橙色交替单胞菌PB1对海洋细菌的拮抗作用及其胞外产物抑菌活性的研究
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摘要
有益菌(probiotics)作为生物控制的手段用于水产养殖中，可以调节和改善养殖微生态环境，控制和减少养殖动物暴发病害的发生，在国内外以引起普遍重视。特别是从养殖环境中以及养殖动物体表或体内分离筛选的土著细菌作为潜在有益菌成为研究的热点。
海洋有益菌PB1(Altermonas aurantia)是从健康斑节对虾(Penaeus monodon)幼体中分离的。采用琼脂扩散法和十字划线法证明菌株PB1具有广谱抑菌作用，不仅对水产养殖病原哈维氏弧菌和鳗弧菌有拮抗作用，同时PB1对中华绒螯蟹(Eriocheir sinensis)幼体丝状细菌病病原HL-1(Leucothrix mucor)有抑制作用。采用混合培养的方法对PB1的抑菌作用的进行定量测定。使用纸片法，对病原菌W-1、Z_3G_2和H1-1进行药敏试验，结果表明病原菌对抗菌药物高度敏感的种类少，病原菌的耐药性强。而PB1对病原菌的拮抗作用强，因此PB1可以作为潜在有益菌应用于水产养殖。
选用对PB1敏感的鳗弧菌W-1(Vibrio anguillarum)作为指示菌，采用杯碟法对PB1的发酵产物进行分析，培养基的成份影响PB1对W-1的抑制作用。使用2216E液体连续培养7d，在发酵液中未检测到抑菌活性；改用谷氨酸钠培养基液体培养，第3天在发酵液中检测到活性。将菌株PB1和W-1混和培养，84hr在培养液中检测到抑菌活性。培养基中Fe~(3+)的浓度影响PB1的抑制作用，当培养基中铁离子浓度大于700nM时，PB1对涂布在2216E平板W-1无抑制作用。使用特异性化学方法可以检测PB1在MM9GC缺铁培养基中产生异羟肟酸型铁载体。
Probiotics, which are microorganisms with healthy benefit to the host, have been found to be used in aquaculture as a means of diseases biocontrol.
Marine bacterial strain PBl (Altermonas aurantia) was isolated from healthy juvenile black tiger shrimp (Penaeus monodon). It is non-pathogenic bacterium to most of aquacultural animals. The antagonism of PBl was tested by cross-streaking method, agar diffusion assay and BOD in the liquid media. It has a broad spectrum to inhibit tested bacteria including Gram positive and Gram negative strains, especially vibrio. A filamentous bacterium HL-1.(Leucothrix mucor)can be inhibited by PBl also. So strain PBl could be selected as a potential probiotic.
A pathogenic strain W-1 (Vibrio anguillarum) was selected as a indicator bacteria to test the inhibitory activities of PBl. the results showed that the compositions of the medium affected the inhibitory activities of the supernatants of strain PBl. The antagonism of the supernatants can be detected when PBl was cultured in monosodium glutamate medium for 3 days. It can not be detected when in marine 2216E broth for 7 days. The concentration of Fe3+ can influence the antibacterial activity of PBl. The siderophore can not be detected from supernatants from iron-limited medium of PBl culture by the unspecific method -CAS assay, while can be detected by the specific chemical method -Casey assay.
海洋有益菌PB1(Altermonas aurantia)是从健康斑节对虾(Penaeus monodon)幼体中分离的。采用琼脂扩散法和十字划线法证明菌株PB1具有广谱抑菌作用，不仅对水产养殖病原哈维氏弧菌和鳗弧菌有拮抗作用，同时PB1对中华绒螯蟹(Eriocheir sinensis)幼体丝状细菌病病原HL-1(Leucothrix mucor)有抑制作用。采用混合培养的方法对PB1的抑菌作用的进行定量测定。使用纸片法，对病原菌W-1、Z_3G_2和H1-1进行药敏试验，结果表明病原菌对抗菌药物高度敏感的种类少，病原菌的耐药性强。而PB1对病原菌的拮抗作用强，因此PB1可以作为潜在有益菌应用于水产养殖。
选用对PB1敏感的鳗弧菌W-1(Vibrio anguillarum)作为指示菌，采用杯碟法对PB1的发酵产物进行分析，培养基的成份影响PB1对W-1的抑制作用。使用2216E液体连续培养7d，在发酵液中未检测到抑菌活性；改用谷氨酸钠培养基液体培养，第3天在发酵液中检测到活性。将菌株PB1和W-1混和培养，84hr在培养液中检测到抑菌活性。培养基中Fe~(3+)的浓度影响PB1的抑制作用，当培养基中铁离子浓度大于700nM时，PB1对涂布在2216E平板W-1无抑制作用。使用特异性化学方法可以检测PB1在MM9GC缺铁培养基中产生异羟肟酸型铁载体。
Probiotics, which are microorganisms with healthy benefit to the host, have been found to be used in aquaculture as a means of diseases biocontrol.
Marine bacterial strain PBl (Altermonas aurantia) was isolated from healthy juvenile black tiger shrimp (Penaeus monodon). It is non-pathogenic bacterium to most of aquacultural animals. The antagonism of PBl was tested by cross-streaking method, agar diffusion assay and BOD in the liquid media. It has a broad spectrum to inhibit tested bacteria including Gram positive and Gram negative strains, especially vibrio. A filamentous bacterium HL-1.(Leucothrix mucor)can be inhibited by PBl also. So strain PBl could be selected as a potential probiotic.
A pathogenic strain W-1 (Vibrio anguillarum) was selected as a indicator bacteria to test the inhibitory activities of PBl. the results showed that the compositions of the medium affected the inhibitory activities of the supernatants of strain PBl. The antagonism of the supernatants can be detected when PBl was cultured in monosodium glutamate medium for 3 days. It can not be detected when in marine 2216E broth for 7 days. The concentration of Fe3+ can influence the antibacterial activity of PBl. The siderophore can not be detected from supernatants from iron-limited medium of PBl culture by the unspecific method -CAS assay, while can be detected by the specific chemical method -Casey assay.
引文
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