Efficiency of indoleacetic acid,gibberellic acid and ethylene synthesized in vitro by Fusarium culmorum strains with different effects on cereal growth

Three different Fusarium culmorum strains having a pathogenic, a deleterious (deleterious rhizosphere microorganism), or a promoting (plant growth promoting fungus) effect on plant growth were studied for their ability to synthesize in vitro the phytohormones indoleacetic acid (IAA), gibberellic acid (GA), and ethylene. All the phytohormones tested were synthesized in cultures supplemented with wide concentration ranges of glucose and tryptophan or methionine (precursors of phytohormone synthesis). The amounts of these secondary metabolites synthesized by the particular strains were found to be significantly different. The non-pathogenic PGPF strain (DEMFc2) synthesized the highest amounts of IAA and GA, a fact that could be responsible for the growth-promoting properties of this strain. A pathogenic strain synthesized the highest amount of ethylene, which could be responsible for the negative effect of this strain on plant growth. F. culmorum isolates with a high capacity for IAA synthesis also have a high capacity for GA synthesis and irrespective of the growth conditions, a high positive correlation (R > 0.9) between the concentrations of synthesized IAA and GA in F. culmorum cultures was found. It is worth mentioning that the optimal conditions for the growth of F. culmorum isolates and the synthesis of the individual phytohormones differed from one another. The optimal growth conditions were 1.0% of glucose and 9.9 mM of methionine or 6.0 mM of tryptophan. The optimal conditions for ethylene synthesis were 0.5% of glucose and 6.6 mM of methionine, whereas 1.0% of glucose and 9.0 mM of tryptophan were optimal for IAA and GA synthesis.     

Regulation of the shikimate pathway of carrot cells in suspension culture

The activity of the first enzyme of the shikimate pathway, 3-deoxy-d-arabino-heptulosonate 7-phosphate synthase, is demonstrated in extracts of Daucus carota cells grown in suspension culture. Maximum specific enzyme activity is found midway through the logarithmic growth of the culture; cells in lag and stationary phases of growth have lower enzyme levels. The enzyme is activated by tyrosine and tryptophan. The extent of activation varies during cell growth.     

Prey-Specific Growth Responses of Freshwater Flagellate Communities Induced by Morphologically Distinct Bacteria from the Genus Limnohabitans

Because their large growth potential is counterbalanced with grazing by heterotrophic nanoflagellates (HNF), bacteria of the genus Limnohabitans, which are common in many freshwater habitats, represent a valuable model for examining bacterial carbon flow to the grazer food chain. We conducted experiments with natural HNF communities taken from two distinct habitats, the meso-eutrophic Římov Reservoir and the oligo-mesotrophic Lake Cep (South Bohemia). HNF communities from each habitat at distinct seasonal phases, a late April algal bloom and a late May clear water phase, were each fed 3 Limnohabitans strains of differing cell sizes. Water samples were prefiltered (5 μm) to release natural HNF communities from zooplankton control and then amended with the Limnohabitans strains L. planktonicus II-D5 (medium sized, rod shaped), Limnohabitans sp. strain T6-5 (thin, long, curved rod), and Limnohabitans sp. strain 2KL-3 (large solenoid). Using temporal sampling and prey treatment, we determined HNF growth parameters such as doubling time, growth efficiency, and length of lag phase prior starting to exponential growth. All three Limnohabitans strains supported HNF growth but in significant prey-, site-, and season-dependent fashions. For instance, addition of the moderately large T6-5 strain yielded very rapid HNF growth with a short lag phase. In contrast, the curved morphology and larger cell size of strain 2KL-3 made this prey somewhat protected against grazing by smaller HNF, resulting in slower HNF growth and longer lag phases. These trends were particularly pronounced during the late May clear-water phase, which was dominated by smaller HNF cells. This may indicate a longer “adaptation time” for the flagellate communities toward the large prey size offered.     

Customized media based on miniaturized screening improve growth rate and cell yield of methane-oxidizing bacteria of the genus Methylomonas

The growth of twelve methanotrophic strains within the genus Methylomonas, including the type strains of Methylomonas methanica and Methylomonas koyamae, was evaluated with 40 different variations of standard diluted nitrate mineral salts medium in 96-well microtiter plates. Unique profiles of growth preference were observed for each strain, showing a strong strain dependency for optimal growth conditions, especially with regards to the preferred concentration and nature of the nitrogen source. Based on the miniaturized screening results, a customized medium was designed for each strain, allowing the improvement of the growth of several strains in a batch setup, either by a reduction of the lag phase or by faster biomass accumulation. As such, the maintenance of fastidious strains could be facilitated while the growth of fast-growing Methylomonas strains could be further improved. Methylomonas sp. R-45378 displayed a 50 % increase in cell dry weight when grown in its customized medium and showed the lowest observed nitrogen and oxygen requirement of all tested strains. We demonstrate that the presented miniaturized approach for medium optimization is a simple tool allowing the quick generation of strain-specific growth preference data that can be applied downstream of an isolation campaign. This approach can also be applied as a first step in the search for strains with biotechnological potential, to facilitate cultivation of fastidious strains or to steer future isolation campaigns.     

