Hyphal anastomosis and complementary growth of fused cells inAlternaria alternata

Hyphal anastomosis and complementary growth of fused cells inAlternaria alternata were investigated. Sixty-four experimental isolates were divided into anastomosis-positive (A⁺) and anastomosis-negative (A⁻) groups based on their self-anastomosing ability. Nonself-anastomoses (interisolate) were readily distinguished from self-anastomoses (intraisolate) by using a mixed culture of conidia and hyphal fragments prepared from the respective isolates. Nonself-anastomosis occurred only between the A⁺ isolates irrespective of their pathogenicity and geographic origin. The breakdown of cell walls and the establishment of cytoplasmic continuity between fused cells were microscopically observed only in the self-anastomoses. The frequency of the nonself-anastomosis was, in general, lower than that of the self-anastomosis. For analysis of complementation between the fused cells, mutants doubly marked with auxotrophy and hygromycin B (Hyg) resistance were prepared from wild-type isolates. The identity of the mutants was confirmed by RAPD analysis using three arbitrary primers. Complementary growth occurred only between an A⁺ isolate and its mutant(s) on a minimal medium containing Hyg, demonstrating that the self-anastomoses resulted in perfect cell fusions and the nonself-anastomoses were contact or imperfect fusions.     

Effects of Rhizobium inoculation on growth and nodulation ofAlbizia falcataria (L.) Fosh. andAcacia mangium Willd. in the nursery

Three separate experiments were conducted in the nursery using grassland soil as a growing medium. The first experiment was conducted to assess the nodulation of the two legume trees grown in unamended soil, the second was done to determine the effects of N-fertilizer application on the interaction of the five Rhizobium isolates withA. falcataria and the third experiment was conducted to determine the effects of liming on theRhizobium × A. mangium interaction. Three local Rhizobium isolates, A₁₆, A₁₈, and A₁₄ were effective forA. falcataria with A₁₆ as the most promising strain under the conditions described. In general, application of combined nitrogen suppressed nodulation ofA. falcataria. Nodulation in the absence of combined N exceeded those fertilized with 30 kg N ha⁻¹ by 114.0%, 60 kg N ha⁻¹ by 209.6% and 100 kg N ha⁻¹ by 237.1%. Two local isolates, Am₁₀ and Am₂, and an introduced strain, NA 1533 from Australia were promising forA. mangium. Unlike inA. falcataria, the application of combined nitrogen at the rate of 100 kg N ha⁻¹ did not suppress nodulation inA. mangium. Liming the soil to pH 6.5 regardless of nitrogen fertilizer application improved the performance of the Rhizobium—A. mangium symbiosis.     

Distribution of Phytophthora cinnamomi-Supprcssive Soil in Nature

Phytophthora cinnamomi-suppressivc soils were found to be widely distributed in nature. About 40 % of soil samples collected from locations with different vegetation, soil type or elevation throughout the island of Hawaii were suppressive to chlamydospore germination of Phytophthora cinnamomi. Soil samples collected from the same general areas varied greatly in degree of suppressive-ness to P. cinnamomi, ranging from conducive to strongly suppressive. Among the 155 soil samples tested, those with pH close to 4 or 8 tend to be more suppressive to P. cinnamomi than those with pH close to 6.     

Mechanisms of salt tolerance in transgenic Arabidopsis thaliana constitutively overexpressing the tomato 14-3-3 protein TFT7

