KHOA HOC CONG NGHE
DAIMH G I A K H A IXIAIMG P H O M G BEIXJH H O A I T O GAIXI TLJY C A P ( A H RIM D) CIJA Streptomyces X e S S TREIXJ
T O M T H E CHAIXJ T R A I X I G [Litopenaeus t/annameii
Vo Hong Phupng^, Lfi Hdng PhudcS Hodng I h i Nhung', Nguyfin Thi Ngpc Tinh', D$ng Ngpc Thuy'. T r ^ Minh Tning^, VO Bich Xodn'. Pham Th| Huyfin Difiu^, Cao Vinh Nguyfin' TOMTAT
Vibno. parahaemolyUcus la tac nhan gay ra benh hoai tii gan tuy cap Unh (AHPND), b^nh co thi' la nguyfin nhan gay chfit torn len den 100%. Kiem soat dich benh bang giai phap sinh hoc la mot Irong nhung each tifip can can thiet. Muc lifiu cua nghien cuu nay la danh gia hieu qua ciia Streptomyces Irong ki^m soal ty le chet qua rniic cua tfim the chan trang do bfinh AHPND. Ket qua nghien cini cho thay Streptomyces X285 dupc phan lap thanh cong tir bun day ao nu6i torn tiia lai dong bang song Cihi Long Chung vi khuan nay cfi kha nang doi khang voi V. parahaemolyticus tren dia thach hai Idp voi vong khang khuan la 49 mm. Ngoai ra, trong dieu kien thi nghiem, chung X285 (15 h^it den 40 hal/50 mL) co kha niSng irc chfi su phat tnfin ciia
V. parahaemolyticus a mai do 10'' CFU/mL sau 36 gio ddng nuoi cay. Ngoai ra, khi lom dupc cho an thuc an phdi trpn X285 voi mat do ICV CFU/g ben luc Irong 15 ngay. Xy le torn chfit giam vin ly le bao hfi (RPS%) tren 50% sau 10 ngay gay nhi^m bdng V. parahaemolyticus. Hon nua. X2«.") co the tifit ra mpt s6 enzyme nhu amylase, protease, lipase va cellulose. Chinh vi thfi, Streptomyces'X2SS dupc xem la vi khudn co liem nang su- dung lam chiing vi sinh trong san xuat chfi pham sinh hpc nhim han che benh AHPND.
T&khda; .\lll'.\l>. Streptomyces .\2S5. vong khang khuan. dongnuoicay, RPS (%).
1. SAT VAN B l
Bfinh hoai tii gan tuy cdp (AHPND) la mdt trong nhung bfinh nguy hiem hdng ddu boi kha nang lay nhifim nhanh chdng Irong thdi gian tir 10 - 30 ngay sau khi tha tdm vao ao moi [1]. Nguyfin nhan bfinh dupe xdc dinh la do vi khuan 1: parahaemolyticus da bi nhifim mdt toai thfi thuc kliuan cd chiia plasmid pVPA3- mang gen gay ddc PirA va PirB cd the gay ty tfi chet lfin dfin 100% trfin tdm [2j.
Cac bifin phap thdng thudng dupc sir dung de kifim soat dich benh la cac hda chdt tong hpp va khang sinh. Tuy nhien, bfin canh nhirng tdc ddng tich cue mang Iai chung cdn cd nhung tac dpng dfiu cue nhu kha nang tich luy trong mdi trudng sfing cung nhu trong ca thit cua san pham thuy san. Tir do, gay hien tuong khdng thudc ddi vdi cac chung vi sinh vat, dnh huong sue khoe ngudi tifiu dung va cung ta rao can thuong mai xudt khau. Trai lai, cac san phdm probiotic mang lai nhtmg loi ich tich cue da duoe chiing mmh voi kha nang toai trir cac vi sinh vat gay bfinh thdng qua canh tranh bam dinh trong duong rudt, canh ti-anh dinh dudng. san xuat cac hpp
' Vi?n Nghien cuu N'ufii trdng iliuj van 2 ' Tnrong Dai hpc Su pham ihanh phd Ho Chi Minh Email' [email protected]
NONG NGHIEP VA PHAT TRIEN NONG THON
chdt uc chfi, tang cudng he mifin dich va cdi thifin chat luong nubc [3). Cac chi Streptomyces cd lpi cd thfi dupc coi ta cac vi Idiuan probiotic tifim nang trong nudi trdng thuy sdn voi khd nang chuyen hda thli cap. smh tdng hpp khdng sinh, cdc chdt khdng tchudn, tao ra mpt so emzym ngo^i bao, ho trp tang trudng cua cdc vi sinh vgt va dam bdo chat luong nuoe [4]. You va ctv (2007) [5] cung da chung minh Streptomyces albus cd khd nang san xuat cac hop chat lie chfi va cac chdt chuyfin hda lifin quan den su hinh thanh mang smh hpc cila cac tdc nhan gay bfinh nhu Vibrio harveyi, V. vulnificus va V. anguillarum.