Regulation of the Pathway for the Degradation of Anthranilate in Aspergillus niger

Studies were carried out to determine the factors governing the induction of anthranilate hydroxylase and other enzymes in the pathway for the dissimilation of anthranilate by Aspergillus niger (UBC 814). The enzyme was induced by growth in the presence of tryptophan, kynurenine, anthranilate, and, surprisingly, by 3-hydroxyanthranilate, which was not an intermediate in the conversion of anthranilate to 2,3-dihydroxybenzoate. There was an initial lag in the synthesis of anthranilate hydroxylase when induced by tryptophan, anthranilate, and 3-hydroxyanthranilate. Cycloheximide inhibited the enzyme induction. Comparative studies on anthranilate hydroxylase, 2,3-dihydroxybenzoate carboxy-lyase, and catechol 1:2-oxygenase revealed that these enzymes were not coordinately induced by either anthranilate or 3-hydroxyanthranilate. Structural requirements for the induction of anthranilate hydroxylase were determined by using various analogues of anthranilate. The activity of the constitutive catechol oxygenase was increased threefold by exposure to anthranilate, 2,3-dihydroxybenzoate, or catechol. 3-Hydroxyanthranilate did not enhance the levels of catechol oxygenase activity.     

Extracellular trehalose utilization by Saccharomyces cerevisiae

Two haploid strains of Saccharomyces cerevisiae viz. MATα and MATa were grown in glucose and trehalose medium and growth patterns were compared. Both strains show similar growth, except for an extended lag phase in trehalose grown cells. In both trehalose grown strains increase in activities of both extracellular trehalase activities and simultaneous decrease in extracellular trehalose level was seen. This coincided with a sharp increase in extracellular glucose level and beginning of log phase of growth. Alcohol production was also observed. Secreted trehalase activity was detected, in addition to periplasmic activity. It appeared that extracellular trehalose was hydrolyzed into glucose by extracellular trehalase activity. This glucose was utilized by the cells for growth. The alcohol formation was due to the fermentation of glucose. Addition of extracellular trehalase caused reduction in the lag phase when grown in trehalose medium, supporting our hypothesis of extracellular utilization of trehalose.     

A study of the growth kinetics of Yersinia enterocolitica serotype O:3 in pure and meat culture systems

The growth kinetics of a virulence plasmid-bearing (P+) and a plasmid-cured (P−) strain of Yersinia enterocolitica serotype O:3 in pure and meat culture were investigated. Growth studies were carried out at 25 and 37 °C in supplemented phosphate-buffered saline, buffered peptone water , cefsulodin-irgasan-novobiocin broth base or supplemented broth base (CIN). The lag phase durations and growth rates under these conditions were determined by linear regression analysis. In pure culture, under most sets of equivalent conditions, P+ and P− strains had similar lag phase durations. However, under one set of conditions, i.e. CIN broth at 37 °C, the lag phase duration of the P+ strain was significantly longer than P−. In all but the most selective medium, P+ strains had slower growth rates than P− strains at 37 °C, probably due to the increased metabolic burden entailed in the maintenance of the virulence plasmid. In the most selective medium, i.e. CIN broth, P+ strains grew significantly faster than P−. This finding suggests that possession of virulence plasmid confers an enhanced ability to grow in the presence of selective agents. In meat cultures, both strains had longer lag phases than in equivalent pure cultures, with longer lag phases noted at 37 than at 25 °C. No significant differences were observed between the length of lag phases of P+ and P− strains in meat culture. Both strains of Y. enterocolitica displayed faster growth rates in meat cultures than in pure cultures, indicating that one or more components of meat enhanced the growth of this organism. The effects and interaction of incubation temperature, enrichment broth and meat on the growth kinetics of plasmid-bearing and plasmid-cured Y. enterocolitica strains are discussed.     

