The growth of external AM fungal mycelium in sand dunes and in experimental systems

We estimated the biomass and growth of arbuscular mycorrhizal (AM) mycelium in sand dunes using signature fatty acids. Mesh bags and tubes, containing initially mycelium-free sand, were buried in the field near the roots of the dune grass Ammophila arenaria L. AM fungal mycelia were detected at a distance of about 8.5 cm from the roots after 68 days of growth by use of neutral lipid fatty acid (NLFA) 16:1ω5. The average rate of mycelium extension during September and October was estimated as 1.2 mm day⁻¹. The lipid and fatty acid compositions of AM fungal mycelia of isolates and from sand dunes were analysed and showed all to be of a similar composition. Phospholipid fatty acids (PLFAs) can be used as indicators of microbial biomass. The mycelium of G. intraradices growing in glass beads contained 8.3 nmol PLFAs per mg dry biomass, and about 15% of the PLFAs in G. intraradices, G. claroideum and AM fungal mycelium extracted from sand dunes, consisted of the signature PLFA 16:1ω5. We thus suggest a conversion factor of 1.2 nmol PLFA 16:1ω5 per mg dry biomass. Calculations using this conversion factor indicated up to 34 μg dry AM fungal biomass per g sand in the sand dunes, which was less than one tenth of that found in an experimental system with Glomus spp. growing with cucumber as plant associate in agricultural soil. The PLFA results from different systems indicated that the biomass of the AM fungi constitutes a considerable part of the total soil microbial biomass. Calculations based on ATP of AM fungi in an experimental growth system indicated that the biomass of the AM fungi constituted approximately 30% of the total microbial biomass. This revised version was published online in June 2006 with corrections to the Cover Date.     

Cloning and sequencing of a novel glutaryl acylase β-subunit gene ofPseudomonas cepacia BY21 from bioinformatics

Pseudomonas cepacia BY21 was found to produce glutaryl acylase that is capable of deacylating glutaryl-7-aminocephalosporanic acid (glutaryl-7-ACA) to 7-aminocephalosporanic acid (7-ACA), which is a starting material for semi-synthetic cephalosporin antibiotics. Amino acids of the reported glutaryl acylases from variousPseudomonas sp. strains show a high similarity (>93% identity). Thus, with the known nucleotide sequences ofPseudomonas glutaryl acylases in GenBank, PCR primers were designed to clone a glutaryl acylase gene fromP. cepacia BY21. The unknown β-subunit gene of glutaryl acylase from chromosomal DNA ofP. cepacia BY21 was cloned successfully by PCR. The β-subunit amino acids ofP. cepacia BY21 acylase (GenBank accession number AY948547) were similar to those ofPseudomonas diminuta KAC-1 acylase except that Asn408 ofP. diminuta KAC-1 acylase was changed to Leu408.     

Screening tomato-associated bacteria for biological control of grey mold on tomato

Aiming at discovering effective biocontrol agents (BCAs) against grey mold on tomato caused by Botrytis cinerea Pers., we selected 819 bacterial isolates from the surface as well as the interior of the roots, stems, and leaves of tomato plants grown in B. cinerea-infested fields. In a dual-culture assay, 116 isolates (14.16%) showed antagonism against B. cinerea and fewer ones against five additional tomato-associated fungal pathogens – Pythium ultimum, Phytophthora capsici, Fusarium oxysporum f. sp. lycopersici, Sclerotinia sclerotiorum and Ralstonia solanacearum. Thirty-one isolates with antagonism to B. cinerea and at least one of the five additional pathogens were assessed for their efficacy in controlling grey mold on tomato in a greenhouse test. Thirteen of them attained the efficacy over 50% and were subjected to the second greenhouse test, in which 12 isolates consistently accomplished the biocontrol efficacy over 50%, with isolates ABc28 and ABc22 achieving the efficacy of 66.71% and 64.90%, respectively. Under greenhouse conditions, the above two as well as isolates ABc2, ABc11 and ABc17 increased tomato biomass by more than 20% in comparison with the control. The 12 antagonistic isolates accomplishing the biocontrol efficacy over 50% in both greenhouse tests were considered potential BCAs against grey mold, which were identified as Pseudomonas spp., Pantoea spp., Bacillus spp. and Chryseobacterium spp. Ten of them were found to produce at least one of the three hydrolytic enzymes (protease, cellulase and chitinase) and/or siderophore, which might be involved in their mechanisms of suppressing the disease. Based on the origin of these 12 strains, the leaf tissue, especially the leaf interior, of tomato plants grown in a B. cinerea-infested field appears to be a good source of potential BCAs against grey mold.     

