海洋赤杆菌科菌株的类胡萝卜素合成基因分析

赤杆菌科(Erythrobacteraceae)菌株广泛分布于海洋环境,可合成类胡萝卜素等多种色素。类胡萝卜素具有光保护和抗氧化能力,对于赤杆菌科菌株适应海洋生态系统具有重要作用。本研究收集了107个海洋来源的赤杆菌科细菌基因组,分析了类胡萝卜素合成途径中关键酶的编码基因crtEBIGYZW在基因组中的分布特征,构建了基于氨基酸序列的不同编码基因系统发育树,阐明了深海和浅海来源的赤杆菌科菌株中类胡萝卜素合成基因的存在和分布规律,探究了不同的类胡萝卜素合成基因在近海与深海来源的赤杆菌科菌株进化过程中的遗传模式。研究结果显示,海洋来源的赤杆菌科菌株均含有crtEIGZ基因,约98.1%的菌株含有crtBY基因,约43.9%的菌株中存在crtW基因,其类胡萝卜素合成基因在菌株间存在分布差异,但在深海与浅海来源上并未显示特异性。此外,通过比较不同编码基因的系统发育树拓扑结构发现,赤杆菌科菌株的类胡萝卜素合成基因crtBYZW与系统发育密切相关,而crtEIG基因多通过水平基因转移获得,这有助于更好地评估类胡萝卜素合成基因家族在赤杆菌科中的进化,也为其他海洋细菌中的类胡萝卜素合成途径及基因研究提供科学依据,从而有助于开发海洋来源的产类胡萝卜素菌株。

对番茄枯萎病具防效的海洋源赤杆菌YH-07的发酵条件探究

一株分离自连云港海域的赤杆菌Erythrobacter sp.YH-07对番茄枯萎病病原菌有较好的平板抑菌效果。通过单因子水平试验、正交试验对该菌株的培养基配方及摇瓶发酵条件进行优化,最优培养基配方为酵母浸膏0.25%、蔗糖1%、蛋白胨1%、Na Cl 3%和Mg SO4·7H2O 0.03%;最优发酵条件组合为培养温度30℃,140 r/min摇床培养,接种量1%,装液量50 m L/250 m L,初始p H 7.0。优化后的平板抑菌能力提高了9.4%。温室盆栽试验表明,赤杆菌YH-07对番茄枯萎病有显著抑制作用,防治效果保持在50%以上。同时显著降低番茄根际土的真菌数量,增加细菌和放线菌数量。综上所述,赤杆菌YH-07能有效防治番茄枯萎病害,改善根际微生物数量结构,具有进一步开发应用的前景。

赤杆菌科微生物分类研究进展

赤杆菌科微生物隶属于细菌域(Bacteria)、变形菌门(Proteobacteria)、α-变形菌纲(Alphaproteobacteria)、鞘脂单胞菌目(Sphingomonadales)。依据16S rRNA基因序列的系统发育分析结果,2005年赤杆菌科被提议建立,成立之初由3个属5个种组成;在过去10年中,该科的物种数量增加迅速;截止2015年10月,赤杆菌科合格发表6个属44个种。赤杆菌科微生物在土壤、热泉、海水和沉积物等环境中广泛分布,其在环境修复、食品工程及生物医药等领域具有重要的应用前景。此外,部分菌株具有细菌叶绿素a,营好氧异养生长兼具光合作用功能,是好养不产氧光合异养菌的一个重要类群,近年来其在全球海洋碳循环和能量代谢过程中的重要作用日益凸显,得到了国内外学者的关注。本文综述了赤杆菌科微生物的分类特征、生态分布和应用基础等方向的研究进展,并对其分类研究趋势做了分析。

印度洋深海沉积物可培养放线菌多样性及抗革兰阴性菌活性物质研究

目的从印度洋深海沉积物中分离可培养海洋放线菌,对获得的一株海洋微生物进行抗革兰阴性菌活性物质研究。方法设计11种分离培养基进行海洋放线菌分离,通过16S r RNA基因序列比对分析确定菌株分类;利用多种色谱方法,对菌株IMB16-059产生的活性物质进行分离纯化,通过波谱学和化学方法鉴定其结构。结果根据菌落表型共分离得68株微生物,经16S r RNA分析表明,其中34株属于放线菌门,覆盖5科5属;其余属于厚壁菌门和变形菌门。25株发酵产物显示出抗革兰阴性菌活性。从赤杆菌IMB16-059中分离鉴定了6个化合物,其结构分别确定为L,L-环(脯氨酸-丙氨酸)(1),L,L-环(脯氨酸-酪氨酸(2)),D,L-环(脯氨酸-酪氨酸)(3),邻氨基苯甲酸(4),胸腺嘧啶(5),1H-喹啉-4-酮(6)。化合物1和4显示出抗铜绿假单胞菌活性,在样品浓度为20μg/片时,抑菌圈直径分别为13和20mm。结论印度洋深海沉积物蕴含丰富的微生物类群,其发酵产物显示出较好的抗革兰阴性菌活性,可为新抗生素发现提供重要的菌种资源。

