01书!"#$%&’()*+,-./*01!"#,$%&,’((...

5
!"#$%&'()*+,-./*01 !"#$%&'(( !"#$%&#$'&()*&+,-#$./012345!"67 316022 23 )*+,-./0123456789:;<=>?@ABCD6E*FGHIJBKLMN#OPQRABCDSAB TNUVQWAB*8XYZ[\]34^_@AB6E*B_`abcde。[ #Ofg89hijk+,-lm01234 567n<IJBBopqOr:sn<tuvwxyzBStaphylococcusaureus ?{Q|B}L*Bo~OQPQ|B CDO:n<WABoKLBEscherichiacoli )、 wzBMoniliaalbican SBBacillus subtilis }[BPQRABT。[ tBo GP57 S GT16 HHi ¡¢ 16SrDNA8£VQR¤[¥¦B §Streptomyces ), ¨©ª8«[ 99%S 100%R7 GT16 VQ[¬-¥¦BStreptomycesgriseoplanus )。[ ®34FGH¯H673 4IJB°±²{X³´*@ABCD6*B_µ<¶{·*\]456 0123456FGH¯H6ABCD:; 78+9: S917  ;<=>? A  ;@A: 0517-6611 2018 21-0001-04 SeparationofMarineActinomycetesandScreeningofAntagonisticBacteria TIANYunfang FUZeqin YEYingying NationalEngineeringResearchCenterforMarineAquaculture MarineScienceandTechnology Colleage ZhejiangOceanUniversity Zhoushan Zhejiang316022 Abstract Objective Symbioticorepiphyticactinomyceteswithantibacterialactivesubstanceswereisolatedandscreenedfromcommoninter tidalzonemarineanimalsinZhoushan.Differentmethodswereusedfordeterminingtheantimicrobialactivityandantimicrobialspectrum the classificationstatusofantagonisticbacteriawasalsoidentifiedforprovidingbasicdataforstudyingthesourceofmarineantibacterialsubstances. Method Throughseparationandpurificationtechnology strainswereobtainedfromseveralintertidalzonemarineanimalsinZhoushan.Strains withcertainbacteriostaticeffectsonStaphylococcusaureus wereobtainedafterinitialscreeningbyspotinoculation.Antimicrobialactivitywas quantitativelydeterminedbyenzymeassay.Thehighefficiencyantagonisticbacteriastrainswereobtainedbyfilterpaperdiffusionmethod. Esche richiacoli Moniliaalbican andBacillussubtilis wereusedasindicatorstodeterminetheirantimicrobialspectrum. Result Thetypicalstrains GP57andGT16wereidentifiedbyphysiological morphologicaland16SrDNAmolecularidentification.Allofthemwere Streptomyces withasim ilarityof99%and100%respectively GT16wasidentifiedasa Streptomycesgriseoplanus. Conclution Amongmarinesymbioticorepiphyticmi crobes marineactinomycetesmaybeaverygoodsourceofbacteriaproducingantibacterialactivesanddeservefurtherstudy. Keywords Intertidalzone Marineanimals Symbioticorepiphyticmicrobe Antimicrobialactivity Screening BCDE !"#1990 —), ¸¹º»¼½¾¿\]H\]#À34H6fÁ}ÂÃ\]ÄÅ¿+Æ8£Ç ÈÉq\]FGHI 2018-04-02 #$89:;<=#$>?@A4BCDEF;G: <HIJEKLMNOPQRSTUB=>?@A4;V WXYZ[\]^%_`EabXcG;d#$@Ae 89:Ff;OPIEghXc #$i;IEjkH lmU#$@AEnCHSE=#opqr;<SE= st#uvwIx#yEz{l|}~Ss(#$@ A4EIU;;x|;Ei; IZ|;i;I |;Ee x M; I=;<=s;|; t E|;i;IjU#$IE ¡ ¢ ¡£¤_¥¦%§¨©ª#$I «K¬j :89:-:®|;¯j°SF; ;I<HEf±OPFI 3-5 EKL#$|;i;Ie #$i;IEgh²°SF;;I<HIJE³% ´L µ¶X[\·#¸¹º®#$@AI|;i ;IE»¼½j»¼z¾tI¿ÀÁEi;I ÂjBi;IOPF;E<HIJYÃZÄj'&-»¼ EÅÆÇÈÉÊ#$|;i;I4ËÌj°SÍ Î;I<HEIJÏ;dÐdG;dÑÒÓÔ ÕÖNÖ×ØÙGÚÛ<HIJÜd®tab< HIJ°SÝKÞßàáâãä[åæç®CH£Êi;I4]èÌEOP<HIJéÍUIXcê ë¯;Fì3íîÑïÑðñò;ó@Aô¯ãU2õ ö;FÇÈ©ªEst#$;I<HIJ÷er(Iø ù;Ei;I¿F;E 7-10 |;i;IEÃ; I<HÑï(øùSô¯(B¿Eúû@ASü ýÇÈÊúû@Aþÿ»¼B·i;I!(;I<H ®"{#$E%&'â (X(jE»¼z¾#$89 :°SF;))kCDE;I<HOPIÖE´Lô t°S%E'&»¼*+,áE-./0}1 ;I&"234,á`E50675089Ñ :©;;Iö&Ø<®é=>?@A&'E©;Z #$89:F;E<HOPFIE»¼B©]^CÎS LE)*DErUÇÈ·#$89:EFG50HV =j)*EIJKHbS%¤_E#$89:L K50MáNOM67®úPÁ#$IZ»¼·QFáÑR50SËTöUVWÌ 152 X°SG:<HE :XYW°SÝÞ×:<HE:XÂZ1[\S×: àáE:XM]Z»¼PÁ#$I|;i;I]^ S^1 JK-LM 1.1 JK 1.1.1 k_`a%bcMetopograpsusquadridentatus )、 µ NOPQRSJ.AnhuiAgric.Sci.2018 46 21 ): 1-4 17

