Biotransformation of benzonitrile herbicides via the nitrile hydratase–amidase pathway in rhodococci

The aim of this work was to determine the ability of rhodococci to transform 3,5-dichloro-4-hydroxybenzonitrile (chloroxynil), 3,5-dibromo-4-hydroxybenzonitrile (bromoxynil), 3,5-diiodo-4-hydroxybenzonitrile (ioxynil) and 2,6-dichlorobenzonitrile (dichlobenil); to identify the products and determine their acute toxicities. Rhodococcus erythropolis A4 and Rhodococcus rhodochrous PA-34 converted benzonitrile herbicides into amides, but only the former strain was able to hydrolyze 2,6-dichlorobenzamide into 2,6-dichlorobenzoic acid, and produced also more of the carboxylic acids from the other herbicides compared to strain PA-34. Transformation of nitriles into amides decreased acute toxicities for chloroxynil and dichlobenil, but increased them for bromoxynil and ioxynil. The amides inhibited root growth in Lactuca sativa less than the nitriles but more than the acids. The conversion of the nitrile group may be the first step in the mineralization of benzonitrile herbicides but cannot be itself considered to be a detoxification.     

Effect of four common herbicides on the population of microorganisms in Red Sandy soil

Summary A study was conducted to know the effect of four common herbicidesviz, Lasso, Nitrofen, Propinol, and Simazine on soil microorganisms in Red Sandy soil. In general the application of different herbicides did not adversely affect the microbial population. The bacterial population was decreased by the application of propinol and nitrofen. Simazine and lasso stimulated the fungal population. The stimulation in Azotobacter population was observed in lasso, nitrofen and simazine treated soil.     

Tolerance of the nitrogen-fixing system ofAzotobacter vinelandii for four commonly used pesticides

The effects of two insecticides, DDT and lindane, and two herbicides, Dalapon-Na and 2, 4, 5-T, upon the growth and nitrogen-fixing capacity ofAzotobacter vinelandii in pure culture were investigated using concentrations of the pesticides equivalent to the usual field application rates, and fifty times these levels. The acetylene reduction technique was used to assay nitrogen fixation. No significant effects of the pesticides at either concentration were detected upon growth or acetylene reduction. This study was supported by a grant from the Rural Credits Development Fund of the Reserve Bank of Australia.     

Interactions between Azotobacter chroococcum and vesicular-arbuscular mycorrhiza and their effects on plant growth

B rown , M. E. & G arr , G. R. 1984 Interactions between Azotobacter chroo-coccum and vesicular-arbuscular mycorrhiza and their effects on plant growth. Journal of Applied Bacteriology 56 , 429–437.
The effects of simultaneous inoculation of roots of lettuce seedlings with vesicular-arbuscular endophytes and Azotobacter chroococcum are described. Endophytes alone increased yields of lettuce grown in partially sterilized P-deficient soil, but not in the same unsterile soil. Dual inoculation with endophytes and Azotobacter in both soils produced larger plants than either inoculum alone. In another poorly structured unsterile soil with adequate P for plant growth, endophytes depressed yields of lettuce. There was no association between endophytes and Azotobacter . In none of the experiments did Azotobacter influence the level of endophyte infection in the roots; but numbers of Azotobacter on the root systems were decreased in the presence of the endophytes. Azotobacter alone always increased growth of young plants before the effects of the endophytes were observed.     

自生固氮菌的生态分布及其对农药抗性的研究

将不同地点的土样均匀地撒在无氮培养基平板上,分离并鉴定自生固氮菌。同时采用抑菌圈法和分光光度计测透光率的方法,检测6种常用的田间农药对自生固氮菌的生长影响;还运用茚三酮显色反应检测农药对自生固氮菌固氮效果的影响。结果表明,不同农药及其不同浓度对自生固氮菌生长及固氮量的影响有所不同。为添加固氮菌以提高农作物产量,监测固氮菌含量来评估作物产量,以及田间正确使用农药提供了理论依据。     