Combinative effects of a bacterial type-III effector and a biocontrol bacterium on rice growth and disease resistance

Expression of HpaG_Xoo, a bacterial type-III effector, in transgenic plants induces disease resistance. Resistance also can be elicited by biocontrol bacteria. In both cases, plant growth is often promoted. Here we address whether biocontrol bacteria and HpaG_Xoo can act together to provide better results in crop improvement. We studied effects ofPseudomonas cepacia on the rice variety R109 and the hpaG_Xoo-expressing rice line HER1. Compared to R109, HER1 showed increased growth, grain yield, and defense responses toward diseases and salinity stress. Colonization of roots byP. cepacia caused 20% and 13% increase, in contrast to controls, in root growth of R109 and HER1. Growth of leaves and stems also increased in R109 but that of HER 1 was inhibited. WhenP. cepacia colonization was subsequent to plant inoculation withRhizoctonia solani, a pathogen that causes sheath blight, the disease was less severe than controls in both R109 and HER1; HER1, nevertheless, was more resistant, suggesting thatP.cepacia and HpaG_Xoo cooperate in inducing disease resistance. Several genes that critically regulate growth and defense behaved differentially in HER1 and R109 while responding toP. cepacia. In R109 leaves, theOsARF1 gene, which regulates plant growth, was expressed in consistence with growth promotion byP. cepacia. Inversely,OsARF1 expression was coincident with inhibition in growth of HER1 leaves. In both plants, the expression ofOsEXP1, which encodes an expansin protein involved in plant growth, was concomitant with growth promotion in leaves instead of roots, in response toP. cepacia. We also studiedOsMAPK, a gene that encodes a mitogen-activated protein kinase and controls defense responses toward salinity and infection by pathogens in rice. In response toP. cepacia, an early expression ofOsMAPK was coincident with R109 resistance to the disease, while HER1 expressed the gene similarly whetherP. cepacia was present or not. Evidently,P. cepacia and G_Xoo-gene mediated resistance may act differently in rice growth and resistance. Whereas combinative effectsof P. cepacia and HpaG_Xoo in disease resistance have a great potential in agricultural use, it is interesting to study mechanisms that underlie interactions involving biocontrol bacteria, type-III effectors and pathogens. These authors contributed equally to this paper.     

Biological Control of Fusarium Wilt in Tomato Caused by Fusarium oxysporum f. sp. lycopersici by AMF Glomus intraradices and some Rhizobacteria

In the present study, the effects of the arbuscular mycorrhizal fungus (AMF) Glomus intraradices Schenck & Smith and four rhizobacteria (RB; 58/1 and D/2: Pseudomonas fluorescens biovar II; 17: P. putida; 21: Enterobacter cloacae), which are the important members of the rhizosphere microflora and biological control agents against plant diseases, were examined in the pathosystem of Fusarium oxysporum f. sp. lycopersici [(Sacc) Syd. et Hans] (FOL) and tomato with respect to morphological parameters (fresh and dry root weight) and phosphorous (P) concentration in the roots. Treatments with single and dual inoculation with G. intraradices and RB strains reduced disease severity by 8.6–58.6%. Individual bacteria inoculations were more effective than both the single AMF and dual (G. intraradices + RB) inoculations. In addition, the RB and G. intraradices enhanced dry root weight effectively. Significant increases in root weights were recorded particularly in the triple inoculations compared with single or dual inoculations. Compared with the non‐treated controls all biological control agents increased P‐content of treated roots of plants. Colonization with RB increased especially in triple (FOL + G. intraradices + RB) inoculations whereas colonization of G. intraradices was significantly decreased in treatment of FOL + G. intraradices compared with triple inoculations. The results suggest that suitable combinations of these biocontrol agents may ameliorate plant growth and health.     

