinomycetes, S variabilis
'Authorfor correspondence: Tel; +84-4-38363257; E-mail: legiahvcnsh(giemailcom
