Tìm kiếm các chủng vi sinh vật có hoạt tính sinh học nhằm tạo chế phẩm làm sạch nước nuôi tôm

3.

^

DE TAI KHOA HOC TRONG DIEM

TIM KIEM CAC CHUNG VI SINH VAT CO HOAT TINH
SINH HOC N H A M T A O CHE PHAM LAM SACH NlTdC
NUOI TOM
MA SO: QG - TD 05-05

Chu tri de tai:

GS.TS. Pham Van Ty

Cdc thanh vien:

GS. TS. Nguyen Lan Dung
PGS. TS. Nguyin Thanh Du^crng
TS. Bui Viet Ha
ThS. Dao Thj Lu-ong
ThS. Tr§n Thj Thanh Huyin
ThS. Nguyin buy Thjnh
CN. Pham Dii-c Ngoc
KS. Nguyen Thj Thuy Hang

HA NOI - 2007

DAI HOC OUOC GIA HA NOI
rPUNG TAM THONG TIN THU VIEN

0006000666^


CAO TOM TAT
a, Tin di tdi: Tim kilm cac chung vi sinh vat c6 hoat tinh sinh hoc nham tao che
pham lam sach nude nudi torn.
b, Chu tri de tdi: GS. TS. Pham Van Ty
c, Cdc can bg tham gia: GS. TS. Nguyin Lan Dung, PGS. TS. Nguyen Thanh
Duong, TS. Bui Viet Ha, ThS. Dao Thi Luang, ThS. Trkn Thi Thanh Huyin, ThS.
Nguyen Duy Thinh, CN. Pham Due Ngoc, KS. Nguyin Thi Thuy Hang.
d, Muc tiiu vd ngi dung nghien cuu
- Muc tieu:
•

Phan lap va tuyen chon bo vi sinh vat c6 kha nang sinh cac chat cd hoat
tinh sinh hoc cao, giup phan giai nhanh cac chat huu co du thua va diet cac
vi sinh vat gay benh co hoi trong ao nuoi.

•

Tu cac chung da lira chon, nghien cuu che tao che pham vi sinh than thien
mdi trudng vdi gia re nham lam sach nude nuoi torn, thay the cho viec su
dung khang sinh.

- Noi dung nghien curu
•

Phan lap va tuyen chon cac chung vi sinh vat co nhung dac diem sau:

+ Cd kha nang sinh cac chat hoat dong sinh hoc (enzym ngoai bao) phan giai

nhanh cac chat hiiu co khong mong muon trong ao torn.
+ Co kha nang sinh chat khang khuan chdng mot so vi khuan gay benh thudng
gap d tdm.
•

Nghien cuu dac diem sinh hoc, phan loai cac chung lira chon.

•

Lira chon dieu kien nuoi cay thich hop cho sinh trudng va sinh tong hop
cac hoat chat sinh hoc.

•

Nghien cuu tao che pham vi sinh (probiotic) de xu ly nude ao nuoi tom.

•

Khao nghiem che pham d phong thi nghiem va trong ao nuoi.

e, Cdc kit qua chinh
Tir 130 chung vi khuan co kha nang sinh cac chat hoat dong sinh hoc phan lap
dugc tir cac mau dat, bun thu thap tir nhieu tinh, da chon dugc 5 chung cd hoat tinh cao
va CO cac uu diem dung trong nudi trong thuy san:
- Cac chung V20 va VI1 co kha nang cung luc sinh cac enzym amylaza, chitinaza,
proteaza, xenlulaza va lipaza.


1 nhu: Proteus sp., E.coli, Pseudomonas spp.,
Aeromonas spp., dac biet Vibrio spp. la vi khuan gay benh phd bien nhSt d tdm.

Ca 5 chiing tren diu sinh trudng trong cac mdi trudng thong thudng va cd
kha nang chiu mudi nen chiu dugc mdi trudng nude bien.
Khi nghien curu cac dac dilm phan loai theo phuong phap truyen thdng
(Bergey's Manual) kit hop vdi phan tich trinh tir rADN 16S cd the ket luan:
- Chung V20 la Bacillus licheniformis
- Cac chung V75 va VI1 la 5. subtilis
- Chung L40 la Lactobacillus plantarum
- Chiing LI I la Enterococcus lactis
Da tien hanh nghien ciru cac yeu to anh hudng den sinh trudng va sinh tdng
hgp cac enzym ngoai bao, kha nang tdng hgp cac chat diet khuan bao gdm: thanh
phan mdi trudng (ngudn cacbon, nito, khoang thich hgp), pH, nhiet do, thdi gian
nudi thich hgp.
Da nghien cuu va che tao thanh cdng che pham probiotic dang bgt va dang
dich mang ten EMUNIV-S tu 5 chiing da lira chon. Quy trinh san xuat bao gdm: lira
chgng nhan gidng cap I, cap 2, len men tren mdi trudng xdp va djch the; lira chgn
chat mang, ty le phdi hgp gidng, thdi gian bao quan.
Da tien hanh khao nghiem hieu qua cua che pham trong phdng thi nghiem va
tren hien trudng:
- Xu ly nude nudi tdm bi d nhiem trong be kinh
- Xii ly biin loang (slime) cua ao nudi d nhiem trong be kinh
Ket qua cho thay che pham cd tac dung cai thien chat lugng nude. Cac chi so
BOD, COD, NH4, NO ", H2S... deu giam so vdi ddi chumg. Bun cd mau xam trong
khi d be ddi chung bun cd mau den va mui thdi.
Thi nghiem nudi tdm trong be kinh cd xu ly bang che pham cung cho ket qua
truong tir
Khao nghiem tai ao gdm 2 loai: 3 ao 20.000 m2 tai Dinh Vu - Hai Phdng
cho thay cac chi so NH4, NO2, H2S, BOD, COD d cac ao thi nghiem ludn thap hon
so vdi ddi chung. Cac dong vat thiiy sinh nhdm Copepoda, Cladocera, Rotatoria
cao hon cac loai vi tao lam thirc an cho tdm nhu Chlorella, Skeletonema,
Chaetoceros cung tang len trong khi tao doc Ceratium. vi khuan lam gay benh ban

vd tdm giam di.


ig vdu uoi uiiuiig lit Hiyo, iiaiig simi tang gap doi (2,6-3 tan/ha) so vdi ddi chung la 1,5
- ha.
Tai Ddng Nam Dien, Nam Dinh da tien hanh khao nghiem tren 20.000 m^ ao nuoi
lim so sanh che pham EMUNIV-S vdi mot so loai che pham dang luu hanh tren thi
idng (Biochie cua My, BRF-2 cua Viet Nam) theo phuong phap so sanh tuong d6i. Kit
la budc dau cho thay:
- Dung EMUNIV-S de xu ly ao nuoi tom su cho mau nude dep hon (mau non
m6i), ben mau hon va cd do trong tot hon so vdi ddi chung va vdi 2 che pham khac tinh
' 45 den 90 ngay nuoi tha.
- Khi su dung EMUNIV-S lugng nito tdng so va photpho tdng so trong bun la
lap nhat.
- Su dung EMUNIV-S cho toe do tang trudng tuong ddi cua tom su tot hon so vdi
ai ohe pham con lai.
Ket qua khao nghiem tai phong thi nghiem tai phdng thi nghiem ciing nhu trong
ilu cd thay EMUNIV-S la che pham co hieu qua lam sach nude ao nudi torn va do do
jm tang nang suat.