Using combinations of biocontrol agents to control Botrytis cinerea on strawberry leaves under fluctuating temperatures

Experiments were conducted with Botrytis cinerea on strawberry leaves to investigate where combinations of commercially available biological control agents (BCAs) might control B. cinerea more effectively than individual BCAs. Specifically, we studied the persistence of biocontrol activities, spread of BCAs among leaves, and biocontrol efficacy in relation to application regimes: mixed versus single BCA, pre-versus post-inoculation application, and sequential versus simultaneous application. Three BCA products (Sentinel, Serenade and Trianum) were used for this study. Overall, Serenade did not significantly reduce sporulation of B. cinerea on strawberry leaf discs whereas Sentinel and Trianum gave a similar and significant biocontrol efficacy. Biocontrol efficacy remained almost unchanged 10 days after application at 20/20°C (day/night) or 24/16°C temperature regimes. In contrast, reduced biocontrol efficacy at 26/14°C suggests BCA survival was reduced under these conditions. Incidence of B. cinerea sporulation on leaf discs was ca. 60% higher on leaves that emerged after the BCA application than on leaves directly exposed to BCA, indicating insufficient amount of the BCA had managed to spread to new leaves. Combinations of BCAs, whether applied simultaneously or sequentially (48 h apart), did not improve disease control over the most effective BCA within the combination applied alone. This indicated possible antagonism or interference between the BCAs. Results suggested that there was significant antagonism for most combinations of the three BCAs tested and the degree of antagonism increased as the time from BCA application to pathogen introduction lengthened.     

Growth response of marigold (Tagetes erecta L.) to inoculation withGlomus mosseae,Trichoderma aureoviride andPythium ultimum in a peat-perlite mixture

The interactions between the mycorrhizal fungusGlomus mosseae, the plant pathogenPythium ultimum, and a pathogen-antagonistTrichoderma aureoviride in the rhizosphere ofTagetes erecta (marigold) were studied for their effects on plant growth in a peat-perlite substrate. Mycorrhizal fungus inoculation protected the plant againstP. ultimum, since both phytomass production and foliar development were higher in mycorrhizal plants.T. aureoviride had no effect on nonmycorrhizal plants in the presence or absence ofP. ultimum. However, more biomass was produced by mycorrhizal plants whenT. aureoviride was present, whether or not soil was infested withP. ultimum. ei]R Rodriguez-Kabana     

Effect of<Emphasis Type="Italic">Glomus intraradices</Emphasis> isolated from Pb-contaminated soil on Pb uptake by<Emphasis Type="Italic">Agrostis capillaris</Emphasis> is changed by its cultivation in a metal-free substrate