A hydroponics experiment was conducted to investigate the effects of iron plaque on root surfaces with respect to selenite uptake and translocation within the seedlings of two cultivars of rice (Oryza sativa L. cv Xiushui48 and Bing9652). Different amounts of iron plaque were formed by adding 0, 10, 30, 50, 70 mg Fe l⁻¹ in the nutrient solution. After 24 h of growth, the amount of iron plaque was positively correlated with the Fe²⁺ addition to the nutrient solution. These concentrations of Fe, inducing plaque, had no significant effect on the shoot and root growth of rice plants in 50 μg Se l⁻¹ nutrient solution. The amount of Se accumulated in iron plaque was positively correlated to the amount of iron plaque. Increasing iron plaque decreased the selenium concentration in shoots and in roots. At the same time, the translocation of Se from roots to shoots was reduced with increasing amounts of iron plaque. At both the shorter and longer exposure times, the ratio of root- to-shoot selenium was higher than in the controls. More Se stayed in the roots at the longer exposure time than at the shorter time. The concentration of selenium in the xylem sap was sharply decreased with increasing amount of iron plaque on the rice roots. The DCB (dithionite-citrate-bicarbonate)-extracted Se was up to 89.9–91.1% of the total Se when roots with iron plaque (Fe 70) were incubated in 50 μg Se l⁻¹ solution for 30 min. This DCB-extracted Se, however, accounted for only 21.9–28.7% of total Se when roots with iron plaque were incubated in the same solution for 3 days. Se adsorbed in iron plaque can be desorbed by low-molecular-weight organic acids, similar to the desorption of Se from ferrihydrite. These results suggest that iron plaque might act as a ‘buffer’ for Se in the rhizosphere.     

Evidence of adaptive tolerance to nickel in isolates of Cenococcum geophilum from serpentine soils

Selection for metal-tolerant ecotypes of ectomycorrhizal (ECM) fungi has been reported in instances of metal contamination of soils as a result of human activities. However, no study has yet provided evidence that natural metalliferous soils, such as serpentine soils, can drive the evolution of metal tolerance in ECM fungi. We examined in vitro Ni tolerance in isolates of Cenococcum geophilum from serpentine and non-serpentine soils to assess whether isolates from serpentine soils exhibited patterns consistent with adaptation to elevated levels of Ni, a typical feature of serpentine. A second objective was to investigate the relationship between Ni tolerance and specific growth rates (μ) among isolates to increase our understanding of possible tolerance/growth trade-offs. Isolates from both soil types were screened for Ni tolerance by measuring biomass production in liquid media with increasing Ni concentrations, so that the effective concentration of Ni inhibiting fungal growth by 50% (EC₅₀) could be determined. Isolates of C. geophilum from serpentine soils exhibited significantly higher tolerance to Ni than non-serpentine isolates. The mean Ni EC₅₀ value for serpentine isolates (23.4 μg ml⁻¹) was approximately seven times higher than the estimated value for non-serpentine isolates (3.38 μg ml⁻¹). Although there was still a considerable variation in Ni sensitivity among the isolates, none of the serpentine isolates had EC₅₀ values for Ni within the range found for non-serpentine isolates. We found a negative correlation between EC₅₀ and μ values among isolates (r = −0.555). This trend, albeit only marginally significant (P = 0.06), indicates a potential trade-off between tolerance and growth, in agreement with selection against Ni tolerance in “normal” habitats. Overall, these results suggest that Ni tolerance arose among serpentine isolates of C. geophilum as an adaptive response to Ni exposure in serpentine soils. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Induction of Sexual Reproduction in Single A<Superscript>2</Superscript> Isolates of <Emphasis Type="Italic">Phytophthora</Emphasis> Species by <Emphasis Type="Italic">Trichoderma viride</Emphasis>

ABOUT half the species of Phytophthora, including many important plant pathogens such as P. palmivora, P. cinnamomi and P. infestans, are heterothallic. Consistent induction of sexual reproduction in agar cultures of heterothallic species requires the pairing of isolates of the two compatibility groups, A¹ and A² (refs. 1–3). Although sex organs are occasionally formed in cultures of single isolates, such isolates though intrinsically bisexual are normally self-sterile^2–3. In consequence it has been assumed that in nature the presence of both compatibility types is required for the formation of oospores.     