CAC CONG TRINH CONG BO VA SAN PHAM DAO TAO
I. Cong bo 2 cong trinh
1. Tran Thi Thanh Huyen, Pham Van Ty, Dao Thi Luang, 2007. Tuyen chon vi
chuan co kha nang ddi khang vdi mot so tac nhan gay benh thudng gap d torn. Gui dang
Fap chi Khoa hoc , Dai hoc Quoc gia Ha Noi
2. Nguyen Duy Thinh, Dao Thi Luang, Bui Viet Ha, Pham Due Ngoc, Tran Thi
Thanh Huyen, Pham Van Ty, 2007. Thu nghiem che pham vi sinh vat cai thien nude
nuoi torn. Gui dang Tap chi Khoa hoc , Dai hoc Quoc gia Ha Ngi
II. Dao tao: 2 thac sT bao ve nam 2006.
1. Tran Thi Thanh Huyen vdi de tai: 'Than lap, tuyen chgn cac chiing vi sinh vat

CO hoat tinh sinh hoc cao de tao che pham dung trong xu ly 6 nhiem moi trudng nuoi
torn"
2. Nguyen Duy Thinh vdi de tai: "Su dung che pham vi sinh vat de xii ly 6 nhiem
moi trudng nude nudi tdm"
3. Hai khoa luan cu nhan sinh hoc, 2006


/ . itnn nmn su aung kmn phi cua de tdi
Kinh phi dugc c^'p: 300 trieu dong
Kinh phi da sii dung: 300 tri6u dong

KHOA QUAN L t
(Ky va ghi rd ho ten)

CHU TRI DE TAI
(Ky va ghi rd ho ten)

^

PHO CHU NHIEM KHOA

.OSTS. I>'>4/» Sa^

TS VAsUY^ C''^^ ^j

Jf^Ma.

TRl/C)NG DAI HOC KHOA HOC TlTNHIEN

HO HiEu THUONC

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PROJECT

SCREENING OF BIO ACTIVE COMPOUNDPRODUCING MICROORGANISMS FOR
TREATMENT OF SHRIMP CULTURE WATER

CODE NUMBER: QG - TD 05-05

COORDINATOR: PROF.DR. PHAM VAN TY

HANOI - 2007


PROJECT

SCREENING OF BIOACTIVE COMPOUNDPRODUCING MICROORGANISMS FOR TREATMENT
OF SHRIMP CULTURE WATER
CODE NUMBER: QG - TD 05-05

Coordinator:

Prof. Dr. Pham Van Ty

Members:

Prof. Dr. Nguyen Lan Dung
Assoc. Prof. Dr. Nguyen Thanh Duong
Dr. Bui Viet Ha
MSc. Dao Thi Luong
MSc. Tran Thi Thanh Huyen
MSc. Nguyen Duy Thinh
BSc. Pham Due Ngoc
Eng. Nguyen Thi Thuy Hang

HANOI - 2007


SUMMARY
a. Project title: Screening of bioactive compound-producing microorganisms for
treatment of shrimp culture water.
b, Coordinator: Prof Dr. Pham Van Ty
c. Members: Prof Dr. Nguyen Lan Dung, Assoc. Prof Dr. Nguyen Thanh Duong,
Dr. Bui Viet Ha, MSc. Tran Thi Thanh Huyen, MSc. Dao Thi Luong, MSc. Nguyen Duy
Thinh, BSc. Pham Due Ngoc, Eng. Nguyen Thi Thuy Hang.
d, Objectives and main contents
- Objectives:
•

Isolation and screening of microorganisms, which are capable of rapidly
degrading undesirable organic substances in shrimp culture pond and
possess antibacterial properties against common diseases in shrimp.

•

Production of cheap and friendly environmental-friendly microbial
preparation using the selected strains to improve shrimp culture water. This
preparation can be used as a replacement of the traditional method
(antibiotics)

- Main contents
Isolated and screened
characteristics:

microbial

strains

processing

the

following

•

Isolated and screened microbial strains processing the following
characteristics: (i) capable of secreting bioactive substances (enzymes) to
rapidly decompose undesirable organic substances in shrimp culture pond;
(ii) capable of producing anti-bacterial substances against common
pathogenic bacteria in shrimps.

•

Investigated biological and classification characteristics of the selected
strains

•

Optimized the culture conditions to obtain maximal growth and bioactive
compound production

•

Studied on manufacture of microbial preparation (probiotic) by using
selected strains for treatment of shrimp culture pond


•

Tested the effectiveness of this preparation using shrimp culture tanks
and ponds.

e. Key results
- 130 bioactive compound-producing bacterial strains were isolated from soil
and slime samples from different provinces. Among these, five strains were selected
basing on their high activity in producing bioactive compound and their potential
application to shrimp culture.
- Two strains V20 and VI1 were capable of secreting multiple enzymes,
including amylase, protease, cellulose, chitinase and lipase.
- Three strains V75, L40, and VI1 exhibited antibacterial activity against
shrimp pathogens, including Proteus sp., E.coli, Pseudomonas spp., Aeromonas
spp., especially Vibrio spp. (the most common pathogen).
- All five above strains could grow in regular medium and in high salt

medium, indicating that these strains can grow in seawater.
- Results of classification study using traditional method (Bergey's Manual)
and I6S ribosomal DNA sequence analysis revealed that:
+ V20 is Bacillus licheniformis
+ V75 and VI1 are B. subtilis
+ L40 is Lactobacillus plantarum
+ LI I is Enterococcus lactis
- Affects of culture condition on growth, exogenous enzyme synthesis and
anti-bacterial substance synthesis, including media (carbon, nitrate, mineral), pH,
temperature, and culture duration were investigated.
- Liquid and power forms of Probiotic preparation were successfully
developed and called EMUNIV-S. EMUNIV-S is a mixture of 5 selected strains.
- The effectiveness of the preparation was tested in tanks and in culture pond:
+ Treated the contaminated shrimp culture water in tanks with the
preparation
+ Treated the contaminated slime in tanks with the preparation
- The results revealed that the preparation improves the water quality. There
was a decrease in indicators such as BOD, COD, NH4, NO2.. H2S... in treated tanks
when compared to untreated control. The slime was gray in treated tanks while in
untreated control, it was dark and stink.