Upload: others

Post on 04-Oct-2020

4 views

Category:

Documents


0 download

TRANSCRIPT

Page 1: 01书!"#$%&’()*+,-./*01!"#,$%&,’(( (!"#$%&#$’&()*&+,-#$./012345,!"67 316022)23 [)*]+,-./0123456789:;?@abcd6e*fghijb,klmn#opqrabcdsab t,nuvqwab

书书书

!"#$%&'()*+,-./*01

!"#

,$%&

,'((

 (!"#$%&#$'&()*&+,-#$./012345

,!"67

316022)

23

 [)*

]+,-./0123456789:;<=>?@ABCD6E*FGHIJB

,KLMN#OPQRABCDSAB

T

,NUVQWAB*8XYZ

,[\]34^_@AB6E*B_`abcde

。[#O

]fg89hijk

,+,-lm01234

567n<IJBBo

。pqOr:sn<tuvwxyzB

(Staphylococcusaureus)?{Q|B}L*Bo

;~��OQ�PQ|B

CD

;�����O�:n<��WABo

;KL���B

(Escherichiacoli)、�w��zB

(Moniliaalbican)S�����B

(Bacillussubtilis)

}[��BPQRABT

。[��

]t��Bo

GP57S

GT16��H�Hi

、��� ¡¢

16SrDNA8£VQ

,R¤[¥¦B

§

(Streptomyces),̈©ª8«[

99%S

100%,R7

GT16VQ[¬­¥¦B

(Streptomycesgriseoplanus)。[�®

]34FGH¯H67

,3

4IJB°±²{X³´*@ABCD6*B_

,µ<¶�{·*\]

456

 012

;3456

;FGH¯H6

;ABCD

;:;

78+9:

 S917  ;<=>?

 A  ;@A:

 0517-6611(2018)21-0001-04

SeparationofMarineActinomycetesandScreeningofAntagonisticBacteriaTIANYunfang,FUZeqin,YEYingying (NationalEngineeringResearchCenterforMarineAquaculture,MarineScienceandTechnologyColleage,ZhejiangOceanUniversity,Zhoushan,Zhejiang316022)Abstract [Objective]SymbioticorepiphyticactinomyceteswithantibacterialactivesubstanceswereisolatedandscreenedfromcommonintertidalzonemarineanimalsinZhoushan.Differentmethodswereusedfordeterminingtheantimicrobialactivityandantimicrobialspectrum,theclassificationstatusofantagonisticbacteriawasalsoidentifiedforprovidingbasicdataforstudyingthesourceofmarineantibacterialsubstances.[Method]Throughseparationandpurificationtechnology,strainswereobtainedfromseveralintertidalzonemarineanimalsinZhoushan.StrainswithcertainbacteriostaticeffectsonStaphylococcusaureuswereobtainedafterinitialscreeningbyspotinoculation.Antimicrobialactivitywasquantitativelydeterminedbyenzymeassay.Thehighefficiencyantagonisticbacteriastrainswereobtainedbyfilterpaperdiffusionmethod.Escherichiacoli,MoniliaalbicanandBacillussubtiliswereusedasindicatorstodeterminetheirantimicrobialspectrum.[Result]ThetypicalstrainsGP57andGT16wereidentifiedbyphysiological,morphologicaland16SrDNAmolecularidentification.AllofthemwereStreptomyceswithasimilarityof99%and100%respectively,GT16wasidentifiedasaStreptomycesgriseoplanus.[Conclution]Amongmarinesymbioticorepiphyticmicrobes,marineactinomycetesmaybeaverygoodsourceofbacteriaproducingantibacterialactivesanddeservefurtherstudy.Keywords Intertidalzone;Marineanimals;Symbioticorepiphyticmicrobe;Antimicrobialactivity;Screening

