Isolation and identification of Fusarium oxysporum f. sp. cubense in Fujian Province, China

Fusarium wilt, caused by Fusarium oxyporum f. sp. cubense(Foc), is the most serious disease affecting banana production.To clarify the distribution of the Foc races in Fujian Province of China, 79 soil samples were collected from four regions of Zhangzhou City, the primary banana production area in Fujian. We isolated and identified 12 Foc strains based on internal transcribed spacer(ITS) sequence analysis, PCR amplification by using Foc-specific primers and pathogenicity assays.Our analysis indicated that 11 isolates belong to Foc race 1, and 1 isolate belongs to the Foc tropical species race 4(TR4).Although TR4 has previously been reported to occur in primary banana-producing provinces, such as Hainan, Guangxi,and Guangdong of China, this is the first report of TR4 isolated from the soil in Fujian Province. Monitoring the presence of Foc, in particular, the TR4 strains in the soil, is the basic strategy to prevent and control Fusarium wilt.

Identification of Biocontrol Strain HJX1 and Determination of Its Antifungal Spectrum

A biocontrol strain HJX1,which had good inhibitory effect against Fusarium oxysporum f. sp. cubense,was identified as Bacillus subtilis based on traditional morphology,16 S rRNA gene sequence and fatty acid profiles. The sequence of 16 S rRNA gene had 99% homology to B. subtilis according to the DNA fragment of 1 521 bp amplified with a pair of universal primers,P0 and P6,of bacteria. Confront culture with pathogenic fungi showed that this strain had good inhibitory effect against 10 pathogenic fungi.

Isolation and Preliminary Identification of the Pathogen of Anthracnose in Camellia sp.

The pathogens of anthraenose sampled from leaves and fruits of Camellia oleifera in Cenxi City and C. vietnamensis in Ningming country, Guangxi Prov- ince, were isolated via tissue methods and four isolates were obtained. Morphological description and phylogenetic analysis were made based on rDNA ITS se- quences. The results showed that the strains 742, GW2 and CR1 isolated from leaves were clustered into one group, and the strain CRF1 isolated from fruits was clustered into another group. All the four strains belonged to Colletotrichum gloeosporioides complex.

Selection and Molecular Biological Identification of a Strain of Bacillus sp. Inhibiting the Growth of Saprolegnia ferax

Based on the theory of biological control of Saprolegnia ferax,antagonism test of nine strains of Bacillus sp. to S. ferax JL was carried out. Bacillus sp.BA1 was screened to have significantly inhibitory effects on the growth of S. ferax JL( P 【 0. 05). Then,the effects of Bacillus sp. BA1 on different sources of S. ferax were carried out. Results showed that BA1 also had significantly inhibitory effects on S. ferax 6#,10# and S2( P 【 0. 05). Sequence of 16 S r DNA of BA1 was analyzed; and homologous alignment analysis showed that BA1 had more than 99% similarity with Bacillus cereus. Therefore,it could be concluded that strain BA1 was B. cereus,which significantly inhibited the growth of S. ferax and could be used as the biological control agent for S. ferax diseases in aquaculture.

n-Hexadecane and pyrene biodegradation and metabolization by Rhodococcus sp. T1 isolated from oil contaminated soil

The high-molecular weight polycyclic aromatic hydrocarbons(PAHs) pyrene and typical long chain alkane nhexadecane are both difficult to degrade. In this study, n-hexadecane and pyrene degrading strain Rhodococcus sp. T1 was isolated from oil contaminated soil. Strain T1 could remove 90.81% n-hexadecane(2 vol%) and 42.79% pyrene(200 mg·L^(-1)) as a single carbon within 5 days, respectively. Comparatively, the degradation of pyrene increased to 60.63%, but the degradation of n-hexadecane decreased to 87.55% when these compounds were mixed. Additionally, identification and analysis of degradation metabolites of Rhodococcus sp. T1 in the above experiments showed that there were significant changes in alanine, methylamine, citric acid and heptadecanoic acid between sole and dual substrate degradation. The optimal conditions for degradation were then determined based on analysis of the pH, salinity, additional nutrient sources and liquid surface activity.Under the optimal conditions of pH 7.0, 35 °C, 0.5% NaCl, 5 mg·L^(-1) of yeast extract and 90 mg·L^(-1) of surfactant,the degradation increased in single or dual carbon sources. To our knowledge, this is the first study to discuss metabolite changes in Rhodococcus sp. T1 using sole substrate and dual substrate to enhance the long-chain alkanes and PAHs degradation potential.