Microbial Responses to Environmentally Toxic Cadmium

Abstract We analyzed the soil microbial communities from one uncontaminated and two metal-impacted soils and found that while cadmium adversely affected the numbers of culturable bacteria in all soils, cadmium-resistant isolates were found from each of the soils. With exposure to 24 and 48 μg ml^-1 soluble cadmium, the metal-contaminated soil communities were more resistant than the uncontaminated soil community. In addition, in one metal-stressed soil, the resistant population became more resistant with increased cadmium levels. Ribosomal 16S DNA sequencing identified the isolates as Arthrobacter, Bacillus, or Pseudomonas spp. Further characterization demonstrated that two of the isolates were highly resistant to soluble cadmium with maximum resistance at 275 μg ml^-1 cadmium. These isolates were also resistant to a variety of antibiotics, namely ampicillin, gentamicin, penicillin, and streptomycin, but no overall correlation was found between enhanced antibiotic resistance and cadmium resistance. One Pseudomonas isolate H1 did become more resistant with increasing cadmium levels, suggesting a different resistance mechanism at high cadmium concentrations. Received: 29 April 1999; Accepted: 7 July 1999; Online Publication: 30 November 1999     

Soil fumigation and phosphorus supply affect the formation of Pisolithus-Eucalyptus urophylla ectomycorrhizas in two acid Philippine soils

To examine the effects of microbial populations and external phosphorus supply of two Philippine soils on mycorrhizal formation, Eucalyptus urophylla seedlings were inoculated with two Pisolithus isolates and grown in fumigated, reinfested and unfumigated soil fertilized with four rates of phosphorus. The Pisolithus isolates used were collected from under eucalypts in Australia and in the Philippines. Soils were infertile acid silty loams collected from field sites in Pangasinan, Luzon and Surigao, Mindanao.Significant interaction was observed between inoculation, soil fumigation and phosphorus supply on mycorrhizal formation by the Australian isolate in Surigao soil but not in Pangasinan soil. Soil fumigation enhanced mycorrhizal formation by the Australian isolate but did not affect root colonization by the Philippine isolate. Root colonization by the Australian isolate was highest in the reinfested soil while for the Philippine isolate it was highest in the unfumigated soil. The Australian isolate was more effective than the Philippine isolate in promoting growth and P uptake of E. urophylla seedlings in both soils. Total dry weight and P uptake of E. urophylla seedlings inoculated with the Australian isolate were maximum in fumigated and in the reinfested Pangasinan and Surigao soils supplied with 8 mg P kg^-1 soil. In the unfumigated soil, growth of seedlings inoculated with the Australian isolate was significantly reduced. Seedlings inoculated with the Philippine isolate had the largest dry weights and P contents in unfumigated Pangasinan and Surigao soils supplied with 8 mg P kg^-1 soil.These results indicate that the performance of the Australian Pisolithus isolate was markedly affected by biological factors in unfumigated soil. Thus, its potential use in the Philippines needs to be thoroughly tested in a variety of unfumigated soils before its widespread use in any inoculation programme.     

Serotyping of <Emphasis Type="Italic">Bacillus thuringiensis</Emphasis> isolates,their distribution in different Jordanian habitats and pathogenicity in <Emphasis Type="Italic">Drosophila melanogaster</Emphasis>

The investigation of Bacillus thuringiensisin 40 different samples collected from 12 different Jordanian habitats involved the isolation of 80 Bacillus thuringiensis isolates. Out of these isolates, 47 were pathogenic to the third instar larvae of Drosophila melanogaster. The highest viable count of Bacillus thuringiensis was estimated among soil samples contaminated with decomposed animal bodies (14.25 × 10⁷ c.f.u./g), and the lowest viable count was obtained from soils contaminated with engine oil (0.17 × 10⁷c.f.u./g). Serotyping of the 80 isolates against 55 antisera indicated the presence of 13 serotypes, 12 were identical or cross-reacted withaizawai, higo,israelensis, kenyae, kumamotoensis, kurstaki, malaysiensis, morrisoni, pakistani,sooncheon,tohokuensis, andthuringiensis, whereas the remaining one reacted negatively with the 55 tested antisera indicating the presence of an unknown serotype. Israelensis was the dominant serotype among all the samples except those from decomposed animal and olive-cultivated soils. The pathogenic isolates were found to be in 11 of the 13 serotypes. Spherical parasporal crystals were the most common and toxic crystal types.     

