Ectomycorrhizal fungi: A new source of atmospheric methyl halides?

Incomplete source budgets for methyl halides – compounds that release inorganic chlorine and bromine radicals which, in turn, catalyze atmospheric ozone depletion – limit our ability to predict the fate of the stratospheric ozone layer. We report here the first measured emissions of methyl chloride, methyl bromide, and methyl iodide from ectomycorrhizal fungi. We grew nine fungal isolates on growth media containing halide concentrations similar to those found in soils and plant tissues. The observed range of emissions was 0.003–65 μg methyl chloride, 0.001–3 μg methyl bromide, and 0.02–12 μg methyl iodide g^?1 dry weight fungi day^?1. Species varied in production rates of methyl chloride vs. methyl bromide vs. methyl iodide. Cenococcum geophilum, a widespread ectomycorrhizal fungus, was further tested to investigate the effects of halide substrate concentration in growth media. Emissions from this species increased linearly with increasing concentrations of both bromide and iodide. In addition, a subset of four fungi was studied with two media concentrations each of chloride, bromide, and iodide (0.2 or 20 mm ). These fungi had similar responses to halide concentration, despite 1000‐fold differences in baseline emission rates between isolates. Finally, high chloride concentrations (20 mm ) in media did not appear to inhibit emissions of methyl bromide or methyl iodide. Overall, ectomycorrhizal fungi might be an important source of methyl halides to the atmosphere, and substrate concentrations and community composition may influence production levels in ecosystems.     

Activity of Plodia interpunctella (Lepidoptera: Pyralidae) in and around flour mills

Studies were conducted at two flour mills where male Indian meal moths, Plodia interpunctella (Hübner), were captured using pheromone-baited traps. Objectives were to determine the distribution of male P. interpunctella at different locations in and around the mills throughout the season, and to monitor moth activity before and after one of the mills was fumigated with methyl bromide to assess efficacy of treatment. Commercially available sticky traps baited with the P. interpunctella sex pheromone were placed at various locations outside and within the larger of the two mills (mill 1). Moths were captured inside mill 1 after methyl bromide fumigations. The highest numbers of P. interpunctella were caught outside the facility and at ground floor locations near outside openings. Additional traps placed in the rooms above the concrete stored-wheat silos at mill 1 during the second year captured more moths than did traps within the mill's production and warehouse areas. In another study, moths were trapped at various distances from a smaller flour mill (mill 2) to determine the distribution of moths outdoors relative to the mill. There was a negative correlation between moth capture and distance from the facility, which suggested that moth activity was concentrated at or near the flour mill. The effectiveness of the methyl bromide fumigations in suppressing moth populations could not be assessed with certainty because moths captured after fumigation may have immigrated from outside through opened loading bay warehouse doors. This study documents high levels of P. interpunctella outdoors relative to those recorded inside a food processing facility. Potential for immigration of P. interpunctella into flour mills and other stored product facilities from other sources may be greater than previously recognized. Moth entry into a food processing facility after fumigation is a problem that should be addressed by pest managers.     

Susceptibility of Maconellicoccus hirsutus (Homoptera: Pseudococcidae) to methyl bromide

Eggs, crawlers, early nymphs, late nymphs, and adults of the pink hibiscus mealybug, Maconellicoccus hirsutus (Green), were tested for their susceptibility to methyl bromide in 2-h laboratory fumigations at ambient conditions (25 degrees C, 95% RH). Dose-response tests indicated that the egg was the most susceptible life stage with an LC99 of 20.2 mg/liter. Based on probit analysis of dose-response data, no significant differences were observed among susceptibilities of the crawler, early stage or late stage nymphs, or adults at either the LC50 or LC99 level, but late stage nymphs were more tolerant than early stage nymphs in a separate paired comparison test. Confirmatory tests showed that a dose of 48 mg/liter methyl bromide, the USDA-Animal Plant Health Inspection Service treatment dose schedule for mealybugs at 21-26 degrees C, produced 100% mortality of all life stages. On the basis of these results, we conclude that the methyl bromide treatment schedule for mealybugs will provide quarantine security for M. hirsutus infesting commodities for export or import.     

