养殖场底泥中芽孢杆菌属细菌的生态学研究

对近海养殖场底泥细菌的研究，发现养殖池底泥中芽孢菌为优势菌群。应用形态学、生理生化和16S rDNA序列测定，对芽孢杆菌属细菌做进一步分类的结果表明，该属细菌在底泥中的主要种群为蜡状芽孢杆菌、巨大芽孢杆菌、短小芽孢杆菌、坚强芽孢杆菌、Bacillus baekryungensis等。同时，还分析了这些芽孢杆菌属细菌在底泥元素的地球化学循环中的作用，认为这类细菌应属于陆生微生物，在养殖环境中代谢有机化合物产生氨、亚硝酸盐和H2S，对养殖环境产生不利的作用和影响。     

噬菌弧菌N1对淡水和海水养殖水体细菌群落影响PCR-DGGE分析

【目的】研究噬菌弧菌Bacteriovorax sp. N1在一个投放周期内对淡水和海水养殖环境中细菌、弧菌总数及细菌群落结构的影响。【方法】自市售微生态制剂分离蛭弧菌N1并进行分子鉴定,测定其裂解效果,制备高浓度N1菌液分别投放至淡水红鲤鱼(red carp)和海水仿刺参(Apostichopus japonicus)养殖水体,采用细菌平板计数法及PCR-变性梯度凝胶电泳(PCR-DGGE)技术分析48 h内水体环境中细菌、弧菌总数及细菌群落结构变化。【结果】经鉴定蛭弧菌N1为噬菌弧菌,其对大肠杆菌(Escherichia coli)、溶藻弧菌(Vibrio alginolyticus)、黄海希瓦氏菌(Shewanella smarisflavi)、灿烂弧菌(Vibrio splendidus)、哈氏弧菌(Vibrio harveyi)和金黄色葡萄球菌(Staphylococcus aureus)均有裂解效果。将噬菌弧菌N1投放进淡水和海水养殖环境的12～24 h,其能显著降低两种环境中弧菌含量(P 0.05)。DGGE分析显示添加噬菌弧菌N1后淡水组优势菌群弧菌属(Vibrio,a1)和不可培养杆菌属(Uncultured bacterium, a5)在12 h以后含量有明显的减少,假单胞菌属(Pseudomonas, a3)和红杆菌科Shimia属(Shimia, a6)菌含量增加。海水组优势菌群不可培养杆菌属(c2)菌在12 h时变成非优势菌群,而白杆菌属(Albirhodobacter,c1)增加成为优势菌群。噬菌弧菌N1在水中含量在24 h时降至最低。【结论】噬菌弧菌N1对海水和淡水环境中的弧菌属和不可培养杆菌属菌群有明显的裂解作用导致其含量下降,但也使假单胞菌属(Pseudomonas),红杆菌科Shimia属和白杆菌属(Albirhodobacter)菌群含量增加,但生物效应不明。为维持噬菌弧菌N1对弧菌的控制,需要24 h左右重新补充投放。     

芽孢杆菌胞外产物的研究进展

芽孢杆菌大多能分泌高活性的胞外产物（extracellular products），如蛋白酶、淀粉酶、脂肪酶等水解酶和果胶酶、葡聚糖酶、纤维素酶等分解植物性饵料中非淀粉多糖的酶类。近年来芽孢杆菌被作为益生菌广泛应用于水产养殖业，它可提高养殖对虾的消化机能，促进其对营养物质的吸收，同时还可降解水体中的有机碎屑，将之迅速转化为无机物，从而有效地促进养殖水体中生态良性循环；还有报道芽孢杆菌能分泌一些抗菌类物质，抑制部分病原菌，提高机体的免疫力。在微生物工业生产中芽孢杆菌也常用于生产某些酶制剂，为满足生产的需要，学者多将芽孢杆菌胞外产物中的某一特定成分进行提纯分析。笔者就芽孢杆菌胞外产物中的一些主要成分的研究进展作一系统综述。     