Bfin canh dd, mpt so Streptomyces dupc sii dung don hoac kfit hpp vdi cdc chi khac nhu Bacillus va Lactobacillus cd khd nang cdi thifin cdc thdng sd tang trudng. dieu hda ky chii va hfi vi sinh vat trong nudc cung nhu tang khd nang dfi khang vdi mam bfinh.
Cdc nhdm Streptomyces RL8 va BMbi-StrepMix cho ty le song trfin tom gan 95% khi cdng cam nhifim vdi vi khudn Vibno parahaemolyticus CAIM 170 [6).
Xginen Xuan Cdnh va ctv (2016) da phan lap xa khuan 25.2 dfii khang vol V. parahaemolyticus vdi duong kinh vdng Idiang 15 mm [71. Trong khi dd, Ngo Thj Tucmg Chau va ctv (2016) cho rang chung Streptomyces sp. Al dupc phan tap lu cdc ao nuoi tdm Thira Thifin - Hue cd kha nang ddi khang ven
KYI -THANG 11/2019 109
KHOA HOC COMO MftH^
chiing vi khudn Vibno sp. V7 va VIO la nguyen nhan gay bfinh d cdc ao tom nudi Thira Thifin - Hufi vdi dirdng khang khuan lan lupt la 40 mm va 36 mm [81.
Ngodi ra, tdc gid nay cung danh gia chiing Streptomyces A8 cd khd nang uc chfi manh vdi vi khuan V paraJiaemolyticus V6 la tdc nhan gay bfinh AHPND trfin tom Iai Thira Thifin - Hufi [9].
Cac nghifin ciiu trong va ngoai mroc chu yfiu tap trung trong phan lap va danh gia Idid nang doi khang vdi V. parahaemolyticus bdng phuong phap khufich tan trfin dia thach va chua cd nhifiu cdng trinh nghifin cuu danh gia hifiu qua cua chi Streptomyces trong phong bfinh AHPND frfin tdm. Do dd, vific phan lap, thu nghifim chiing Streptomyces X285 lic chfi su phdt trien cua V. parahaemolyticus gay bfinh h o ^ lli gan tuy cdp trfin tdm ta mpt nghien cuu nhdm sang lpc va cung cap thfim cdc chiing gidng vi sinh cd tpi cd khd nang han chfi bfinh AHPND.
2. VAT LJ£U VA PHUDNG PHAP 2.1. V$t lifiu nghifin ciiu
Chiing Stieptomyces X2S5 duoc phan lap tir biin ao nudi tdm Ida tai tinh Bac Lieu. Vi khuan V.
paraliaemolyticus gay bfinh AHPND dupc phan tap va dinh danh trong khudn kho de tai "Nghien cuu tao chf' pham vi smh ddi khang Vibrio spp. gay bfnh hoai tu gan tuy cap ti^n tom su va tom the chan tidng".
2.2. Phuong phdp nghifin cuu
2.2.1. Khao sat kha nang doi khang cua X^S.} voi V. parahaemolyticus tivng dieu kif n in vitro
Phuong phap thach hai ldp
Kha nang ddi khdng cua X285 vdi chung V.
parahaemolyticus dupe xdc dinh bang phuong phdp thach hai top [10]. Kich thudc dudng kinh vdng khang khuan (D) duoe chia thanh 4 muc dp voi D - 0 mm: khong uc chfi; D < 5 mm: cho vung lie chfi yfiu;
5 mm < D < 10 mm: cho viing uc chfi hung binh va D
> 10 mm: cho viing tic chfi tdt [11].