BCDE

 !"#

(1990—),̧ ,¹º»¼½

,¾¿\]H

,\]#À

34H6�

。fÁ}Â

,Ã�\]Ä

,Å¿

,+Æ8£Ç

ÈÉq\]

FGHI

 2018-04-02

  #$89:;<=#$>?@A4

,BCDEF;G:

<HIJEKLMNOPQR

,STUB=>?@A4;V

WX

,YZ[\]^%_`EabXcG;d

。#$@Ae

89:Ff;OPIEghXc

[1]。#$i;IEjkH

lmU#$@AEnCH

,SE=#opqr;<

,SE=

st#uvwIx#yEz{l|}

,~Ss��(#$@

A4E��I�U�;

、��

、�;x|;E��

,��i;

I�Z�|;i;I

[2]。�|;�Ee

2�x

2�M�;

I=�����;<=s�

,���;�|;

2�

。�t�

�E�|;i;I�j��U#$����IE�� ¡

¢��� ¡£

,¤_¥¦%

。§¨©ª#$����I�

«�K�¬j :

、89:

、­:®�|;

,̄�j°SF;

;I<HEf±OPFI

[3-5]EKL

。#$�|;i;Ie

#$i;IEgh�²

,°SF;;I<HIJE³%

´L

[6]。

µ¶X

,[\·#¸

、¹º®#$@A��I�|;i

;IE»¼½j

,»¼z¾�t��I¿ÀÁEi;I��

Âj

,Bi;IOPF;E<HIJYÃZÄj'&-»¼

EÅÆ

。ÇÈÉÊ#$�|;i;I4�ËÌj�°SÍ

Î;I<HEIJ

,��Ï;d

、Ðd

、G;d

、ÑÒ�ÓÔ

Õ

、Ö�NÖ×ØÙ

、GÚÛ<HIJ

、Üd®

。�tab<

HIJ°SÝKÞ

、ßàáâ

、ãä[åæç®CH

。�£

Êi;I4]èÌEOP<HIJ

,éÍU��I

,Xcê

ë

,̄;Fì3íî

,ÑïÑðñò;ó@A

,ô¯ãU2õ

ö;F

ÇÈ©ªEst#$;I<HIJ÷er(��Iø

ù�;Ei;I¿F;E

[7-10]。�|;i;IE�Ã�;

I<HÑï(øùS�

,ô¯(B¿�Eúû@AS�ü�

,ýÇÈÊúû@Aþÿ»¼B·i;I�!(;I<H

®"{#$E%&'â

[2]。(X(jE»¼z¾

:#$89

:°SF;)�)kCDE;I<HOPI�ÖE´L

,ô

�t°S�%E'&»¼�*+,áE-.

。/0}�1

;I&"2Eë34��,á

,̀E5067�5089Ñ

:©;

,;I

、ö&

、Ø<®é=>?�@A&'E©;

,Z

#$89:F;E<HOPFIE»¼�B©]^CÎS

LE)*DE

。rUÇÈ·#$89:EFG�50HV

=�j)*�EIJ

,K�HbS�%¤_E#$89:L

K50Má

。NOM67®úP�Á#$�IZ»¼·Q

FáÑR50S�ËTöUVWÌ

152X°SG:<HE

:X

,YW°SÝÞ×:<HE:X

,ÂZ�1[\S×:

àáE:X

,M]Z»¼P�Á#$�I�|;i;I]^

S^

1 JK-LM

1.1 JK

1.1.1 k_

。`a%bc

(Metopograpsusquadridentatus)、µ

NOPQRS

,J.AnhuiAgric.Sci.2018,46(21):1-4,17    

Page 2: 01书!"#$%&’()*+,-./*01!"#,$%&,’(( (!"#$%&#$’&()*&+,-#$./012345,!"67 316022)23 [)*]+,-./0123456789:;?@abcd6e*fghijb,klmn#opqrabcdsab t,nuvqwab

"de

(OstrearivularisGould)、fgh

(Sebastiscusmarmoratus)、

ijkl

(Tetraclitasquamosa)�mnopq

(Siphonariajaponica),

rFGU67®úP�Ást

1.1.2 �u:

。vwÜxyz:

(Staphylococcusaureus)、%

{|:

(Escherichiacoli)、}~��|:

(Bacillussubtilis)�

���:

(Moniliaalbican)。1.2 

TUV

 89:�Ë50S

:�s���50S

[11]。

1.3 WXY%&'Z'$[+,\]

 k_���û

,�

ÿ��

,á�!2���ÿZ

10-2~10-4E����!UÝ

�s���50S�

,g�

3���

,30℃50

,·:X��

��

、��

,Má���9�Ë2�èî:�

,jf�9�Ë

。.�

3�·��

,·��50SÑ�!

1.4 ./*^[_0

 vwÜxyz:àZ�u:

,áÆ

�2·k_���U

。��ËTöî:XÆ�U� �u

:E50S�

,3���

。30℃50

,3、5、7d)¡¢

1f

UVS×:£E:X¤��¥�

1.5 `=Mabca

 M

630nm�æ¦ÿZ

0.01Evw

Üxyz:�u:50�Z·�

,áÖ§¨2

[12]©ª

2h\

_«\:XEæ¦ÿ

,MWÌ:XE×:Þ¬

1.6 d*ecb

 ��U:X©­50

6d,©­�®¯Ë

5

,á°±²³´2�©­��©­�µ�¶UvwÜxy

z:����

,3���

,30℃50

,«_¤·¸×:£

1.7 fg/*hi*^/*jkc

 M%{|:

、�Ü�

�:�}~��|:Z�u:

,�º«:X©­��©­�

µ�á°±²³´2�»¼2½«

,30℃50

,¡¢·¸

1.8 /*hi*^[la

1.8.1 ¾ó&¡¢

。¿À

《i;I&*Á

》[13]4E²2

�ò502¡¢

4XÝÞÄG:XE:Å

、�1Å��1

¾óCÆ

,¤��;û;ö[\

1.8.2 �1[\

。á

MPÇÙÈ]è

4Xº«:XÉ

DNA,bZÊì

16SrDNAËÌ�Í

,Fá�ÎÏI

341F(5′CCTACGGGAGGCAGCAG3′)

� W Î Ï I

907r(5′CCGTCAATTCCTTTRAGTTT3′)

��

PCR³Ð

,áÑÒó ¡Ó

Ô

、Õö

、�Ö×

,ØJÙ]èÁÚSÛÜ£cÂݲ9EÕ

ö1

,MÃå�EÊáÏIàZ«ËÏIËÌ

,Þß�#à

á;I«Ë

。¿À«Ëâ¬á

Blastãä(

GenBank¤Àå

4E

16SrDNAËÌæ·

,á

MEGA5.0ãä��ËÌ�Í

¤çè��©éê

2 mno+p

2.1 qr*s'tuv

 À¡¢

,Ý��!ëì�í

,îb

��4Sstoïð#$Cî:�E;ñ

,òó%²:�¾

Ã:ô

,ý:�EÉå;ñëìÍí

,õU�V

(z

1)。

w

1 qrTUV*s'txv

Table1 Colonygrowthoncoatculturemedium

k_

Sample��ÿ

Dilution :�¤

Thenumberofcolony∥X

µ"de 10-2 8 26 8(+2:ô

OstrearivularisGould 10-3 0 0 010-4 1 0 0

fgh 10-2 38(+14:ô

) 38(+13:ô

) 64(+13:ô

Sebastiscusmarmoratus 10-3 2(+7:ô

) 1 310-4 0 0 0

mnopq 10-2 52(+3:ô

) 20(+9:ô

) 39(+6:ô

Siphonariajaponica 10-3 8 3 110-4 0 0 1

`a%bc 10-2 81(+14:ô

) 103(+11:ô

) 64(+13:ô

Metopograpsusquadridentatus 10-3 58 24 3110-4 1 0 0

ijkl 10-2 49(+15:ô

) 24(+%²:ô

) 116(+10:ô

Tetraclitasquamosa 10-3 10 13 5(+2:ô

10-4 �

0 0 2·�

Control 0 ö

:M÷øù¡¢ÌEîX:�Z

1X:�

 Note:Asinglecolonyobservedwiththeeyesisonecolony

2.2 +,*^_y01-\]TU

 Ø�ËTö

,Êk_

4�YW#$89:

152X

,B4k_`a%bcúSE8

9:¤_Âj

(77X

),k_fgh�mnopq4YWE

89:é·Íâ

,µ"de4YWEÂâ

(4X

)。�:X�

�§�

,Gzu�s���50S

;Pzu`a%bc

;Hzufgh

;Mzuµ"de

;Tzuûjkl

;Jzumn

opq

,¤üOz:X�

2.3 `=zM{_0*^|0

 Ö§¨2\_«\:X=

ýþ�u:Z;ñs9ÿ�Eæ¦ÿ

,â¬z¾:Xæ¦ÿ

r=

0.2!"

,°SÍÎE×:àá

,·vwÜxyz:E

;ñSÍÎE×Øàá

(#

1)。ØìUV

,V$:X

GY11、GP17、GM2、GY7、GP48、GP6、GP57、GP71、GT16

9Xæ¦

ÿ=

0.1!"E:X

2.4 |0*^/*hikc

 ��UZE:XáÆ�2Ô

�U���u:E50S�

,½«B×:£%R

,â¬z¾

B4

4XrSÍÎE×:àá

。«_©ª

GY11:X×:£

%RZ

(9.590±0.001)mm,B&

3X89:rZ

9mmMW

ÑR:X'�E×:Þ¬ÑR

(#

2)。

2           ÊËÌÍÎ�

                         2018}

Page 3: 01书!"#$%&’()*+,-./*01!"#,$%&,’(( (!"#$%&#$’&()*&+,-#$./012345,!"67 316022)23 [)*]+,-./0123456789:;?@abcd6e*fghijb,klmn#opqrabcdsab t,nuvqwab

8

1 ~t

630nm�*^[���

Fig.1 Absorbanceofstrainsatthewavelengthof630nm

8

2 ��*^{������*[d*gn

Fig.2 ThebacteriostaticeffectofdifferentstrainsonStaphylo

coccusaureus

2.5 ./*/*jca

 G:(«\⬩ª:X

GP57

·

3��u:rSGH

,:X

GP71、GT16·�Ü��z:�

%{|:°SGH

,GY11·�Ü��z:�}~��|:

°SGH

(z

2)。

w

2 4̂%&'()*[/*hi

Table2 Antibacterialactivityof4symbioticorepiphyticactinomyce

tes

:X

Strain���z:

Moniliaalbican%{|:

Escherichiacoli}~��|:

BacillussubtilisGP71 ++ + -GP57 + ++ +GY11 + - ++GT16 + ++ - ö

:+zuSGH

;++zuSÍÎEGH

;-zu�GH

 Note:+indicatedresistance;++indicatedstrongresistance;-indicatednoresistance

2.6 +9��la

2.6.1 :X�1[\

。M

GP57N

GT16Z)�Í*�:X

E�1¾óCÆ

,�1¾ó+,-

(#

3)。

8

3 *^���Z��

Fig.3 Microscopicmorphologicalcharacteristicsofstrainsspore

2.6.2 ;û;ö[\

。¾.�ö

:4X:rK/¾.©;0

1�öªQ

APIÇÁâ¬

:APIÇÁ4©ª:X

GT16、GY11rKM

á

19�2c

,2cMá3Íí

  4iW¡¢ð;:Å�S¢:žóNB5Ü

,⬩

ª

4X:r4u*�E89::�CÆ

(z

3)。2.6.3 

�1[\

。¿À��©éê

,GP57�

GT16r(,

6:7RcHÍÝ

,B4:X

GT16(8�,6:RcHÂ

Zéµ

,Z

100%。K�

,�:X

GT16[\Z8�,6:

(Streptomycesgriseoplanus);ô:X

GP57ÎËUê£

,ý9Ë

͵

,â ¾ó&CÆ�;û;öâ¬

,[\Z,6:7

346Ï

21Ð

                 !"#Ñ

 3456FGHIJB89¢WAB:;