有益微生物对水质因子的调控效应

随着我国水产养殖业的发展 ,集约化、半集约化的养虾模式随之迅猛发展 ,养殖规模不断扩大。在养殖过程中 ,因残存饵料腐烂 ,生物代谢 ,以及生物残体沉积 ,致使养殖水体理化环境和生态环境恶化 ,从而影响生物的生长、发育和行为活动。为改善养殖环境 ,促进水产养殖业健康可持续性地发展 ,近年来微生物技术在水产养殖业中得到了广泛应用[1,2 ] ,1) 。利用芽孢杆菌和光合细菌等有益微生物能调节和稳定养殖水体中的水质因子 ,分解、转化养殖水体的有机物的特性 ,达到改良、净化和营造养殖生态环境的效果 ,在生产上取得了明显的成效。为此 ,从 1 …     

马氏珠母贝肠道及其养殖水体可培养细菌群落结构

【目的】了解马氏珠母贝(Pinctada fucata martensii)肠道及其养殖水体可培养细菌的群落组成。【方法】采用2216E平板涂布法研究海区养殖马氏珠母贝肠道与养殖水体的可培养菌群种类及丰度。【结果与结论】马氏珠母贝肠道及其养殖水体的可培养细菌归属于2门(变形菌门和厚壁菌门)3纲7目10科23属56种。属水平上,肠道中以弧菌属(74.7%)和假交替单胞菌属(18.7%)为主;养殖水体中α-变形菌纲的FJ943236_g属(40.7%)丰度最大,弧菌属(16.7%)相对肠道丰度较低。样品共有菌属为弧菌属、假交替单胞菌属、发光杆菌属和芽孢杆菌属;肠道特有菌属为希瓦氏菌属和盐单胞菌属;养殖水体特有菌属主要为FJ943236_g、鲁杰氏菌属和Nautella。在种水平上,7个种为二者共有;马氏珠母贝肠道和养殖水体特异性菌种分别为18个和31个。虽然门水平上马氏珠母贝肠道中可培养细菌群落与其养殖水体中的细菌群落大致相似,但在属、种水平上二者差异明显。     

水产养殖用芽孢杆菌制剂的检测

通过平板菌落计数法检测编号为GD1、GD2、GD3、GD4、GD5、JS、JX、SD1和SD2等9种水产养殖用芽孢杆菌制剂的有效活菌数,结合形态、生理生化和16S rRNA基因序列分析对各制剂中分离的产芽孢细菌代表菌株进行鉴定。结果显示:除GD1、GD2和GD5制剂未标明芽孢数量外,其余制剂检测到的芽孢数量均低于或远低于标注数量;经鉴定,各制剂中最优势类型,GD2、GD3、GD5、SD1和SD2制剂为地衣芽孢杆菌(Bacillus licheniformis),GD4、JS和JX制剂为解淀粉芽孢杆菌(B.amyloliquefaciens),GD1制剂为枯草芽孢杆菌(B.subtilis)。除GD4、GD5和SD1制剂未标明具体菌种以及GD2的标注菌种与检测结果不一致外,其他制剂的标注菌种与被鉴定的优势类型芽孢杆菌基本一致。     