Methyl benzoate fumigation for control of post-harvest pests and its effects on apple quality

Methyl benzoate is a newly discovered natural insecticide. In this study, we evaluated methyl benzoate as a potential fumigant for controlling four pests including western flower thrips (WFT), Frankliniella occidentalis, lettuce aphid, Nasonovia ribisnigri, rice weevil, Sitophilus oryzae, and bulb mites, Rhizoglyphus spp., at different temperatures and effects on post-harvest quality of apples. Methyl benzoate fumigations were effective against all four pests. Complete control of WFT and lettuce aphid was both achieved in 8, 16 and 24 hr at 25, 13 and 2°C, respectively. For rice weevil, complete control was achieved in 16 and 72 hr with and without rice at 25°C, respectively. Complete control of bulb mites was achieved in 64 hr at 25°C on peanut. In addition, a 24 hr methyl benzoate fumigation at 2°C had complete control of WFT but had no negative impact on visual quality of apples of three varieties 4 weeks after fumigation. These results suggest that methyl benzoate fumigation has potential to be an alternative treatment to methyl bromide fumigation, which has been phased out, for post-harvest pest control for stored products and may also have potential to control other pests on fresh products.     

Adenylyl cyclase activation by halide anions other than fluoride

Adenylyl cyclase of rat liver and fat cells is activated by chloride, bromide, and iodide in addition to fluoride, previously believed to be uniquely effective among the halide anions. Liver homogenates are activated approximately 6 fold by fluoride while chloride and bromide increase cyclase by 3 fold and iodide about 2 fold. Optimal concentrations of chloride, bromide and iodide are about 100 times higher than those required for activation by fluoride. The cyclase of fat cell ghosts is activated some 9 fold by fluoride, but the other halide anions produced effects very similar in magnitude to those seen with liver, although for fat the optimally effective concentrations were lower. These observations appear to relate adenylate cyclase to a number of other anion activated enzymes, some of which have already been studied in pure form by a number of physico-chemical techniques, and which may serve as models for understanding the action of fluoride and other anions on adenylyl cyclase.     

Phytotoxicity of methyl bromide on chrysanthemum cuttings

The phytotoxicity of methyl bromide gas on chrysanthemum cuttings was investigated in relation to gas concentration, length of exposure, temperature, and variety and age of cuttings. Concentrations of 10.5 to 13.5 g/m³ for 4 to 5 h produced a concentration time product (CTP) of 54 g h/m³ which is suitable for safe fumigation of most varieties of chrysanthemum. Length of exposure was not critical up to 5 h. However, fumigations of cuttings which were more than 3 wk old generally caused excessive phytotoxicity and therefore cannot be recommended. There was less damage at temperatures of between 14 and 16°C than at higher temperatures. A table of predictions from a fitted regression equation is included to demonstrate the increased susceptibility of chrysanthemums at higher temperatures and ages of cuttings.     

The oxidation of reduced nicotinamide nucleotides by hydrogen peroxide in the presence of lactoperoxidase and thiocyanate, iodide or bromide

Lactoperoxidase (EC 1.11.1.7) catalysed the oxidation of NADH by hydrogen peroxide in the presence of either thiocyanate, iodide or bromide. In the presence of thiocyanate, net oxidation of thiocyanate occurred simultaneously with the oxidation of NADH, but in the presence of iodide or bromide, only the oxidation of NADH occurred to a significant extent. In the presence of thiocyanate or bromide, NADH was oxidized to NAD(+) but in the presence of iodide, an oxidation product with spectral and chemical properties distinct from NAD(+) was formed. Thiocyanate, iodide and bromide appeared to function in the oxidation of NADH by themselves being oxidized to products which in turn oxidized NADH, rather than by activating the enzyme. Iodine, which oxidized NADH non-enzymically, appeared to be an intermediate in the oxidation of NADH in the presence of iodide. NADPH was oxidized similarly under the same conditions. An assessment was made of the rates of these oxidation reactions, together with the rates of other lactoperoxidase-catalysed reactions, at physiological concentrations of thiocyanate, iodide and bromide. The results indicated that in milk and saliva the oxidation of thiocyanate to a bacterial inhibitor was likely to predominate over the oxidation of NADH.     