Development of Competence in the Bacillus subtilis Transformation System

Competence in Bacillus subtilis, assayed by the ability of cells to be transformed with bacterial deoxyribonucleic acid (DNA) or transfected by phage DNA, has been shown to occur in a single semisynthetic medium with peak activity occurring 3 hr after the cessation of logarithmic growth. No step-down conditions or culture manipulations were necessary for routine transfection of 1% of the population. The results demonstrate that bacteriophage DNA is a valid assay for studying the development of competence in B. subtilis. Predictions of workers using transforming bacterial DNA, who have suggested that competence in B. subtilis is associated with a specific phase of growth, are substantiated. The peak of competence is not affected by marked differences in the rate of growth during the logarithmic phase. The effect on development of competence by this procedure of some components (including casein hydrolysate, tryptophan, and histidine) which were routinely included in the transformation medium by other investigators has been determined by use of infectious phage DNA as an assay. We have demonstrated that tryptophan, as well as histidine, increases the transformation frequency—even in strains which do not have auxotrophic demands for these components. Glutamic acid and alanine depress optimal levels of transfection.     

Mathematical description of the growth of Lactobacillus sake and Lactobacillus pentosus under conditions prevailing in fermented sausages

The growth behaviour of Lactobacillus sake and Lactobacillus pentosus was determined in a model system simulating the conditions of fermenting sausages. Minor effects on the growth were observed by varying the concentrations of glucose, peptone, manganese and sodium nitrite. Temperature and sodium chloride concentration were found to have most effect on the growth. The measured data were used for a mathematical model describing the growth response of L. sake and L. pentosus sufficiently well to estimate the behaviour of the investigated strains. In all combinations relevant to sausage fermentation L. sake proved to be more competitive. It exhibited a shorter lag phase, higher maximal growth rate and higher final cell yield than L. pentosus.     

On-off regulation of the tryptophan promoter in fed-batch culture

Fundamental properties of the trp promoter were investigated in fed-batch culture using a recombinant containing the lacZ gene controlled by this promoter. In tryptophan-deficient conditions, the amount of β-galactosidase accumulated in the cell was 10% of total cellular proteins. In the presence of the amino acid, it was repressed at a lower level, but considerable expression was observed in the later stages of cultivation. Although increasing concentration of tryptophan seemed to repress the promoter more completely, it strongly inhibited the bacterial growth. On-off regulation of the promoter was achieved by controlling the tryptophan level during fed-batch culture.     

Characterization of marine bacteria highly resistant to mercury exhibiting multiple resistances to toxic chemicals

Several strains of bacteria unusually highly resistant to mercury were isolated from seawater and marine sediment samples and identified by 16S rDNA sequencing and were also characterized by a battery of biochemical and morphological tests. The bacterial isolates were identified to belong to the genera Pseudomonas, Alcaligenes, Brevibacterium and Bacillus. Many of the chosen isolates were tested for growth in the presence of different heavy metals and a variety of xenobiotics. Growth curves of all six bacteria highly resistant to mercury examined for growth at different concentrations of Hg exhibited prolonged lag phase, during which time necessary physiological adaptations to toxic milieu were undergone. All the strains tested for antibiotic resistance showed little to no effect of antibiotics on their normal growth. Results of this study demonstrate the occurrence of diverse groups of marine prokaryotes capable of high tolerance to mercury with a potential to degrade a variety of toxic heavy metals and xenobiotics.     

Comparison of growth,nutritional utilisation patterns,and niche overlap indices of toxigenic and atoxigenic Aspergillus flavus strains

The effects of temperatures (20–30 °C) and water activity (0.90–0.99 a_w) on the lag phase duration, mycelial growth, and nutritional utilisation patterns of two toxigenic (AFL1⁺ & AFL2⁺) and three atoxigenic (AFL1⁻, AFL2⁻, & AFL3⁻) Aspergillus flavus strains were evaluated in vitro. Both temperature and a_w and their interactions had a significant influence on the growth and nutritional utilisation patterns (p < 0.05). There were no significant differences between toxigenic and atoxigenic strains in terms of lag phase prior to growth and mycelial growth rates. Based on carbon source (CS) utilisation patterns, toxigenic and atoxigenic strains' niche size was greater at higher temperatures and in wetter conditions. Additionally, based on niche overlap indices (NOIs), regardless of temperature, when water was freely available, atoxigenic and toxigenic strains co-existed. However, under moisture stress, the nutritional competitiveness was variable. Temporal carbon utilisation sequences (TCUS) of toxigenic and atoxigenic strains were compared. At 0.99 a_w most CS sources were utilised by the strains and the time to detection (TTD) of each strain was shortest on monosaccharides at the same level of a_w. Conversely, under moisture stress the least number of CS was utilised. The current study has demonstrated that carbon utilisation patterns are equally important as are other determinants of competitiveness and that growth rate alone is not a key attribute which determines competitiveness.     