Diversity of Secondary Metabolites from Two Antarctic Microbes Rhodococcus sp. NJ-008 and <i>Pseudomonas</i>sp. NJ-011

Diversity analysis on secondary metabolites of Antarctic microbes, Rhodococcus sp. NJ-008 and Pseudomonas sp. NJ-011, together with the structural elucidation of some purified compounds, has been carried out for understanding of their chemical constituents. The methanol extracts of Rhodococcus sp. NJ-008 and Pseudomonas sp. NJ-011 were subjected to HPLC-TOF MS test for diversity analysis on secondary metabolites, respectively. The chemical constituents of NJ-011 are mainly N-containing compounds including some alkaloids and short polypeptides, while those of NJ-008 are not N-containing ones. Three compounds were also isolated and identified from extract of NJ011 by different column chromatography and preparative HPLC, and their structures were elucidated as b-carboline (1), 3-benzylhexahydropyrrolo[1,2-a]pyrazine-1,4-dione (2) and 3-isobutylhexahydropyrrolo[1,2-a]pyrazine-1,4-dione (3) by comparison of TOF MS, 1Hand 13C-NMR data with those reported. More microbial material of Pseudomonas sp. NJ-011 should be needed for exploration of the minor constituents with complicated structures.

Inducing Expression and Reaction Characteristic of Nitrile Hydratase from Rhodococcus sp. SHZ-1

Inducing expression and the reaction characteristic of nitrile hydratase （NHase） from Rhodococcus sp. SHZ-1 were investigated. The results showed that the expression of NHase was greatly enhanced with the cooperation of acrylonitrile and ammonium chloride as inducer in the medium and the specific activity of NHase was increased of 44%. Then the temperature, pH, concentration of acrylonitrile and acrylamide were evaluated, which affected the activity and reaction characteristic of NHase. It was found that the temperature and concentration of acrylarnide were the most important factors for the catalyzation of NHase. The optimal catalysis temperature of NHase from Rhodococcus sp. SHZ-1 was 30℃, and the activation energy of the hydration of NHase was 90.2kJ·mol^-1 in the temperature range from 5℃ to 30℃. Kmof NHase was 0.095mol·L^-1 using acrylonitrile（AN） as substrate, and NHase activity was inhibited seriously when acrylonitrile concentration was up to 40g·L^-1, the substrate inhibition constant Ki is 0.283mol·L^-1. Moreover, the NHase from Rhodococcus sp. SHZ-1 had very strong tolerance to acrylamide, in which the final concentration of acrylamide reached to 642g·L^-1 and the residual activity of NHase still maintained 8.6% of the initial enzyme activity.

Microbial Removal of Phenol and P-Chlorophenol from Industrial Waste Water Using Rhodococcus sp.RSP8 and Its Growth Kinetic Modeling

A phenol-degrading microorganism, Rhodococcus sp.RSP8, was used to study the substrate interactions during cell growth on phenol and p-chlorophenol dual substrates. Both phenol and p-chlorophenol could be utilized by the bacteria as the sole carbon and energy sources. When cells grew on the mixture of phenol and p-chlorophenol, strong substrate interactions were observed. The p-chlorophenol inhibited the degradation of phenol, on the other hand, phenol also inhibited the utilization of p-chlorophenol. The overall cell growth rate depends on the co-actions of phenol and p-chlorophenol. In addition, the cell growth and substrate degradation kinetics of phenol, p-chlorophenol as single and mixed substrates for Rhodococcus sp.RSP8 in batch cultures were also investigated over a wide range of initial phenol concentrations (5-1600 mg.L–1) and initial p-chlorophenol concentrations (5 – 250 mg.L–1). The single-substrate kinetics was described well using the Haldane-type kinetic models, with model constants of μm1 = 0.15 h–1, KS1 = 2.22 mg.L–1 and Ki1 = 245.37 mg.L–1 for cell growth on phenol and of μm2 = 0.0782 h–1, KS2 = 1.30 mg.L–1 and Ki2 = 71.77 mg.L–1, K′i2 = 5480 (mg.L–1)2 for cell growth on p-chlorophenol. Proposed cell growth kinetic model was used to characterize the substrates interactions in the dual substrates system.

Study on Characteristics in the Removal Process of Ammonia Nitrogen and Nitrate Nitrogen by an Isolated Heterotrophic Nitrification-Aerobic Denitrification Strain Rhodococcus Sp.