- Application of the preparation to ponds (three 20,000-m^ ponds at Dinh
Vu-Hai Phong) gave similar results. Moreover, there was an increase in favorable
microflora including Copepoda, Cladocera, Rotatoria, Chlorella, Skeletonema,
Chaetoceros. In contrast, there was a decrease in bad microflora including
Ceratium and cyanobacteria.
- Population of Vibrio parahaemolyticus was reduced 10 times. Survival of
shrimp in treated and untreated pond was 64-70% and 41%, respectively. Shrimp
production in treated pond was 2.6-3 ton/ha, double the production in untreated

pond (1,5 ton/ha).
In Dong Nam Dien, Hai Phong province, an experiment was designed to
compare the effectiveness of EMUNIV-S with two conunercial preparations
(Biochie-USA and BRF-2-Vietnam) (Biochie cua My, BRF-2 ciia Viet Nam). The
preliminary results were as follows (in comparison to the two commercial
preparations):
- Water in shrimp culture pond treated with EMUNIV-S was clearer and
greener.
- Nitrate and Phosphate amount in slime from EMUNIV-S treated pond was
lower.
- Growth rate of shrimp in EMUNIV-S treated pond was higher.
In conclusion, all above results suggest that treatment with EMUNIV-S leads to
improved culture water and enhanced shrimp production.

PUBLICATIONS AND TRAINING ACTIVITY
I. Two publications
1. Tran Thi Thanh Huyen, Pham Van Ty, Dao Thi Luong. Screening of
bacteria producing bioactive compound against common pathogens in shrimps.
Science magazine. 5/2007.
2. Nguyen Duy Thinh, Dao Thi Luong, Bui Viet Ha, Pham Due Ngoc, Tran
Thi Thanh Huyen and Pham Van Ty. An experiment on using microbial preparation
for water quality improvement of shrimpculture ponds. Vietnam National
University Hanoi. Journal of Science. 2007.
II. Training activity: 2 MSc completed in 2006.


1. Tran Thi Thanh Huyen, project: "Isolation and screening of bioactive
compound-producing microorganisms for treatment of contaminated shrimp culture
ponds."
2. Nguyen Duy Thinh, project: "Application of microbial preparation for

treatment of contaminated shrimp culture ponds"

PROJECT COORDINATOR

FACULTY OF BIOLOGY

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TucfH A^ht'^

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PROJECT IMPLEMENTING ORGANIZATION
HANOI UNIVERSITY OF SCIENCE, VNU


Lcn NOI D A U
La mot trong nhung nuoc co dieu kien thien nhien im dai, nganh nuoi trong
thuy san cua Viet Nam trong nhung nam qua da phat trien vugt bac, dat dien tich
tren 700 nghin ha nuoi trong va vuon len thuoc nhom cac quoc gia co san lugng
nuoi tr6ng thuy san dung dau the gioi. Nam 2006 tong kim ngach xuat khau thuy
san dat tren 3 ty USD, trong do mot nua la tu tom, chi dung sau Trung Quoc va
Thai Lan. Nganh nuoi trong thiiy san thirc sir da giai quyet van de viec lam cho
ngudi lao dong, tang thu nhap, gop phan xoa doi giam ngheo va tang trudng kinh
t6 cho dat nuoc.
Ngay 11/1/2006 Thii tuong Chinh phu da ky Quy^ dinh s6 10/2006/QDTTG phe duyet Quy hoach tong the phat trien nganh thuy san den nam 2010 va djnh
huong den nam 2020. Theo do nganh thuy san phai phan dau tro thanh nganh san
xuat hang hoa Ion, san pham phai co chat luong tot, co sue canh tranh cao tren thi

truong the gioi de dua nganh nay tiep tuc phat trien mot each ben vmig.
De nghj quyet ciia chinh phii tro thanh hien thirc la mot nhiem vu day kho
khan, do chiing ta dang phai doi mat voi nhung thach thuc Ion ve 6 nhiem moi
truong, lam cho djch benh ngay cang nhieu va ngay cang phuc tap, khien cho riii ro
ciia nganh nuoi torn ngay cang Ion, nhieu noi co nguy co mat trang do tom chet co
khi den 100%, gay thiet hai nang ne cho nguoi dan.
Truoc day de phong chong benh cho tom, nguoi dan thuong dung hoa chat
va chat khang sinh. Ngay nay cac bien phap nay khong con dugc ap dung nua do du
lugng hoa chat va chat khang sinh ton du trong san pham. EU da ra thong bao se
kiem tra 100% tom nhap khau vao cac nuoc EU tir Viet Nam. Nam 2006 Nhat Ban ban hang Ion ciia Viet Nam, ciing da canh bao ve du lugng khang sinh trong torn
Viet Nam, ma neu khong cai thien thi se co nguy co mat thi truong nay,
Moi truong va benh la hai khia canh ciia mot van de, chiing co tac dong qua
lai voi nhau. Phan Ion dich benh 6 torn xuat phat tir 6 nhiem moi trudng. Do vay,
viec tao ra mot che pham sinh hoc de xu ly 6 nhiem moi trudng co y nghla song con
doi vdi nganh nuoi trong thuy san noi chung va nganh nuoi tom noi rieng. Hien tai
chiing ta khong kiem soat het dugc cac che pham sinh hoc dang luu hanh tren thi


trudng. Cac che pham cua nude ngoai khong nhimg dugc ban vdi gia qua dat ma
con chua nhieu loai vi sinh vat ma mot khi tung vao thien nhien chiing ta khong
kiem soat dugc.
Xuat phat tu nhung ly do neu tren, muc tieu ciia de tai nghien curu cua chiing
toi nham:
- Tuyen chgn bo vi sinh vat cd kha nang sinh chat cd hoat tinh sinh hgc cao
giiip giiip phan giai nhanh cac chat hiru co du thira va diet cac vi sinh vat gay benh
CO hoi trong ao nuoi.
- Tu cac chiing da lira chon, tao che pham vi sinh than thien moi trudng vdi
gia re dung xu ly 6 nhiem moi trudng thay the cho viec su dung khang sinh trong
nghe nuoi torn.
Trong ban bao cao nay, ten de tai cung da dugc nit ggn, mot so ngi dung

nghien curu da dugc chinh sua (bo phan che pham thiic an, thu ggn dien tich thi
nghiem) theo y kien cua "Hoi dong danh gia de cuong de tai trgng diem" ngay
9/4/2005. Nhan sir tham gia de tai cung thay doi do thuyen chuyen cong tac.
Cuoi Cling chiing toi muon nhan co hoi nay giri Idi cam on chan thanh tdi
Dai hoc Quoc gia Ha noi da cap kinh phi, Ban lanh dao trudng Dai hgc Khoa hgc
Tu nhien, Ban Khoa hgc cong nghe. Ban Ke hoach - Tai chinh, phdng Khoa hgc Cong nghe. Ban Chu nhiem Khoa Sinh hgc da tao dieu kien thuan Igi de chiing toi
trien khai de tai.
Trong qua trinh thuc hien de tai chiing toi cung nhan dugc sir giiip d5 cua
nhieu can bg khoa hgc va dong nghiep, nhan dip nay chiing toi xin chan thanh cam
on nhiing su giiip do quy bau do
Hd noi, thdng 4 nam 2007
Tap the can bo tham gia de tai