侧孢短芽孢杆菌对铜绿微囊藻藻际微生物群落的影响

【目的】探究铜绿微囊藻(Microcystis aeruginosa)藻际微生物群落组成以及与溶藻细菌侧孢短芽孢杆菌(Brevibacillus laterosporus)Bl-zj共培养对藻际微生物群落的影响。【方法】将侧孢短芽孢杆菌Bl-zj与铜绿微囊藻共培养4 d后(BL组),收集样品,以单独培养的铜绿微囊藻(BG组)为对照,提取样品总基因组DNA后进行16S rDNA扩增、构建文库、测序及生物信息学分析,比较BL组与BG组间藻际微生物群落的差异。【结果】测序平均获得高质量序列85390条,946个OUT(Operational Taxonomic Unit),BL组特有OTU 339个。物种组成分析显示,BG组微生物群落主要以微囊藻属(Microcystis)为主,其余相对丰度由高到低分别为假单胞菌属(Pseudomonas)、中慢生根瘤菌属(Mesorhizobium)、短芽孢杆菌属(Brevibacillus)和红杆菌属(Rhodobacter)。BL组微生物群落主要以短芽孢杆菌属(Brevibacillus)为主。BL组的微生物多样性相较于BG组显著下降(P<0.05)。经预测,BG组微生物功能集中在能量代谢与外源生物降解与代谢,BL组微生物功能集中在碳水化合物代谢和氨基酸代谢功能。【结论】侧孢短芽孢杆菌Bl-zj与铜绿微囊藻共培养,打破了铜绿微囊藻藻际微生物群落的平衡,与Bl-zj的溶藻进程有关。     

大口黑鲈肠道益生菌的分离与鉴定

【目的】分离鉴定大口黑鲈（Micropterus salmoides）肠道益生菌,评估分离菌用作饲料添加剂的益生潜力。【方法】采集大口黑鲈肠道样本,采用稀释涂布法分离细菌,经16S rRNA序列比对和生化试验鉴定种属,并进行抗逆性能、黏附能力、溶血活性和药敏特性等鉴定。【结果】从大口黑鲈肠道分离菌株84株,经测序、blast比对,筛选出3株同源性最高的菌株MS-1、MS-2、MS-3,分别鉴定为地衣芽孢杆菌（Bacillus licheniformis）、解淀粉芽孢杆菌（Bacillus amyloliquefaciens）和贝莱斯芽孢杆菌（Bacillus velezensis）。3株分离菌均为γ-溶血,不具有生物膜,疏水性50%～93.3%,自凝集性43.6%～69.2%;可在pH 2.0～12.0环境和5.0 g/L胆盐中存活;经高温处理后仍可生长;3株菌对大多数抗生素敏感。【结论】芽孢杆菌MS-1、MS-2和MS-3抗逆性高,黏附性能及饲料学安全性高,可作为大口黑鲈养殖中潜在益生菌。     

中草药复合益生菌制剂对凡纳滨对虾生长、抗病力及水质的影响

将枯草芽孢杆菌、纳豆芽孢杆菌、地衣芽孢杆菌等比例混合,制成芽孢杆菌制剂;将芽孢杆菌制剂与粪肠球菌、嗜酸乳杆菌等比例混合制成复合益生菌制剂;将分别用3种芽孢杆菌发酵的中草药等比例混合制成中草药芽孢杆菌制剂;将分别用5种益生菌发酵的中草药等比例混合,制成中草药复合益生菌制剂。在饲料中分别添加4种益生菌制剂(活菌为2×10~7 cfu/g),研究4种制剂对凡纳滨对虾(Litopenaeus vannamei)生长、抗病力及水质的影响。结果表明:1)4种益生菌制剂均可提高凡纳滨对虾的成活率、增重率和饲料利用率(P0.05),中草药益生菌制剂组的促生长效果优于益生菌制剂组,以中草药复合益生菌制剂组促生长效果为最佳(P0.05);2)4种益生菌制剂均可维持对虾养殖水体p H值、氨态氮和亚硝酸盐含量的稳定(P0.05),中草药益生菌制剂对水质的改良效果优于益生菌制剂(P0.05);3)用1×10~8 cfu/m L的哈维氏弧菌(Vibro harveyi)菌液浴浸泡凡纳滨对虾10 d,凡纳滨对虾的累计死亡率由大到小依次为中草药复合益生菌制剂组(31.11%)、复合益生菌制剂组(35.56%)、中草药芽孢杆菌制剂组(37.78%)、芽孢杆菌制剂组(44.44%)、对照组(93.33%)。在饲料中添加一定比例的中草药复合益生菌制剂可提高凡纳滨对虾的生长指标、改善养殖水体环境、提高对虾的抗病力。     