Endophytic fungi of teak leaves Tectona grandis L. and rain tree leaves Samanea saman Merr.

Summary In order to study the foliar endophytes from teak (Tectona grandis L.) and rain tree (Samanea saman Merr.) growing in the campus of Chulalongkorn University, healthy leaves were collected at two-monthly intervals during January to December. The number of genera and species, together with their colonization frequency (CF%) in mature teak and rain tree leaves were greater than those in the young leaves. More endophytic isolates in the leaves of both trees were recovered during the rainy season. The fungal genera found in both young and mature teak leaves were Alternaria, Colletotrichum, Nigrospora, Phomopsis and mycelia sterilia. Phomopsis was the dominant genus in both young (newly emerged) and mature leaves. Fusarium, Penicillium, Schizophyllum commune and members of the Xylariaceae were found only in mature leaves. For the rain tree leaves, species of Phomopsis and mycelia sterilia were found in both young newly emerged and mature leaves. Colletotrichum and Penicillium were found only in mature leaves, whereas Nigrospora was found only in young newly emerged leaves. In this study, Phomopsis was the dominant genus in the leaves of both tree species. A total of 37 isolates of endophytic fungi isolated from teak and rain tree leaves were tested for the production of antimicrobial activities. Out of these, 18 isolates could produce inhibitory substances effective against Bacillus subtilis, Staphylococcus aureus and Escherichia coli and 3 isolates inhibited growth of Candida albicans in vitro.     

Isolation and Molecular Characterization of Heavy Metal-Resistant Azotobacter chroococcum from Agricultural Soil and Their Potential Application in Bioremediation

Pollution of soil with heavy metals, herbicides, antibiotics and other chemicals is known to have a negative effect on microbial activities. Therefore, the aim of this study was to isolate cultures of Azotobacter sp. from polluted and unpolluted soils and to study the effect of these pollutants on their growth. A total of 120 Azotobacter sp. were isolated from soils irrigated with wastewater (contaminated soils) and groundwater (uncontaminated soils). These isolates were screened for resistance to heavy metals, herbicide and antibiotics. Also, the soils from which the cultures were isolated were analyzed for the concentrations of Zn²⁺, Cd²⁺, Cu²⁺, Pb²⁺ and Mn²⁺ they contained. Contaminated soil showed high levels of heavy metals as compared to uncontaminated soil. The size of the Azotobacter population in contaminated soil was lower than that in uncontaminated soil. Of the Azotobacter isolates, 64 that were recovered from contaminated soil exhibited high resistance to heavy metals (Hg²⁺, Cd²⁺, Cu²⁺, Cr³⁺, Co²⁺, Ni²⁺, Zn²⁺ and Pb²⁺) and herbicide 2,4-D compared to 56 isolates from uncontaminated soil. Also, isolates from contaminated soil showed high resistance to chloramphenicol, nitrofurantoin and co-trimoxazole compared to those isolated from uncontaminated soil. The majority of Azotobacter isolates from contaminated soil showed multiple-resistance to different metal ions and antibiotics. All isolates failed to grow at pH less than 6. Salt concentration (5%) was found to be inhibitory to all isolates. The most potent isolates from contaminated soil that showed multiresistance to all substances tested were identified on the basis of morphological and biochemical characteristics, and 16S rRNA as A. chroococcum. These resistant isolates could be employed in contaminated soils and/or bioremediation.     