Genetic control of methyl halide production in Arabidopsis

Methyl chloride (CH(3)Cl) and methyl bromide (CH(3)Br) are the primary carriers of natural chlorine and bromine, respectively, to the stratosphere, where they catalyze the destruction of ozone, whereas methyl iodide (CH(3)I) influences aerosol formation and ozone loss in the boundary layer. CH(3)Br is also an agricultural pesticide whose use is regulated by international agreement. Despite the economic and environmental importance of these methyl halides, their natural sources and biological production mechanisms are poorly understood. Besides CH(3)Br fumigation, important sources include oceans, biomass burning, tropical plants, salt marshes, and certain crops and fungi. Here, we demonstrate that the model plant Arabidopsis thaliana produces and emits methyl halides and that the enzyme primarily responsible for the production is encoded by the HARMLESS TO OZONE LAYER (HOL) gene. The encoded protein belongs to a group of methyltransferases capable of catalyzing the S-adenosyl-L-methionine (SAM)-dependent methylation of chloride (Cl(-)), bromide (Br(-)), and iodide (I(-)) to produce methyl halides. In mutant plants with the HOL gene disrupted, methyl halide production is largely eliminated. A phylogenetic analysis with the HOL gene suggests that the ability to produce methyl halides is widespread among vascular plants. This approach provides a genetic basis for understanding and predicting patterns of methyl halide production by plants.     

陆地生态系统卤甲烷释放特点及其生态意义

大气卤甲烷与平流层臭氧破坏密切相关,并参与光化学反应,还具有一定的．温室效应和污染毒害作用。研究发现：(1)大气CH3Cl和CH3Br存在巨大的未知源,它们的已知源分别仅占已知汇的大约1／2～2／3和60％。而CH3I的源和汇还都不确切;(2)陆地生态系统有可能是最大的卤甲烷自然释放源;(3)生物合成和土壤非生物生产是陆地生态系统卤甲烷生产的两个主要途径;(4)沿海湿地、水稻田、热带森林等陆地生态系统是卤甲烷主要释放源;(5)陆地生态系统卤甲烷的自然释放可能在生物竞争、生物代谢和大气环境污染方面具有重要的生态意义;(6)随着大气卤甲烷人为释放源的控制,其自然释放源的相对重要性将更加突出。提出了当前陆地生态系统卤甲烷释放研究的重点方向以及我国开展相关研究的重要意义。     

Sulfuryl fluoride as a quarantine treatment for Anoplophora glabripennis (Coleoptera: Cerambycidae) in regulated wood packing material

The Asian longhorned beetle, Anoplophora glabripennis (Motschulsky) (Coleoptera: Cerambycidae), was probably introduced into the United States from China with solid wood packing and dunnage during the 1980s, and it has recently become established in limited infestations near several major cities in the United States. Regulated wood packing material (RWPM) arriving in the United States from China is required to undergo fumigation with methyl bromide (MeBr), to be heat treated, or kiln dried. Sulfuryl fluoride (SF) is a candidate fumigant to replace MeBr under certain conditions. SF fumigations were conducted in 432-liter Lexan chambers held in a 6.1-m (20-foot) refrigerated container for temperature control. Each fumigation consisted of 12 Populus spp. 10- by 10- by 115-cm timbers, of high moisture content, naturally infested with Asian longhorned beetle. During 2001, we fumigated wood for 24 h at a range of doses (20-112 g/m3) and temperatures (4.4, 10.0, 15.6, and 21.1 degrees C) and subjected the data to probit analysis. Confirmatory fumigations were conducted at doses of 120 and 104 g/m3 at temperatures of 10.0 and 15.6 or 21.1 degrees C, respectively, which resulted in complete kill of all larvae. Pupae that became available later in the year as temperatures warmed were fumigated at 15.6 and 21.1 degrees C with 104 g/m3, which resulted in complete pupal mortality. The next year (2002), we conducted 24-h fumigations with doses of 116 g/m3 at 4.4 and 10.0 degrees C with cold-harvested wood infested with cold-acclimated larvae. Cold-acclimated larvae required much higher concentration times time (CxT) product for control at 4.4 and 10.0 degrees C compared with nonacclimated larvae. Sulfuryl fluoride treatments at a dose of 104 g/m3 and temperature of 15.6 degrees C and above and that achieved a CxT product of 1,095 g-h/m3 or above are recommended for RWPM infested with Asian longhorned beetle larvae and pupae.     

