Wine phenolic compounds influence the production of volatile phenols by wine-related lactic acid bacteria

Aims: To evaluate the effect of wine phenolic compounds on the production of volatile phenols (4‐vinylphenol [4VP] and 4‐ethylphenol [4EP]) from the metabolism of p‐coumaric acid by lactic acid bacteria (LAB). Methods and Results: Lactobacillus plantarum, Lactobacillus collinoides and Pediococcus pentosaceus were grown in MRS medium supplemented with p‐coumaric acid, in the presence of different phenolic compounds: nonflavonoids (hydroxycinnamic and benzoic acids) and flavonoids (flavonols and flavanols). The inducibility of the enzymes involved in the p‐coumaric acid metabolism was studied in resting cells. The hydroxycinnamic acids tested stimulated the capacity of LAB to synthesize volatile phenols. Growth in the presence of hydroxycinnamic acids, especially caffeic acid, induced the production of 4VP by resting cells. The hydroxybenzoic acids did not significantly affect the behaviour of the studied strains. Some of the flavonoids showed an effect on the production of volatile phenols, although strongly dependent on the bacterial species. Relatively high concentrations (1 g l^?1) of tannins inhibited the synthesis of 4VP by Lact. plantarum. Conclusions: Hydroxycinnamic acids were the main compounds stimulating the production of volatile phenols by LAB. The results suggest that caffeic and ferulic acids induce the synthesis of the cinnamate decarboxylase involved in the metabolism of p‐coumaric acid. On the other hand, tannins exert an inhibitory effect. Significance and Impact of the Study: This study highlights the capacity of LAB to produce volatile phenols and that this activity is markedly influenced by the phenolic composition of the medium.     

紫茎泽兰根区土壤酚酸类物质组成及其对土传病菌的影响

紫茎泽兰根系释放的化感物质是导致其土壤环境发生变化的重要因素,可为其成功入侵提供有利条件.以空白土样为对照,提取健康和感染棉蚜的紫茎泽兰根区土壤酚酸类化感物质,采用GC-MS分析其组分和相对含量.结果表明：不同处理根区土壤酚酸类化感物质的种类没有变化,但相对含量发生了改变.其中,虫害处理土样中苯甲酸和对羟基苯甲酸的相对含量均显著高于对照,但对羟基肉桂酸的相对含量显著低于对照,且苯甲酸、对羟基苯甲酸和对羟基肉桂酸的相对含量在3种处理土样中的比例存在差异.用上述3种物质单体及它们在3个处理土样的比例复配进行抑菌试验发现：在较低浓度（50～150 mg·L^-1）下,3种化感物质对5种土传性病菌有显著的抑制作用,但复配后对5种病菌的抑制作用各不相同,虫害处理的复配对其中3种病原菌的抑制作用均低于对照和健康植株.     

除草剂氟磺胺草醚降解菌FB8的分离鉴定与土壤修复

【目的】从氟磺胺草醚污染土壤分离高效降解菌株,进行分类学鉴定、降解特性及土壤修复能力初步研究,为氟磺胺草醚污染土壤微生物修复提供新的菌株。【方法】通过形态特征、生理生化特征和16S rRNA序列分析方法进行菌株鉴定;通过农药初始浓度、pH值、温度等环境因素的研究得到菌株的最适生长条件;通过敏感作物和靶标杂草的盆栽生测试验,验证菌株对氟磺胺草醚污染土壤的修复能力。【结果】本试验从黑龙江省长期施用氟磺胺草醚的大豆田地中分离出一株能以氟磺胺草醚为唯一碳源生长的细菌FB8,初步鉴定为假单胞菌属(Pseudomonas),该菌株在96 h内对500 mg/L氟磺胺草醚的降解率高达86.75%,其最适生长条件为500 mg/L农药初始浓度、初始pH6.0-8.0、35-37℃,该菌株处理30 d能够显著恢复敏感作物玉米和高粱的各项生物量指标,对氟磺胺草醚浓度为5 mg/kg的土壤修复效果明显。【结论】从黑龙江省污染土壤中筛选得到的高效降解氟磺胺草醚的门多萨假单胞菌Pseudomonas mendocina FB8,盆栽生测试验表明该菌株具有很好的土壤修复作用,可为氟磺胺草醚的生物修复研究提供适宜的菌种资源。     