Development and heavy metal tolerance of two cultivation lineages of the indigenous isolate of arbuscular mycorrhizal fungus (AMF)Glomus intraradices PH5 were compared in a pot experiment in soil from lead (Pb) smelter waste deposits. One lineage was sub-cultured in original Pb-contaminated soil; the second one was maintained for 13 months in an inert substrate (river sand) without Pb stress. The contribution of these cultivation lineages to the Pb uptake and accumulation by the host plantAgrostis capillaris was investigated. The experiment was conducted in a compartmented system where the lateral compartments withAgrostis seedlings were separated from the central pot containing 4-week olderAgrostis plants by a nylon mesh for allowing out-growing of extraradical mycelium (ERM) from the pot. No differences in mycorrhizal colonization, ERM length and viability were observed between the two lineages ofG. intraradices PH5 in the soil of the isolate origin. However, the ability to support plant growth and Pb uptake differed between the lineages and also between the plants in the central pots and the lateral compartments. The growth of the plants in the central pots was positively affected by AMF inoculation. The plants inoculated with the lineage maintained in original soil showed larger shoot biomass and higher shoot P content as compared to the other inoculation treatments. The shoot Pb concentration of these plants was lower when compared to the plants inoculated with the lineage sub-cultured in the inert substrate. However the concentration did not differ from non-mycorrhizal control or from the reference isolateG. intraradices BEG75 from non-contaminated soil. Also shoot Pb contents were similar for all inoculation treatments. The development ofG. intraradices BEG75 in the contaminated soil was very poor; this isolate was not able to initiate colonization of seedlings in lateral compartments. In lateral compartments, growth of seedlings in contaminated soil was inhibited by theG. intraradices PH5 lineage maintained in the inert substrate. Pb translocation from the seedling roots to shoots was increased for plants inoculated with either lineage as compared to the non-mycorrhizal control; however, the increase for the lineage cultivated in the inert substrate was significantly higher in comparison with that maintained in the original soil. After 13 months of cultivation in a metal free substrate, theG. intraradices isolate from Pb contaminated soil did not lose its tolerance to Pb as regards colonization of plant roots and growth of ERM in the soil of its origin. However, its ability to support plant growth and to prevent Pb translocation from the roots to the shoots was decreased.     

Suppression of the Biocontrol Agent Trichoderma harzianum by Mycelium of the Arbuscular Mycorrhizal Fungus Glomus intraradices in Root-Free Soil

Trichoderma harzianum is an effective biocontrol agent against several fungal soilborne plant pathogens. However, possible adverse effects of this fungus on arbuscular mycorrhizal fungi might be a drawback in its use in plant protection. The objective of the present work was to examine the interaction between Glomus intraradices and T. harzianum in soil. The use of a compartmented growth system with root-free soil compartments enabled us to study fungal interactions without the interfering effects of roots. Growth of the fungi was monitored by measuring hyphal length and population densities, while specific fatty acid signatures were used as indicators of living fungal biomass. Hyphal ³³P transport and β-glucuronidase (GUS) activity were used to monitor activity of G. intraradices and a GUS-transformed strain of T. harzianum, respectively. As growth and metabolism of T. harzianum are requirements for antagonism, the impact of wheat bran, added as an organic nutrient source for T. harzianum, was investigated. The presence of T. harzianum in root-free soil reduced root colonization by G. intraradices. The external hyphal length density of G. intraradices was reduced by the presence of T. harzianum in combination with wheat bran, but the living hyphal biomass, measured as the content of a membrane fatty acid, was not reduced. Hyphal ³³P transport by G. intraradices also was not affected by T. harzianum. This suggests that T. harzianum exploited the dead mycelium but not the living biomass of G. intraradices. The presence of external mycelium of G. intraradices suppressed T. harzianum population development and GUS activity. Stimulation of the hyphal biomass of G. intraradices by organic amendment suggests that nutrient competition is a likely means of interaction. In conclusion, it seemed that growth of and phosphorus uptake by the external mycelium of G. intraradices were not affected by the antagonistic fungus T. harzianum; in contrast, T. harzianum was adversely affected by G. intraradices.     

Laboratory investigations of Aphodius beetles (Scarabaeidae) as biological control agents of the Cattle Lungworm,Dictyocaulus viviparus (Nematoda: Dictyocaulidae)

The effect of Aphodius fossor, A. luridus and A. foetidus adults on Dictyocaulus viviparus larvae in cattle faeces was investigated. In chambers containing A. fossor and A. luridus, a significant decrease (at 5% level) in the median number of D. viviparus larvae occurred after 90 h at a density of one beetle per g of faeces. The possibility of using dung beetles as biocontrol agents for D. viviparus is discussed.     