Chloride-sensitive fluorescent indicators

Three fluorescent halide-sensitive quinolinium dyes have been produced by the reaction of the 6-methylquinoline heterocyclic nitrogen base with methyl bromide, methyl iodide, and 3-bromo-1-propanol. The quaternary salts, unlike the precursor molecule, are readily water soluble and the fluorescence intensity of these salts is reduced in the presence of aqueous chloride, bromide, and iodide ions, allowing halide solution concentrations to be determined using well-known Stern-Volmer kinetics. One of the dyes, dye 1, has a chloride Stern-Volmer constant of 255 mol(-1) dm(3) which is more than twice that of SPQ [6-methoxy-N-(3-sulfopropyl)quinolinium] used in recent physiological measurements to measure intracellular chloride levels. The dyes have been characterized using steady-state fluorescence spectroscopy and are compared to three similar dyes based on the 6-methoxyquinoline nucleus, reported earlier by the authors, and also to dyes reported by Krapf et al. (Anal. Biochem. 169, 142-150, 1988). The interference of aqueous anions and the potential for using these dyes in biological halide-sensing applications are discussed.     

海洋中溴甲烷的研究进展

溴甲烷是大气中重要的痕量温室气体,对全球变暖和大气化学具有重要作用.海洋在溴甲烷的生物地球化学循环中作用复杂,海洋既是大气溴甲烷的源也是其汇,开展海洋环境中溶存溴甲烷的化学研究,对大气臭氧层的保护工作具有一定的指导意义,也可为在全球尺度上估算海洋环境中溴甲烷对全球环境变化的贡献提供理论依据.本文从溴甲烷的生物地球化学循环、分析测定方法、浓度分布、海-气通量、源汇平衡等方面综述了国内外海洋环境中溴甲烷的研究进展,探讨了目前该领域研究中存在的不足,并对未来的研究进行了展望.
     

Susceptibility of western flower thrips, Frankliniella occidentalis (Pergande) to fumigation with methyl bromide

All life stages of the western flower thrips (Frankliniella occidentalis Pergande) were tested for their response to the fumigant methyl bromide and dose response curves were established. Pseudo-pupae were the most tolerant stage with a calculated LD99 of 53.1 mg litre“¹ h at 15^oC with a gas concentration of 13.5 mg litre”¹, but no survivors were found in any test above a concentration time product of 40 mg litre“¹ h. Standard 4 h fumigations achieving a CTP of 54 mg litre”¹ h that are used to control other pests are therefore likely to give very high levels of control of western flower thrips.     

Physiological assay of liposome-mediated transport of a drug across Xenopus intestine: cell-liposome interaction

(1) The antagonistic effect of atropine methyl bromide entrapped in liposomes on contraction of Xenopus intestine in vitro induced by acetylcholine was studied. The results provided some insight into cell-liposome interaction. (2) Acetylcholine (0.1 mM) was added to the medium in the bath (serosal solution), while liposomes containing atropine methyl bromide in their internal and external phases were added on the mucosal side of the intestine. Large multilamellar liposomes were prepared from egg lecithin (phosphatidylcholine, PC) and cholesterol in various molar ratios. Atropine methyl bromide had most effect in liposomes composed of PC and cholesterol in a ratio of 7:3, less in those with a ratio of 4:5, and none in those with a ratio of 9:1. These effects were parallel with the sizes of these liposomes, determined by quasi-elastic light-scattering; that is, the larger the liposomes, the greater was their effect. Addition (to the liposomes) of phosphatidic acid, the negative charge of which increases the distance between the lamellar layers, increased the effect, indicating that atropine methyl bromide in the space between lamellar layers was effective. Another type of liposomes in which atropine methyl bromide was present only in the external phase of liposomes was as effective as liposomes in which atropine methyl bromide was present in both the internal and external phases. (3) From these results the following new model for liposome-mediated stimulation of transport of atropine methyl bromide is proposed. Large multilamellar liposomes have structural defects in their external lamellae through which atropine methyl bromide in the mucosal solution can penetrate into the space between the external lamellar layers and move into intestinal cells through regions of fusion between the outermost layers of the liposomes and the cell membrane.     