Phuang phap ddng nuoi cay tiong moi tnrdng long (co-cutulre)
Thl nghifim ddng nudi cay dupc tien hanh dua trfin phupng phap cua Augustme vd ctv (2015) [12]
CO chtnh siia nhu sau: \' parahaemolyticus va bao hi Sinpiomyccs X285 dupe tang sinh neng bifit trong mdi tiudng dinh dudng Nutrient Broth (NB) bd sung 1,5% NaCI a 2S"C. Dua vdo phuong trinh tuong quan giira mat dp quang 0D5,s,)n,n va mat dp vi khuan V.
110 NONG NGHIEP VA PHAT
parahaemolyticus 6\iac khao sdt trong thi nghifim tin 1 O D s a ^ - 5.58 X l(V te bao/mL Thai gkui nudi cay tren may ldc vdi tdc dd 150 vdng/phut hr 3-5 gid ^^^
vdi i: parahaemolyticus vd tir 3 - 5 ngay ddi vm Sti-eptomyces X285 den khi kich thudc liat vi khuan X285 dat 0,5- 1 mm. Difin bien mai dp '•
parahaemolyticus duac ghi nhan sau nim sau gidva kfio ddi dfin 48 gid. Cac nghifim thuc duoe thiel kd lap lai 2 Ian. Bd hi thi nghiem eac nghifim Ihur dupe thue tiifin theo bang 1.
Bdng 1. Thi nghifim ddng nudi cdy ciia chiing X285 vd V. parahaemolyticus Nghifim Uiuc
X285 15-VP 10'' X285 25-VP IO*"' X285 40-VP 10"
DC VP 10^
DC VP 10**
DCVPIO'-
Mat dd V.
parahaemolyticus (CFU/mL)
10«
10**
Mat dd X285 (hat/50 mL)
2.2.2. Nghien cuu mot sd dac diSm sinh hoc ciia chiing Stieptomyces X285
Kiem tra kha nang ddng hoa cac ngudn cacbon cua Stieptomyces X285 ti-fin mdi ti-ucmg khoang ISP 9 cd bo sung 1%: glucose, rhamnose, fructose, raffinose, manitol, inositol, cellulose vdi moi trudng chua glucose ldm ddi chiing duong vd khong chiia glucose Iam dot chiing am. Chung X285 duoc cay trfin cac dia thach, li d 28 - SOX quan sat khuan lac sau 10 -16 ngay [13].
Xac dinh kha nang su dung cac ngudn nito tren mdi trudng starch niti-ate [7].
Kha nang sinh cac emzym ngoai bdo cua Chung X285 nhu amylase, lipase duac xdc dinh theo phuong phap cua Hartey va c-*v (2001) [14].'
Hoal tinh sinh cellulose dmx ,.ac c!mu bang each cay chung xa khudn X285 trfin , ,u, trubZ carboxymethylcellulose (CMC) sau do u ' 8 T ^ C theo Kasana va ctv (2008) [15].
Hoat Unh protease duoe quan sat tren ,»-i tiirdng skim milk agar kel hop vdi tluific th^ T P A
25% [141. '^^UiirTCA 22.3 Phmm^rphap dinh danh sn,., },or ni. - •
chungX2K-> '^^Phan tu Tach chifii
phuong phap ( j .DN dia chl, -burg va ctv TRIEN N 6 N G THON - KY 1 - TH/
''^D. Kh:
UK^P luech
KHOA HOC CONG NGHE
d^i trinh t y 16S r R N A b d n g p h d n l i n g PCR vdi cdp mdi CO tiinh t u 2 7 F 5'- AGA G T T T G A T S M T G G CTC AG - 3 ' va 1492R Id 5' - T A C GGY T A C C T T G T T ACG A C T T -3'. C h u trinh luan nhifit d u p c t h u c hifin t h e o W e i s b u r g vd ctv (1991) [16]. T r i n h tu RNA d u p e p h a n tich t h d n g q u a p h a n . mfim BioEdit va BLAST trfin ngdn h a n g g e n d e x a c d i n h s u t u o n g d d n g cdc toai ti-fin n g a n h a n g G e n b a n k .