CHrCfNG 1. TONG QUAN
VAI TRO CUA CHE PHAM SINH HOC TRONG NUOI TRONG THUY SAN
1. DAT VAN DE
Viet Nam la nude cd tiem nang Idn trong nudi trong thiiy san, vdi bd bien dai
3260 km, khi hau am ap quanh nam, Viet Nam co tren 30 tinh nudi trong thiiy san.
Tinh den nam 2003, ca nude cd 530.000 ha nuoi tdm, nam 2005 tong dien tich nudi
trong da len den 750.000 ha. Tuy nhien, sir phat trien 6 at nghe nudi trong thuy san,
dac biet la nghe nudi tom d cac tinh ven bien dan den sir biing no dich benh tren
dien rgng, gay thiet hai nang ne cho ngudi dan. Sir pha san cua nhimg ngudi dan d
Kien Thuy - Thai Binh cung nhu d nhieu dia phuong khac la bai hoc dau xdt do chi
chay theo quy mo nudi trong ma khong quan tam tdi yeu to mdi trudng. Sir 6 nhiem
xay ra rat nghiem trgng trong chinh ao nudi va moi trudng sinh thai xung quanh do
thai ra mot lugng Idn cac chat hiju co tir thiic an du thua, tir phan va cac chat thai
khac cung nhu sir ton dong ciia cac loai hoa chat, chat khang sinh dugc sir dung
trong qua trinh nuoi trong ma khong dugc xir ly.
Benh cua thuy san thu&ng bat ngudn tir chinh moi tru&ng ao nudi. Viec

hinh thanh Idp biin day do sir tich tu lau ngay ciia cac chat hiru co, chat can va la noi
sinh song cua cac vi sinh vat gay thoi, cac vi sinh vat sinh cac khi doc nhu amoniac,
nitrit, hydrosulflia...
2. VI SINH VAT GAY BENH
Cac vi sinh vat gay benh cho tom rat da dang, cd the la virut, vi khuAn, n4m
sgi va nguyen sinh dong vat. Mot so nhiem tir tdm gidng, cdn phAn Idn cac tac nhan
nay cd san trong ao ho nudi torn,
Benh do virut co khoang 15-16 loai, thudng lay lan nhanh va lam cho ty le
tdm chet cao.
Cac benh pho bien la:
- Benh BMV (Monodon Baculovirus) thugc ho Baculoviridae
- Benh dom trang (WSSV - White spot syndrome virus) do virut thuoc ho
Nimaviridae gay nen


- B?nh dau vang (YHV - Yellow head virus) do virut tuong tu Coronaviridae
gay nen.
- Benh hoai tir dudi vd va co quan tao mau (IHHNV - Infectious hypodermal
and hematoietic necrosis virus)
- Benh teo gan tuy do virut HPV - Hepatopancreatic parvolike virus thuoc
hg Parvoviridae.
Bfnh do vi khuan rat da dang bao gom cac benh nhu den vd, den mang,
phat sang, nau mang, rung dudi, ddm den, phan trang. Vi khuan gay benh cd the
thugc chi Vibrio, Aeromonas. Pseudomonas... Tuy nhien phd bien nhat van la cac
loai thuoc chi Vibrio nhu: V. harveyi. V. alginolyticus, V. parahaemolyticus, V.
vulnificus, trong do V. harveyi la loai chu yeu gay benh phat sang. Hau het vi khuan
gay benh la bgn co hoi, chiing thudng gay nhiem trong cac trudng hgp tdm da hi
nhiem tac nhan khac, bi stress do moi trudng, bj Id loet tir trudc hoac khi thirc an
thieu can doi ve mat dinh dudng.
Benh do nam: Nam Muco bam vao mang, Fusarium lam den mang, mot so

nam khac nhu Lagenidium, Haliphtoros phat trien luon dudi Idp vd chitin. Nam cdn
gay ra benh dd than do tiet ra aflatoxin.
Benh do nguyen sinh dong vat bao gdm Gregarina ky sinh trong rugt lam
tom cham Idn, benh tom bong do Microsporea bam vao co van lam due co. Mot
so ky sinh phia ngoai nhu Zoothammium spp., Epistylis spp. gay can trd chirc nang
ho hap...
Phan lan cdc vi sinh vat gay benh ke tren Id mot phan cua he vi sinh vat
binh thu&ng trong moi tru&ng (nude bien, ao hd, song rach). Chiing dugc xem la
tac nhan gay benh thu cap hoac gay benh ca hot Mot khi sir can bang cua he vi
sinh vat trong ao nudi bi pha vd, cac vi sinh vat cd hai se phat trien d at va sdm
cong hudng vdi cac yeu to cd hai khac de gay benh. Sir d nhiem mdi trudng trong
thdi gian ngan se tac dong tdi he than kinh, trong thdi gian dai se lam tang stress
dan den giam kha nang tieu thu thirc an, giam tang trudng, lam tang man cam doi
vdi cac tac nhan gay benh dac biet la cac virut. Tat ca cac yeu to neu tren se lam cho
thuy san chet hang loat chi trong mot thdi gian ngan.


Sir canh bao tir phia Nhat Ban cuoi nam 2006 khien chiing ta can phai xem
xet lai mot each nghiem tuc ve ky thuat nudi trong de tir do dua ra giai phap hgp ly,
dam bao ve sinh an toan thuc pham neu nhu khdng mudn sau Nhat Ban, theo phan
ung Domino se la cac nude khac. Va nhu the khd Idng day manh xuat khau va thirc
hien thanh cdng nghi quyet cua Chinh phu.
Neu khdng sir dung hoa chat vd chat khdng sinh thi giai phap him hieu de
giai quyet van de 5 nhiem moi tru&ng trong ao nuoi chinh Id viec su dung cdc
che pham sinh hgc. Bien phap nay dd vd dang su dung pho bien tai cdc nude
nuoi trong thuy san & Dong vd Tay Ban cdu nhu My, Braxin, Australia, Ixrael,
Thdi Lan, Ddi Loan, Trung Quoc, Malaysia..
3. CHE P H A M SINH HQC PROBIOTIC
Probiotic (trg sinh) la thuat ngir cd nguon gdc tir tieng Hy lap, bao gdm hai
tir: Pro co nghla la vi va biosis co nghTa la sir song, la thuat ngir timg xuat hien nam

