泰州市奶牛乳房炎调查及其病原菌的分离

奶牛乳房炎是奶牛场的常发病和难以防治的疾病,为进一步了解引起奶牛乳房炎的病原微生物,本研究通过对泰州市3个奶牛场572头泌乳牛的乳房炎发病率进行了调查并对乳房炎主要病原菌进行分离和鉴定。结果表明,奶牛临床型乳房炎的发病率为7.69%,隐性型乳房炎发病率为56.64%,乳区阳性率为33.8%,其中临床型：隐性=1：7.36。从65份确定患有乳房炎的奶样中共分离获得8种细菌、156株分离株,且引起乳房炎的主要病原菌是金黄色葡萄球菌和表皮葡萄球菌,其次是停乳链球菌、大肠杆菌、乳房链球菌和无乳链球菌,其检出率分别为42.31%、23.08%、8.97%、6.41%、5.13%和1.28%。该项调查结果初步明确了泰州市乳房炎发病情况,同时为进一步综合防治奶牛乳房炎和研制乳房炎治疗药物提供了科学依据。     

猕猴桃软腐病主要病原菌的分离、鉴定及其生长特性研究

为探究猕猴桃果实采后软腐病害的主要病原菌及其生长特性,2017年9月于湖南凤凰县猕猴桃基地采集具有软腐病症状的100个果实病样,取患病猕猴桃病健交界处果肉,进行病原菌的分离纯化、菌株形态学观察、病原菌的分子生物学鉴定试验,同时分析不同培养基、碳源、氮源、温度和pH值对病原菌生长的影响。结果表明,菌种鉴定为葡萄座腔菌(Botryosphaeria dothidea)和间座壳菌(拟茎点霉菌有性态,Diaporthe phaseolorum)。两种菌在PSA、YPGA、PDA、SDA培养基的生长速度均较快,葡萄座腔菌和间座壳菌生长速率分别在16.1～18.6 mm·d^-1和13.9～16.5 mm·d^-1之间,且菌丝密度均较稠密。淀粉、蔗糖和麦芽糖作为碳源时,病原菌生长效果较好,葡萄座腔菌和间座壳菌生长速率分别在15.9～17.7 mm·d^-1和12.4～16.4 mm·d^-1之间。酵母粉、蛋白胨、甘氨酸作为氮源时,较适宜葡萄座腔菌和间座壳菌生长,两病原菌生长速率分别在11.0 mm·d^-1     

一株耐辐射考克氏菌的分离与鉴定

从10kGy60Coγ射线辐照处理的新疆戈壁土壤中分离到一株耐辐射微生物I-7R,该菌菌落为橙红色革兰氏阳性球菌。最适生长温度为30℃,(G+C)mol%含量为67.5%,不具有氨苄青霉素、氯霉素、四环素、卡那霉素、潮霉素、利福平及壮观霉素等抗性。UV辐射生存曲线显示,I-7R菌株与大肠杆菌K-12相比,具有较强的紫外辐射抗性。16S rDNA序列比较分析表明I-7R菌株与Kocuria rosea、K.erythromyxa以及K.polaris同源性达到99%。结合I-7R菌株生理生化试验结果,该株菌归属于考克氏菌属(Kocuria),并与Kocuria rosea最为相近,暂命名为K.rosea I-7R。     

橄榄黑斑病病原菌的分离鉴定及防治药剂筛选

为明确引起潮州市橄榄黑斑病的病原菌种类，筛选有效防治药剂，对发病果实采用组织分离法进行了病原菌的分离与纯化，并结合形态学和分子生物学技术对病原菌进行鉴定。鉴定结果表明，引起橄榄黑斑病的病原菌为链格孢菌(Alternaria alternata)。通过菌丝生长速率法测定了14种杀菌剂对病原菌的抑菌效果，结果表明14种杀菌剂对链格孢菌均有抑制作用，其中3%中生菌素可湿性粉剂、80%克菌丹水分散粒剂、8%氟硅唑微乳剂和40%苯甲·嘧菌酯悬浮剂的抑菌效果较好，EC50分别为1.267、1.207、1.100、0.944 mg·L^-1。田间防效测定结果表明，40%苯甲·嘧菌酯悬浮剂、8%氟硅唑微乳剂的防治效果显著，防效为68.58%、65.44%；而30%王铜、2%春雷霉素的防效较差，仅为10%左右。综上，推荐将8%氟硅唑微乳剂和40%苯甲·嘧菌酯悬浮剂2种药剂作为防控橄榄黑斑病的首选药剂。本研究可为橄榄黑斑病高效杀菌剂的筛选提供数据参考，并为其防控工作提供科学依据。     