Isolation and characterization of phorate degrading soil bacteria of environmental and agronomic significance

Phorate [O,O-diethyl-S-(ethylthio)methyl phosphoradiothioate] degrading bacteria were isolated from agricultural soil and characterized based on their morphological and biochemical characteristics. The selected isolates PS-1, PS-2 and PS-3 were presumptively identified as Rhizobium, Pseudomonas and Proteous species, respectively. The HPLC analysis of phorate in bioaugmented soil revealed its complete disappearance within 40 days. The degradation isotherms of the isolates PS-1, PS-2 and PS-3 suggested time-dependent disappearance of phorate following the first order rate kinetics at the corresponding rate constants of 0.04, 0.05 and 0.04 days-1. Besides, the isolates concurrently exhibited substantial phosphate solubilization, indole acetic acid, and siderophore production. The isolate PS-3 also showed anti-fungal activity against a phytopathogen Fusarium oxysporum. As a result of the multifarious biological properties, the isolates have been suggested to be important bioresource for efficient bioinoculant development.     

Genome‐wide identification of tolerance mechanisms toward p‐coumaric acid in Pseudomonas putida

The soil bacterium Pseudomonas putida KT2440 has gained increasing biotechnological interest due to its ability to tolerate different types of stress. Here, the tolerance of P. putida KT2440 toward eleven toxic chemical compounds was investigated. P. putida was found to be significantly more tolerant toward three of the eleven compounds when compared to Escherichia coli. Increased tolerance was for example found toward p‐coumaric acid, an interesting precursor for polymerization with a significant industrial relevance. The tolerance mechanism was therefore investigated using the genome‐wide approach, Tn‐seq. Libraries containing a large number of miniTn5‐Km transposon insertion mutants were grown in the presence and absence of p‐coumaric acid, and the enrichment or depletion of mutants was quantified by high‐throughput sequencing. Several genes, including the ABC transporter Ttg2ABC and the cytochrome c maturation system (ccm), were identified to play an important role in the tolerance toward p‐coumaric acid of this bacterium. Most of the identified genes were involved in membrane stability, suggesting that tolerance toward p‐coumaric acid is related to transport and membrane integrity.

     

p‐Coumaric acid decarboxylase from Lactobacillus plantarum: Structural insights into the active site and decarboxylation catalytic mechanism

p‐Coumaric acid decarboxylases (PDCs) catalyze the nonoxidative decarboxylation of hydroxycinnamic acids to generate the corresponding vinyl derivatives. Despite the biotechnological relevance of PDCs in food industry, their catalytic mechanism remains largely unknown. Here, we report insights into the structural basis of catalysis for the homodimeric PDC from Lactobacillus plantarum (LpPDC). The global fold of LpPDC is based on a flattened β‐barrel surrounding an internal cavity. Crystallographic and functional analyses of single‐point mutants of residues located within this cavity have permitted identifying a potential substrate‐binding pocket and also to provide structural evidences for rearrangements of surface loops so that they can modulate the accessibility to the active site. Finally, combination of the structural and functional data with in silico results enables us to propose a two‐step catalytic mechanism for decarboxylation of p‐coumaric acid by PDCs where Glu71 is involved in proton transfer, and Tyr18 and Tyr20 are involved in the proper substrate orientation and in the release of the CO₂ product. Proteins 2010. © 2009 Wiley‐Liss, Inc.     