1965 nhung dugc nhac tdi nhieu trong nhung nam gan day do tinh uu viet ciia
chung. Probiotic dugc dinh nghla la che pham chira cdc vi sinh vat song da chgn Igc
ma khi dua vao ca the qua duang mieng vai lugng du lan se dnh hu&ng co lai den
sire khoe vat chu.
Ddi vdi ngudi va dong vat (gia siic, gia cam), cac vi khuan probiotic thudng
la vi khuan lactic, dugc tuyen chgn cong phu vi chung dac trung cho timg loai. Tinh
hieu qua cua probiotic da dugc chung minh bao gdm kha nang chdng nhiem triing do
vi khuan va virut (vi du virut rota gay tieu chay), kha nang chdng ung thu, kich thich
hoat dong ciia he mien dich, lam giam cholesterol, cai thien sir dung lactoza d nhung
ca the khong dung nap lactoza (khdng udng dugc sira), giam tac dung phu ciia chat
khang sinh... Sd dT probiotic cd dugc hieu qua nhu vay la vi chiing tac ddng lam on
djnh khu he vi sinh vat dudng ruot, lam tang cac vi khuan cd ich (cac vi khuan sinh
vitamin, sinh chat khang khuan, vi khuan phan giai dudng, tinh bgt.,..), lam giam cac
vi khuan co hai (cac vi khuan canh tranh thirc an, sinh chat doc...).
Trong nudi trong thdy san dinh nghia ve probiotic dugc hieu vdi nghla
rgng hon, lien quan den cd mdi tru&ng nuoi trong. Khdc vai dong vat tren can, a
ddng vat thiiy sinh, khu he vi sinh vat duang rugt khong ton tai nhu Id mot quhn


the dgc lap dac thii cho timg loai, md cd su tuong tac thu&ng xuyen vdi mdi
tru&ng va v&i ho^t dong cua vat chu. Vi sinh vat cd mat trong mdi trudng nude
anh hudng true tiep den thanh phan khu he vi sinh vat dudng rugt va ngugc lai. Cac
vi sinh vat cd mat trong dudng ruot hau het co ngudn gdc tir moi trudng. Thay cho
muc dich chu yeu la tieu diet cac bao tu vi khuan, che pham sinh hgc dugc san xuat
va dua vao sir dung vdi muc dich chu yeu la kich thich sir gia tang cua cac vi sinh
vat CO Igi trong ao nudi.
Ngay nay, che pham sinh hoc dugc coi la cong cu him hieu, tao nen tang
vung chac cho phan Idn hoat ddng nudi trong thuy san tren the gioi. Che pham sinh
hgc da dugc chap nhan rgng rai de khong che djch benh, tang sire de khang. Khac
vdi bien phap hda hoc va khang sinh, che pham sinh hgc cung cap mot phuong thirc

an toan ben vung ddi vdi ngudi nuoi va tieu diing.
7. CHE P H A M SINH HOC HOAT DONG THEO PHUONG THlTC N A O ?
Mac dii nhieu loai probiotic dugc dua vao sir dung trong nuoi trdng thuy san
trong vai thap nien qua nhung viec sir dung cac che pham nay chu yeu theo kinh
nghiem, mdi chi ap dung cac ket qua nghien cim ve probiotic tren ngudi va ddng
vat d can cho ddng vat thiiy sinh ma chua thuc su hieu mot each tudng tan co che
tac dong cua probiotic ddi vdi nhdm dong vat nay. Tuy nhien ngudi ta cho rang, bat
ky mot che pham sinh hoc nao cung phai dat dugc 3 qua trinh sau:
- Khdng che sinh hgc: nhimg ddng vi khuan cd ich trong che pham cd kha
nang sinh cac chat khang khuan, vi du bacteriocin de tieu diet cac vi khuan gay
benh trong ao.
- Tao sire song mdi: Cac vi khuan trong che pham khi dua vao ao se phat
trien manh me ca ve so lugng va hoat tinh, cd kha nang tdn tai trong mdi trudng va
trong dudng ruot, anh hudng co Igi doi vdi vat nudi.
- Xir ly sinh hgc: Kha nang phan giai cac chat hiru co trong nude giai
phong cac axit amin, glucoza, cung cap thirc an cho cac vi sinh vat cd ich, giam
thieu thanh phan nito vd co nhu: NH3, NO2, NO3'..., giam miii hoi thoi, cai thien
chat lugng nude.


8. VI SINH VAT Sir DUNG TRONG PROBIOTIC
•

•

Viec sir dung cac vi sinh vat huu ich nham canh tranh vdi cac vi khuan gay
benh da dugc img dung rgng rai trong chan nudi nhu la mot bien phap tdi uu thay
thS cho viec sir dung khang sinh, va ngay nay dang dan dugc ddn nhan nhu mot giai
phap quan trgng kiem soat benh trong nuoi trdng thuy san. Sir canh tranh cua vi sinh
vat la mot trong cac qua trinh sinh thai cd the dan den sir thay ddi thanh phan loai

cua bat ky he sinh thai nao, cho dii la dat, nude, trong co the ddng vat hoac trong
mdi trudng khac. Nhung thay doi nhd co the anh hudng tdi toe do phat trien hoac tir
vong dan den sir thay doi ve loai uu the trong quan xa. Mac dii chung ta con chua
biet het tat ca cac nhan td kiem soat toe do tang trudng ciia mot loai nao dd nhung
ciing du de co the thay ddi thanh phan loai theo quy luat ire che canh tranh. Vi sinh
vat CO the tiet ra cac chat khang khuan, tuy khong diet toan bg loai canh tranh
nhung tieu diet dii de thiet lap lai the can bang, lam giam sd lugng loai canh tranh
xuong dudi ngudng gay hai kinh te.
Thanh phan ciia che pham probiotic thudng la mot tap hgp cac chiing vi sinh
vat song, dugc tuyen chon, tdi uu hoa, lam kho bang phun say, ddng khd hoac bgc
trong alginat. Moi nha san xuat co the chon cac loai khac nhau, tuy nhien pho bien
nhat van la cac loai sau day:
- Vi sinh vat sinh phirc he enzym amylaza, proteaza, xenlulaza, lipaza... phan
giai chat hiru co khong mong muon cd trong ao nuoi nhu thuc an thira, v6 tdm, xac
tao va ddng vat thiiy sinh. Cac vi sinh vat nay bao gom: Bacillus subtilis, B.
licheniformis, B. megaterium, B. polymyxa....
- Cac vi sinh vat sinh chat khang khuan (bacteriocin, pediocin,..) gdm
Lactobacillus plantarum, L. acidophilus..., Bifidobacterium sp., Pediococcus sp.
- Ngoai ra mot so che pham con chiia nam men Saccharomyces cerevisiae, S.
boulardii, cac vi khuan nitrat hoa nhu Nitrosomonas, Nitrobacter.
Mot thanh phan khac ciing thudng dugc thay trong che pham probiotic do la
tap hgp cac enzym cd nguon goc vi sinh vat nhu amylaza, proteaza, lipaza,
xenlulaza, chitinaza, mot so vitamin thiet yeu hoac axit amin va chat khoang...
nham kich thich hoat tinh ban dau ciia vi sinh vat trong che pham va xiic tac cho sir


hoat dgng cua enzym trong mdi trudng. Cac vi sinh vat dugc lira chgn phai cd cac
d^c dilm sau:
- Khdng sinh dgc td, khdng gay benh cho vat chu va khdng anh hudng \ku
d6n he sinh thai mdi trudng.