Page 4: 01书!"#$%&’()*+,-./*01!"#,$%&,’(( (!"#$%&#$’&()*&+,-#$./012345,!"67 316022)23 [)*]+,-./0123456789:;?@abcd6e*fghijb,klmn#opqrabcdsab t,nuvqwab

(Streptomyces)。

w

3 4̂./*^[TU��

Table3 Theculturecharacteristicsof4antagonisticstrains

:X

Strain:�¾ó

Colonialmorphology

ð;:Å

Aerialmycelium

S¢:Å

Substratemycelium

GP71:;

,<=Ñ>? 8@Ü

,A- BCÜ

GP57:;

,<=Ñ>? �Ü

,A- wÜ

GY11:;

,<=Ñ>? �Ü wÜ

GT16:;

,<=Ñ>? �Ü CÜ

3 m�o��

NOFG67stP�Á�

5�k_

,�ËTö$

152X:X

,ÇÁk_D�XcUst

。=·øùk_EVE�

ùMêþFG

,Ñïù=st�Fk

,ô¯ù=yt�Ht

�Fk

,M]YWêFGE�|;i;IIc

=UVÄG:XEì34JáÆ�2

、°±²2�Ö§

2

,©ªéR:X=ÑRUV"2W¿4uE×:àáÑ

R

,ÂZV$

9X×:Þ¬é·ÍíE:X

。ôZá°±²

2�Uÿ

,KnS×:àáE:X

GP48、GP16、GY7、GP6�

GM2LS×:£F;

,ÂMUV$

4X×:Þ¬ÍíE:X

ZÇE:X

。µ¶X

,NO"®

[14]»¼©ª@APQðR

ò ¡©;S,HTU

,VWK:EG<H

、2õ:XES

,H�[å ¡EXU®

。K�

,=·ÄG:X��UV

ÿ

,hFáÑRE"2��jfUV

,Y_Zâ

CO2®·i

;IE#$

8

4 *^�����V�

16SrDNAV���

Fig.4 PhylogeneticanalysisofStreptomycesbasedonthe16SrDNAgenesequences

  ÇÁ©ªÑR:XEG:(eÑRE

,B4:X

GP57·�Ü��z:

、%{|:�}~��|:[Ss\EG

H

;ô

GY11·�Ü��z:�}~��|:SGH

;GP71�

GT16·�Ü��z:�%{|:SGH

,ô·}~��

|:�GH

。\]^®

[15]_=

1998¶÷]$i;IEG:

<Hz`(a(:XE50SÃ��50bä

(V

pH、c

ÿ

、dÿ®

)S�

。¿Me«

,:XG:Þ¬ÑRùK(50

bäYSs\��

。À%&

,Taechowisan®

[16]Êf4�Ë

Ì¢;89:

,B4

3�:XKÎg×Øhi�

(Colletotrichummusae),5

�Kj×Øk��lm:

(Fusariumoxysporum),

ôS

2�K×Ø�

2�^Ç­:

。r�ùMW$ÑR

:XEG:noV=s\pq

4X::�¾óS¿ÑR

,ýeÑKíîúÊ:�¾ó

r::XE��ús

[17]。Øì¾ó&¡¢

、;û;ö[\

16SrDNA�1[\

,Ü\:X

GT16Z8�,6:

;:X

GP57Z,6:7

。Sun®

[18]Ê#¸4�ËWÌj��|;

#$89:

,B4÷��,6:7

。Nithyanand®

[19-20]~Ê

¹ºEt�4�ËÌj�<H89:

,��u¹|:7

(Curtobacterium)、,6:7®

。2010¶

,Goodfellow®

[21]�

�CÇÈÉÊ#$@A4�ËE89:S

50�7

,ùv#

$89:es�°S�%B©´LEi;IIc

。#$�

|;i;I4

,#$89:ùKes��íEFG:<HI

E:c

,wWê�sxE»¼

� ;<

[1]!"#

.$%&'()*+,-

[J]../01

(23456

),2011,32(5):219-222.

[2]78

.9:3$%;3<2(=><?)*+

[J].@AB*+

,2008(2):227-230.

[3]HOLMSTR?MC,KJELLEBERGS.MarinePseudoalteromonasspeciesareassociatedwithhigherorganismsandproducebiologicallyactiveextracellularagents[J].FEMSMicrobiologyEcology,1999,30(4):285-293.