Magnesium requirement of some of the principal rumen cellulolytic bacteria

Information available on the role of Mg for growth and cellulose degradation by rumen bacteria is both limited and inconsistent. In this study, the Mg requirements for two strains each of the cellulolytic rumen species Fibrobacter succinogenes (A3c and S85), Ruminococcus albus (7 and 8) and Ruminococcus flavefaciens (B34b and C94) were investigated. Maximum growth, rate of growth and lag time were all measured using a complete factorial design, 2(3)×6; factors were: strains (2), within species (3) and Mg concentrations (6). R. flavefaciens was the only species that did not grow when Mg was singly deleted from the media, and both strains exhibited a linear growth response to increasing Mg concentrations (P<0.001). The requirement for R. flavefaciens B34b was estimated as 0.54 mM; whereas the requirement for R. flavefaciens C94 was >0.82 as there was no plateau in growth. Although not an absolute requirement for growth, strains of the two other species of cellulolytic bacteria all responded to increasing Mg concentrations. For F. succinogenes S85, R. albus 7 and R. albus 8, their requirement estimated from maximum growth was 0.56, 0.52 and 0.51, respectively. A requirement for F. succinogenes A3c could not be calculated because there was no solution for contrasts. Whether R. flavefaciens had a Mg requirement for cellulose degradation was determined in NH₃-free cellulose media, using a 2×4 factorial design, 2 strains and 4 treatments. Both strains of R. flavefaciens were found to have an absolute Mg requirement for cellulose degradation. Based on reported concentrations of Mg in the rumen, 1.0 to 10.1 mM, it seems unlikely that an in vivo deficiency of this element would occur.     

Comparison of the adaptive potential of the Arthrobacter oxydans and Acinetobacter lwoffii isolates from permafrost sedimentary rock and the analogous collection strains

A comparative study was conducted on the adaptive mechanisms of the strains Arthrobacter oxydans K14 and Acinetobacter lwoffii EK30A isolated from permafrost subsoil sediments and of those of the analogous collection strains (Ac-1114 Type and BSW-27, respectively). In each pair of the strains compared, the strains differed in terms of (i) growth-related, physiological, and biochemical properties; (ii) resistance to stress factors; (iii) capacity for generation of dormant forms (DFs) under growth arrest conditions, and (iv) intrapopulation production of phase variants. The strains isolated from permafrost displayed a lower growth rate but were more resistant to repeated freezing-thawing treatment than the collection strains. Under the same growth conditions, the permafrost strains formed larger numbers of cystlike anabiotic DFs, extraordinarily small cells, and forms that became nonculturable during long-term storage. Resuscitation of the nonculturable forms resulted in a 2- to-7-fold increase in the percentage of FISH-detectable metabolically active cells. The permafrost strains were also distinguished by increased genome lability. This facilitated their dissociation into intrapopulation variants with phenotypically distinct colonial and morphological properties and different antibiotic resistance. The phenotypic variability was more prominent in Arthrobacter (for which it was not reported previously) than in Acinetobacter. In the populations produced by plating the dormant bacterial forms, the qualitative and quantitative characteristics of the phase variant spectra varied depending on the formation conditions and the composition of the solid media used for the plating. Thus, the permafrost isolates of A. oxydans and Ac. lwoffii were distinguished from their collection analogs by a more manifest adaptive potential including stress resistance, the intensity of DF generation under growth arrest conditions, and increased intrapopulation variability.     

Physiological and enzymatic aspects of histidine-mediated control of the tryptophan pathway

Summary It would thus appear that in Saccharomyces cerevisiae there are two forms of histidine-mediated control on the tryptophan pathway. In some strains histidine increases anthranilate synthetase and indole glycerol phosphate synthetase activities, while tryptophan synthetase decreases. In other strains histidine affects coordinately all enzymatic activities involved in tryptophan biosynthesis. The two groups of strains also differ in the formation, during the growth of the enzymatic activities involved in tryptophan biosynthesis. This difference in the relative rates at which the two enzymes are formed may explain the accumulation of intermediates in the cultural media of some strains. The derepression of anthranilate synthetase and indole glycerol phosphate synthetase activities by histidine is particularly manifest in the auxotrophic his₃ strains that show these activities very depressed in histidine starvation; large amounts of this amino acid stimulate them to a considerably greater extent than in prototrophic strains.Abbreviations IGP imidazole glycerol phosphate - InGP indole glycerol phosphate - ASase anthranilate synthetase - InGPase indole-3-glycerol phosphate synthetase - TSase tryptophan synthetase - Tris tris (hydroxymethyl)-aminomethane This investigation was supported by a research grant of C.N.R. (Consiglio Nazionale delle Ricerche, Roma).     