Pesticide tolerant Azotobacter isolates from paddy growing areas of northern Karnataka, India

A total of 14 Azotobacter strains were isolated from different paddy cultivating soils with pH ranging from 6.5 to 9.5 by using serial dilution agar plate method. The strains were Gram negative, rod shaped, cyst forming and developed brown to black colored colonies, which were glistening, smooth, slimy on Ashby’s agar plates. Biochemically they were positive for biochemical tests namely, indole production, citrate, catalase, carbohydrate fermentation and Voges–Proskauer test. Further, sequence analysis of PCR amplicons obtained from these cultures revealed the presence of five different Azotobacter species viz., Azotobacter vinelandii, Azotobacter salinestris, Azotobacter sp., Azotobacter nigricans subsp. nigricans and Azotobacter tropicalis. Phylogenetically these strains were grouped into two distinct clusters. These strains were tested for their ability to grow on a media containing four different pesticides such as pendimethalin, glyphosate, chloropyrifos and phorate, which are commonly used for the paddy. Out of 14 strains tested, 13 strains were able to grow on a media containing herbicides such as pendimethalin, glyphosate and insecticides like chloropyrifos and phorate. However, five Azotobacter strains were able to grow at higher concentration of 5 % pesticides, without affecting their growth rate. Further, the effect of pesticides on the indole acetic acid (IAA) production by Azotobacter strains was also estimated. Azotobacter-16 strain was found to produce 34.4 μg ml^?l of IAA in a media supplemented with 1,000 mg of tryptophan and 5 % of pendimethalin. Present study reveals that species of Azotobacter are able to grow and survive in the presence of pesticides and no significant effects were observed on the metabolic activities of Azotobacter species.     

Flow of microbially fixed nitrogen in a model ecosystem

Summary The preceding observations showed that both Azotobacter and a mixed flora with Azotobacter are about equally capable of fixing atmospheric nitrogen. During subsequent transfer of the nitrogen there was a distinct differentiation between movement of the newly fixed nitrogen and the nitrogen that was already present in tissues introduced in the beginning of the experiment. Pathways and transfer coefficients of both N¹⁴ and N¹⁵ were different in the presence of the different microbial populations. Turnover was more complex and in general slower in the presence of a mixed population than with pure Azotobacter. The lower transfer coefficients in the mixed populations may reflect more recycling within these populations than in pure cultures of Azotobacter. Denitrification losses upon acidification were negligible for N¹⁴ but appreciable for N¹⁵. The effect of acidification tended to be greater in systems with pure cultures of Azotobacter than in systems with a mixed culture.Research sponsored by the U.S. Atomic Energy Commission under contract with the Union Carbide Corporation.Research sponsored by the U.S. Atomic Energy Commission under contract with the Union Carbide Corporation.     

Effect of several herbicides on bacterial populations and activity and the persistence of these herbicides in soil