Expression of Exogenous Gene in Prokaryotes and Eukaryotes

[Objective] This study aimed to investigate the expression of exogenous gene in prokaryotes and eukaryotes. [Method] The expression vector pTYB2-WF harboring target gene was transformed into E. coli ER2566. IPTG was employed to induce the expression of phytase gene. The expression of phytase fusion protein was detected by SDS-PAGE, and the fusion protein was further purified. Phytase gene phyA was expressed in Pichia pastoris expression system. Yeast recombinant vector pPIC9K-phyA was constructed and transformed into P. pastoris GS115 to construct engineering strain GS115-pPIC9K-phyA. [Result] Phytase protein was ex-pressed under methanol induction. Enzyme activity assay indicated that the activity of phytase was 7.3 U/ml. P. pastoris engineering strain GS115-pPIC9K-phyA was successful y constructed. [Conclusion] Methanol yeast expression mechanisms play a certain role in molecular biology and industrial applications.

Isolation of an acid producing Bacillus sp. EEEL02: Potential for bauxite residue neutralization

Bauxite residue deposit area(BRDA)is a typical abandoned mining wasteland representing extreme hostile environment with increased alkalinity.Microbially-driven neutralization of bauxite residue,based on the microbial acid producing metabolisms,is a novel strategy for achieving rapid pH neutralization and thus improving its environmental outcomes.The hypothesis was that these extreme conditions promote microbial communities which are capable of novel ecologically relevant functions.Several alkaliphilic acid producing bacteria were isolated in this study.One strain was selected for its superior growth pattern and acid metabolism(termed EEEL02).Based on the phylogenetic analysis,this strain was identified as Bacillus thuringiensis.The optimized fermentation conditions were as follows:pH 10;NaCl concentration 5%;temperature 25℃;EEEL02 preferred glucose and peptone as carbon and nitrogen sources,respectively.Based on optimal fermentation conditions,EEEL02 induced a significant pH reduction from 10.26 to 5.62 in 5-day incubation test.Acetic acid,propionic acid and CO2(g)were the major acid metabolites of fermentation,suggesting that the pH reduction in bauxite residue may be caused by acid neutralization derived from microbial metabolism.This finding provided the basis of a novel strategy for achieving rapid pH neutralization of bauxite residue.

The Impact of Beauveria bassiana, Bacillus thuringiensis （Bt） Spray, Trichogramma pretiosum and Spinosad on the Lepidoptera： Crambidae, European Corn Borer （Ostrinia nubilalis）

The research cxamined the effects of microbial and novel insecticidal control strategies and also Irichogramma pretiosum on the I,epidoptera： Crambidae, the European corn borer （ECB）, （Ostrinia nubilalis）. The treatments included ~ino^d, from bacteria the entomopathogenic fungus Beauveria bassiana, the egg parasitoid Trichogramma pretiosum, and foliar applied Bt spray, and t^vo combinations Bearveria bassiana plus Bt spray and Trichogramma pretiosum and spinosad. The infestations by the European corn borer larvae on stalks of corn were very＂ severe in all the control treatments both 2006 and 2007. The microbial treatments caused various levels of European corn borer mortality. In all the parameters there was a significant difference between the control and other treatments with a P 〈 0.00 l. In this research, treatment with spinosad emerged as the most effective biological agent in the control of the European corn borer. The treatments had no significant effects on the relative abundance and composition of non-target arthropods （P 〉 0.05）. This research is relevant in boosting underutilized control strategies and increasing stakeholder adoption of integrated pest management practices and thereby＇ reducing the use of conventional insecticides especially for orgamc thrmers. This is very important especially in certain countries where the commumities are skeptical about consuming transgenic crops.

Efficacy of Trichogramma evanescens and Bacillus thuringiensis kurstaki to Control Lesser Date Moth Batrachedra amydraula Merck

Efficacy of the egg parasitoid, Trichogramma evanescens and the bio-pesticide bacteria, Bacillus thuringiensis kurstaki （Btk） in reduction of lesser date moth, Batrachedra amydraula infestations were studied during seasons of 2011 and 2012. The result of 2011 showed that the release of egg parasitoid by rate of 500 and 1,000 parasitoids/palm tree achieved 55.06% and 67.45% reduction in infestation percentage, respectively. Meanwhile high reduction in infestation was achieved by Btk （78.65%） at dose rate of 6 g Btk powder/kg talc powder. The result of 2012 showed that the highest control efficacy （58.78%） was achieved at the first time of the control process, using Btk at dose rate of 6 g/kg talc powder. This result was not different significantly from the release of egg parasitoid by rate of 1,000 parasitoids/palm tree which achieved 53% reduction in infestation percentage. At the second time （after one week from the first application）, the efficacy of Btk at dose rate of 6g/kg talc powder was increased to 78.78%, which differ significantly with the release of egg parasitoid by the same rate of 1,000 parasitoids/palm tree, having 61.79% efficacy. Reapplication of the control method achieved an increase in efficacy of the control agents, such increase was significant for both the release of the parasitoid at rate of 1,000/palm tree and for Btk at a dose of 6 g/kg talc powder.