Microorganisms and Maillard reaction products: a review of the literature and recent findings

Research on the impact of Maillard reaction products (MRPs) on microorganisms has been reported in the literature for the last 60 years. In the current study, the impact of an MRP-rich medium on the growth of three strains of Escherichia coli was measured by comparing two classic methods for studying the growth of bacteria (plate counting and optical density at 600 nm) and by tracing MRP utilisation. Early stage and advanced MRPs in the culture media were assessed by quantifying furosine and N ^ε -carboxymethyllysine (CML) levels, respectively, using chromatographic methods. These measures were performed prior to and during bacterial growth to estimate the potential use of these MRPs by Escherichia coli CIP 54.8. Glucose and lysine, the two MRP precursors used in the MRP-rich medium, were also quantified by chromatographic means. Compared to control media, increased lag phases and decreased growth rates were observed in the MRP-rich medium for two out of the three Escherichia coli strains tested. In contrast, one strain isolated from the faeces of a piglet fed on a MRP-rich diet was not influenced by the presence of MRPs in the medium. Overall, CML as well as the products obtained by the thermal degradation of glucose and lysine, regardless of the Maillard reaction, did not affect the growth of the three strains tested. In addition, no degradation of fructoselysine or CML was found in the presence of Escherichia coli CIP 54.8.     

Fitness of Mycobacterium tuberculosis Strains of the W-Beijing and Non-W-Beijing Genotype

Background

Multidrug resistant tuberculosis (MDR-TB) is a major threat for global tuberculosis control. The W-Beijing Mycobacterium tuberculosis genotype has been associated with drug resistance. Elucidation of the mechanisms underlying this epidemiological finding may have an important role in the control of MDR-TB. The aim of this study was to evaluate the fitness of drug-susceptible and MDR M. tuberculosis strains of the W-Beijing genotype compared with that of Non-W-Beijing strains.

Methodology/Principal Findings

Fitness of M. tuberculosis strains was determined by evaluating the difference in the growth curves obtained in the MGIT960 automated system and assessing the competitive growth capacity between W-Beijing and non-W-Beijing strains. The W-Beijing MDR strains had a significant longer lag phase duration compared to the other strains but did not present a significant fitness cost. When grown in competition they had an advantage only in medium containing 0.1% Tween 80.

Conclusions/Significance

It was not possible to confirm a selective advantage of W-Beijing strains to grow, except for differences in their resistance to Tween 80. Further studies are needed to elucidate the putative advantage of W-Beijing strains compared to other genotypes.     

Taurine catabolism: III. Evidence for the participation of the glyoxylate cycle

The metabolic fate of acetate, produced during taurine catabolism in Pseudomonas aeruginosa TAU-5, appear to involve the glyoxylate cycle. Organisms grown on taurine have significantly higher levels of malate synthetase and isocitrate lyase than cells grown on nutrient broth, but were comparable to the levels found in acetate-grown organisms. Itaconate, an isocitrate lyase inhibitor, produced a prolonged lag phase and reduced the growth rate of organisms when it was present in the taurine or acetate growth medium. Ethylmethanesulfonate treatment of TAU-5 yielded mutant strains unable to grow on taurine or acetate as sole carbon sources, due to a lack of either malate synthetase or isocitrate lyase. Spontaneous revertants derived from these mutant strains regained the missing enzyme activity and the ability to grow on taurine or acetate.     

Molecular analysis of intragenic recombination at the tryptophan synthetase locus in Neurospora crassa

Fifteen different classically generated and mapped mutations at the tryptophan synthetase locus in Neurospora crassa have been characterized to the level of the primary sequence of the gene. This sequence analysis has demonstrated that intragenic recombination is accurate to order mutations within one open reading frame. While classic genetic analysis correctly ordered the mutations, the position of mutations characterized by gene sequence analysis was more accurate. A leaky mutation was found to have a wild-type primary sequence. The presence of unique polymorphisms in the primary sequence of the trp-3 gene from strain 861 confirms that it has a unique history relative to the other strains studied. Most strains that were previously shown to be immunologically nonreactive with antibody preparations raised against tryptophan synthetase protein were shown to have nonsense mutations. This work defines 14 alleles of the N. crassa trp-3 gene.