Summary Samples of a sandy loam soil were supplied with normal, 10-fold and 100-fold rates of ioxynil, dalapon, mecoprop, dichlorprop, MCPA + dichlorprop, picloram, and amitrole-T and incubated at 29°C at 65 per cent of the waterholding capacity. Treated soil samples were compared with untreated samples. Samples supplied with (NH₄)₂SO₄ and herbicides were used to investigate the effect of the herbicides on the rate of nitrification and the production of nitrite. In several cases higher numbers of bacteria were found for a longer or shorter period in soil treated with herbicides. There was some evidence that certain groups of bacteria had adapted to ioxynil in a soil sample supplied with the 100-fold rate of this chemical. After 2 or 4 weeks lower numbers ofAzotobacter chroococcum were found at the normal rate of ioxynil, dalapon, mecoprop, and dichlorprop. At the 100-fold rate of application the numbers of Azotobacter were unfavourably affected by all herbicides. The production of mineral nitrogen was hardly affected by the normal and 10-fold rates of application. In the first week the rate of nitrification was slightly depressed in soil samples treated with the normal rates of dichlorprop and amitrole-T and with the 10-fold rates of dalapon, mecoprop, and MCPA + dichlorprop. Strong inhibition of the nitrification for at least 7 weeks was shown by the 10-fold rate of amitrole-T. At the 100-fold rate all herbicides, with the exception of picloram, depressed the rate of nitrification for a longer or shorter period. During the second week a very small increase of nitrite was found in the samples treated with the normal and 10-fold rates of dalapon, mecoprop, dichlorprop, and amitrole-T. A small increase of nitrite was noted for 26 weeks in samples treated with the 100-fold rates of amitrole-T. A highly significant depression of CO₂ evolution was found in the first week in samples treated with the normal rates of ioxynil, dalapon, mecroprop, dichlorprop, and amitrole-T, also in samples treated with the 10-fold rates of dalapon, mecoprop, dichlorprop, MCPA + dichlorprop, picloram, and amitrole-T. A highly significant depression of CO₂ production was found after 8 weeks in all the samples treated with herbicides at the 100-fold rate with the exception of the sample treated with picloram. The decomposition of the herbicides was studied in soil samples treated with the 100-fold rates of herbicides. Only traces of dalapon and mecoprop were found after 9 months, but 7.2% ioxynil, 29.8% dichlorprop, 39% (MCPA + dichlorprop), 52.1% picloram and 52.2% amitrole-T were still present in active form.     

Effect of non-ionic tensides on the growth of some soil bacteria

Summary The effect of some nonylphenyl-ethylene oxide polymers on the growth of Bacillus megaterium, B. cereus var. mycoides, B. polymyxa, B. subtilis, Pseudomonas fluorescence and Azotobacter chroococcum was investigated in the concentration range 20–800 ppm with the agar diffusion method. The zones of inhibition, restricted growth and eventual stimulation were determined with a Shimadzu C-930 dual wavelength TLC scanner. The data matrix was evaluated by principal component analysis. A. chroococcum was insensitive to each tenside at each concentration. The growth of the other microorganisms was inhibited by the tensides. With B. megaterium and B. cereus var. mycoides stimulation was also observed. The effect of the non-ionic tensides decreased with increasing length of the hydrophilic ethylene oxide chain. This phenomenon can be explained by the assumption that the activity of tensides depends on their membrane-damaging effect. The bulky nonylphenyl group inserts between the apolar fatty acid chains disorganizing the membrane structure. The longer hydrophilic ethylene oxide chain modifies the distribution of tenside between the apolar and polar regions of the membrane, preferring the aqueous phase. This results in the decrease or loss of biological activity. Offprint requests to: T. Cserháti     

Vegetative growth and sporulation ofBipolaris sorokiniana on sequentially older infected leaves ofPoa pratensis exposed to postemergence herbicides

Poa pratensis was exposed to the postemergence herbicides 2,4-D, MCPP, and dicamba at concentrations of 10^–6 M applied to the soil surface. Sequentially older leaf blades were inoculated withBipolaris sorokiniana and detached to determine the effect of the herbicides on saprophytic hyphal growth and sporulation on the tissue of each older leaf. 2,4-D and dicamba stimulated hyphal growth in vitro from leaf tissue of all ages; MCPP had no effect on hyphal growth. All herbicides increased sporulation byB. sorokiniana on leaf tissue of all ages. Both hyphal growth and sporulation increased progressively on tissue from sequentially older leaves ofP. pratensis exposed to dicamba.Journal Paper No. J-14985 of the Iowa Agriculture and Home Economics Experiment Station, Ames, Iowa, USA, Project No. 2616.     