基于构建微生物传感器的甲基对硫磷降解菌的分离鉴定及其降解特性研究

有机磷农药是目前环境中残留量最多的农药之一,对其残留量的检测及降解机制的研究对于环境污染及生态修复具有重要意义。微生物传感器由生物学元件与换能器构成,因具有成本低廉、易于微型化及选择性高等特点而被广泛应用于各种生化物质的分析和检测。本文从长期受农药污染的土壤中分离出4株能以甲基对硫磷为碳源生长的菌株,根据形态特征和16S r RNA基因序列同源性分析,对4株降解菌进行鉴定,利用高效液相色谱测定降解率,选取降解率最高的1株菌进行降解机制研究,以期将其应用于测定环境中甲基对硫磷残留的电位型微生物传感器的构建。结果表明,在甲基对硫磷初始浓度50 mg·L-1、30℃、p H 7.0的培养条件下培养7 d,4株菌对甲基对硫磷的降解率均在78%以上,其中1株菌的降解效率可达100%。16S r RNA基因序列测定表明,该菌株属于克雷伯氏菌属,命名为Klebsiella sp.MP-6。利用液相色谱-质谱联用对其降解产物的研究表明,菌株MP-6水解甲基对硫磷主要产生二甲基硫代磷酸(dimethyl thiophosphoric acid,DMTP)和对硝基苯酚(p-nitrophenol,PNP),极少部分PNP通过产生4-硝基邻苯二酚(4-nitrocatechol,4-NC)和1,2,4-苯三酚(1,2,4-BT)进一步代谢。结果表明,基于测定中间产物对硝基苯酚(p-nitrophenol,PNP)的电位响应信号,该菌株适用于构建测定海水及土壤等环境中有机磷农药的微生物传感器。     

白茶中金花菌的分离与鉴定

金花白茶是白茶中的一种新兴产品.研究金花白茶中金花菌的类型有利于为白茶产业的创新发展提供一定的参考依据.本研究以产自2018年的湖南湘丰牌桑植白茶作为材料,通过平板涂布法及三点接种法分离纯化金花菌,采用形态学观察并联合分子生物学手段对所获得的分离株进行鉴定.分离获得一株金花菌BC-1,经菌落特征、显微形态观察、ITS测...     

黄颡鱼源维氏气单胞菌的分离、鉴定及特性分析

为确定导致江西南昌地区黄颡鱼腹水病暴发的病原,进行了病鱼的临床解剖观察、病原菌分离鉴定和回归感染、病原菌株药敏及毒力基因检测等系统性试验和分析。结果显示,患病鱼体鳃、肝脏、肾脏以及肠道等组织均发生不同程度的病变;由病鱼体内分离获得一株优势菌PL1,经过16S rDNA序列比对和系统进化树分析,鉴定为维氏气单胞菌（Aeromonas veronii）。回归感染结果表明,菌株PL1能够引起黄颡鱼与自然患病相同的典型腹水病病症,具有较强的致病性;药敏试验结果显示,菌株PL1对30种抗菌药物中多粘菌素、复方新诺明、呋喃唑酮等12种药物敏感,对氧氟沙星、红霉素和哌拉西林3种药物中度敏感,而对诺氟沙星、环丙沙星、万古霉素等15种抗生素具有耐药性,呈现出多重耐药性。毒力基因检测结果显示,菌株PL1携带有aerA、act、gcaT、ser、fla、ascV等6种毒力基因,表明菌株PL1具有较强的毒力。本研究为黄颡鱼腹水病的防治提供了科学指导,同时为深入研究患病机制提供了基础数据。     

一株百合内生细菌 Burkholderia sp. FJb-2的分离鉴定及其体外抑菌促生效应

从北京市房山区百合资源圃采集红芯百合(Fangio)植株样品,通过组织块分离法分离内生细菌;进一步以百合灰霉病菌—灰葡萄孢菌( Botrytis cinerea)ACCC36423、百合叶尖干枯病菌—葡萄座腔菌( Botryosphaeria dothidea)ACCC37263和百合枯萎病菌—尖孢镰刀菌( Fusarium oxysporum)F01139等几种主要百合病原真菌为指示菌,采用平板对峙法筛选具有拮抗活性的拮抗菌株;并对拮抗菌株进行产 1-氨基环丙烷 -1-羧酸(ACC)脱氨酶活性、产生长素(IAA)、溶磷、产铁载体等多种植物促生特性测定;最后通过形态观察及 16S rRNA基因序列分析,初步鉴定其种属。结果从百合茎部分离获得一株内生细菌菌株 FJb-2;该菌株对灰葡萄孢菌、葡萄座腔菌和尖孢镰刀菌均有较高拮抗活性,对病原真菌生长的抑制率范围为 60%～ 81%;该菌株产 ACC脱氨酶活性为     