2.2.4 Kha nang phong benh AHPND cua Streptomyces )C285 tifn tdm the chan ti-dng khi phdi tion vdi thuc dn
Stieptomyces\2%^ dupc phdi trpn vao thiic an vdi 3 li^u khdc nhau. Thiic an duoc kiem tra mat dp Sti-eptomyces X285 trude khi cho tom an khoang 10' CFU/g, 5x10' CFU/g, 10'* CFU/g cho an hfin tuc 15 ngay trudc khi gay nhifim vdi V.
parahaemolyticus (10^ CFU/mL). Bd tri thi nghifim dup^ the hifin trfin bang 2. Tdm sau khi gSy nhifim dupc theo doi vd ghi nhan ty lfi chet va kha nang bat mot trong 10 ngay lien tuc. Ty tfi bdo hp cua tdm cho an thiic an phdi trpn Streptomyces X285 sau khi gay nhifim V. parahaemolyticus dupc Iinh loan di;a theo cong thiic cua Amend (1981) [17] RPS % (Realtive Percentage Survival) = 1 - (A/B) X 100%. Trong dd A la phan tram tdm chet sau khi gay nhifim b nhdm cho an thtic an cd phdi trpn Streptomyces X285 va B Id phdn tram tdm chet a nhdm dfii chiing duong.
Bdng 2. Bd tri thi nghifim phdng bfinh cua Streptomyces s
Nghiem thiic
X286 10' CFU/g X286 5x10" CFU/g X286 10' CFU/g Ddi chung (DC)
V.
parahaemolvticus (VP)
Ddi c h u n g NB*
p. s a u k h i p h o i trOn tl
iVIatdo V.
parahaemolyticus (CFU/mL)
10^
10^
lie an Matdd bdo tli xa
khuan trong thue an (CFU/g) 10' 5x10'
W
thuc theo phuang p h ^ phan Uch One Way ANC)\A vdi phfip thir DLCAN bang phdn mfin SPSS.
3. KFT QUA VA THAO LUAN
3.1. K h a o sdt k h d n a n g ddi k h d n g ciia 5£n?j5toin>«esX285 ddi vdi V. parahaemolyticus
3.1.1. Phuongphap thach hai ldp
Nghifin cmi khd nang ddi Idiang cua Streptomyces X285 vdi I: parahaemolyticus cho thay thoi gian phat trien tren mdi truong dinh dirdng NA (Nuhient Agar ed bd sung 1,5% NaCt) cua X285 tir 3 -5 ngay nhung khd nang ddi khang vdi V. parahaemolyticus a mat dd 10^
CFU/mL rdt tdt va dn dinh trong 48 gid u d nhifit dp 30°C vdi dudng Idnh khdng Ichuan dao dpng trong khoang 49+0,2 mm (Hinh 1).
Hinh 1. Khd ning ddi khdng cua chiing X285 v6i V. parahaemolyticus 3.1.2. Phuongphap ddng nuoi cay (co<ulture)
2.2.5. Phuong phap xuly solieu
Sd lifiu thu thap duoc tinh loan gia tri trung binh, dd Ifich chuan, so sanh su khac biel giiia cdc nghifim
Hinh 2. Kha nang ddi khang cua X285 vdi mdt dfi V. parahaemolyticus IA 10^ CFU/mL Difin bifin mat dd V. parahaemolyticus trong cdc nghiem thiic dfing nudi cay bdt ddu V.
parahaemolyticus mat do 10'' CFU/mL duoc the hifin trfin hinh 2. Ttr thoi diem 12 gid den 24 gid, mat dd V. parahaemolyticus a nghifim thiic doi chirng dat khoang 10^ CFU/mL. mat do cao nhal tai thdi diem 18 gip 2.25 + 0,36 x 10^ CFU/mL. Tuy nhifin, sau 30 gid trd dl mat dp V. parahaemolyticus giam nhe va dat Ichoang 10' CFU/mL dfin 48 gio Ichao sat
NONG NGHIEP VA PHAT TRIEN NONG THON - KY 1 - THANG 11/2019 111
KHOA HOC C6lltt MttHi Trong khi do, mat do I'. parahaemolyticus (VP)
cac nghidm thuc dong nuoi c^y X285 - 40 hat - VPIO^.
X285 - 25 hat - VP10«. X285 -15 hat - VP10« cao nhat tai thoi diem 12 giir voi 1.45 ± 0.64 x 10» CFU/mL.