13种茎叶处理除草剂对不同生育期白花鬼针草的生物活性

[目的] 白花鬼针草是一种恶性外来入侵杂草,近年来已侵入农田,对农业生产及生态系统带来严重危害。为筛选防治白花鬼针草的有效除草剂,分析评价了13种常见茎叶处理除草剂对幼苗期和成株期白花鬼针草的防治效果。[方法] 采用整株盆栽法,在白花鬼针草幼苗期（2~3对叶期）和成株期（6~7对叶期）分别进行茎叶喷雾处理,每种除草剂设置3个剂量。[结果] 供试的13种除草剂中,灭生性除草剂草甘膦、草铵膦和敌草快对幼苗期和成株期的白花鬼针草防效达到100%。选择性除草剂中,麦草畏和辛酰溴苯腈对幼苗期和成株期的白花鬼针草均有较好的防效,三氯吡氧乙酸、乙羧氟草醚和氯吡嘧磺隆在高剂量下对幼苗期的白花鬼针草有较好的防除效果,但对成株期的白花鬼针草防效较差,氯氟吡氧乙酸、乳氟禾草灵、灭草松、二氯吡啶酸、乙氧氟草醚对幼苗期和成株期白花鬼针草防效均较差。[结论] 白花鬼针草对多种化学除草剂具有较强的耐药性,生育期对除草剂防除白花鬼针草的效果有较大影响。灭生性除草剂草甘膦、草铵膦、敌草快及选择性除草剂辛酰溴苯腈和麦草畏适用于防除白花鬼针草。     

Cyanuric acid--a s-triazine derivative as a nitrogen source for some soil microorganisms

Cyanuric acid was not toxic for soil microorganisms examined and was even observed to stimulate the growth of Azotobacter in chernozem. Some isolated fungi were capable of cleaving the ring of cyanuric acid. With the use of 15N-labeled cyanuric acid it was found that the nitrogen taken from this compound by Aspergillus minutus and Pseudogymnoascus sp. was incorporated into their proteins. About 70-90% of 15N derived from cyanuric acid was detected in the biomass of the examined fungi. The ability of soil microorganisms to cleave the triazine ring is of importance in the detoxication of soils treated with triazine herbicides.     

三种除草剂对外来入侵植物大狼耙草的防除效果

大狼耙草是危害生物多样性的一种恶性杂草。使用使它隆、二甲四氯、草甘膦3种除草剂对大狼耙草进行不同浓度、不同防治时期防除效果研究,结果表明:使它隆对大狼耙草平均致死率为92%,二甲四氯为85%,草甘膦为82%,但三者差异不显著;高、中、低3种浓度(使它隆分别为0.47、0.40、0.33g·L-1,二甲四氯分别为2.80、2.33、1.87g·L-1,草甘膦分别为5.18、3.89、2.59g·L-1)对大狼耙草致死率差异极显著,高浓度溶液防除效果较好;大狼耙草植株在幼苗期(植株高20cm)、生长盛期(植株高40cm)、花期(植株高60cm)3个防治时期致死率差异极显著,幼苗期防除效果理想。研究表明,在大狼耙草幼苗期使用0.47g·L-1浓度的使它隆防除效果最好。     

Potentiated susceptibility of ascites tumor to acyl derivatives of ascorbate caused by balanced hydrophobicity in the molecule

Orders of susceptibility of Ehrlich ascites tumor to L-ascorbic acid (Asc), its 6-stearoyl (6S), 6-palmitoyl (6P) and 2,6-dipalmitoyl (DP) derivatives were assessed in vitro and in vivo: 6P (a 50% growth inhibitory concentration (IC50) for cultured cells, 12 microM; an increased life-span of treated mice, 283%) greater than 6S (61 microM; 240%) much greater than Asc (430 microM; 122%) greater than or equal to DP (greater than 200 microM; 89%), indicating that the enhanced susceptibility was due to acyl moiety substituted at C6-hydroxyl group of Asc, but was retracted by further substitution at C2-hydroxyl group. Equimolar mixture of Asc and palmitic acid, stearic acid or their methyl esters was much less cytotoxic than 6P or 6S. Thus the enhanced susceptibility was not primarily due to an additive cytotoxic effect of ascorbyl and acyl moieties, but to a balanced hydrophobicity introduced into the molecule by a poorly cytotoxic acyl moiety.     

