油茶壳堆肥微生物的培养及功能芽孢杆菌的筛选和初步应用

【目的】从自然状态下的油茶壳堆肥产品中筛选功能芽孢杆菌,为油茶壳堆肥高效生产、饲料发酵等应用提供菌剂。【方法】利用高通量测序技术测定广西某油茶壳堆肥中微生物群落分布,筛选适宜堆肥微生物生长的培养基以获得多样性较丰富的堆肥芽孢杆菌菌群,经平板稀释涂布法分离纯化菌株,使用水解圈法和酶活测定方法进行水解酶功能分析,利用菌株形态特征观察、16S rDNA分子鉴定法明确菌株的种属,并运用分子生物学软件MEGAX构建系统发育树。使用重铬酸钾法测定腐殖酸含量。【结果】芽孢杆菌科(Bacillaceae)是优势菌科,占比达55.58%;分离到15株芽孢杆菌,同时具有淀粉酶、纤维素酶、蛋白酶3种水解酶活性的菌株有6株;同时具有淀粉酶和纤维素酶活性的有1株;仅具有蛋白酶或纤维素酶一种酶活性的有2株。其中菌株Bacillus sp. YX11的蛋白酶活力达到27.07±3.28 U·mL~(-1),淀粉酶活力达123.97±3.19 U·mL~(-1),纤维素酶活力达15.75±0.23 U·mL~(-1)。含有这15株菌的复合菌制剂有利于腐殖酸的生成,提高了堆肥品质。【结论】鉴定出的Bacillus cereus YX02和Bacillus flexus FYF01等菌株具有进一步研究开发的价值,可用于开发油茶壳堆肥微生物菌制剂。     

生活垃圾处理机生物降解生活垃圾的微生物研究

[目的]采用生活垃圾处理机生物降解生活垃圾中的微生物。[方法]收集厨余垃圾,对降解混合垃圾、分类垃圾的优势菌群中的菌株和复合菌剂中的菌株进行培养和鉴定。[结果]鉴定出原菌剂中微生物11株;降解淀粉类垃圾的优势菌群微生物3株,分别为枯草芽孢杆菌（Bacillus subtili,s BDh9）、杂菌（sta1）和淀粉液化芽孢杆菌（Bacillus amyloliquefaciens B,BDh1）;降解蛋白质类生活垃圾优势菌群微生物3株,分别为枯草芽孢杆菌（Bacillus subtili,s BDh9）、杂菌（sta1）和肉食杆菌属（Carnobacterium divergens,BD5）;降解纤维素类垃圾的优势菌群微生物5株,分别为枯草芽孢杆菌（Bacillus subtilis,BDh9）、杂菌（sta1）、伯克霍尔德菌（Burkholderia multivoran,s BDh6）、环状芽孢杆菌（Bacillus circulans,BD1）和杂菌（sta2）。[结论]该研究可为建立生物降解生活垃圾自主技术打下基础。     

辣椒枯萎病主要致病菌的分离鉴定及农用拮抗菌筛选

[目的]分离鉴定辣椒枯萎病主要致病菌,探寻能够有效抑制该致病菌的生防菌株.[方法]采用植物病害组织致病菌划线分离方法分离致病菌,经形态学以及ITS区域PCR检测,确定菌株属性,通过平板拮抗筛选,确定对该致病菌具有拮抗作用的农用生防菌株.[结果]初步鉴定该致病菌为尖孢镰刀(Fusarium oxysporum),将该致病菌与11株农用微生物芽孢杆菌和3株农用微生物木霉菌进行拮抗试验,发现枯草芽孢杆菌(Bacillus subtilis)、苏云金芽孢杆菌(Bacillus thuringiensis)、解淀粉芽孢杆菌(Bacillus amyloliquefaciens)、海洋芽孢杆菌(Bacillus marinus)、坚强芽孢杆菌(Bacillus firmus)、甲基营养型芽孢杆菌(Bacillus methylotrophicus)6株芽孢杆菌对该致病菌拮抗效果明显,长枝木霉(Trichoderma longibrachiatum)与哈茨木霉(Trichoderma harzianum)2株木霉菌对该致病菌具有明显的拮抗效果.[结论]筛选到了8株能够有效抑制该病原菌的生防菌株,为以后研究生防肥料奠定了基础.     

土壤微生物菌肥在盐碱地造林中的作用

从我国内蒙古磴口地区的甘草、紫花苜蓿、樟子松、苦豆、刺槐、紫穗槐、沙棘等盐碱地植物根际土壤中分离出固氮微生物10株,从中筛选出3株固氮芽孢杆菌.经鉴定,1株(96-10)为巨大芽孢杆菌(Bacillus megaterium),2株(96-02;96-05)为蜂房芽孢杆菌(Baeillus alve).以草炭为载体制备成微生物菌剂,分别在内蒙古磴口地区和陕西榆林试验区,对10种树种进行了田间施肥试验,获得一定的效果.     