Role of phenolic acids and enzymes of phenylpropanoid pathway in resistance of chayote fruit (Sechium edule) against infestation by melon fly,Bactrocera cucurbitae

Melon fly is a serious pest of cucurbits all over the world causing huge losses to yield. However, the only exception is the chayote fruit (Sechium edule) that shows resistance to melon fly infestation. Studies on culture of melon fly indicated the absence of plant traits resisting oviposition on chayote fruit. However, the melon fly was unable to complete its life cycle successfully on chayote showing that factors inhibiting larval development in melon fly could be attributed to biochemical constituents. Studies were, therefore, carried out to compare the biochemical responses of chayote, a melon fly resistant species and bitter gourd, a susceptible species to melon fly infestation with regard to the levels of phenolic acids and activities of the enzymes of phenylpropanoid pathway (PPP) leading to synthesis of lignin. The resistant chayote exhibited significantly higher accumulation of lignin associated with higher activities of phenylalanine ammonia‐lyase (PAL), tyrosine ammonia‐lyase (TAL), cinnamyl alcohol dehydrogenase (CAD) and peroxidase (POD). On the contrary, the susceptible bitter gourd recorded lower activities of PAL, CAD and POD and a decreasing trend of TAL during infestation associated with a lower lignin content. The monomer composition of lignin in the resistant chayote showed twofold higher level of guaiacyl (G) and syringyl (S) units compared to susceptible bitter gourd and the G/S ratio during infestation increased in chayote while decreasing in bitter gourd. The levels of PPP intermediates, p‐coumaric acid was higher in chayote while p‐hydroxy benzoic acid, a chemo‐attractant, was higher in bitter gourd. Incorporation of p‐coumaric acid in the larval diet strongly inhibited larval growth even as p‐hydroxy benzoic acid promoted growth confirming the direct role of p‐coumaric acid in conferring resistance to chayote. The level of salicylic acid, a signal molecule involved in induction of defence response, was higher in chayote compared to bitter gourd. Chayote also exhibited higher level of activity of POD in the phloem exudates compared to bitter gourd. The higher concentration of sugars in exudates of chayote might act like signalling molecules causing activation of plant genes, especially of the phenylpropanoid biosynthesis pathway and possibly produce an osmotic effect inducing resistance against the melon fly. Thus, the study revealed that the resistance in chayote to melon fly infestation is a complex, multi‐layered process in which the activities of PPP enzymes generating phenolic intermediates leading to lignin biosynthesis and the composition of exudates appear to play significant roles. Besides, the study also indicated that different forms of lignin might play a role in the resistance of chayote against melon fly infestation.     

Isolation and characterization of 2,4-dichlorophenoxyacetic acid-catabolizing bacteria and their biodegradation efficiency in soil

Bacterial isolates (NJ 10 and NJ 15) capable of degrading the herbicide 2,4-dichlorophenoxyacetic acid (2,4-D) were isolated from agricultural soil by enrichment culture technique. The isolates exhibited substantial growth in mineral salt medium supplemented with 0.1–0.5% of 2,4-D as a sole source of carbon and energy. Based on their morphological, cultural and biochemical characteristics, the isolates NJ 10 and NJ 15 have been identified as Pseudomonas species and Pseudomonas aeruginosa, respectively. Biodegradation studies in a soil microcosm enriched with pure cultures of the isolates demonstrated a time-dependent disappearance of 2,4-D from the 100 mg/kg herbicide-amended soil. The HPLC data analysis revealed 96.6 and 99.8% degradation in the soil inoculated with the pure cultures of isolates NJ 10 and NJ 15, respectively with in 20 days of incubation at 30 °C. Both the isolates showed significant solubilization of inorganic phosphate [Ca₃(PO₄)₂] on the specific Pikovskaya's medium.     

氯氰菊酯降解菌株L12的分离鉴定及降解特性

[目的]通过分离纯化氯氰菊酯降解菌,研究降解特性为实际应用提供理论依据.[方法]利用富集驯化培养技术分离菌株,通过形态、生理生化特征及16S rRNA基因序列分析鉴定菌株.测定培养液中氯氰菊酯的浓度和代谢产物以及菌体细胞的密度和细胞表面疏水性,研究菌株的降解特性.[结果]从生产氯氰菊酯的农药厂污水曝气池中,分离到1株能...     

