Isolation and partial characterization of culturable endophytic Arthrobacter spp. from leaves of maize (Zea mays L.).

The presence and diversity of the endophytic Arthrobacter species are relatively uninvestigated. We determined the genotypic and physiological diversity of the endophytic Arthrobacter spp. isolated from the interior leaf tissues of four out of six maize cultivars tested, all of which were collected under field conditions. Strains A7 and A9 isolated from Król cultivar and strain A23 isolated from KB1902 cultivar were identified as A. nicotinovorans. Strains A18, A22 and A34 identified as A. nitroguajacolicus were isolated from KB1903, KB1902 and Cyrkon cultivars respectively. Isolated strains tolerate the presence of o-coumaric, p-coumaric, chlorogenic, and ferulic acids in a concentration of at least two times higher than naturally occurring in maize tissue. The presence of the nitrogenase reductase gene nifH, a marker of biological nitrogen fixation, was determined in only two strains of A. nitroguajacolicus A18 and A34. None of the tested strains released compounds inhibiting in vitro the growth of the fungi Fusarium moniliforme and F. graminearum, pathogenic to maize. Seed inoculations with the A. nicotinovorans strain A7 suppressed the development of seedlings of four maize cultivars, which were not colonized by this species of bacteria, but stimulated the development of seedlings of the host-cultivar Król. Also, the A. nitroguajacolicus strain A18, which was applied as seed treatment, suppressed the development of seedlings of three maize cultivars, which were not colonized by this species of bacteria. The occurrence of endophytic A. nicotinovorans and A. nitroguajacolicus inhabiting interior tissues of maize leaves of specific maize cultivars is a first report in literature.

Lack of nifH gene in some endophytic bacteria isolated on RC solid medium.

Presence of endophytic bacteria was reported in many crops including maize (Zea mays L.). Endophytes play a significant role in plant nutrient and pesticide uptake. Application of endophytic bacteria is a goal of sustainable agriculture. Occurrence of Azospirillum strains is often reported as tissue inhabiting bacteria of maize. The biological N2-fixation is one of most important processes assigned to this bacteria. The objective of this study was to examine the biodiversity of Azospirillum spp. isolated from the leaves of 6 cultivars of Zea mays L.. They were cultivated on two experimental fields at Smolice and Kobierzyce, (Poland). Strains of Azospirillum spp. were isolated on the solid RC medium. Forty four isolates grown as a small intensive red colonies were selected. To verify ability to N2-fixation isolates were analyzed based on nifH gene presence. Presence of nifH gene was tested using PCR method with PolF and PolR universal degenerate primers. The presence of nifH gene was found in 6 tested strains isolated from leaves of 3 cultivars (Cyrkon, Kosmo230, KB2704) from Smolice location, only. Our results suggest that selection of Azospirillum-like strains on RC solid medium based on appearance of colony is not correlated with theirs ability to nitrogen fixation or used degenerated primers (PolF, PolR) are not universal enough.

A Methylobacterium-like organism from algal crusts covering silicone rubber electric insulators in Africa.

AIMS: The primary goals of this study were to isolate, identify and characterize culturable bacteria living in a close association with microalgae within green crusts covering silicone rubber electric insulators in Tanzania. METHODS AND RESULTS: Twenty-four bacterial colonies were isolated from an Apatococcus crust. Characterization by statistical analyses of total cellular protein profiles demonstrated that they were highly similar to one another. Final identification was achieved using 16S rDNA sequencing and fatty acid methyl ester profiling. These analyses revealed the presence of microbes with high similarity to Methylobacterium radiotolerans. The selected isolate, A1, displayed strong inhibitory activity against Rhizoctonia solani and was found to be resistant to relatively high concentrations of zinc in the growth medium. CONCLUSIONS: This study revealed the presence of M. radiotolerans bacteria in a novel environment--within algal crusts formed on electrical insulators in Africa. Moreover, this bacterium was found to be a predominant culturable species within those complex algal-microbial associations. The isolate also shared some traits of biotechnological importance with other members of the Methylobacterium genus. SIGNIFICANCE AND IMPACT OF THE STUDY: The data presented provide a valuable contribution concerning the formation and function of associations between green microalgae and bacteria. This study also provides some information about the utility of bacteria from the genus Methylobacterium in biotechnological applications, such as biocontrol of rhizoctoniosis and bioremediation of heavy metal-contaminated soils.

Protein fingerprinting as a complementary tool for the classification of Pseudomonas bacteria.

A collection of total 42 bacterial strains belonging to the genus Pseudomonas were characterised based on protein fingerprinting using sodium dodecyl sulphate polyacrylamide gel electrophoregrams of cell-free extracts. Densitometrical analysis revealed unique and distinct profiles characteristic of the studied species. This comparison differentiated the isolates into four main clusters and twelve subclusters. The obtained protein patterns have proved to be an effective and reliable method both for the classification of bacteria and for showing similarities and variability among them.

Production and secretion of 5-n-alkylresorcinols by Fusarium culmorum.