[4]ELAMRAOUIB,ELAMRAOUIM,COHENN,etal.Antifungalandantibacterialactivityofmarinemicroorganisms[J].Annalespharmaceutiquesfrancaises,2014,72(2):107-111.

[5]SCOPELM,DOSSANTOSO,FRASSONAP,etal.AntiTrichomonasvaginalisactivityofmarineassociatedfungifromtheSouthBrazilianCoast[J].Experimentalparasitology,2013,133(2):211-216.

(ÒÓÔ

17Õ

4           ÊËÌÍÎ�

                         2018}

Page 5: 01书!"#$%&’()*+,-./*01!"#,$%&,’(( (!"#$%&#$’&()*&+,-#$./012345,!"67 316022)23 [)*]+,-./0123456789:;?@abcd6e*fghijb,klmn#opqrabcdsab t,nuvqwab

w

2 äåÝãæç-mn

Table2 Orthogonalexperimentdesignandresult

�

No.���

Potato(A)

­�A

Yeastpowder(B)

�Õ>

Magnesiumsulfate(C)

ûÕ<�=

Dipotassiumphosphate(D)

:Å:g

Mycelialdryweightg/L

1 1 1 1 1 8.762 1 2 2 2 7.643 1 3 3 3 5.344 2 1 2 3 5.895 2 2 3 1 10.376 2 3 1 2 7.377 3 1 3 2 7.158 3 2 1 3 8.189 3 3 2 1 7.45k1 7.247 7.267 8.103 8.860k2 7.877 8.730 6.993 7.387k3 7.593 6.720 7.620 6.470R 0.630 2.010 1.110 2.390

3 m�

50SE�ÃÃ�MN)Ã�EÞÿe#$�ë:Å

Õö3EghKd

,%Ô#$}:Å:gE%¹

。Ä»¼M

�ëZÇI

,M:Å:gZ½«�§

,Êì·¦á2c

、�c

��UV

,Ü\C©­50SEÂç2c��c

,���w

AZÂç2c

,­�AZÂç�c

,ÊìîKdÇÁ�Û>

ÇÁ

,»¼C^Ç�ë:XEÂç©­50Sê"

:���

wA

4%、­�A

0.100%、�Õ>

0.15%、ûÕ<�=

02%。ÄÇÁYWE50Sê"°SêØíî

、F:Å_jEç

Æ

,ùàZ2�ö:�;FEùhê"��Lë

� ;<

[1]CDE

,FGH

,IDJ

,K

.LMNOPQRSTUVW>5X

[J].YZ[\

,2015(4):149-151.[2]

]^_

,̀a

,bcd

.LMNO

(ef

)(g

CCEF89)3h3B*+

[J].(<ij

,2015,34(4):621-631.[3]

klm

,nop

,qrs

,K

.NOtu(vwxy)*+z{

[J].(

<ij

,2015,34(4):541-552.[4]RADZKIW,ZIAJASOTYSM,NOWAKJ,etal.EffectofprocessingonthecontentandbiologicalactivityofpolysaccharidesfromPleurotusostreatusmushroom[J].LWTFoodScienceandTechnology,2016,66:27-33.

[5]|}~

,��3

,|��

,K

.ef���t���������*+

[J].����

,2015,36(10):223-226.[6]

���

,n��

,��

,K

.������ef� d5)¡¢

[J].�

�£i

,2015,36(4):86-91.[7]

|¤7

.¥g¦3§¨©(ª«¬«���­QZ®¯°

[J].YZ[

\

,2015(21):152-154.[8]OOIVEC,LIUF.Areviewofpharmacologicalactivitiesofmushroompolysaccharides[J].Internationaljournalofmedicinalmushrooms,1999,1(3):195-206.

[9]ZHANGJJ,MENGGY,ZHAIGY,etal.Extraction,characterizationandantioxidantactivityofpolysaccharidesofspentmushroomcompostofGanodermalucidum[J].Internationaljournalofbiologicalmacromolecules,2016,82:432-439.

[10]F±²

,³r´

.¢µ¶5X®·Bef¬«(¸���)*+

[J].7¹º»i¼ij

,2015,32(5):129-133.[11]

�½¾

,b¿

,À±Á

,K

.ef¬«¸Â���·B

[J].ä@�Ä

iij

,2008,30(5):915-918.[12]

ÅÆÇ

,nÈÉ

.�y(¬«(¸ÊË)*+z{

[J].Ì.�y(

,2003,22(6):15-17.