几株有益芽孢杆菌对温度、制粒工艺及pH值的耐受性

报道了实验室分离、筛选的 5株有益芽孢杆菌对温度、对虾饲料制粒工艺流程和pH值的耐受性。结果表明 ,5株芽孢杆菌经 80℃水浴 4 0min后全部存活 ,90℃水浴 2 0min有 35 %～ 70 %存活 ,10 0℃水浴 10min仍有 30 %～ 5 0 %存活 ,显示所筛选的 5株芽孢杆菌对高温有较强的耐受力 ;在对虾饲料中添加 5株芽孢杆菌 ,经整个生产工艺流程后芽孢杆菌存活 95 %,烘干后芽孢杆菌存活 93%,说明杆菌能够承受饲料制粒生产中压力、温度和水分的变化 ;当pH值为 3.8、4 .6、5 .2时 ,分别有 1、3、5株菌能繁殖 ,pH值 6 .0～ 8.5时 ,5株菌生长旺盛 ,将 5株菌经pH值 2 .2～ 4 .6处理1h ,再接种在pH值 7.2的培养基可以良好生长 ,说明杆菌在酸性环境中生长性能较差 ,在弱酸性和碱性环境中生长旺盛 ;能耐受酸性环境而存活 ,再给予适宜环境即能良好生长。 5株芽孢杆菌具有作为水产硬颗粒饲料专用益生素菌株的优势。     

Phylogenetic analysis of cultivable bacteria isolated from Arctic sea-ice

Phylogenetic analysis based on 16S rDNA of 8 strains of cultivable bacteria isolated from Arctic sea-ice was studied.The results showed that strain BJ1 belonged to genus Planococcus,which was a genus of low mole percent G C gram-positive bacteria;strain BJ6 belonged to genus Burkholderia of β-proteobacteria and the rest 6 strain all belonged to γ-proteobacteria,of which strain BJ8 was a species of Pseudoalteromonas,strain BJ2-BJ5 and BJ7 were members of genus Psychrobacter.Phylogenetic analysis also indicated that bacteria of genus Psychrobacter of the isolates formed a relatively independent phylogenetic cluster in comparison with other bacteria belonged to genus Psychrobacter.     

具有多种胞外酶的对虾肠道黏附菌的分离和鉴定

用对虾饲料培养基从健康凡纳滨对虾肠道分离出500株黏附细菌,以产淀粉酶、脂肪酶和蛋白酶能力为指标,筛选出产该3种消化酶的细菌90株,占总菌株的18%.对其中生长较快的69株进行16S rDNA 基因测序,确定其分类地位.结果显示,69株菌分别属于不动杆菌属(Acinetobacter)、芽孢杆菌属(Bacillus)、葡萄球菌属(Staphylococcus)、假交替单胞菌属(Pseudoalteromonas)、气单胞菌属(Aeromonas)、嗜盐单胞菌属(Halomonas)、利斯顿氏菌属(Listonella)、莫拉氏菌属(Moraxella)等,其中数量最多是芽胞杆菌属,占鉴定细菌总数的53.62%,数量最少是气单胞菌属和嗜盐单胞菌属,均占鉴定细菌总数的2.90%.表明对虾肠道黏附菌群中具有较多能分泌多种消化酶的细菌,可进一步开发为促进对虾消化功能的益生菌     