一株氯霉素降解细菌的分离鉴定与代谢特性研究

微生物是介导环境中氯霉素降解转化的主要驱动者,但高效降解矿化菌株资源匮乏,氧化反应介导的代谢途径不清。为研究微生物介导下氯霉素的环境归趋过程,为氯霉素污染环境强化修复提供菌株资源,文中以受氯霉素污染的活性污泥为接种源,首先富集获得一个由红球菌Rhodococcus主导 (相对丰度>70%) 的氯霉素高效降解菌群,并从中分离获得一株能够高效降解氯霉素的菌株CAP-2,通过16S rRNA基因分析鉴定为红球菌Rhodococcus sp.。菌株CAP-2能在不同营养条件下高效降解氯霉素。基于菌株CAP-2对检测到的代谢产物对硝基苯甲酸和已报道的代谢产物对硝基苯甲醛和原儿茶酸的生物转化特征,提出其降解途径是由氯霉素侧链氧化断裂生成对硝基苯甲醛,进一步氧化为对硝基苯甲酸的新型氧化降解途径。该菌株对于氯霉素分解代谢的分子机制研究以及受氯霉素污染环境的原位生物修复应用具有巨大潜力。     

一株红壤溶磷菌的分离、鉴定及溶磷特性

【目的】为了提高红壤磷素利用率,探讨溶磷菌溶磷机理。【方法】利用难溶性无机盐培养基从花生根际土壤样品中分离到一株溶磷菌C5-A,结合菌落形态特征、生理生化和16S rRNA序列确定该菌株的系统发育地位;通过菌株C5-A在NBRIP液体培养基培养过程中培养液pH变化确定其溶磷能力;利用液体发酵实验测定不同的碳源、氮源对菌株C5-A溶磷的影响;通过高效液相色谱检测C5-A在不同氮源培养液中有机酸的种类和浓度。【结果】菌株C5-A鉴定为洋葱伯克霍尔德氏菌(Burkholderia cepacia),遗传稳定性较好。在FePO4和AlPO4培养液中,菌株C5-A的溶磷量和pH变化呈显著负相关;菌株C5-A对磷酸三钙、磷酸铝、磷酸铁、磷矿粉均有较强的溶解能力,最高溶磷量分别为125.79、227.34、60.02和321.15 mg/L;菌株C5-A对不同浓度的两种磷矿粉有较强的溶解能力;分别以麦芽糖和草酸铵为碳源和氮源时溶磷量最高。高效液相色谱检测出10种有机酸,分别为草酸(葡萄糖酸)、乙酸、苹果酸、琥珀酸和5种未知有机酸,然而,乙酸而非草酸似乎是影响C5-A溶磷的重要有机酸。【结论】从红壤花生根际土壤中筛选到一株对难溶性无机盐具有较强溶解能力溶的菌株C5-A,有望为开发高效红壤微生物磷肥提供种质资源。     

Effect of soil type, salinity, and allelochemicals on germination and seedling growth of a medicinal plant Lepidium sativum L.

The present work was conducted to elucidate the effect of soil type, salinity stress and allelochemicals from alfalfa on the germination efficiency, seedling growth and photosynthetic pigments of Medicago sativa L. The highest germination rate (69.9%) was recorded for seeds cultivated in sandy soil followed by that of clay soil (42%) and then loamy soil (19%). The maximum germination rate was attained at 50 mM NaCl concentration level, after that the values were decreased as the salinity levels increased. The maximum lengths (cm) of plumule and radicle and their growth rates (cm/day) were obtained at control level and found to decline significantly as the salinity increased. Aqueous extracts of alfalfa reduced the germination rate, radicle and plumule growth and seedling dry weight of L. sativum. However, chlorophyll and carotenoid content was increased, their maxima were recorded at 50% concentration level. The chlorophyll a/b ratio attained its maximum at the full strength concentration level.     