Characterization of Phytophthora cinnamomi populations from ornamental plants in South Carolina,USA

Abstract

Fifty-one isolates of Phytophthora cinnamomi isolated from ornamental plants in South Carolina, USA, between 1995 and 2000 were characterized by sporangium morphology, mating type, sensitivity to the fungicide mefenoxam, fatty acid methyl ester (FAME) profile analysis, and amplified fragment length polymorphism (AFLP) analysis. Sporangium shapes were predominantly ovoid to ellipsoid, and size averaged 65.5×40.3 μm (length×breadth) with average length/breadth ratio of 1.6. Forty-nine isolates were the A2 mating type with only two A1 isolates found. This is the first report of the A1 mating type of P. cinnamomi in South Carolina. All isolates were sensitive to mefenoxam and EC₅₀ values for all isolates were less than 0.2 μg ml^?1. FAMEs of each isolate were analysed by gas chromatography and revealed five major fatty acids: myristic (14:0), palmitic (16:0), linoleic (18:2ω6c), oleic (18:1ω9c), and eicosapentaenoic (20:5ω3c) acids. These five fatty acids accounted for more than 80% of FAME profiles. Cluster analysis of FAME profiles showed that individual isolates had unique pattern that could be divided into four major clusters. AFLP analysis based on 200 informative loci also separated isolates into four major clusters. A1 isolates were different from all A2 isolates. The percentage of polymorphic loci (10.5%) and Nei's gene diversity (0.0435) were much higher for the two A1 isolates than for any cluster of A2 isolates even though A2 isolates had more isolates within a cluster. A2 isolates exhibited relatively little genetic variation overall, which suggests that these isolates may have come from a common source.     

Multiplicity and control of alcohol dehydrogenase isozymes in various strains of wine yeasts

An investigation has been made of selffertile (homothallic) progeny which are frequently encountered in matings of heterothallic Phytophthora species. The pattern of segregation of self-sterility from these homothallic isolates during vegetative growth, asexual and sexual reproduction has been studied in some detail.Heterokaryosis was shown not be the cause of this secondary homothallism in a number of cases investigated since homothallism could be transmitted by single uninucleate zoospores.The homothallic cultures (A¹ A²) showed different degrees of stability. One culture, obtained as a very fertile sector from an original A¹ A² gave only A¹ A² phenotypes on repeated single zoospore analysis, whilst two other sectors from the same A¹ A² culture gave in addition to A¹ A², some A² and a few A¹ phenotypes. The A² types derived from this homothallic were also unstable, giving some A¹ A² and occasional A¹ types as well as A² types in single zoospore cultures. This contrasts with the stability of the parental A² culture.Self-fertility was transmitted through the oospore to the sexual progeny. Most progeny were A¹ A² but heterothallic A¹ and A² types segregated.It is suggested that the self-fertile condition was due to the presence of an extra chromosome containing the mating type locus, the A¹ A² type being Aaa. A cytological investigation to test this trisomic hypothesis is being reported separately.     

Intraspecific variability of flavonoid glycosides and styrylpyrones from leaves of Cryptocarya mandioccana Meisner (Lauraceae)

Chemical variability in populations of Cryptocarya mandioccana Meisner was investigated using chromatographic profiles of flavonoid glycosides and styrylpyrones from leaves of 57 trees from three sites in the Atlantic rain forest of São Paulo State – Brazil. The flavonoid profiles of the leaves from all trees were similar. However, four chemotypes were recognized by qualitative and quantitative differences in styrylpyrones. Chemotype F showed lower contents of these polyketides than chemotypes FS1, FS2 and FS3, which were characterized by styrylpyrones formed from different numbers of acetate units. Chemotypes with higher levels of styrylpyrones were more frequent in southern regions, with soils of higher K⁺, Ca²⁺ and Mg²⁺ content. Soil and climate could indicate a genetic or environmental influence on styrylpyrones in leaves of C. mandioccana.     

Centrin association with the flagellar apparatus in spores ofPhytophthora cinnamomi

Summary Antibodies raised against the calcium-binding protein centrin, were used to identify and localise centrin containing structures in the flagellar apparatus of zoospores and cysts of the oomycetePhytophthora cinnamomi. Immunoblotting of extracts from zoospores indicates that theP. cinnamomi centrin homologue is a 20 kDa protein. Immunofluorescence microscopy with anti-centrin antibodies reveals labelling in the flagella, the basal body connector and co-localisation along the microtubular R1 root (formerly called AR3) that runs from the right side of the basal body of the anterior flagellum into the anterior of the zoospore close to the ventral surface. The centrin (R1^cen) and tubulin components of the R1 root split into four loops on the right hand side of the ventral groove and rejoin along the left hand side of the groove. The R1 root continues down the left hand side of the zoospore past the basal bodies and parallel to the R4 root. We propose that at least inP. cinnamomi there is no R2 root. Immunogold labelling confirms that centrin is a component of the basal body connector complex. When the zoospores become spherical during encystment, the R1^cen pivots by approximately 90 ° with respect to the nucleus.     