Metabolism of bromide and its interference with the metabolism of iodine

The present knowledge about the metabolism of bromide with respect to its goitrogenic effects, including some conclusions drawn from our recent research on this subject, is reviewed. Firstly, the biological behavior of bromide ion is compared with that of chloride and iodide. Secondly, the details about distribution and kinetics of bromide ions in the body and in 15 different organs and tissues of the rat are given. Significant correlation between the values of the steady-state concentration of bromide in the respective tissue and of the corresponding biological half-life was found in most tissues examined. A remarkably high concentration of radiobromide was found in the skin, which represents, due to its large mass, the most abundant depot of bromide in the body of the rat. Thirdly, the effects of excessive bromide on the rat thyroid are summarized, along with the interference of exogenous bromide with the whole-body metabolism of iodine. It is suggested that high levels of bromide in the organism of experimental animals can influence their iodine metabolism in two parallel ways: by a decrease in iodide accumulation in the thyroid and skin (and in the mammary glands in lactating dams), and by a rise in iodide excretion by kidneys. By accelerating the renal excretion of iodide, excessive bromide can also influence the pool of exchangeable iodide in the thyroid. Finally, our recent results concerning the influence of high bromide intake in the lactating rat dam on iodine and bromide transfer to the suckling, and the impact of seriously decreased iodine content and increased bromide concentration in mother's milk on the young are discussed. We must state, however, that the virtue of the toxic effects of excessive bromide on the thyroid gland and its interference with the biosynthesis of thyroid hormones, as well as the exact mechanism of bromide interference with postnatal developmental processes remains to be elucidated.     

Oxidation of methyl halides by the facultative methylotroph strain IMB-1.

Washed cell suspensions of the facultative methylotroph strain IMB-1 grown on methyl bromide (MeBr) were able to consume methyl chloride (MeCl) and methyl iodide (MeI) as well as MeBr. Consumption of >100 microM MeBr by cells grown on glucose, acetate, or monomethylamine required induction. Induction was inhibited by chloramphenicol. However, cells had a constitutive ability to consume low concentrations (<20 nM) of MeBr. Glucose-grown cells were able to readily oxidize [(14)C]formaldehyde to (14)CO(2) but had only a small capacity for oxidation of [(14)C]methanol. Preincubation of cells with MeBr did not affect either activity, but MeBr-induced cells had a greater capacity for [(14)C]MeBr oxidation than did cells without preincubation. Consumption of MeBr was inhibited by MeI, and MeCl consumption was inhibited by MeBr. No inhibition of MeBr consumption occurred with methyl fluoride, propyl iodide, dibromomethane, dichloromethane, or difluoromethane, and in addition cells did not oxidize any of these compounds. Cells displayed Michaelis-Menten kinetics for the various methyl halides, with apparent K(s) values of 190, 280, and 6,100 nM for MeBr, MeI, and MeCl, respectively. These results suggest the presence of a single oxidation enzyme system specific for methyl halides (other than methyl fluoride) which runs through formaldehyde to CO(2). The ease of induction of methyl halide oxidation in strain IMB-1 should facilitate its mass culture for the purpose of reducing MeBr emissions to the atmosphere from fumigated soils.     

Effect of chaotropic anions on the sodium transport by the Na,K-ATPase

The effect of choline iodide, bromide and chloride on the kinetics of the electrogenic sodium transport by the Na,K-ATPase was investigated in a model system of ATPase-containing membrane fragments adsorbed on the lipid bilayer membrane. The kinetic parameters of Na⁺ transport were determined from short circuit currents after fast release of ATP from its caged precursor. The falling phase of the current transients could be fitted by a single exponential with the time constant, τ ₂. Its temperature dependence allowed an estimation of the activation energy of the rate-limiting reaction step, the conformation transition E₁/E₂. Choline iodide and bromide caused a decrease of the activation energy as well as the overall rate of the process expressed as the pre-exponential factor A of the Arrhenius equation. If choline iodide or bromide were present on the cytoplasmic and extracellular sides of the protein, the temperature dependent changes were more pronounced than when present on the cytoplasmic side only. These results can be explained by an effect of the anions on water structure on the extracellular surface of the protein, where a deep access channel connects the ion-binding sites with the solution. Chloride ions also caused a deceleration of the electrogenic transport, however, in contrast to iodide or bromide, they did not affect the activation energy, and were more effective when added on the cytoplasmic side. This effect can be explained by asymmetric screening of the negative surface charges which leads to a transmembrane electric potential that modifies the ion transfer.     