生物滤料对翘嘴红鲌生长、水质及细菌变化的影响

在实验池投放一定数量的生物刷和生物球等生物滤料,研究其对翘嘴红鲌生长、池塘水质及细菌变化等的影响。结果表明:与对照池相比,实验池翘嘴红鲌生长有增加趋势,对池塘水温、pH、DO、TP、H2S等指标无显著影响(P>0.05),但是在30 d时水体COD显著降低(P<0.05);在90 d时NH4-N显著降低(P<0.05);在60 d和90 d时NO2-N显著降低(P<0.05);与对照池相比,在30 d时实验池水样与生物球中异养菌的数量显著增加(P<0.05);在30、60 d时,实验池生物刷中异养菌的数量显著增加(P<0.05);在90 d时实验池水样、生物球、生物刷氨化细菌、反硝化细菌的数量显著增加(P<0.05);实验池水样、生物球分别在30、90 d时弧菌数量显著降低(P<0.05)。表明生物滤料可增加池塘的异养菌、氨化细菌、反硝化细菌数量,降低水体弧菌数量,一定程度上改善养殖水体,促进鱼体生长。     

Variation of alkaline phosphatase activity in sediments of shrimp culture ponds and its relationship with the contents of C,N and P

1Introduction Activities of extracellular enzymes can serve as theindicators of organic matter content.Enzyme assayinghas been used in the analyses of dissolved organic ma-terials in aquatic environments(Chrost,1991),massloss of plant litter in terrestrial systems(Sinsabaughet al.,1992;Sinsabaugh et al.,1994),processing ofpaniculate and dissolved organic carbon(POC,DOC)in streams(Sinsabaugh and Findlay,1995;Findlayet al.,1998),and processing of particulate organicmatter(POM)in wetlands(J…     

Effect of Zinc Oxide Nanoparticles on Denitrification and Denitrifying Bacteria Communities in Typical Estuarine Sediments

For revealing the effects of increasing of zinc oxide nanoparticles(ZnO NPs) on denitrification and denitrifying bacteria communities in estuarine sediments, the surface sediments of two typical estuaries(the Yangtze River Estuary and the Yellow River Estuary) were added with medium concentration(170 mg L~(-1)) and high concentration(1700 mg L~(-1)) of ZnO NPs for anaerobic culture in laboratory. The concentration of NO_3~-and NO_2~-, the reductase activity and denitrification rate were measured by physicochemical analysis, nir S gene abundance and denitrifying bacteria communities by molecular biological methods. The results showed that ZnO NPs inhibited NO_3~-, NO_2~-reduction process and NO_3~-, NO_2~-reductase activity, and a stronger inhibition effect resulting from the higher ZnO NPs concentration. ZnO NPs decreased nirS gene abundance and community diversity of denitrifying bacteria. In addition, the inhibition degree of ZnO NPs on the denitrification process of sediments in different estuaries was different. These results were of great significance for evaluating the potential ecological toxicity and risks of nanomaterials in estuaries.     

Screening and molecular classification of low-temperature protease from Antarctic microorganism and its characterization

107 strains producing protease were screened from 260 strains of Antarctic psychrophilic bacteria, among which proteolytic activity of five strains was more than 45 U ml^-1. The 16S rRNA gcne sequences homology and phylogcnetic analysis of five Antarctic psychrophillc bacteria showed that NJ276, NJS-9, NJ16-70,NJ345 belonged tO the described genus Pseudoalteromonas and NJ341 belonged to the genus Colwellia. The growth and the protease characteristic of four Antarctic psychrophilic bacteria had been studied, and the result showed that the 6ptimal temperature for growth and protease-produeing of four strains was about 10℃. Their growth and protease-produeing were still high during incubatlng 2-5 days. The maximum proteolytic activity occurred at pH 9 for four Antarctic psychrophilic bacteria. The optimal temperature of protease action of both strains NJ276 and NJ5-9 was about 50℃, however, the optimal temperature of protease aetlon of both strains NJ341 and NJ345 was about 40 ℃, and their proteolytic activity under 0℃ exhibited nearly 30% of the maximum activity, but their thermal stabilities were weaker. These results indicated that proteases from NJ341 and NJ345 were low-temperature proteases.     