Isolation and characterization of nickel-resistant microflora from serpentine soils of Andaman

Serpentine soils of Andaman Islands, India characteristically contain high levels of nickel, cobalt and chromium and are colonized by indigenous nickel-hyperaccumulating plants. Attempts have been made to isolate and characterize nickel-resistant microorganisms from these hitherto unexplored naturally nickel-percolated soils. The majority of the nickel-resistant organisms showed a minimum inhibitory concentration (MIC) of Ni²⁺ ranging from 300 to 400 mg/l and about 3.4% of the total 89 isolates representing bacterial strains were able to grow at 400 mg/l Ni²⁺. The potent Ni²⁺-resistant strains AND305 and AND603 were tentatively identified as Pseudomonas spp. and strain AND408 as Bacillus sp. following detailed analysis of morphological and physio-biochemical characteristics. Growth kinetics of these Ni²⁺-resistant bacteria showed a prolonged lag phase in Ni²⁺-containing media, which extended with increasing nickel concentration. In addition to Ni²⁺, these isolates were also resistant to Co²⁺, Cd²⁺, Cr⁶⁺, Fe³⁺, Cu²⁺, Mg²⁺, Mn²⁺(50–200 mg/l) and Hg²⁺ (0.5–2.0 mg/l) and the multiple metal-resistance of the isolates were also associated with the resistance to antibiotics ampicillin, cycloserine and penicillin G.     

Soil P-status and cultivar maturity effects on pea-Rhizobium symbiosis

In a green-house experiment, five cultivars of Pisum sativum L. grown on soils from 10 different locations in Tunisia, showed significant differences in nodulation, shoot dry matter (shDM) yield and shoot nitrogen content (shNC). The effect of soil on biological nitrogen fixation, as evidenced by the number and weight of nodules, was mainly attributable to the available phosphorus content. Cate-Nelson ANOVA analysis established a critical value of soil test phosphorus (STP) of 20 mg P kg^–1 soil for nodule weight and number for the majority of cultivars. Within cultivars, nodulation varied with maturation period and was correlated with shoot NC. Thus, the overall interaction of soil-P content and cultivar-maturation period were correlated positively with nodulation and to symbiotic effectiveness of strains of Rhizobium leguminosarum bv. viceae indigenous to these soils. Based on an antibiotic susceptibility test and main variable factor analysis of the data obtained, 70 isolates of Rhizobia that nodulate pea, obtained from soils from agricultural sites throughout Tunisia, were identified as belonging to 18 distinct strains. These classes were identified on the basis of symbiotic efficiency parameters (shoot DM yield and shoot NC) as: ineffective (33 isolates), moderately effective (27 isolates), and efficient strains (10 isolates). This study shows that the Mateur site, an agricultural area for millennia in the northern region of Tunisia, harbors rhizobial strains that are highly efficient in fixing N₂ with peas. These results also indicate the importance of strain-cultivar interrelationships and specificity.     

Combined effects of soil properties and Phytophthora cinnamomi infections on Quercus ilex decline

Aims

The importance of soil properties as determinants of tree vitality and Phytophthora cinnamomi root infections was analysed.

Methods

The study comprised 96 declining stands in western Spain, where declining and non-declining holm oak (Quercus ilex L.) trees were sampled. Soil properties (soil depth, Ah horizon thickness, texture, pH, redox potential, soil bulk density and N-NH₄ ⁺ and N-NO₃ ^? concentrations) and P. cinnamomi infections were assessed.