- Cd kha nang bam dinh len niem mac tieu hda va cac md khac ciia vat chii,
canh tranh vi tri bam vdi cac vi sinh vat gay benh, khdng cho chiing tiep xiic true
tiep vdi cac co quan cua co the.
- Co kha nang sinh cac ch§t ire ch6, ngan can su sinh trudng manh me ciia
cac vi sinh vat gay benh. Cac chat nay gdm nhieu loai cd the tac ddng don le hoac
phdi hgp vdi nhau, bao gom cac chat khang sinh, bacteriocin, siderophore, lysozym,
protease, hydroperoxit...
- Co kha nang sinh trudng nhanh, canh tranh thuc an, hda chat, nang lugng
vdi cac vi sinh vat cd hai. Vi du vi khuan probiotic sinh siderophore co kha nang
lien k6t vdi ion sat, lam cho vi sinh vat gay hai khong sinh trudng dugc vi thieu sat.
- Tang cudng kha nang mien dich, tang cudng dap img mien djch tu nhien d
tdm va kha nang tao thanh khang the d ca.
- Co kha nang cai thien chat lugng nude ao nuoi do sir hinh thanh hang loat
enzym phan giai cac chat hiru co, lam giam ham lugng BOD, giam cac khi dgc nhu
H2S, NH3... Khong nhimg the, san pham trao ddi chat ciia vi sinh vat trong probiotic
cdn cung cap cac enzym, cac nguyen td da, vi lugng cho vat chii, giiip chiing sir
dung thirc an hieu qua hon va do do tang trudng tot hon,
9. KET LUAN
Ro rang mdi lo lang ve sir xuat hien cac vi khuan khang thuoc tir ao nudi
trong thiiy san do sir dung chat khang sinh, cd the truyen gen khang thudc cho cac
vi khuan gay benh d ngudi (ton tai ngay trong ao nudi tdm), lam cho khang sinh
khong cdn hieu nghiem de dieu tri benh cho ngudi nira se dugc giai tda neu thay the
bien phap khang sinh bang sir dung che pham sinh hgc. Trong thdi diem hien nay,
viec sir dung che pham sinh hoc la giai phap uu viet nhat de cd dugc nang suat, chat
lugng, hieu qua va sir phat trien ben virng ciia nganh nuoi trdng thiiy san ndi chung
va nganh nudi tom ndi rieng.


2.1. NGUYEN LIEU
2.1.1. Vi sinh vat

- Cac chung vi khuan phan lap tir dat thugc mot sd tinh khu virc phia Bac, nude
dua mudi, nude biin.
- Bg vi khuan kiem dinh: Proteus mirabilis. Pseudomonas sp., Escherichia coli.
Vibrio parahaemolyticus, Aeromonas hydrophyla....do Bao tang Gidng chuan Vi sinh
vat- Trung tam Cdng nghe Sinh hgc- Dai hgc Quoc gia Ha ndi cung cap.
2.1.2. Hoa chat
Cac hda chat lam moi trudng: -Pepton, cazein, gelatin, (Merck-Dire), Tinh bgt
tan (Anh), CMC, Kitin....
Cac hoa chat diing trong phan loai va nghien ctm: Tris-HCl; EDTA; SDS;
isoamyl alcohol; agaroza; ethilium bromid (Sigma), sorensen, axeton.. (Trung Qudc)
2.1.3. Thiet bj, dung cu
May lac on nhiet (Nhat), noi len men (Nhat), may do pH (My), may Ii tam, may
do OD, may GeneAmp PCR System, may dgc trinh tu ABI PRISM 3100 Avant
Genetic Analyzer (My); may ly tam lanh (Sigma, Dire); may soi gel (My); may do
quang phd UV3000; may chay dien di...
2.1.4. Moi tru*dng (xem phu luc)
2.2. PHUOTVG PHAP
2.2.1. Phirong phap phan lap va tuyen chon vi sinh vat
- Phan lap vi sinh vat
Can 10 g mau dat cho bao binh ndn dung tich 250 ml chira 90 ml nude vd
triing, lac deu 30 phiit tren may lac. Pha loang mlu vdi ndng do giam dSn theo co sd
10. Hiit 0,05 ml djch da pha loang d tir 10^-10^ cSy gat d8u tren moi trudng
Hutchinson-Clayton u d 30'^C. Sau 48 gid, cSy vi khu4n da mgc vao mdi trudng thach
NA de giir giong. Doi vdi cac chung lactic, phan lap tren mdi trudng MRS, ii d 30V
trong 48 gid.
- Tuyen chon
Cac chiing vi khuan dugc so tuyen bang phuong phap cay vach tren moi trudng
Hutchinson-Clayton cd bd sung cac co chat: CMC, tinh bot, kitin va tren moi trudng
thach cazein/tween. De tuyen chgn cac chiing sinh vi khu^n sinh khang sinh, vi khuan
dugc nudi cSy tren mdi trudng thach NA. Sau 72 gid, xac djnh hoat tinh khang sinh so

rong tu ddi vdi cac chiing vi khuan lactic, kiem tra
sir xuat hien cac khuan lac cd vdng phan giai CaCOa bao quanh.
- Phirorng phap xac djnh boat tinh enzym va hoat tinh khang sinh
*Phuang phap dat thdi thach
* Phuong phap nhd dich.
Hoat tinh enzym/hoat tinh khang sinh (HTE/HTKS) xac dinh theo cdng thirc:
HTE/HTKS = D - d (mm).
D: Dudng kinh vdng phan giai/vdng vd khuan + dudng kinh Id khoan/dudng
kinh thdi thach.
d: Dudng kinh 16 khoan/dudng kinh thdi thach,
- Xac djnh sinh khoi bang phuong phap do mat do quang hgc-OD d budc sdng 620nm
- Phuong phap dem so lu-ong te bao
Djch nudi cay vi khuan d cac dieu kien thich hgp dugc pha loang vdi mdi ndng
do ke ti^p giam 10 lan, Sau dd cay gat 0,05 ml djch pha loang tren dla thach mdi
trudng, giir trong tii am 30^C. Sau 24 gid d^m sd lugng khuin lac mgc tren dla Petri.
Cdng thuc tinh sd lugng t6 bao:

So lu'ong te bao = ax 1/Kxl/V

a: sd khuan lac trung binh moc tren dTa thach.
K: do pha loang.
V: the tich dich pha loang dugc cay gat tren dTa thach.
- Phu-ong phap xac djnh vi khuSn G(+) hay G(-)
Cd dinh tieu ban vi khuan bang ngon lua rdi nhudm b^ng dung dich tim kk tinh
trong khoang mot phut. Rira bang nude. Nhudm tidp bang dung dich lot (dung djch
Lugol) trong mot phut. Rira b^ng nude. Phii len \k boi dung dich etanol 95%: axeton
(1:1) trong mot phut va lai rira bang nude. Sau dd nhudm tilp b^ng thu6c nhudm mau
dd Saft-anin trong 30-60 giay. Rua qua nude, dh kho rdi soi kinh. Nhom G(+) se khong

bi dung moi hiru co (etanol, axeton) xky phirc ch^t mau giira tim kit tinh va idt nen se
bat mau tim. Nhom G(-) bj dung mdi tay mau thu6c nhudm mau diu nen se bit mau
d6(Safranin)[15].
- Phu-ong phap chuSn do Therner
De xac djnh ham lugng axit ciia cac chung vi khuAn lactic, \iy 10 ml djch sau
nudi cky b6 sung 20 ml nude ck va 1-2 gipt chi thj phenolphtalein 1%. Chuin do bang
NaOH 0,1N den khi xuat hien mau hong nhat b^n trong 30 giay thi dimg lai. Xac dinh
so ml NaOH da chuan do. Do axit dugc tinh theo cong thirc:
°T = VwaOH X 10
"^rr^rxn AA ^ T 1Q Ar\ T h o r n *

% Axit lactic - ^T x 0,009

"• ' '^o Therner tuong duong vdi 9 mg axit lactic.
10


bacterioxin
Ly tam djch nudi cky vi khuSn lactic bang may ly tam lanh vdi toe do 10.000
vdng trong 15 phut, loai bd kk tiia, thu iky phkn dich trong. Bo sung axetol lanh vao
djch nay theo ty le thi tich 1:1. Sau khi giir lanh 4^C qua dem, ly tam thu ket tiia, lam
khd tua d nhiet do phdng va hda tan tua bang dem Sorensen 0,02M dSn pH 7. Thir hoat
tinh khang sinh theo phuong phap khuSch tan tren thach [58].
- Phirong phap djnh lirong kitinaza
Kitin la mot polyme dugc cku thanh tir cac don phan la cac phan tu N-acetyl-Dglucosamin (GlcNAc) n6i vdi nhau qua lien kk (3-1-4. Ham lugng kitinaza dugc xac
dinh d budc song 585 nm thdng qua ham lugng N-acetyl-D-glucosamin (GlcNAc) giai
phdng ra dudi tac dung ciia enzym nay.
Phuangphdp tien hanh: tron nhirng th6 tich bang nhau (0,2 ml) djch enzym tho
vdi 0,5% keo kitin trong dem axetat 0,05M, pH 6. Phan img dugc thuc hien d 37^C
trong 30 phut. Ham lugng GlcNAc dugc giai phong ra dudi tac dung ciia enzym dugc

do d budc sdng 585 nm bang may quang ph6. GlcNAc (Sigma, My) dugc diing lam
tieu chuan, vdi he sd hap phu la 2,4 x 10 [52].
Dua vao dinh luat Lambert-Beer ta tinh dugc ndng do ciia GlcNAc giai phong
ra theo phuong trinh:
Trong do:

OD585 = KCul (*)

K la he s6 hkp phu (K=2,4xl03)
Cu la nong do chat hap phu (GlcNAc)
I la chieu dai anh sang qua dung dich (1=1 cm).

- Phirotig phap xac djnh anh hudng cua cac dieu kien nuoi cay den kha nang tong
hop enzym va khang sinh
a. Lua chgn moi tru&ng nuoi cay thich hgp
Vi khuan dugc nudi tren mdi trudng thich hgp d 28-30^C hoac 40-45°C. Sau 48
gid nudi cay xac djnh hoat tinh enzym, pH sau nuoi cay, va sinh khdi te bao. Sau 72
gid nudi cay, xac dinh hoat tinh khang sinh, pH, va sinh khoi te bao.
b. Anh huong cua thdi gian
Vi khuan dugc nuoi cay tren mdi trudng thich hgp d 28-30'^C hoac 40-45V. De
xac dinh thdi gian thich hgp cho sir tdng hgp enzym, chat khang sinh va ddng thdi xac
djnh sinh khdi. Nuoi cay trong vong 72 gid, sau mdi 12 gid xac djnh hoat tinh enzym,
sau mdi 24 gid xac djnh hoat tinh khang sinh va xac djnh sinh khoi bang phuong phap
do mat do quang.
c. Anh huong cua pH nuoi cay

11


mdi trudng thich hgp d 28-30°C hoac 40-45X, pH

tir 4 dfin 9. Sau dd xac djnh pH sau nuoi cky, sinh khdi, hoat tinh enzym (sau 48 gid)
va khang sinh (sau 72 gid).
r

d. Anh hu&ng cua nhiet do nuoi cay
Vi khukn dugc nudi cky d cac nhiet do: 20; 25; 30; 35; 40; 45; 50^C,.. Sau 48
gid xac djnh pH sau nudi cky, sinh khdi, hoat tinh enzym. Sau 72 gid xac djnh pH sau
nudi cky, sinh khoi, hoat tinh khang sinh,
e. Anh hu&ng cua nguon cacbon vd nito
Vi khuan dugc nudi cay tren moi trudng thich hgp cd bo sung 1% cac ngudn
cacbon hoac nito hiru co, v6 co nhu: tinh bgt, glucoza, saccaroza, CMC, kitin, cam,
hot dau tuong, cao men, pepton... Sau 48-72 gid, xac djnh kha nang sinh trudng va kha
nang sinh enzym, khang sinh ciia cac chiing.
- Anh hudng cua mot so yeu to den boat tinh enzym
Cac chung vi khukn cd kha nang sinh enzym dugc nuoi cay lac 220 vong/phiit d
diSu kien toi uu. Sau 48 gid, ly tam 8000 vong/phiit trong 15 phiit d6 thu djch enzym
tho.
a, Anh huong ciia nhiet dp den hoat tinh enzym
TiSn hanh theo phuong phap nho djch. Sau dd dat cac dTa Petri vao thang nhiet
dp: 20^ 30^ 40^ 50^ 60^ Sau 24 gid, do vdng phan giai co chkt
b, Anh hu&ng cuapH den hoat tinh enzym
Tikn hanh theo phuong phap nho djch, nhung tren co chat dugc pha trong dung
djch dem tao ra gia trj pH 3, 4, 5, 6, 7, 8, 9. Sau 24 gid do vong phan giai co chat.
c, Anh huong ciia thdi gian bao quan den hoat tinh enzym
Djch enzym tho dugc bao quan d nhiet do -4^C trong 5 tuan. Tien hanh thir hoat
tinh theo phuong phap nhd djch sau mdi tuan.
- Xac djnh kha nang chju man
Vi sinh vat dugc cay tren mdi trudng thich hgp cd ndng do mudi tang dan tir
0% den 12%. Sau 48-72 gid, pha loang djch nudi d nong do thich hgp xac djnh kha
nang sinh trudng, hinh dang va kich thudc khuan lac theo phuong phap dem sd lugng