除草剂对黄河三角洲入侵植物互花米草的影响

互花米草(Spartina alterniflora)作为我国危害最严重的外来入侵植物之一,严重威胁滨海湿地生态系统安全。筛选可高效灭除互花米草的除草剂,为互花米草防治提供技术支持。2017年7月在黄河三角洲潮间带对互花米草茎叶喷施不同除草剂,在施药当年和次年对互花米草生长状况和大型底栖动物密度进行跟踪调查。研究结果表明:1)高效氟吡甲禾灵对互花米草的灭除效果最好,既能杀死互花米草地上部分从而完全抑制有性繁殖,又可完全抑制次年的无性繁殖;2)氰氟草酯对互花米草的灭除效果也比较好,可以完全抑制互花米草的生长和结穗,但抑制根状茎无性繁殖能力的效果稍差,草甘膦可以完全抑制互花米草的有性繁殖,但无法抑制次年互花米草的无性繁殖;3)施用除草剂在短期内会毒害某些底栖动物,但在1年后底栖动物种群数量与对照处理无显著差异。除草剂对环境的影响程度与除草剂用量和施用时间息息相关,未来研究中,应在保证灭草效果的前提下,探索最佳用药时间和最低用量以最大限度地降低环境影响。     

Micro-organisms in the rhizosphere of plants inoculated withAzotobacter chroococcum

Summary The hypothesis that inoculation withAzotobacter chroococcum affects the growth of plants indirectly through changing the rhizosphere microflora was investigated. Inoculated and uninoculated wheat and tomato plants were grown in the glasshouse in two different soils, and total bacteria, chitinolytic bacteria, actinomycetes, glucosefermenting bacteria, aerobic cellulose-decomposing bacteria, and anaerobes were determined in intervals in the rhizosphere and in the soil. Root-surface fungi were studied using the Harley and Waid's root-washing technique¹⁰. Azotobacter became established in the rhizosphere of wheat and tomato plants and stimulated their growth. All the bacterial groups examined were more abundant in the rhizosphere than in the soil. Inoculation with Azotobacter delayed the colonization of roots by bacteria, actinomycetes, and fungi in the rhizosphere, but had no effect on other organisms. Inoculation did not affect the dominant root-surface fungi, and minor changes were not consistent.Part of a thesis accepted by the University of London for the degree of Ph.D. in Microbiology.     

Hydrolysis of benzonitrile herbicides by soil actinobacteria and metabolite toxicity

The soil actinobacteria Rhodococcus rhodochrous PA-34, Rhodococcus sp. NDB 1165 and Nocardia globerula NHB-2 grown in the presence of isobutyronitrile exhibited nitrilase activities towards benzonitrile (approx. 1.1–1.9 U mg^?1 dry cell weight). The resting cell suspensions eliminated benzonitrile and the benzonitrile analogues chloroxynil (3,5-dichloro-4-hydroxybenzonitrile), bromoxynil (3,5-dibromo-4-hydroxybenzonitrile) and ioxynil (3,5-diiodo-4-hydroxybenzonitrile) (0.5 mM each) from reaction mixtures at 30°C and pH 8.0. The products were isolated and identified as the corresponding substituted benzoic acids. The reaction rates decreased in the order benzonitrile ? chloroxynil > bromoxynil > ioxynil in all strains. Depending on the strain, 92–100, 70–90 and 30–51% of chloroxynil, bromoxynil and ioxynil, respectively, was hydrolyzed after 5 h. After a 20-h incubation, almost full conversion of chloroxynil and bromoxynil was observed in all strains, while only about 60% of the added ioxynil was converted into carboxylic acid. The product of ioxynil was not metabolized any further, and those of the other two herbicides very slowly. None of the nitrilase-producing strains hydrolyzed dichlobenil (2,6-dichlorobenzonitrile). 3,5-Dibromo-4-hydroxybenzoic acid exhibited less inhibitory effect than bromoxynil both on luminescent bacteria and germinating seeds of Lactuca sativa. 3,5-Diiodo-4-hydroxybenzoic acid only exhibited lower toxicity than ioxynil in the latter test.     