2.65 ± 0.78 X 10* CFU/mL. 2,9 + 0.56 x 10» CFU/mL theo thu tu tren. Thai diem 24 gio dcln 42 gio, mat dp
V. parahaemolyticus a cac nghiem thuc X285 - 40 hat - WIO' X285 - 25 hat • VPIO*, X285 - 15 hat • VP10«
lan lu<7t giam tu 2,05 ± 1.34 x 10* CFU/mL, 2,00 + 1.41 X 10' CFU/mL. 2.86 ± 0.37 x 10' CFU/mL xudng con 20.0 ±14,1 CFU/mL, 12,5 ±3,5 CFU/mL, 20.00 ± 14.10 CFU/mL tuong ling. Tai thoi diem 48 gio, mat do V. parahaemolyticus khong con phat hien 6 nghiem thiic X285 • 15 hat - VpVf va X285 - 25 hat - VplO". Tuy nhien a nghiem thiic X285 - 40 hat - VPIO'. mat dp V. parahaemolyticus con khong dang k^ (2,50 ± 3.53 CFU/mL) va co su khac biet vm nhom ddi chiing DCVPIO' CFU/mL voi p < 0,05 (Hinh 2).
Phan lap. sang lpc cac chiing Streptomyces ddi khang voi V. parahaemolyticus dupc nghien cuu boi mpt SP tac gia nhu Nguydn Xuan Canh (2016) [7] va Ngd Thl Tudng Chau (2016) 18). [91. AugusUne va ctv (2015) [12] cung da ghi nhan trong thi nghiem ddng nudi cay ciia chiing Streptomyces rubrolavendulae M56 vdi su suy giam manh ciia mat dd V. parahaemolyticus tai cac thdi diem 24 gid voi mat dp V. parahaemolyticus mat dp 10* CFU/mL xudng cdn 10- CFU/mL, tai thdi didm 48 gid giam
cdn 10' CFU/mL va khdng cdn \hiy xual hi^" ™ thdi di^m 72 gid. Trong khi do, mSc du mjl i<> '•
parahaemolyticus tiong thi nghidm ciia .1 in.'.'"" *'"' 24 gid giam tir Iff CFU/mL xudng cdn lu CFU/i"'- d ca 3 nghiem thiic nudi chung va giam xudng cdn 10> CFU/mLdnghidm thiic X2S,5.ir>\TlM cho thay kha nang ddi khang I; parahaemolyticus ciia chiing X285 tuong tu so vdi chung .s'm'/*"".'''"^*
rubrolavendulae M56 dupc phan l$p tir n'am tich cCia Vinh Bengal. Kha nang lic che su phat tiien cua vi khuan gay benh V. parahaemolyticus cd the dupc giai thich thdng qua kha nang sinh khang sinh c i a xa khuan hoac sinh tdng hpp cac hpp chat thiT cap [6.
18, 191.
3.2. S J c 6iim sinh li, sinh hda cua Stieptomyces X285 trSn mdi trudng ISP (Intemational Streptomyces Project)
Chiing X285 phat trien tdt tren mdi trudng ISP 2.
ISP 3, ISP 4, ISP 6 cho thay su phat trien nhanh cila khuan ty co chat (KTCC) cho hinh dang khuan lac ldn tli 5 - 7 mm, mau vang ddn vang nau, bd mat xu xi, tam nhd. Trong khi dd khuan ty khi sinh (KTKS) tir mau trang den xam. Trai lai, X285 phat trien tuong ddi kem trdn mdi trudng ISP 5 va ISP 7 hinh dang khuan lac nhd 1 - 3 mm, mau vang. Su thay dpi mau sac cua X285 trong vdng tir 7 ngay den 21 ngiy cho mau sdc dam dan va chuyen sang vang nau nhung khdng chuyen mau sang mau nau den hoac den nen chung X285 khdng sinh sic td melanin (Hinh 3).
Hinh 3, M5t tnrtc chiing X285 nudi ciy trdn mdi trudng ISP trong 14 ngiy {A) .Mdi hirdng ISP2: (B) Moi tiirdng 1SP3: (C) Mdi tiirdng ISP4. (!» ,/„, ihudng !•
trudng ISPlr. IF) Mdi trudng ISP7.