Response to cadmium of Daucus carota hairy roots dual cultures with Glomus intraradices or Gigaspora margarita

Ri T-DNA-transformed carrot roots were cultivated in two experiments either non-inoculated or inoculated with the arbuscular mycorrhizal (AM) fungi Glomus intraradices or Gigaspora margarita. The influence of two concentrations of cadmium (Cd) in the medium (2 mg l^–1, 4 mg l^–1) on both root and mycelium growth was tested. Both parameters were estimated at 10-day intervals for 70 or 100 days for G. intraradices and Gi. margarita, respectively. In the first experiment, G. intraradices showed a rapid spread of extraradical mycelium (ERM) and reached average densities per treatment of about 90 cm cm^–2 agar medium after 70 days. At the higher Cd level, the growth of ERM was delayed in comparison to the treatment without Cd addition. Root growth was inhibited by both Cd levels; the inhibition was, however, significantly lower in the treatments inoculated with G. intraradices compared to the non-inoculated control. In the second experiment, the ERM of Gi. margarita started to grow after a period of 50 days and reached average densities per treatment of only up to 27 cm cm^–2 by the end of the cultivation. The growth of Gi. margarita mycelium was not inhibited by Cd. No differences in root growth were observed between the Gi. margarita inoculated and non-inoculated treatments. The inhibitory effect of Cd on root growth differed between the non-inoculated treatments in both experiments. The study has shown that the AM fungus Glomus intraradices can alleviate Cd-induced growth inhibition to carrot hairy roots. The potential and limits of the monoxenic system in studying the interaction between AM fungi and heavy metals are discussed.     

An Indigenous Drought-Tolerant Strain of Glomus intraradices Associated with a Native Bacterium Improves Water Transport and Root Development in Retama sphaerocarpa

The effects of interactions between Bacillus thuringiensis, a drought-adapted bacterium, and two isolates of Glomus intraradices, an arbuscular mycorrhizal (AM) fungus, on Retama sphaerocarpa, a drought-adapted legume, were investigated. The fungal isolates were an indigenous drought-tolerant and a nonindigenous drought-sensitive isolate. Shoot length and root growth, symbiotic parameters, water transport (in terms of percent relative plant water uptake), and volumetric soil moisture and soil enzymatic activities in response to microbial inoculations were evaluated. Retama plants colonized by G. intraradices plus Bacillus possessed similar shoot length after 30 days from sowing compared with noninoculated Retama plants after 150 days. Inoculation with drought-adapted bacterium increased root growth by 201%, but maximum root development was obtained by co-inoculation of B. thuringiensis and the indigenous G. intraradices. Nodules were formed only in plants colonized by autochthonous AM fungi. Relative water uptake was higher in inoculated than in noninoculated Retama plants, and these inoculants depleted soil water content concomitantly. G. intraradices-colonized Retama reached similar shoot length irrespective of the fungal origin, but there were strong differences in relative water uptake by plants colonized by each one of the fungi. Indigenous G. intraradices-colonized roots (evaluated as functional alkaline phosphatase staining) showed the highest intensity and arbuscule richness when associated with B. thuringiensis. The interactive microbial effects on Retama plants were more relevant when indigenous microorganisms were involved. Co-inoculation of autochthonous microorganisms reduced by 42% the water required to produce 1 mg of shoot biomass. This is the first evidence of the effectiveness of rhizosphere bacterium, singly or associated with AM fungus, in increasing plant water uptake, which represents a positive microbial effect on plants grown under drought environments.     

Pseudomonas cepacia lipase: Studies on aggregation,purification and on the cleavage of olive oil

Summary To purify the extracellular lipase ofPseudomonas cepacia DSM 50181 and to disintegrate molecular aggregation, the presence of i-propanol (70% per volume) was necessary. The molecular weight was determined to be 33,000 D. Concerning the hydrolytic activity of the pure enzyme towards olive oil additional free fatty acids inhibited the reaction.     