Removal of ammonia nitrogen and nitrate nitrogen by an heterotrophic nitrification-aerobic denitrification strain is an economical and effective method. In this article, a kind of heterotrophic nitrification-aerobic denitrification strain which has aerobic denitrification and heterotrophic nitrification ability was selected, and then was identified as rhodococcus sp. by 16S rRNA sequencing analysis and morphological observation. After that, carbon source utilization and nitrification- denitrification activity of this strain in different C/N, initial nitrogen concentration were studied. In addition, the assimilation and denitrification activities of ammonia and nitrate were also researched under the condition of nitrate and ammonia coexisted in the solution. The results show that the strain can grow in sodium acetate, glucose, sodium succinate and sodium citrate solutions, and it can not survive in sodium oxalate, sucrose and soluble starch solutions. Initial concentration and C/N were important for nitrogen removal rate. This strain can completely remove nitrate/ammonia when nitrate/ammonia concentration was lower than 15 mg l-1/80 mg l-1. the C/N of 10 and of 12 were the optimum C/N ratio in the nitrate and ammonia removal process respectively. pH value rose up sharply in the denitrification process and it increased relatively slowly in the nitrification process, which shows that pH is one of the most important factor inhibiting the denitrification removal process. Nitrite concentration was much higher in denitrification process than in nitrification process. In addition, this strain gave priority to utilizing ammonia as nitrogen source when ammonia and nitrate coexisted in the solution.

The growth of the Rhodococcus sp. on diesel fuel under the effect of heavy metals and different concentrations of zinc

Co-contamination of diesel fuel and heavy metals can be challenging for microbial remediation due to the complex composition of the fuel and the inhibitory effect of heavy metals.There is an urgent need to study this interaction to improve the pollutant removal efficiency in the Polar Regions.The growth of an Antarctic bacterium,Rhodococcus sp.was studied by comparing the growth at the logarithmic phase under the effect of selected heavy metals(Pb,Cr,As,Cd,Cu,Zn,Ni,Hg and Co).The selected heavy metals inhibited the growth of the Rhodococcus sp.on diesel fuel in an order from highest to lowest of:Hg>Zn>Cd>Cu>Co>Ni>As>Pb>Cr.Growth on diesel fuel co-contaminated with Hg and Zn were 2.95%and 5.71%,respectively compared to the no-metal control.A further experiment with various Zn concentrations was conducted.The specific growth rate of Rhodococcus sp.co-contaminated with different concentrations of Zn showed a correlation coefficient(r)of 0.916,and was modelled with an exponential decay model.Additional investigation is needed to determine the effect of low concentration of Zn on hydrocarbon degradation.It is important to understand the relationships between microbes,hydrocarbons and heavy metals,especially in the Polar Regions because this interaction might be promising in treating hydrocarbon-polluted sites containing heavy metals.The data and results also provide baseline tools of bioremediation processes at low temperatures and the knowledge of the ecological roles of Rhodococcus sp.in Antarctica.

Screening and Identification of a Bacterial Strain Antagonistic to Vibrio parahaemolyticus

In the present study, bacterial strains were isolated from the shrimp （Penaeus orientalis） ponds in different regions of Hainan Province, and tested for their antagonistic activity against Vibrio species on 2216E plates, using V. parahaemolyticus, V. harveyi and V. anguillarum as the indicator bacteria. As a result, an antagonistic strain was screened out and numbered 20160522Z-10 （referred to as Z-10）. Then, the antibacterial activity and minimum inhibitory concentration （MIG） of Z-10 against the indicator bacteria, and its antibacterial effect against other six Vibrio species were determined. The results proved that Z-10 was antagonistic to all the three indicator bacteria, and its MIC against V. parahaemolyticus, V. harveyi and V. anguillarum was 2.5× 10^4, 2.5× 10^5 and 2.5× 10^5 CFU/ml. Additionally, Z-10 had a certain antibacterial effect against other Vibrio species. Finally, the strain was identified as a Pseudoalteromonas sp. by 16S rDNA sequence analysis. The bacterium as a new probiotic, is expected to be used in disease prevention and control in marine aquaculture.