Oxygen enhances phosphine toxicity for postharvest pest control

Phosphine fumigations under superatmospheric oxygen levels (oxygenated phosphine fumigations) were significantly more effective than the fumigations under the normal 20.9% atmospheric oxygen level against western flower thrips [Frankliniella occidentalis (Pergande)] adults and larvae, leafminer Liriomyza langei Frick pupae, grape mealybug [Pseudococcus maritimus (Ehrhorn)] eggs, and Indianmeal moth [Plodia interpunctella (Hübner)] eggs and pupae. In 5-h fumigations with 1,000 ppm phosphine at 5 degrees C, mortalities of western flower thrips increased significantly from 79.5 to 97.7% when oxygen was increased from 20.9 to 40% and reached 99.3% under 80% O2. Survivorships of leafminer pupae decreased significantly from 71.2% under 20.9% O2 to 16.2% under 40% O2 and reached 1.1% under 80% O2 in 24-h fumigations with 500 ppm phosphine at 5 degrees C. Complete control of leafminer pupae was achieved in 24-h fumigations with 1,000 ppm phosphine at 5 degrees C under 60% O2 or higher. Survivorships of grape mealybug eggs also decreased significantly in 48-h fumigations with 1,000 ppm phosphine at 2 degrees C under 60% O2 compared with the fumigations under 20.9% O2. Indian meal moth egg survivorships decreased significantly from 17.4 to 0.5% in responses to an oxygen level increase from 20.9 to 40% in 48-h fumigations with 1,000 ppm phosphine at 10 degrees C and reached 0.2% in fumigations under 80% O2. When the oxygen level was reduced from 20.9 to 15 and 10% in fumigations, survivorships of Indianmeal moth eggs increased significantly from 17.4 to 32.9 and 39.9%, respectively. Increased O2 levels also resulted in significantly lower survival rates of Indianmeal moth pupae in response to 24-h fumigations with 500 and 1,000 ppm phosphine at 10 degrees C and a complete control was achieved in the 1,000 ppm phosphine fumigations under 60% O2. Oxygenated phosphine fumigations have marked potential to improve insecticidal efficacy. Advantages and limitations of oxygenated phosphine fumigation are discussed.     

Preparation of methyl ester precursors of biologically active agents

This method enables scientists to easily convert biologically active carboxylic acids into their methyl esters ("pro-drugs" generally having improved ability to penetrate cell membranes) using only equipment commonly found in a biology laboratory. An ion-exchange resin is used to convert the acid into its salt, which is thereby sequestered on the resin. The addition of methyl iodide converts the salt to the ester, which has no affinity for the resin and is readily eluted. Evaporation of the liquid phase provides the pure methyl ester. The preparation in good chemical yields of methyl esters of bioactive agents in excellent purity and 10-20 mg quantities can be achieved using this method. The method can be completed in 1 day.     

EFFECTS OF BROMIDE AND IODIDE ON STALK SECRETION IN THE BIOFOULING DIATOM ACHNANTHES LONGIPES (BACILLARIOPHYCEAE)1

Extracellular polymeric substance (EPS) secretion was examined in the stalked marine diatom Achnanthes longipes Ag. in defined medium. This common biofouling diatom exhibited an absolute requirement for bromide for stalk production and substratum attachment, whereas elevated iodide concentrations in the growth medium inhibited stalk formation and adhesion. Varying EPS morphologtes resulted from altering bromide and iodide levels: pads, stalk-pads, stalks, and no EPS. Cells showed no differences in growth with bromide or iodide concentrations, indicating that they were not physiologically stressed under conditions that impaired EPS secretion. Cells grown in elevated iodide secreted significantly more soluble extracellular carbohydrate into the medium, suggesting that the EPS was soluble and unable to be polymerized into a morphologically distinct gel. By replacing sulfate with methionine, the diatom lost its ability to form stalks even in the presence of bromide, indicating that free sulphate may be required for proper cross-linking of stalk polymers. Lotus-FITC, a fluorescent-tagged lectin, preferentially labeled the EPS and, thus, was used to visualize and quantify EPS secretion along a bromide gradient in conjunction with an image analysis system. This technique demonstrated a direct correlation between the amount of bromide present in the medium and the specific EPS morphology formed.