Results

Tree mortality rates increased with low soil bulk densities, which were also associated with more P. cinnamomi-infected trees. Occurrence of infected trees was higher in fine textured soils and in thick Ah horizons. Fine textured soils favoured trees, but with the presence of P. cinnamomi their health status deteriorated. Soil under declining trees had higher N-NO₃ ^?/N-NH₄ ⁺ ratio values than under non-declining trees. Additional soil properties changes associated to grazing were not related to decline and P. cinnamomi infections.

Conclusions

The implications of P. cinnamomi in holm oak decline and the influence of soil properties as contributors to pathogen activity were demonstrated. Fine soil textures and thick Ah horizons, usually favourable for vigour and vitality of trees growing in the Mediterranean climate, were shown to be disadvantageous soil properties if P. cinnamomi was present. Fine soil textures and thick Ah horizons are frequently related with higher levels of soil moisture, which increase the inoculum of the pathogen and favours root infection. Grazing does not seem to be directly linked to Q. ilex health status or P. cinnamomi root rot.     

Enhanced rates of P700<Superscript>+</Superscript> dark-reduction in leaves of<Emphasis Type="Italic"> Cucumis sativus</Emphasis> L. photoinhibited at chilling temperature

The changes in electron transport within photosystem I (PSI) were studied in detached leaves of Cucumis sativus L. during the course of irradiation with moderate white light (300 mol photons m^–2 s^–1) at 4°C. When intact leaves were exposed to the combination of moderate light and low temperature, the amplitude of far-red light-induced P700 absorbance changes at 820 nm (A₈₂₀), a relative measure of PSI, progressively decreased as the light treatment time increased. Almost no oxidation of P700 was noticeable after 5 h. Methyl viologen accelerated the oxidation of P700 to a steady-state level and also increased the magnitudes of A₈₂₀ changes in photoinhibited leaves, reflecting the rapid removal of electrons from native carriers. Photoinhibition under moderate light and chilling temperature also accelerated the rate of P700⁺ reduction after far-red light excitation as the half-times of the two exponential components of P700⁺ decay curves decreased relative to the control ones. A detailed analysis of the kinetics of P700⁺ reduction using diuron alone or the combination of diuron and methyl viologen strongly favours an increased rate of electron donation from stromal reductants to PSI through the plastoquinone pool following photoinhibitory treatment. Importantly, the marked acceleration of P700⁺ re-reduction is the consequence of the irradiation of leaf segments at low temperature and not caused by chilling stress alone.Abbreviations A ₀ and A ₁ Primary acceptor chlorophyll and secondary electron acceptor phylloquinone - FR Far-red light - F _X , F _A , and F _B Iron–sulfur centers - MT Multiple-turnover flash - MV Methyl viologen - Ndh NAD(P)H-dehydrogenase - PQ Plastoquinone - PS Photosystem - P700 Reaction-center chlorophyll of PSI - ST Single-turnover flash     

Denitrification activity of Bradyrhizobium sp. isolated from Argentine soybean cultivated soils

Two hundred and fifty strains, all of them representatives of native Bradyrhizobium sp., isolated from soils cultivated with soybean have been characterized by their denitrification activity. In addition, the denitrification potential of those soils was also measured by evaluating the most-probable-number (MPN) of denitrifying bacteria and the denitrification enzyme assay (DEA). Of the 250 isolates tested, 73 were scored as probable denitrifiers by a preliminary screening method. Only 41 were considered denitrifiers because they produced gas bubbles in Durham tubes, cultures reached an absorbance of more than 0.1 and NO³⁻ and NO²⁻ were not present. Ten of these 41 were selected to confirm denitrification and to study denitrification genes. According to N₂O production and cell protein concentration with NO³⁻, the isolates could be differentiated in three categories of denitrifiers. The presence of the napA, nirK, norC and nosZ genes was detected by production of a diagnostic PCR product using specific primers. RFLP from the 16S-23S rDNA spacer region (IGS) revealed that denitrifiers strains could be characterized as Bradyrhizobium japonicum and strains which were non-respiratory denitrifiers as B. elkanii.