链霉菌Streptomyces sp.P325的化学成分研究

对Streptomyces sp.P325的化学成分进行研究。采用正相硅胶柱色谱、ODS反相柱色谱、凝胶柱色谱及半制备高效液相色谱等分离技术对Streptomyces sp.P325发酵产物的乙酸乙酯萃取部分进行分离纯化。从中共分离得到6个化合物,通过波谱数据分析和文献比对鉴定其结构,其中包括2个新化合物:四羟基十八碳二酸I(1)和四羟基十六碳二酸I(2),以及4个已知化合物:iso-frenolicin B(3)、(R)-7-acetyl-3,6-dihydroxy-8-propyl-3,4-dihydronaphthalen-1(2H)-one(4)、对羟基苯甲酸甲酯(5)和isonicotinaldehyde(6)。对分离得到的化合物1~6进行细胞毒活性评价。结果显示,化合物3对HepG2细胞系表现出显著的抑制活性(IC50为13.11μmol/L),对HeLa细胞的抑制活性为中等(IC50为25.04μmol/L)。

一株3,5,6-三氯-2-吡啶酚降解菌Delftia sp.DFT的分离鉴定及降解途径研究

微生物降解是消除农药残留物3,5,6-三氯-2-吡啶酚(TCP)的有效方法,但其降解机制并不清楚。为探究TCP完整的代谢途径和降解机制。采用富集分离法从农田土壤中分离了一株TCP降解细菌,并分别通过高效液相色谱和液相色谱-质谱(LC-MS)技术检测了菌株的降解性能,并鉴定了其代谢产物。该菌株为Delftia sp.DFT菌,它能在48 h内对50 mg/L的TCP降解率达34.28%,同时也能降解3,5-二氯-2-吡啶醇、6-氯-2-吡啶醇和2-羟基吡啶等其他氯代吡啶类化合物。在降解TCP过程中,共鉴定了12种不同的TCP代谢产物。初步推测水解-氧化脱氯途径、还原脱氯途径、反硝化及烷基化途径可能共同参与了菌株DFT对TCP的降解。据文献调研,这是首次在Delftia sp.菌中发现有四种可能的TCP代谢途径的报道,这为纯培养物中的TCP降解机制研究提供了参考信息。

喹啉降解菌Rhodococcus sp.QL2的分离鉴定及降解特性

从某焦化厂生物处理系统的活性污泥中驯化、分离出1株能以喹啉为唯一碳、氮、能源生长代谢的菌株QL2.经过对其形态特征、生理生化特征和16S rRNA序列分析鉴定该菌株为红球菌属（Rhodococcus sp.）.研究表明,菌株QL2利用喹啉生长的适宜温度为35～42℃,培养基初始pH为8～9,摇床转速为150 r/min.外加氮源能促进菌株的生长,其中无机氮比有机氮、铵态氮比硝态氮更利于细菌的生长.在喹啉初始浓度为60～680 mg/L范围内菌株QL2降解喹啉符合零级动力学方程.喹啉初始浓度为150 mg/L时在8 h内完全降解,TOC去除率14 h内可达到70%.降解过程中产生有颜色的物质,且杂环上的氮原子以氨氮的形式被释放.通过HPLC及GC/MS分析出喹啉降解过程中的主要中间产物为2-羟基喹啉.该菌底物利用范围广,能降解苯酚、萘、吡啶等多种芳香族化合物.

Rhodococcus sp. Ns对硝基苯酚的好氧生物降解

通过驯化富集培养,从红树林底泥中分离出6株硝基苯酚降解菌,其中Rhodococcussp.Ns为对硝基苯酚(PNP)与邻硝基苯酚(ONP)的高效降解菌.在好氧条件下该菌可以耐受小于1.8 mmol/L的PNP,能够利用PNP和ONP为唯一碳源、能源和氮源生长并将其完全矿化.研究了Rhodococcussp.Ns在不同pH、盐度与浓度范围下,PNP的降解特性并探讨了该菌降解PNP的途径.实验得出该菌在盐度<5‰、pH>5的条件下能较快生长,1.5 mmol/L的PNP在96h内被完全降解,并检测到至少2种中间产物4-硝基儿茶酚(4-nitrocatechol)和1,2,4-苯三酚(1,2,4-benzenetriol).红树林底泥中固有的细菌对PNP和ONP具有高效降解作用.