Fusarium culmorum F1 was found to produce and secrete into the culture medium several of 5-n-alkylresorcinols. The amount of resorcinolic lipids was 5.3 microg/g and 0.9 microg/l in mycelium and in post-culture liquid, respectively. First of all F. culmorum F1 produces saturated homologues with C15 to C25 side chains. The extract from the medium contained only homologues with shorter carbon chains (C13 to C17).

5-n-Alkylresorcinols from the green microalga Apatococcus constipatus.

Mixtures of six 5-n-alkylresorcinol (ARs) homologues were isolated from acetone extracts of four isolates of the unicellular green microalga Apataococcus constipatus. The pattern of homologues in different algal isolates was diverse. The predominant compounds were 1,3-dihydroxy-5-n-heneicosylbenzene (AR C(21:0)) and 1,3-dihydroxy-5-n-tricosylbenzene (AR C(23:0)) or 1,3-dihydroxy-nonadecylbenzene (AR C(19:0)), depending on the strain. ARs were identified by chromatographic and spectroscopic means.

Alkylresorcinols are abundant lipid components in different strains of Azotobacter chroococcum and Pseudomonas spp.

The occurrence of various amounts of 5-n-alkylresorcinols was shown in lipids extracted from 14 bacterial strains of Azotobacter chroococcum as well as from strains of Pseudomonas aureofaciens, P. chlororapsis, and P. fluorescens. The amount of alkylresorcinols found varied from 2.3 to 56.2 microg/mg (dry weight) of cells in A. chroococum and from 0.2 to 0.8 microg/mg (dry weight) of cells in Pseudomonas spp. Strains of both genera produce saturated homologs with C13 to C27 side chains. C19, C21, and C23 homologs are predominant in and characteristic for A. chroococum strains, the C15 homolog is predominant in and characteristic for P. chlororapsis and P. fluorescens, and the C17 homolog is predominant in and characteristic for P. aureofaciens. The presence of 5-n-(2-ketoalkyl)resorcinols, not previously observed, was demonstrated in lipids isolated from the cells of A. chroococum Az5.

Studies on antifungal properties of phenol complexes with secondary amines. 1. The effect of complex formation of nitro- and chlorophenols with dicyclohexylamine on the antifungal activity in vitro.

Inhibitory doses (ID50) of phenols complexed with dicyclohexylamine are lower (21.4-47.7%) than the inhibitory doses of free phenols. This is demonstrated in vitro by experiments with Aspergillus oryzae and Penicillium expansum on solid medium.

Studies on antifungal properties of phenol complexes with secondary amines. 2. Antifungal properties in vitro of the complex of 2,4,5-trichlorophenol with dicyclohexylamine.

In studies in vitro it has been found that 2,4,5-trichlorophenol in the form of a complex with dicyclohexylamine is two times more active an antifungal agent than free chlorophenol.

Interaction between bacterial metabolites and some pesticides. II. Change of phytotoxicity of the herbicide Roneet by the phenolic metabolites of Arthrobacter sp.

The bacteria from Arthrobacter genus isolated from sugar beet rhizosphere were found to produce gallic, protocatechuic, p-hydroxybenzoic, syryngic, vanillic, veratric acids, p-quinone and two unidentified phenolic compounds. The mixture of the bacterial phenolic compounds increased the phytotoxicity of Roneet, inhibiting the germination of wheat. Model experiments showed that the phenolic acids used with the herbicide Roneet increased its phytotoxicity and p-hydroxybenzoic acid was found to be the most active. It was found that in the presence of the phenolic acids, particularly of p-hydroxybenzoic acid, the herbicide content in the seeds of wheat was reduced. The UV and IR spectra of equimolar amounts of p-hydroxybenzoic acid and Roneet showed the hydrogen bond between H of carboxylic group of acid and O of C=O group of the Roneet molecule. It could be concluded that Roneet conjugated with p-hydroxybenzoic acid was more phytotoxic than the initial compounds.

Herbicidal effect on some metabolic processes of Azotobacter chroococcum.

The effect of pyramin (1-phenyl-4-amino-5-chloro-piridazone-6) and venzar (3-cyclohexyl-5,6-trimethyleneuracil) on growth, oxygen uptake and nitrogenase activity was examined. The possibilities of herbicide accumulation in cells and binding by mucopolisaccharides were also tested. Stimulation of growth was observed in the presence of pyramin (20 and 100 ppm) but no influence of venzar was found. Venzar was found to penetrate into the cells. Pyramin did not penetrate into the cells but its content decreased in cultures of Azotobacter strains. However, no degradation products of pyramin were found. Pyramin was detected in mucopolisaccharide fractions in range 0.2-47 micrograms of Pyrazon per 100 mg. Pyramin and venzar reduced the acetylene reduction rate of crude extracts by 50% when doses of active substances were equal to the weight of crude extract protein contents. Doses required for similar effect in living cells were however more than fourteen times higher. Addition of polivinylpyrrolidone and albumin protected the N2-ase from negative effect of pyramin and to a lesser degree from that of venzar.