产胞外多糖菌株的分离鉴定及其功能研究

微生物产生的胞外多糖(exopolysaccharides, EPS)可促进大粒径土壤团聚体形成,高产EPS的菌株在土壤改良、促进作物生长方面具有较好的应用前景。【目的】从土壤样品中筛选高产胞外多糖的细菌,研究其在土壤改良、环境适应性、广谱抗病等方面的功能,为制备土壤改良型功能菌剂提供候选菌株。【方法】采用蒽酮硫酸法测定菌株胞外多糖的产量,通过形态学观察、生理生化试验及16S rRNA基因序列测定确定其分类地位,结合土壤培养试验研究菌株对土壤团聚体形成的影响。【结果】获得3株胞外多糖产量大于500 mg/L的细菌,经鉴定A-5为地衣芽孢杆菌(Bacillus licheniformis),XJ-3为萎缩芽孢杆菌(Bacillus atrophaeus),KW3-10为耐盐芽孢杆菌(Bacillus halotolerans)。菌株A-5、XJ-3、KW3-10处理后,土壤大团聚体(>0.25 mm)含量较对照分别提高了4.07、2.14和3.16倍。3株菌株对疮痂链霉菌(Streptomyces scabies)、尖孢镰刀菌(Fusarium oxysporum)、茄链格孢菌(Alternaria solani)和立枯丝核菌(Rhizoctonia solani)等多种植物病原菌具有明显的抑制效果,可耐受pH为5-9和NaCl含量1%‒9%的盐碱环境,促进植物生长,其中KW3-10的代谢产物中IAA含量为25.58 mg/L。【结论】菌株A-5、XJ-3、KW3-10可显著促进土壤团粒结构形成,具有较好的广谱抗病性和促生长特性,可作为高效复合功能菌剂的候选菌株。     

环渤海湾地区连作苹果园土壤中酚酸类物质变化

分析了山东昌邑、栖霞、蓬莱,辽宁大连、抚宁、绥远,河北昌黎、青县等地苹果园连作土壤中酚酸物质的组成和含量,结果表明:连作障碍发生的苹果园土壤中酚酸类物质的组成和含量在不同地区、不同土层厚度间存在显著差异。苹果园土壤中酚酸类物质的含量从春季到秋季随时间的延长逐渐减少,但过程缓慢。且非连作园土壤中酚酸类物质含量显著少于连作土。连作土壤中酚酸物质的组成和含量在不同季节间差异显著,尤其春季土壤中酚酸物质的种类与夏季和秋季显著不同。这可能是因为植物在不同季节分泌的酚酸类物质种类和含量有差别.不同土层中酚酸物质的分布因季节不同而有显著差异。夏季土壤中酚酸物质主要分布在浅层土壤中,而秋季则主要分布在深层土壤中。这可能是由于浇水等果园管理措施和自然降水对土壤中的酚酸物质产生的淋溶作用,使得大量的酚酸物质向深层土壤运动,最终造成了秋季果园连作土中随土层加深而酚酸物质含量增加的现象。不同地区苹果园连作土中,对羟基苯甲酸、(+)-儿茶素、咖啡酸、阿魏酸含量与非连作土无显著差异,而焦性没食子酸、绿原酸和根皮苷显著高于非连作土。焦性没食子酸、绿原酸和根皮苷可能是引起山东、辽宁、河北地区苹果园连作障碍的关键酚酸物质。     

Structural and functional characterization of solute binding proteins for aromatic compounds derived from lignin: p‐Coumaric acid and related aromatic acids

Lignin comprises 15–25% of plant biomass and represents a major environmental carbon source for utilization by soil microorganisms. Access to this energy resource requires the action of fungal and bacterial enzymes to break down the lignin polymer into a complex assortment of aromatic compounds that can be transported into the cells. To improve our understanding of the utilization of lignin by microorganisms, we characterized the molecular properties of solute binding proteins of ATP‐binding cassette transporter proteins that interact with these compounds. A combination of functional screens and structural studies characterized the binding specificity of the solute binding proteins for aromatic compounds derived from lignin such as p‐coumarate, 3‐phenylpropionic acid and compounds with more complex ring substitutions. A ligand screen based on thermal stabilization identified several binding protein clusters that exhibit preferences based on the size or number of aromatic ring substituents. Multiple X‐ray crystal structures of protein–ligand complexes for these clusters identified the molecular basis of the binding specificity for the lignin‐derived aromatic compounds. The screens and structural data provide new functional assignments for these solute‐binding proteins which can be used to infer their transport specificity. This knowledge of the functional roles and molecular binding specificity of these proteins will support the identification of the specific enzymes and regulatory proteins of peripheral pathways that funnel these compounds to central metabolic pathways and will improve the predictive power of sequence‐based functional annotation methods for this family of proteins.Proteins 2013; 81:1709–1726. © 2013 Wiley Periodicals, Inc.     