[13]|ÍÎ

,Ï$Ð

.������ef(ª«3Ñ)¡¢*+

[J].ÒÓ

£iÔÕi¼ij

(Ö×£iØ

),2006,24(1):74-76.[14]

`a

,]^_

,]Ù

,K

.��ÚÛ�ÜÛ��ÝfÞßà?áâ4ã

=>)¡¢

[J].(<ij

,2015,34(4):761-771.[15]

,åæç

,èéê

,K

.ëìíî<)3ïÔÕz{

[J].ðS��

£Ô

,2013,29(7):1747-1750,1701.

檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪檪(»ÔÔ

4Õ

[6]JARUCHOKTAWEECHAIC,SUWANBORIRUXK,TANASUPAWATTS,etal.NewmacrolactinsfromamarineBacillussp.Sc026[J].JNatProd,2000,63(12):984-986.

[7]HEINDLH,WIESEJ,THIELV,etal.PhylogeneticdiversityandantimicrobialactivitiesofbryozoanassociatedbacteriaisolatedfromMediterraneanandBalticSeahabitats[J].Systematicandappliedmicrobiology,2010,33(2):94-104.

[8]ROMANENKOLA,UCHINOM,KALINOVSKAYANI,etal.Isolationphylogeneticanalysisandscreeningofmarinemolluscassociatedbacteriaforantimicrobial,hemolyticandsurfaceativities[J].Microbiologicalresearch,2008,163(3):633-644.

[9]SANTOSOCS,PONTESPVML,SANTOSJFM,etal.Isolation,characterizationandphylogenyofspongeassociatedbacteriawithantimicrobialactivitiesfromBrazil[J].Researchinmicrobiology,2010,161(7):604-612.

[10]ñòÌ

,�ó

.$%;3<ôÛõöï33<=>S÷ï<)*+

[J].3<�øz{

,2000,20(5):28-31.[11]

ñ¡ù

.���ú6

[M].ÑD

:ûù£iÔÕüØý

,1991:370.[12]

þÿ!

,�"#

,nÇ

.¥¸$yã%&'()*+(4,-)Z®

[J].Ì.;3.iPr

,2009,21(8):685-687.[13]

�/0

.;3<i12

[M].Y3

:Bi��üØý

,2010:77-86.[14]

4ùÃ

,ñ5

.6789:;)+<=µ>>?

[J].TU

,2014,36

(4):327-335.[15]

@ÁA

,��B

,åCD

,K

.EF$GHIJ$%KL<9:3;3

<)2(=>

[J].MN$O

,1998,17(4):439-444.[16]TAECHOWISANT,PEBERDYJF,LUNYONGS.Isolationofendophytic

actinomycetesfromselectedplantsandtheirantifungalactivity[J].Worldjournalofmicrobiologyandbiotechnology,2003,19(4):381-385.

[17]BRASIERCM.Fungalspeciesinpractice:Identifyingspeciesunitsinfungi[M]//CLARIDGEMF,DAWAHHA,WILSONMR,etal.Species:Theunitsofbiodiversity.London:Chapman&Hall,1997:135.

[18]SUNW,DAISK,JIANGSM,etal.CulturedependentandcultureindependentdiversityofActinobacteriaassociatedwiththemarinespongeHymeniacidonperlevefromtheSouthChinaSea[J].Antonievanleeuwenhoek,2010,98(1):65-75.

[19]NITHYANANDP,INDHUMATHIT,RAVIAV,etal.CultureindependentcharacterizationofbacteriaassociatedwiththemucusofthecoralAcroporadigitiferafromtheGulfofMannar[J].Worldjournalofmicrobiologyandbiotechnology,2011,27(6):1399-1406.

[20]NITHYANANDP,MANJUS,PANDIANSK.PhylogeneticcharacterizationofculturableActinomycetesassociatedwiththemucusofthecoralAcroporadigitiferafromGulfofMannar[J].FEMSMicrobiologyLetters,2011,314(2):112-118.

[21]GOODFELLOWM,FIEDLERHP.Aguidetosuccessfulbioprospecting:Informedbyactinobacterialsystematics[J].Antonievanleeuwenhoek,2010,98(2):119-142.

7146Ï

21Ð

                   ¦§¨Ñ

 ­±B��8��b*�i