U : NONG NGHIEP VA PHAT TRIEN NdNGTHOr' KY 1 - TH^
'•• (E) Mdi
KHOA HOC CdNG NGHl
Bdng 3. Khd ndng sir dung ngudn carbon vd nguon nita khdc nhau cua chung X285
Nguon carbon
Glucose Inositol Mannilot Fructose Rhamnose
Kha nang phat tnen sau 7 ngdy nudi cay
+ ++
++
++
++
Ngudn mto
NaNO-, Beef extract
Kha nang phat trien sau 7 ngay nudicav
+ ++
KNO, -M- NH,,NOi NH^Cl (NHJ,S(),
Pepton ++
an) sau khi gay nhifim voi I'. paraliaemolyticus. Ty lfi chfit trung binh ldn linrt d cac nghifim thue XL'Sfi - 10' CFU/g dao dpng 58,33%, voi ty tfi bao hp (RPS%) trfin tom la 36,36%. Nghifim thuc X2.s:i 5x10' CFU/g, t>- lfi chet khoang 53,33% vd RPS (%) tuong ung la 41.82%. Trong khi dd, nghifim thue X285 -10*
CFU/g cho ty lfi bao ho cao nhat vdi 54,54% (Hinh 4).
Difiu ndy phu hop vdi thi nghifim cua Das va ctv (2006) khi theo doi su tang trudng cua tdm an thiie an chua Streptomyces d cdc mat dp cho an Id 0, 2,5;
5,0; 7,5 va 10 g/kg thiic an trong 1 ngdy, 10 ngay va 20 ngdy. O lieu 10 g/kg thi tdng sd vi Ichuan Vibno gidm cdn 0,2 x W CFU/mL, su tang hirdng ve chieu ddi tdm la 15,79% va khdi lupng la 57,97%.
Ghi chu- (•) chung X2S5 khong cd kha ning phat trien: (+) Chung X2S5 co kha nSng phat trien bing ddi chung duong; (-t-h) chungX285 co kha ndng phat trien tdt hon ddi chung duong.
3.3. K^t qua dinh danh 5fn?pto/n>resX285 bdng phuong phdp sinh hpc phdn tii
Ket qua giai trinh tu va so sanh chung X285 vdi cac lo^i thupc chi Streptomyces trfin ngan hdng Gen hank cho thay co su tuong dong 99% vdi Streptomyces variabilis, S. griseincamatus, S.
erythrogriseus, S. labedae, S. coelicolor, S.
gnseoens. Su tuong dong giua cae loai nay vdi ehung X285 cho thay chung X285 thuoc chi Streptomyces luy nhifin vific su dung khuech dai doan gen IBS RNA cho thay khd phan loai chi Sheptomyces dfin loai [20, 21]. Difiu nay cung difin ra tuong tu voi chung GMN4.003'' khi phan Uch bdng 16S RNA thi muc dd tirong ddng cua chung nay vdi toai duoc so sanh vdi Stieptomyces radiopugnans R97'^ ta 99%
nhung tai cd nhirng dac diem khdc nhau ddn dfin dfi xudt vk su phan loai toai mbi trong chi Streptomyces [22]. Sir xac dinh dfin toai cua Sdetomyces spp. cd Ihfi kfit hpp giiia phan ung khuech dai PCR doan gen 16S RNA va ky thuat RAPD vdi cdc doan moi cd dd nhay va dd dac hifiu cao. Cac chung cd nhiing dac difim Immg dong se tao ra cdc kich thudc ngdu nhifin gidng nhau, cd thfi phan thanh cum vd tir dd cd the tdm gidm thdi gian xac dinh loai 123].
3.4. Ddnh gid kha nSng phfing tri bfinh AHPND trfin tdm cua ^repti}mycesY2.^
Ty tfi tdm ctiel a nghiem thiic phdi trfin X285 vdo thiic an dupe cdi thifin dang ke so vdi nghifim thiic doi chiing duong (khong sir dung X285 trong fhiic
i k~* i * ^ - i *
Hinh 4. Ty Ifi ch6t cfing ddn (90 sau khi gdy nhigm V.