粪臭素高效降解菌YKSW-6的分离、鉴定及降解特性

【背景】粪臭素是畜牧堆肥中有机污染物的主要成分,造成养殖场及周边环境恶化,粪臭素污染问题亟待解决,利用微生物降解粪臭素是一种环保节能的有效方法。【目的】分离鉴定粪臭素高效降解菌株,研究其降解特性,为粪臭素降解提供高效的菌种资源,为该菌株应用于臭味污染环境的净化提供基础。【方法】以粪臭素为唯一碳源的无机盐培养基作为培养基质,从猪粪堆肥样品中分离筛选粪臭素高效降解菌株,通过形态特征和16S rRNA基因序列分析进行分离菌株的初步鉴定,分析其生长规律及粪臭素降解特性,并利用气相色谱质谱联用(GC-MS)对菌株代谢粪臭素的产物进行分析。【结果】从样品中分离获得一株能以粪臭素为唯一碳源的细菌YKSW-6菌株,形态学和16S rRNA基因序列分析初步鉴定该菌株为戈登氏红球菌(Rhodococcus gordoniae)。接种量为10%时,该菌培养14 h对100 mg/L的粪臭素降解率达到100%。其能够利用D-山梨醇、溴-丁二酸等18种碳源,对亚碲酸钾、溴酸钾等13种化学敏感物具有抗性。菌株YKSW-6在5%接种量、温度30-42℃和pH值为6.0-9.0时对100 mg/L的粪臭素降解效率均能达到100%,菌株生长和降解粪臭素的最佳条件为：pH 7.2,温度37℃,转速180 r/min。GC-MS结果表明,粪臭素在菌株的作用下C2先被氧化,转变为3-甲基羟基吲哚,随后进一步被氧化为N-(2-乙酰基苯基)甲酰胺。同时中间产物还有苯乙醛和苯乙酸。【结论】红球菌YKSW-6为目前已报道的降解粪臭素能力较强的菌株,丰富了粪臭素降解菌种的资源库,为实际环境微生物修复应用提供了理论参考。     

The effect of quinoid and phenolic compounds on urease and dehydrogenase activity and nitrification in soil

Summary 1,4-Naphthoquinone; 2-methyl-1,4-napthoquinone; 2,3-dichlorohydroquinone; 4,6-di-tert.butyl-o-benzoquinone; 4,6-di-tert.butylpyrocatechol and 4-tert.butylpyrocatechol at concentrations of 10 and 20 ppm inhibit mineralization of urea N by reducing urease activity and/or nitrification in soils. Coating of urea with these chemicals was more effective than direct application to the soil. There was no adverse effect of any of the chemicals either on germination or on the growth of wheat plants at concentrations of 20 and 50 ppm added to soil.     

苦参腐解物对酚酸类物质胁迫下茄子生长和根际土壤微生物活性的影响

研究了盆栽条件下添加苦参腐解物对酚酸物质(香草醛和肉桂酸)胁迫下茄子生长及根际土壤微生物种群数量和土壤酶活性的影响。与对照相比,苦参腐解物处理促进了酚酸作用下茄子的株高和茎粗生长,增加了茄子土壤微生物的种群数量,尤其增加了细菌和放线菌数量。随着外源香草醛和肉桂酸浓度的增加,细菌和放线菌数量先增大后减小,而真菌数量先降低后增加;同时,茄子根际土壤过氧化氢酶、多酚氧化酶、脲酶的活性均高于对照,呈先增大后降低的变化趋势。