四环素类抗生素降解菌的分离与鉴定

为了获得四环素类抗生素的高效降解菌,从长期受四环素类抗生素污染的土壤中分离、重复驯化筛选得到6株对四环素类抗生素具有降解能力的菌株;采用高效液相色谱法研究了6株降解菌的降解效果,并通过形态、生理生化特征及16S rDNA序列分析对其进行鉴定。结果表明,菌株TJ-2#对土霉素、四环素和金霉素3种四环素类抗生素的降解效果最好,降解率分别为58.3%、63.9%和65.5%,TJ-6#其次;6株降解菌均能以四环素类抗生素作为唯一碳源生长。经形态、生理生化特征及16S rDNA鉴定,菌株TJ-2#为木糖氧化无色杆菌(Achromobacter sp.),TJ-6#为枯草芽胞杆菌(Bacillus sp.)。本研究结果对四环素类抗生素的生物降解具有一定的现实意义。     

Isolation and identification of iron-reducing bacteria from gley soils

Seventy-one facultative anaerobic bacteria, capable of reducing iron oxide in pure culture, were isolated from three differently gleyed subsoils. The bacteria were picked at random from poured plates (10⁻⁵ and 10⁻⁶) inoculated with serially diluted soil samples. An attempt was made to identify these strains by morphological and biochemical tests. Among these 71 iron-reducing bacteria, all except three were capable of reducing nitrate to nitrite and 35 reduced nitrite further into gaseous compounds (denitrification), but only one strain (Bacillus subtilis) produced H₂S. Based upon their physiological and morphological properties, 38 strains were allotted to the genus Pseudomonas, 31 sporeformers to the genus Bacillus and two were regarded to be coryneform (Arthrobacter?) bacteria. Species identified were Ps. denitrificans (23), ps. stutzeri (8) ps. fluorescens-putida (5), Bacillus cereus (6), B. cereus var. mycoides (14) and Bacillus subtilis (9). Two spore-forming bacilli, two non-pigmented pseudomonads and two coryneform type of bacteria could not be identified. The significance of the enzyme nitrate reductase (nitratase) of these bacteria for anaerobic respiration and as a mechanism of iron reduction is discussed.     

Isolation and identification of oxamyl-degrading bacteria from UK agricultural soils

Bacteria capable of utilising oxamyl as the sole carbon source were isolated from seven different agricultural soils that had previously demonstrated enhanced oxamyl degradation in a soil incubation study. Partial sequencing and alignment of the 16S rRNA gene showed little diversity amongst isolates, with 26 of the 27 isolates demonstrating similarity to the genus Aminobacter. The most common species isolated was Aminobacter aminovorans, while a number of the isolates demonstrated an equal degree of similarity to the species Aminobacter niigataensis and Chelatobacter heintzii. One isolate was identified as Mesorhizobium sp. This is the first time that organisms involved in the degradation of oxamyl have been isolated and identified.     

Isolation and identification of boron-accumulating bacteria from contaminated soils and active sludge

Abstract

High levels of boron are toxic to living organisms. The removal of boron from contaminated water is an important way of dealing with this environmental problem. For this purpose, we screened for bacteria that can absorb high levels of toxic boron from the environment. A range of boron-accumulating bacteria was isolated from boron-contaminated soil in Turkey, from uncontaminated soil in Tokyo, Japan, and from active sludge in Tokyo by semi-quantifying the boron accumulation of each single-cultured bacterium. Intracellular boron concentrations ranged from 0.3 to 1.36?nmol?g^?1 dry weight. Nineteen isolates identified by 16S rRNA gene sequence analysis were most closely related to the genera Bacillus, Variovorax, Pseudomonas, Shewanella, Mycobacterium and Rhodococcus. To our knowledge, this is the first study examining a wide range of bacterial strains in terms of boron accumulation.     