parahaemolyticus c^ nghifim thiic Kfit qud trfin cho thay hifiu qua bao hfi tfim ty tfi thuan vol mat dp Sti-eptomyces X285 bd sung vao thiic an. Hifiu qua cao nhat trong nghifim thirc cho an ta 10" CFU/g va hifiu qua nay khac bifit so voi cdc cac nghifim thiic sii dung X285 phdi trdn thiic an mat dp 10" CFU/g (p < 0,05). Tac nhan gay bfinh thuong phat tnen nhanh va kha nang bam dinh cua chung probiotic trong duong rudt cd the cham hon tdc dp ddo thai cua vat chu tam cho mat do probiotic trong dudng rudt thap vd tam gidm kha nang canh tranh voi vi khuan gay bfinh [24]. Chinh vi the, hieu qua bd sung dmh ky hay thudng xuyfin mot lupng vi sinh vat cd tpi trpng thiic an gdp phan duy tn mat do cua chung trong hfi tifiu hda cua vat chu mdt cdch tdt nhat [25]. Theo nghifin curu cua Das va ctv (2010) [26] khi cho an kfit hpp Streptomyces CLS-28, Streptomyces CLS-39, Stieptomyces CLS-45, ty Ifi chet trfin tdm khi gay nhifim vdi V. harveyi a mat dd 10' CFU/mL tdn luot ta 31.33%, 38.00%. 42,00% thdp hon t>' tfi chfit cr nhdm ddi chiing la 58%.
NONG NGHIEP VA PHAT TRIEN NONG THON - KY 1 - THANG 11/2019 113
KHOA HOC C 6 N a M B u t 4. Xn LUM
Sti-eptomyces >a.'<3 su dung glucose, manitol.
inositol, rhanmose, fructose, cao thit bo, KNO3, pepton nhu la ngudn carbon va nito trong qua trinh sinh trudng. Ngoai ra, Streptomyces X285 cd the tiet ra cac emzym ngoai bao nhu amylase, lipase, protease frong giai doan phat trien. Hon nira. X285 ddi khang tdt vdi V. parahaemolyticus vdi vdng khang khuan dao ddng 49 mm. Mai khac, X285 mat dp 15 hat/50 mL cd the lie che dupc V.
parahaemolyticus m^t dp bat dau 10*^CFU/mL sau 36 gid ddng nudi cay.
Ben canh dd, khi phdi trdn X285 10» CFU/g thiic an cho an lidn hac 15 ngay cd kha nang giam ty Id tdm chel cSp tinh vdi ty le RPS dat 54.54% sau 10 ngay gay cam nhidm vdi V. parahaemolyticus. Chinh vi the, chiing X285 cd the duoc xem nhu mdt vi khuan probiotic cd tiem nang trong phdng benh ANPND tren tdm.
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EVALUATION OF Stivpti}myces-}i285 IN PREVENTION OF ACUTE HEPATOPANCREATIC NECROSIS DISEASE (AHPND) IN WHITE LEG SHRIMP {Utopenaeus vannamei)
Vo Hong Phuong, Le Hong Phuoc, Hoang Thi Nhung, Nguyen Thi Ngoc Tinh, Dang Ngoc Thuy, Tran Minh Trimg, Vo Bich Xoan, Pham Thi Huyen Dieu, Cao Vinh Nguyen
Summary
Vibno parahaemolyticus is the causative agent of AHPND (Acule hepatopancreatic necrosis disease) that can cause mortality up to 100% in shrimp. The developing biological solution to control AHPND is urgendy needed. The objective of this study was to determine the effectiveness of SUeptomyces X285 in controlling the mass mortality of while leg shrimp {Dtopenaeus vannamei) due to AHPND. The result shows that Streptomyces X285 was successfully isolated from sediments of the nce-shnmp rotation ponds in Mekong Delta This strain was found to inhibit V. parahaemolyticus with halo zone of 49 mm by double-layer agar method. In in vitro test, X285 strain (15 to 40 bio-granules per 50 mL) inhibited ihe growth of the V.
paraliaemolyticus al the concentration of 10" CFU/mL after 36 hours of co-cultured. In addition, in in vivo test, feeding Litopenaeus vannameimth W CFU of X285/ g feed during 15 days was able to reduce shnmp mortahty and relative percentage survival (RPS %) was above 50% after 10 days of challenging shnmp with V. parahaemolyticus Moreover. X285 strain could secret some enzymes such as amylase, protease, lipase and cellulose. Therefore. Streptomyces X285 has potential to be used as beneficial bacteria in manufacturing probiotic product to reduce AHPND.
Keywords: AHPND. Streptomyces X285, halo zone, co-culture. RPS (%).
Ngudi phan bi^n: PGS.TS. To Long Tlianh Ng^ynHn bii: 4/9/2019
Ngiy th6ng qua phan bi0n: 4/10/2019 Ngay duyet dang: 11/10/2019
NONG NGHIEP VA PHAT TRIEN NONG THON-KYI -THANG 11/2019 115