Inoculated common beans are protected againstMacrophomina phaseolina byBurkholderia cepacia UPR 5C

Ashy stem blight (ASB), caused byMacrophomina phaseolina (Tassi) Goid., is a common severe disease affectingPhaseolus vulgaris in Puerto Rico and the Dominican Republic. The effect of inoculating seeds ofP. vulgaris withBurkholderia cepacia strain UPR 5C on the severity of ASB was studied under greenhouse conditions. Results of this study showed thatB. cepacia reduced the ASB disease severity by 71% and was compatible withRhizobium phaseoli CIAT 632.     

Significance of ecology in the development of biocontrol agents against soil‐borne plant pathogens

Approaches to the development of inoculant biocontrol agents (BCAs) of soil‐borne pathogens are discussed. Based on an analysis of the success of Peniophora (Phlebia) gigantea and Agrobacterium radiobacter it is argued that most subsequent attempts to develop inoculant BCAs have failed because the organisms were selected for in vitro antagonism but were ecologically unsuited to the environments where pathogens grow. The reported modes of action of BCAs are reviewed and in some cases reinterpreted. It is argued that antibiosis and some types of mycoparasitism have not been shown to be direct mechanisms of biocontrol in vivo; they might, instead, facilitate competition for substrates or sites as the primary mechanism of control. The ecology of BCAs in general is reviewed, with emphasis on interactions in microsites and the design of appropriate screening strategies. Specific examples are used to illustrate these points.     

Hexose phosphate metabolism and exopolysaccharide formation inPseudomonas cepacia

When grown on solid medium containing excess glucose, glucose dehydrogenase-deficient (Gcd^–) mutants ofPseudomonas cepacia 249 formed large amounts of an exopolysaccharide comprised of galactose, glucose, mannose, glucuronic acid, and rhamnose. The Gcd⁺ parent strain failed to accumulate comparable amounts of exopolymer from glucose because of its rapid conversion of glucose to gluconic and 2-ketogluconic acids and its lower content of enzymes related to glucose-1-phosphate synthesis. Both Gcd⁺ and Gcd^– strains ofP. cepacia accumulated exopolymer when substrates such as mannitol and glycerol were substituted for glucose. A survey of clinical isolates from patients with cystic fibrosis indicated that there was no correlation between ability ofP. cepacia to colonize the respiratory tracts of such individuals and increased capacity to form exopolymer related to glucose dehydrogenase deficiency.     

Differential effects of <Emphasis Type="Italic">Paenibacillus</Emphasis> spp. on cucumber mycorrhizas

The effects of 17 Paenibacillus strains on root colonization by Glomus intraradices or Glomus mosseae and plant growth parameters (shoot and root weight) of mycorrhizal cucumber plants were examined. The Paenibacillus strains were originally isolated from mycorrhizal (G. intraradices) and non-mycorrhizal cucumber rhizosphere and/or hyphosphere, except for strain EJP73, which originated from a Pinus sylvestris-Lactarius rufus ectomycorrhiza. Root colonization of cucumber plants by G. intraradices or G. mosseae was unaffected by all seven strains of Paenibacillus polymyxa, but was decreased or increased by four strains of Paenibacillus macerans and strain EJP73 of Paenibacillus sp. Overall, shoot dry weight of cucumber grown in symbioses with either G intraradices or G. mosseae was unaffected by inoculation with all of the Paenibacillus strains, except for strain MB02-429 of P. macerans, which increased the shoot dry weight in the cucumber-G. mosseae symbiosis. On the other hand, several Paenibacillus strains caused altered root growth. Three strains of P. polymyxa and four strains of P. macerans increased the root fresh weight of the cucumber–G. intraradices symbiosis, whereas three strains of P. polymyxa and one strain of P. macerans as well as Paenibacillus sp. EJP73, decreased the root fresh weight of the cucumber–G. mosseae symbiosis. In conclusion, our results show that bacteria from several species of Paenibacillus differentially affect cucumber mycorrhizas.     