Effects of some non-pathogenic micro-organisms on the growth of gnotobiotic barley plants

Summary Mated cultures of Blakeslea trispora, which produce trisporic acid, inhibited root and shoot extension of barley plants grown in gnotobiotic cultures, where the carbon from roots was the sole source of carbon available to the microorganism. Unmated cultures of B. trispora and Mucor hiemalis, which do not synthesize trisporic acid, inhibited root extension only. Azotobacter chroococcum had a variable effect in different experiments on both root and shoot extension but could at least partially relieve the inhibitory effect of B. trispora on root growth. Plant growth regulators, applied to roots exogenously, produced effects on plants which, although comparable with some of the inhibitory effects of the micro-organisms, were not comparable with the stimulatory effects of A. chroococcum. re]19751127     

Screening of free-living rhizospheric bacteria for their multiple plant growth promoting activities

Plant growth promoting rhizobacteria (PGPR) are known to influence plant growth by various direct or indirect mechanisms. In search of efficient PGPR strains with multiple activities, a total of 72 bacterial isolates belonging to Azotobacter, fluorescent Pseudomonas, Mesorhizobium and Bacillus were isolated from different rhizospheric soil and plant root nodules in the vicinity of Aligarh. These test isolates were biochemically characterized. These isolates were screened in vitro for their plant growth promoting traits like production of indoleacetic acid (IAA), ammonia (NH(3)), hydrogen cyanide (HCN), siderophore, phosphate solubilization and antifungal activity. More than 80% of the isolates of Azotobacter, fluorescent Pseudomonas and Mesorhizobium ciceri produced IAA, whereas only 20% of Bacillus isolates was IAA producer. Solubilization of phosphate was commonly detected in the isolates of Bacillus (80%) followed by Azotobacter (74.47%), Pseudomonas (55.56%) and Mesorhizobium (16.67%). All test isolates could produce ammonia but none of the isolates hydrolyzed chitin. Siderophore production and antifungal activity of these isolates except Mesorhizobium were exhibited by 10-12.77% isolates. HCN production was more common trait of Pseudomonas (88.89%) and Bacillus (50%). On the basis of multiple plant growth promoting activities, eleven bacterial isolates (seven Azotobacter, three Pseudomonas and one Bacillus) were evaluated for their quantitative IAA production, and broad-spectrum (active against three test fungi) antifungal activity. Almost at all concentration of tryptophan (50-500 microg/ml), IAA production was highest in the Pseudomonas followed by Azotobacter and Bacillus isolates. Azotobacter isolates (AZT(3), AZT(13), AZT(23)), Pseudomonas (Ps(5)) and Bacillus (B(1)) showed broad-spectrum antifungal activity on Muller-Hinton medium against Aspergillus, one or more species of Fusarium and Rhizoctonia bataticola. Further evaluation of the isolates exhibiting multiple plant growth promoting (PGP) traits on soil-plant system is needed to uncover their efficacy as effective PGPR.     

Interactions between Azotobacter and "phosphobacteria" and their establishment in the rhizosphere as affected by soil fertility.

The effects on plant growth of "bacterial fertilizers" prepared from Azotobacter spp. and phosphate-solubilizing bacteria ("phosphobacteria") have been the subject of much controversy. Cases where no plant-growth stimulation occurred may often be accounted for by the failure to establish the bacterial inocula in the rhizosphere. Three factors that may influence inocula establishment, i.e. soil fertility, manuring, and interactions between Azotobacter and "phosphobacteria," were examined in pot experiments, designed for statistical analysis, in two neutral-alkaline soils, using lavender plants (Lavandula spica L.). During the experiments the numbers of Azotobacter and "phosphobacteria" were counted. Dry weights of roots and shoots were recorded after 16 weeks of growth. At the end of the experiments there were always more Azotobacter and "phosphobacteria" in the rhizospheres when plants were inoculated with both groups of organisms together than when they were inoculated singly. Addition of 2% farmyard manure to the richer soil enhanced this effect. Plant growth was greatest when seedlings were inoculated with both Azotobacter and the "phosphobacteria".