高效污泥降解菌种的筛选及在污泥堆肥中的效果

从采集的高温样品中分离筛选出4株污泥堆肥菌株,经鉴定,这4株菌均系芽孢杆菌属(Bacillus)。将这4株菌等比例混合制成微生物菌剂,按0、3%、5%和10%的菌剂添加量接种至堆肥体系中,筛选合适的堆肥接种量,并探究其在污泥堆肥中的应用效果。结果表明:添加微生物菌剂的处理组在各项堆肥指标上明显优于空白组,处理组堆体的升温速度、物料水分去除率和最高温度值均高于空白组,堆肥结束时,处理组均完全腐熟,而空白组未达腐熟标准。各处理组中,以3%菌剂添加量的堆肥效果最好,各项指标最佳:堆体最高温度达63 ℃,堆体高于55 ℃的高温可持续4 d,堆肥后种子发芽指数为134.1%。说明筛选配制的微生物菌剂在污泥堆肥中具有一定的应用潜力。     

降解牛粪纤维素的复合微生物菌剂制备

[目的]制备降解牛粪纤维素的复合微生物菌剂.[方法]通过正交试验,确定地衣芽孢杆菌、枯草芽孢杆菌和放线菌属4株菌的最优配比,制成降解牛粪纤维素的复合微生物菌剂.[结果]复合微生物菌剂中HP2菌的所占比例为主要因素,各菌种最佳配合比例为TG1∶P3∶HP2∶HN1 =3∶1∶1∶1.[结论]该研究为复合微生物菌剂在牛粪堆肥中的应用提供了理论依据.     

不同微生物茵剂处理对鸡粪堆肥发酵的影响

选择3种混合菌剂进行了鸡粪堆肥发酵试验.研究表明,鸡粪堆肥通过接种微生物菌剂,可以明显提高堆肥初期的发酵温度,加快堆肥物料的水分挥发,改变鸡粪中的微生物数量,缩短堆肥发酵周期,促进堆肥快速腐熟,特别是接种菌剂1(乳酸菌、酵母菌、枯草芽孢杆菌、沼泽红假单孢菌混剂)效果最好,与对照相比,堆肥发酵初期温度提高,中期达到55℃以上,高温期持续8 d,水分含量降低8%,细菌、放线菌数量明显降低.     

作物病害高效生防菌株的定向快速筛选及其种类鉴定

[目的]筛选和鉴定能产Bacillomycin D的芽孢杆菌。[方法]通过PCR技术对36株具有抑菌活性的芽孢杆菌进行检测,筛选出具有Bacillomycin D合成酶基因的菌株,并利用快速蛋白液相色谱技术(FPLC)对筛选出的菌株进行检测,最后通过16S rDNA和gyrB基因序列对这些菌株进行分子鉴定。[结果]共筛选出4株能产生Bacillomycin D的菌株,其中2株为萎缩芽孢杆菌(Bacillus atrophaeus),2株为解淀粉芽孢杆菌(B.amyloliquefaciens)。[结论]该研究为高效微生物农药的开发奠定了基础。     

利用PCR-DGGE方法研究添加复合菌剂对堆肥微生物群落的影响

为研究接种复合菌剂对堆肥微生物群落变化的影响,以奶牛粪便和稻草为原料进行堆肥试验,设添加复合菌剂BLD(由3株木质纤维素降解细菌组成,经测序鉴定分别为枯草芽孢杆菌、地衣芽孢杆菌、假单胞菌属中未鉴定种)和不加菌剂两个处理,利用传统平板培养与聚合酶链式反应-变性梯度凝胶电泳(PCR-DGGE)方法相结合研究两种堆肥处理对微生物群落演替的影响.结果表明,基于传统培养方式,微生物数量呈升高-降低-升高-降低的趋势,且细菌数量明显大于真菌数量;DGGE图谱显示,两种堆肥处理的条带数呈现与传统培养相似的变化趋势.接种菌株在堆肥初期成功定殖,高温期成为优势菌株,降温期优势逐渐减弱.接种菌剂增加了堆肥中细菌数量,提高了堆肥微生物群落多样性,从而促进堆肥微生物群落演替,缩短堆肥腐熟时间.     

耐盐碱微生物复合菌剂的选育、复配及其对盐碱地的改良效果

筛选得到4株能够在盐碱环境下生长的假单胞菌（ Pseudomonas sp．）、枯草芽孢杆菌（ Bacillus subtilis）、巨大芽孢杆菌（ Bacillus megaterium）、酵母菌（ Yeast）,并对这4株菌进行He－Ne激光诱变育种,正突变菌株能在高盐和髙碱条件下生长。经过复配后将其作为功能菌发酵生产生物有机肥,应用于盐碱地改良。与对照相比,微生物复合菌剂对降低盐碱地含盐量、pH值效果显著,有机质含量上升明显。在经过改良的盐碱土中种植紫花苜蓿,出苗率、株高、产量均不同程度增加。     