Immunological and biological relationship among flagellin of <Emphasis Type="Italic">Pseudomonas aeruginosa,Burkholderia cepacia</Emphasis>, and <Emphasis Type="Italic">Stenotrophomonas maltophilia</Emphasis>

The flagellar protein (flagellin) was isolated and purified from strains of Pseudomonas aeruginosa, Burkholderia cepacia and Stenotrophomonas maltophilia. A significant difference was observed in the molecular weight of different flagellin preparations obtained from these bacterial isolates. Antiserum prepared against S. maltophilia flagellin did not react with flagellin of P. aeruginosa or/and B. cepacia on Immunoblot or in indirect ELISA. In addition the anti-flagellin did not agglutinate P. aeruginosa and B. cepacia. No inhibition of motility of P. aeruginosa and B. cepacia was observed in presence of antiserum; though the latter inhibited the motility of S. maltophilia. The results of the present study prove that no specific relationship existed among all the studied flagellar proteins obtained from closely related bacteria.     

Seasonal variation of arbuscular mycorrhizal fungi in temperate grasslands along a wide hydrologic gradient

We studied seasonal variation in population attributes of arbuscular mycorrhizal (AM) fungi over 2 years in four sites of temperate grasslands of the Argentinean Flooding Pampas. The sites represent a wide range of soil conditions, hydrologic gradients, and floristic composition. Lotus glaber, a perennial herbaceous legume naturalised in the Flooding Pampas, was dominant at the four plant community sites. Its roots were highly colonised by AM fungi. Temporal variations in spore density, spore type, AM root colonisation, floristic composition and soil chemical characteristics occurred in each site and were different among sites. The duration of flooding had no effect on spore density but depressed AM root colonisation. Eleven different types of spores were recognized and four were identified. Two species dominated at the four sites: Glomus fasciculatum and Glomus intraradices. Spore density was highest in summer (dry season) and lowest in winter (wet season) with intermediate values in autumn and spring. Colonisation of L. glaber roots was highest in summer or spring and lowest in winter or autumn. The relative density of G. fasciculatum and G. intraradices versus Glomus sp. and Acaulospora sp. had distinctive seasonal peaks. These seasonal peaks occurred at all four sites, suggesting differences among AM fungus species with respect to the seasonality of sporulation. Spore density and AM root colonisation when measured at any one time were poorly related to each other. However, spore density was significantly correlated with root colonisation 3 months before, suggesting that high colonisation in one season precedes high sporulation in the next season.     

Long-chain fatty acids and ethanol affect the properties of membranes inDrosophila melanogaster larvae

The larval fatty acid composition of neutral lipids and membrane lipids was determined in three ethanol-tolerant strains ofDrosophila melanogaster. Dietary ethanol promoted a decrease in long-chain fatty acids in neutral lipids along with enhanced alcohol dehydrogenase (EC 1.1.1.1) activity in all of the strains. Dietary ethanol also increased the incorporation of¹⁴C-ethanol into fatty acid ethyl esters (FAEE) by two- to threefold and decreased the incorporation of¹⁴C-ethanol into free fatty acids (FFA). When cultured on sterile, defined media with stearic acid at 0 to 5 mM, stearic acid decreased ADH activity up to 33%. In strains not selected for superior tolerance to ethanol, dietary ethanol promoted a loss of long-chain fatty acids in membrane lipids. The loss of long-chain fatty acids in membranes was strongly correlated with increased fluidity in hydrophobic domains of mitochondrial membranes as determined by electron spin resonance and correlated with a loss of ethanol tolerance. In the ethanol-tolerant E2 strain, which had been exposed to ethanol for many generations, dietary ethanol failed to promote a loss of long-chain fatty acids in membrane lipids. We are grateful for the support of National Institutes of Health Grant AA06702 (B.W.G.) and National Science Foundation Grant CHE-891987 (R.G.K.).