te bao va xac djnh hoat tinh sinh hpc cua chiing,
- Xac djnh tinh doi khang
Cac chiing vi sinh vat dugc tien hanh thir tinh doi khang theo phuong phap nho
djch tuong tu nhu xac djnh hoat tinh khang sinh.
- Phuong phap tao che pham
a, Lua chgn chat mang cho len men xdp: Cay gidng cap hai vao cac moi trudng
cd chira cac loai chat manp khac nhau nhu: Bgt gao, bgt dau tuong, cam, bgt thoc mam
12


m. Bd sung nude den dp am 45-50%. U 48 gid d
30**C, sau dd xac djnh sd lugng ik bao ciia timg loai vi khukn.
b. Ty le gidng cay thich hgp: Gidng ckp hai dugc bd sung vao mdi trudng chira
chat mang thich hgp ddi vdi timg loai vi khuSn theo ty le khac nhau: 0,1-0,5-1-5-1013-15-20 %. Bd sung nude d k do im thich hgp va u 2 ngay, sau dd dim sd lugng ik
bao cua timg loai vi khuSn dk xac dinh kha nang sinh trudng cua cac chiing.
c. Thai gian ii thich hgp: Chat mang sau khi bd sung gidng va lam am dugc u d
nhiet do 30V. Cir sau 24 gid tiln hanh lay mlu xac dinh sd lugng te bao cua timg loai
vi khuan.
d. Thai gian bao quan cua chi phdm: Djch len men sau thdi gian thich hgp
dugc trdn vdi chat mang theo ty le gidng cky thich hgp, sau khi u dem sky. Trdn cac
thanh phan va nghien nhd, ddng vao cac gdi polyme hai Idp 250g. Kiem tra bien dong
s6 lugng timg loai vi khuan va hoat tinh sinh hgc cua che pham moi thang.
- Thir nghif m nuoi tom (xem chuong 3)
a, Nuoi torn sd trong phdng thi nghiem
Tdm su dugc nuoi trong be kinh, the tich mdi be 100 lit, nude lay tir ao lang
nude nudi tom, vdi dp man 25%o, mdi be tha 100 con tdm su vdi trgng lirgng ban dau
0,7- 0,8g va kich thudc 2,5-3cm. Be thi nghiem bo sung 0,5gA che pham vi sinh, be ddi
chung khdng bo sung che pham. Hang ngay cho tdm an thiic an cong nghiep va sue khi
trong qua trinh nuoi. Cir sau 7 ngay thay 1/3 nude va can do trgng lugng ciia tom.
h, Khao nghiem che phdm VSV ngodi ao nuoi torn

Thi nghiem dugc tien hanh d dia diem Nuoi tom cong nghiep: Xi nghiep nuoi
trong thiiy san Dinh Vu, Hai Phdng: Dac diem ao nudi d vu thii 3, dp sau ciia nude dat
tir 1,0-1,2m, cac ao tom xay dung tren nen rimg ngap man, trudc dd la bai cay sii vet
day dac. So do bd tri thi nghiem nhu sau: Tdng 3 ao nudi tdm, 2 ao thi nghiem, ao sd 1
CO dien tich 10.000m-, ao sd 3 cd dien tich 5.000m^, ao sd 2 ddi chung cd dien tich
5.000ml
Ao
1
2
3

Cong thuc
Thi nghiem
Ddi chiing
Thi nghiem

Cbe pham
EMUNIV-S
Khdng sir dung
EMUNIV-S

Mat do
(con/m^)
20
20
20

Nen day
Dat Cling
Dat Cling

Dat cung

+ Tat ca cac ao deu dugc xir ly trudc khi thi nghiem theo dung quy trinh cua ao
nudi cdng nghiep (xir ly phai day, xir ly nude bang chlorin, diet tap bang saponin, khir
f r n n o K a n o v A i hAt^

13


lay nude, han che sir dung hda chat trong chu ky
nudi. Trong cac ao deu cd he thdng quat nude, cung cac che do cham sdc... Theo ddi
cac chi s6 mdi trudng thuy ly, thuy hda ... dinh ky 10 ngay mot Ian.
+ Trong qua trinh nudi tat ca cac ao deu sir dung cung thiic an tdm cdng nghiep.
+ Ao sd 1 va sd 3 bo sung che pham vi sinh chu ky 10 ngay mot lan. Thang thii
nhat 2,5kg/ha/lSn. Thang thii hai 4kg/ha/lSn. Thang thii 3 trd len 6-7kg/ha/ian.
+ Phan tich nude theo tai lieu "Hudng dSn phan tich mSu cho ao nudi trong thiiy
san" cua Claude E. Boyd & Craig Stueker vdi may quang pho UV/vis - Hitachi,
Model: U-2010.
+ Xac dinh cac chi sd thuy ly, thuy hda theo tai lieu [26].
-H Thu mau sinh vat ndi bang ludi vdt hinh chdp ndn, dudng kinh mieng ludi
25cm, chieu dai 90cm. Vai ludi vdt thuc vat ndi cd 75 (75 sgi/cm^), vai ludi ddng vat
ndi CO 49.

+ Mau sinh vat ndi dugc cd djnh trong dung dich formalin 5%.
+ Kiem tra tdc dp sinh trudng cua tdm dinh ky 15 ngay mot lan. Danh gia ty le
sdng va nang suat tdm tinh toan sau khi thu hoach.
+ Kiem tra sinh trudng cua tdm bang phuong phap do chieu dai tdm theo dinh
ky bang thudc chia dp. Can trgng lugng tom bang can dien tir cd dp chinh xaclO'^g,
-h Ty le tdm sdng dugc xac dinh bang phuong phap "sir dung sang an" trong
thang nuoi thii" nhat, phuong phap quang chai va xac dinh sd lugng ca the/m^ tir thang

nudi thir hai den khi thu hoach, ket hgp vdi ket qua can dem sau khi thu hoach.
+ Xac djnh nang suat nuoi tdm sau khi thu hoach, trir cac chi phi thii:c an, con
gidng, hda chat va cac chi phi khac.
+ Thdi gian khao nghiem bat dau tir 28/03/2006 den 30/08/2006.

- Phuong phap xac djnh mot so dac diem sinh hoc cua vi khuan
Xac dinh khd ndng sinh bdo tu
Cay vi khuan tren moi trudng thach thudng. Sau 3 ngay tien hanh xir ly nhiet d
80^C. Diing vi khuan da xir ly nhiet cay lai tren dTa thach thudng dat d 30^C. Sau 24
gid neu xuat hien khuan lac thi ket luan chiing vi khuan cd kha nang sinh bao tir. Neu
khdng thi ket luan ngugc lai.
Xac dinh khd ndng hda long gelatin
Cay vi khuan theo phuong thang dimg, sau vao ong nghiem moi trudng gelatin
dung. Sau 24 gid ii d 30V, lay ong nghiem ra giir d 4 V trong 1 gid. Kiem tra ket qua
neu thay mdi trudng van long thi ket luan co kha nang hoa Idng gelatin. Ngugc lai la
khdng cd kha nang.
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