Chemotaxis in bacteria

Motile Escherichia coli placed at one end of a capillary tube containing an energy source and oxygen migrate out into the tube in one or two bands, which are clearly visible to the naked eye and can also be demonstrated by photography, microscopy, and densitometry and by assaying for bacteria throughout the tube. The formation of two bands is not due to heterogeneity among the bacteria, since the bacteria in each band, when reused, will form two more bands. If an anaerobically utilizable energy source such as galactose is present in excess over the oxygen, the first band consumes all the oxygen and a part of the sugar and the second band uses the residual sugar anaerobically. On the other hand, if oxygen is present in excess over the sugar, the first band oxidizes all the sugar and leaves behind unused oxygen, and the second band uses up the residual oxygen to oxidize an endogenous energy source. The essence of the matter is that the bacteria create a gradient of oxygen or of an energy source, and then they move preferentially in the direction of the higher concentration of the chemical. As a consequence, bands of bacteria (or rings of bacteria in the case of agar plates) form and move out. These results show that E. coli is chemotactic toward oxygen and energy sources such as galactose, glucose, aspartic acid, threonine, or serine. The full repertoire of chemotactic responses by E. coli is no doubt greater than this, and a more complete list remains to be compiled. The studies reported here demonstrate that chemotaxis allows bacteria to find that environment which provides them with the greatest supply of energy. It is clearly an advantage for bacteria to be able to carry out chemotaxis, since by this means they can avoid unfavorable conditions and seek optimum surroundings. Finally, it is necessary to acknowledge the pioneering work of Englemann, Pfeffer, and the other late-19thcentury biologists who discovered chemotaxis in bacteria, and to point out that the studies reported here fully confirm the earlier reports of Beijerinck (4) and Sherris and his collaborators (5,6) on a band of bacteria chemotactic toward oxygen. By using a chemically defined medium instead of a complex broth, it has been possible to study this band more closely and to demonstrate in addition the occurrence of a second band of bacteria chemotactic toward an energy source. Beijerinck (4) did, in fact, sometimes observe a second band, but he did not offer an explanation for it.     

苦豆子内生细菌的分离及拮抗菌的筛选

从塔里木盆地不同地点采集苦豆子,从不同组织中分离内生细菌,并用分离得到的内生细菌对病原真菌做皿内拮抗试验,筛选有生物拮抗活性的内生菌菌株。结果表明,不同来源的苦豆子内生细菌数量上有较大差异,苦豆子不同组织内生细菌的种类和数量也有差异。从苦豆子各种组织中分离得到129株内生细菌,其中5株对多种植物病原真菌具有拮抗作用,这些菌株在苦豆子植株内均可定殖,属于芽孢杆菌。     

Stress-induced mutagenesis in bacteria

The evolutionary significance of stress-induced mutagenesis was evaluated by studying mutagenesis in aging colonies (MAC) of Escherichia coli natural isolates. A large fraction of isolates exhibited a strong MAC, and the high MAC variability reflected the diversity of selective pressures in ecological niches. MAC depends on starvation, oxygen, and RpoS and adenosine 3',5'-monophosphate regulons; thus it may be a by-product of genetic strategies for improving survival under stress. MAC could also be selected through beneficial mutations that it generates, as shown by computer modeling and the patterns of stress-inducible and constitutive mutagenesis. We suggest that irrespective of the causes of their emergence, stress-induced mutations participate in adaptive evolution.     

Cannibalism by sporulating bacteria

Spore formation by the bacterium Bacillus subtilis is an elaborate developmental process that is triggered by nutrient limitation. Here we report that cells that have entered the pathway to sporulate produce and export a killing factor and a signaling protein that act cooperatively to block sister cells from sporulating and to cause them to lyse. The sporulating cells feed on the nutrients thereby released, which allows them to keep growing rather than to complete morphogenesis. We propose that sporulation is a stress-response pathway of last resort and that B. subtilis delays a commitment to spore formation by cannibalizing its siblings.     

禽畜养殖粪便中多重抗生素抗性细菌研究

通过对新乡地区8家养猪场和11家养鸡场饲喂抗生素情况的调研,发现头孢氨苄、阿莫西林、卡那霉素、庆大霉素等4种抗生素是该地区被普遍使用的兽药抗生素。通过多点取样法和微生物培养技术对3家养鸡场和3家养猪场不同养殖时期的粪便进行单一抗生素和多重抗生素抗性细菌的检测,结果表明养鸡场堆置1周的粪便中抗头孢氨苄的细菌比例最高,达到65.90%,对所研究的3种和4种抗生素同时抗性的比例高达8.60%-12.51%和9.73%,明显高于饲喂中药的对照养鸡场样本检测结果(0.02%-2.73%和0.12%)。养猪场堆置1周的粪便中检测到抗头孢氨苄的细菌比例也是最高,达到49.12%上,但养猪场粪便中多重抗生素抗性细菌的比例明显低于养鸡场。同时研究发现,在两种养殖场中,幼龄期粪便中检测到的多重抗性细菌比例明显高于成熟期粪便,这可能与养殖过程中鸡、猪在幼龄期由于防病和促生长等因素而同时大剂量使用多种抗生素有关。