Isolation and identification of boron-accumulating bacteria from contaminated soils and active sludge

High levels of boron are toxic to living organisms. The removal of boron from contaminated water is an important way of dealing with this environmental problem. For this purpose, we screened for bacteria that can absorb high levels of toxic boron from the environment. A range of boron-accumulating bacteria was isolated from boron-contaminated soil in Turkey, from uncontaminated soil in Tokyo, Japan, and from active sludge in Tokyo by semi-quantifying the boron accumulation of each single-cultured bacterium. Intracellular boron concentrations ranged from 0.3 to 1.36 nmol g⁻¹ dry weight. Nineteen isolates identified by 16S rRNA gene sequence analysis were most closely related to the genera Bacillus , Variovorax , Pseudomonas , Shewanella , Mycobacterium and Rhodococcus . To our knowledge, this is the first study examining a wide range of bacterial strains in terms of boron accumulation.     

枸杞鲜果霉变菌种分离鉴定及其生物学特性

为确定引起枸杞霉变的菌种类型,该文以采后鲜枸杞果实为材料,分离导致其霉变的菌种,经形态学特征及分子生物学特征鉴定确定类型,并对与其霉变速率相关的部分生物学特性进行研究。结果表明:从霉变的鲜枸杞中共分离出6株致霉菌,分别鉴定为:菌种1为镰刀菌、菌种2为交链孢霉、菌种6为尖孢镰刀菌、菌种7为青霉菌、菌种11为串珠状赤霉菌及菌种21为草酸青霉。经过回接试验发现6种霉菌均能使健康枸杞鲜果霉变,从这些病斑中再次分离得到相同霉菌。6种菌的致死温度不同,最高致死温度为54℃。菌种1的最适生长温度为40℃,菌种6的最适生长温度为35℃,其余4株菌的最适生长温度均为25℃。6种菌在相对湿度为30%~90%范围内均能生长,最适生长相对湿度为75%~85%。最适p H值范围为4~8,p H值为6时霉菌孢子萌发率最高。通过对6种霉菌与温度、相对湿度、p H值3个与干燥参数相关的生物学特性研究,为今后鲜枸杞干燥过程条件的控制提供理论依据。     

Isolation and identification of effective cellulolytic bacteria in composting process from different sources

Cellulolytic bacteria have a bio-activating role in the composting process. A study was carried out to isolate and identify cellulolytic bacteria from various sources. The isolates were cultured in the carboxymethyl cellulose (CMC) agar medium and incubated at 30°C for 3–7 days. Based on morphological characteristics of the isolates, maximum diameter of a clear zone around the colony and maximum cellulolytic activity, eight isolate were selected for further studies regarding composting experiments.

Molecular tests based on PCR amplification and sequencing of 16S rRNA gene of isolates showed the closest phylogenetic similarity with the species of Stenotrophomonas rhizophila DSM14405 (99.8%), Brevibacterium halotolerans DSM8802 (99.6%), Achromobacter marplatensis B2 (99.8%), Bacillus methylotrophicus CBMB205 (100%), Pseudomonas azotoformans IAM 1603 (99.7%), Bacillus sonorensis NBRC 101234 (99.8%), Bacillus subtilis KCTC 13429 (100%) and Ochrobactrum thiophenivorans DSM 7216 (99.3%). The study of the isolates impact on the composting of palm wastes in a randomized complete block design with 11 treatments in 3 replications showed that strain IB (B. methylotrophicus) caused a significant decrease in C:N ratio (58%). The increasing of microbial respiration compared with control after 30 days incubation at 37°C showed that the B. methylotrophicus strain IB with cellulolytic characteristics can be applied to hydrolysis of cellulosic biomass in the composting processes.     

寒区降解多环芳烃耐冷菌株的分离鉴定及特性

多环芳烃（Polycyclic Aromatic Hydrocarbons,PAHs）是土壤中一种典型的持久性有机污染物。典型寒区东北地区因农业投入品不合理使用、污灌等造成农田土壤中含有大量致癌、致畸与致突变的多环芳烃,针对寒区气候特征致使农田中微生物降解多环芳烃效果不佳,难以改善农田土壤环境并降低食品风险的关键问题,基于目前常用商品化的降解微生物多来自于温暖地区,且难以适应寒区气候的特性,该研究以PAHs典型污染物-菲（PHE）为研究底物,驯化温度15 ℃,筛选分离出适应寒区低温环境的7株菌。经鉴定及降解性能研究,筛选了3株在温度20 ℃、接种量5%、pH值为8、底物浓度500 mg/L,以及外源物质腐殖酸的促进的条件下PHE降解率达到80%的高效降解耐冷菌。以上3株菌两两之间与三者组合均无拮抗关系。对菌株进行碳源的广谱性分析,菌株对2-5环多环芳烃降解率可达15%～85%之间,在将菌株应用至20 ℃土壤环境时,60 d可降解土壤中75%的PHE。该耐冷菌群适应条件符合寒区农田土壤实际环境,研究结果对黑土地区土壤多环芳烃污染的微生物修复提供了一定的基础资料。     