Effects of a Potential Autochthonous Probiotic Bacillus subtilis 2-1 on the Growth and Intestinal Microbiota of Juvenile Sea Cucumber,Apostichopus japonicus Selenka

The effects of Bacillus subtilis 2-1 from the intestine of healthy sea cucumber on the growth, digestive enzyme activities and intestinal microbiota of juvenile sea cucumber(Apostichopus japonicus) were determined in the present study. Sea cucumber was fed with Sargassum thunbergii powder supplemented with B. subtilis 2-1 at different concentrations varying among 0(control), 10~5, 10~7, and 10~9 CFU g~(-1) for 8 weeks. Results showed that the growth performance and intestinal amylase and trypsin activities were significantly increased by dietary B. subtilis 2-1 at 10~9 CFU g~(-1)(P 0.05). However, dietary B. subtilis 2-1 had no significant influence on the lipase activity in sea cucumber(P 0.05). The polymerase chain reaction denaturing gradient gel electrophoresis and 16S rRNA gene sequencing analysis indicated that dietary B. subtilis 2-1 at 105 and 107 CFU g~(-1) inhibited most of the Proteobacteria including those in genus Vibrio. Dietary B. subtilis 2-1 at 10~9 CFU g~(-1) not only decreased the abundance and species of genus Vibrio, but also increased the intensity of genera Psychrobacter and Bacillus. A specific dosage of dietary B. subtilis 2-1 could increase the growth and modulate the intestinal microbiota of sea cucumber; thus it might be a novel probiotic for keeping the health of sea cucumber.     

Characterization and identification of enzyme-producing microflora isolated from the gut of sea cucumber Apostichopus japonicus

Gut microorganisms play an important role in the digestion of their host animals. The purpose of this research was to isolate and assess the enzyme-producing microbes from the Apostichopus japonicus gut. Thirty-nine strains that can produce at least one of the three digestive enzymes(protease,amylase,and cellulase) were qualitatively screened based on their extracellular enzyme-producing abilities. The enzyme-producing strains clustered into eight groups at the genetic similarity level of 100% by analyzing the restriction patterns of 16 S rDNA amplified with Mbo I. Phylogenetic analysis revealed that 37 strains belonged to the genus Bacillus and two were members of the genus Virgibacillus. Enzyme-producing capability results indicate that the main enzyme-producing microflora in the A. japonicus gut was Bacillus,which can produce protease,amylase,and cellulase. Virgibacillus,however,can only produce protease. The high enzyme-producing capability of the isolates suggests that the gut microbiota play an important role in the sea cucumber digestive process.     

Spatiotemporal distributions and environmental drivers of diversity and community structure of nosZ-type denitrifiers and anammox bacteria in sediments of the Bohai Sea and North Yellow Sea,China

Denitrification and anammox processes are major nitrogen removal processes in coastal ecosystems.However,the spatiotemporal dynamics and driving factors of the diversity and community structure of involved functional bacteria have not been well illustrated in coastal environments,especially in human-dominated ecosystems.In this study,we investigated the distributions of denitrifiers and anammox bacteria in the eutrophic Bohai Sea and the northern Yellow Sea of China in May and November of2012 by constructing clone libraries employing nosZ and 16 S rRNA gene biomarkers.The diversity of nosZ-denitrifier was much higher at the coastal sites compared with the central sites,but not significant among basins or seasons.Alphaproteobacteria were predominant and prevalent in the sediments,whereas Betaproteobacteria primarily occurred at the site near the Huanghe(Yellow)River estuary.Anammox bacteria Candidatus Scalindua was predominant in the sediments,and besides,Candidatus Brocadia and Candidatus Kuenenia were also detected at the site near the Huanghe River estuary that received strong riverine and anthropogenic impacts.Salinity was the most important in structuring communities of nosZdenitrifier and anammox bacteria.Additionally,anthropogenic perturbations(e.g.nitrogen overloading and consequent high primary productivity,and heavy metal discharges)contributed significantly to shaping community structures of denitrifier and anammox bacteria,suggesting that anthropogenic activities would influence and even change the ecological function of coastal ecosystems.