石油烃降解菌Rhodococcus sp．15-3的分离鉴定及特性研究

从原油污染土壤中分离筛选到一株石油烃降解菌15-3,根据其形态特征、生理生化特性及16S rDNA序列同源性分析,将其初步鉴定为红球菌(Rhodococcus sp.)。采用室内培养的方法,研究了15-3菌株对原油、原油中的烃类及正十八烷的降解作用。结果表明,15-3菌株能以正十八烷为惟一碳源生长,在48h内对500mg·L-1的正十八烷降解率达96.9%。15-3菌株降解正十八烷的最适温度、pH值和盐浓度(NaCl)分别为30℃、7.0、2%,在低温(10℃)及高盐(4%～5%NaCl)环境下也有良好的降解能力。15-3菌株可以降解原油中C_13～C_32的正构烷烃、芳香烃及姥鲛烷。在含5g·L-1原油的培养基中,30℃培养5d后,菌株15-3对原油的降解率为60.3%,对原油中C_13～C_31烷烃的降解率均大于90%,对原油中芳香烃的降解率为63.6%。该菌株以石蜡为惟一碳源生长时能产生表面活性剂,将发酵液表面张力由58.11mN·m-1降到36.6mN·m-1,表明菌株15-3具有较强的乳化和分散石油的能力。

Rhodococcus sp.T3-1菌株降解乙草胺的特性

以乙草胺为唯一碳源，通过摇瓶培养研究了Rhodococcus sp.T3—1对乙草胺的降解特性．结果表明，菌株T3—1降解乙草胺的最适温度为37℃，且其在pH值6～10的范围内对100mg／L乙草胺的降解率均在96％曲7％之间．该菌株在接种量为5％条件下，14h内可将200mg／L的乙草胺降解95.5％：乙草胺的降解速率与乙草胺初始浓度呈负相关，与菌株T3—1的初始接种量呈正相关．菌株T3-1还可以降解丁草胺，但不能降解丙草胺、异丙草胺和毗草胺．

Rhodococcus sp-1的Mn^2＋生物去除能力及诱导特性

利用传统的细菌分离方法,从地下水生物除锰滤池中分离、纯化出1株细菌,经LBB(leucoberbelin blue)法定性检测其具有Mn2+氧化能力,利用Sherlock细菌鉴定系统初步确定其属于红球菌属(Rhodococcus sp).对红球菌Rhodococcus sp-1菌株的生物除锰能力和诱导特性研究结果表明,Rhodococcus sp-1菌株在达到第1个生长稳定期,细菌浓度2.3×108个/mL时,其Mn2+的去除量可达到35 mg/L,具有很强的Mn2+生物去除能力;同时Fe的存在对Rhodococcus sp-1 Mn2+氧化活性的表达具有诱导作用,在相同培养条件下,Fe3+的诱导性强于Fe2+,其诱导速度是Fe2+的3~4倍;有机Fe3+的诱导性强于无机Fe3+,其诱导速度是无机Fe3+的1.6~2倍.

喹啉降解菌Rhodococcus sp.的降解特性与生物强化作用

以喹啉为唯一碳氮源从某焦化废水处理厂活性污泥中筛选出一株喹啉降解菌菌株红球菌(Rhodococcus sp.)KDQ2,其在24h内能将400mg/L喹啉降解96%,降解的最适条件为37℃和pH6~9,降解动力学符合Haldane方程.KDQ2能利用吡啶,但不能利用苯酚;在喹啉、吡啶和苯酚共存条件下,150mg/L吡啶和400mg/L苯酚不影响150mg/L喹啉在1d时的降解效率.KDQ2能适应含有高浓度苯酚、吡啶和喹啉等污染物的焦化废水环境条件,可以与活性污泥中的微生物共存,提高实际焦化废水中喹啉和TOC的去除能力.

滇池高效聚磷菌Rhodococcus sp.H的分离鉴定及除磷特性研究

采用混合稀释平板法,从富营养化湖泊底泥滇池中分离出1株高效聚磷菌,经分子生物学鉴定,推测其为红球菌属(Rhodococcus sp.),将其命名为Rhodococcus sp.H,并对菌株H在不同理化因素条件下的生长及除磷特性进行了研究。结果表明,该菌株在接种培养24 h后即能完成对数增长期,在pH值为6~7、温度为30℃时能获得较高生物量和除磷率,最佳碳源为乙酸钠和乙醇,最佳氮源为牛肉膏和蛋白胨。当温度高于35℃或低于15℃、p H值高于9或低于5时,菌株的生长会受到明显抑制,除磷率较低。同时研究表明,该菌株的生长会影响其生长环境基质的p H值,在一定范围内具有较强的p H值调节能力。该菌株能够有效利用乙酸钠和乙醇等小分子碳源获得较大生物量和较高的除磷效率,但是不能将铵盐作为唯一的氮源来利用。