盐穗木根际耐盐碱细菌的分离筛选及鉴定

将采集于新疆和田地区于田县盐碱地的2种盐穗木根际土壤样品制备成菌悬液,经适当稀释后,涂布于牛肉膏蛋白胨培养基中,经分离纯化获得20株菌株。经过分别或同时提高培养基的盐、碱浓度进行菌株耐盐碱性能的筛选,获得1株耐盐碱菌株D8,其在pH值12且盐(NaCl)浓度达到120 g/L的培养基中仍可生长,该菌株既属于极端嗜盐微生物又属于极端嗜碱微生物。通过对筛选出来的菌株进行革兰氏染色制片、生理生化检测、电镜观察、Biolog鉴定、16S rRNA基因测序及系统发育树的建立来鉴定种属关系。鉴定结果是该菌为太平洋盐单胞菌(Halomonas pacifica)。     

Isolation and identification of potassium-solubilizing bacteria from tobacco rhizospheric soil and their effect on tobacco plants

Soil potassium supplementation relies heavily on the use of chemical fertilizer, which has a considerable negative impact on the environment. Potassium-solubilizing bacteria (KSB) could serve as inoculants. They convert insoluble potassium in the soil into a form that plants can access. This is a promising strategy for the improvement of plant absorption of potassium and so reducing the use of chemical fertilizer. The objectives of this study were to isolate and characterize tobacco KSB and to evaluate the effects of inoculation with selected KSB strains on tobacco seedlings. Twenty-seven KSB strains were isolated and identified through the comparison of the 16S ribosomal DNA. Among them, 17 strains belonged to Klebsiella variicola, 2 strains belonged to Enterobacter cloacae, 2 strains belonged to Enterobacter asburiae, and the remaining 6 strains belonged to Enterobacter aerogenes, Pantoea agglomerans, Agrobacterium tumefaciens, Microbacterium foliorum, Myroides odoratimimus, and Burkholderia cepacia, respectively. All isolated KSB strains were capable of solubilizing K-feldspar powder in solid and liquid media. K. variicola occurred at the highest frequency with 18 strains. Four isolates, GL7, JM3, XF4, and XF11, were selected for a greenhouse pot experiment because of their pronounced K-solubilizing capabilities. After being treated with the four KSB strains, plant dry weight and uptake of both K and nitrogen (N) by tobacco seedlings increased significantly. These increases were higher with the combination of KSB inoculation and K-feldspar powder addition. Isolate XF11 showed the most pronounced beneficial effect on plant growth and nutrient uptake by tobacco seedlings. Combining the inoculation of KSB and the addition of K-feldspar powder could be a promising alternative to commercial K fertilizer and may help maintain the availability of soil nutrients. Further studies are necessary to determine the effects of these bacterial strains on mobilization of potassium-bearing minerals under field conditions.     

百菌清降解菌的分离鉴定及功能基因分析

百菌清是一种广谱非内源性农药，在土壤中难以降解，已经成为农业环境污染的主要污染源之一。因此，其对环境的污染和被污染土壤的修复技术越来越受到关注。土壤环境中原位降解细菌的多样性对于评价环境毒理学、生物降解性、自净化能力和污染物的修复潜力具有重要价值。该研究从长期被百菌清污染的土壤中收集大量样本，分离到了14种能够明显降解百菌清的细菌。根据菌株形态和rDNA同源性分析，将它们分为假单胞菌属（Pseudomonas sp.）、无色杆菌属（Achromobacter sp.）、苍白杆菌属（Ochrobactrum sp.）、青枯菌属（Ralstonia sp.）和溶杆菌属（Lysobacter sp.）。其中溶杆菌属是该研究中新发现的具有百菌清降解能力的功能菌属，该发现扩大了已知的百菌清降解菌的菌属范围。通过进一步鉴定及生理生化分析，确定了该降解菌的分类地位及理化特征。此外，该研究通过基因文库法克隆到了发挥关键降解作用的水解酶基因，并发现该基因与转座子原件IS-Olup相连，二者组成代谢转座子，具有水平转移的分子基础。通过揭示降解基因在细菌间的漂移机制，为修复百菌清污染土壤的生物技术奠定了理论基础。