The nitrogen requirements of Gluconobacter, Acetobacter and Frateuria.

The physiological role of D-aspartate oxidase (ChDASPO) in the yeast Cryptococcus humicola was analysed through the growth characteristics of a ChDASPO gene-disrupted strain (daspoDelta) and the expression profile of ChDASPO on various combinations of carbon and nitrogen sources. The daspoDelta strain, constructed by homologous integration of the yeast URA3 marker, grew as well as the wild-type strain on ammonium chloride, L-aspartate or D-alanine as the sole nitrogen source. In contrast, the daspoDelta strain did not grow at all on D-aspartate, not only as the sole nitrogen source but also as the sole carbon source or as the sole nitrogen and carbon source, and grew more slowly than the wild-type strain on D-glutamate as the sole nitrogen source. In the wild-type strain, the induction of ChDASPO activity strictly depended on the presence of D-aspartate and was little affected by the co-presence of ammonium chloride, but it was significantly reduced by the co-presence of both glucose and ammonium chloride, which, however, did not abolish the induction, allowing considerable expression of ChDASPO. This expression pattern was consistent with that shown by Northern blot analysis. The daspoDelta strain was more sensitive than the wild-type to the growth retardation by acidic D-amino acids, but not to that by the corresponding L-isomers or D-alanine. These results clearly show that in the yeast, DASPO plays an essential role in the assimilation of D-aspartate and acts as a detoxifying agent for D-aspartate.

Potential involvement of seed storage proteins in susceptibility to aflatoxin contamination was assessed with in vitro tests. Initially, two oilseed storage proteins [cottonseed storage protein (CSP) and zein] were compared with bovine serum albumin (BSA) and collagen. Supplementation of a complete defined medium with either oilseed storage protein resulted in significantly more aflatoxin production by Aspergillus flavus than supplementation with either BSA or collagen. Little or no aflatoxin was produced when either BSA, CSP, or zein was employed (at 0.5%) as both the sole carbon and the sole nitrogen source. Media with collagen (0.5%) as the sole nitrogen and carbon source supported aflatoxin production similar to the complete defined medium. Although lower than levels observed with defined medium, aflatoxin production increased with both increasing CSP concentration (0 to 2.0%) and increasing zein concentration (0 to 6.0%) when these proteins served as both the sole carbon and sole nitrogen source. Denaturing polyacrylamide gel electrophoresis and protease activity assays indicated that fungal acquisition of protein carbon was probably via hydrolysis mediated by the 35 kD metalloprotease of A. flavus. Media lacking nitrogen but containing sucrose (5.0%) and supplemented with either zein (1.7%) or CSP (2.0%) supported three‐ to eightfold more aflatoxin production than the complete defined medium. The results suggest seed storage proteins, when present with an accessible carbon source, may predispose oilseed crops to support production of high levels of aflatoxins by A. flavus during seed infection.

The degradation characteristics of pyridine and phenol by Rhodococcus sp.Chr-9 were studied.The results showed that strain Chr-9 could completely degrade 200 mg L-1 of pyridine or phenol respectively supplemented as sole carbon source within 72 hours in mineral salt medium.The optimal pH and temperature for pyridine degradation by strain Chr-9 were 8.0 and 35 ℃,respectively.The degradation rate of pyridine or phenol by strain Chr-9 was negatively related with its initial concentration in mineral salt medium.Strain Chr-9 could utilize pyridine as sole carbon and nitrogen source for growth,and the supplementation of phenol could enhance the strain growth when there was no other nitrogen source except pyridine in mineral salt media.Fig 7,Ref 12

Past handling practices associated with the manufacturing and processing of the high explosive hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) has resulted in extensive environmental contamination. In-situ biodegradation is a promising technology for remediating RDX contaminated sites but often relies on the addition of a cosubstrate. A sulfate-reducing bacterium isolated from an RDX-degrading enrichment culture was studied for its ability to grow on RDX as a sole source of carbon and nitrogen and for its ability to mineralize RDX in the absence of a cosubstrate. The results showed the isolate degraded 140 muM RDX in 63 days when grown on RDX as a carbon source. Biomass within the carbon limited culture increased 9-fold compared to the RDX unamended controls. When the isolate was incubated with RDX as sole source of nitrogen it degraded 160 muM RDX in 41 days and exhibited a 4-fold increase in biomass compared to RDX unamended controls. Radiolabeled studies under carbon limiting conditions with (14)C-hexahydro-1,3,5-trinitro-1,3,5-triazine confirmed mineralization of the cyclic nitramine. After 60 days incubation 26% of the radiolabel was recovered as (14)CO(2), while in the control bottles less than 1% of the radiolabel was recovered as (14)CO(2). Additionally, approximately 2% of the radiolabeled carbon was found to be associated with the biomass. The 16S rDNA gene was sequenced and identified the isolate as a novel species of Desulfovibrio, having a 95.1% sequence similarity to Desulfovibrio desulfuricans. This is the first known anaerobic bacterium capable of mineralizing RDX when using it as a carbon and energy source for growth.

The utilization of amino acids for growth and their effects on nitrogen fixation differ greatly among the several strains of each species of Azospirillum spp. that were examined. A. brasiliense grew poorly or not at all on glutamate, aspartate, serine, or histidine as the sole nitrogen and carbon sources. Nitrogen fixation by most A. brasiliense strains was inhibited only slightly even by 10 mM concentrations of these amino acids. In contrast, A. lipoferum and A. amazonense grew very well on glutamate, aspartate, serine, or histidine as the sole nitrogen and carbon sources; nitrogen fixation, which was measured in the presence of malate or sucrose, was severely inhibited by these amino acids. It was concluded that growth on histidine as the sole source of nitrogen, carbon, and energy may be used for the taxonomic characterization of Azospirillum spp. and for the selective isolation of A. lipoferum. The different utilization of various amino acids by Azospirillum spp. may be important for their establishment in the rhizosphere and for their associative nitrogen fixation with plants. The physiological basis for the different utilization of glutamate by Azospirillum spp. was investigated further. A. brasiliense and A. lipoferum exhibited a high affinity for glutamate uptake (Km values for uptake were 8 and 40 microM, respectively); the Vmax was 6 times higher in A. lipoferum than in A. brasiliense. At high substrate concentrations (10 mM), the nonsaturable component of glutamate uptake was most active in A. lipoferum and A. amazonense.(ABSTRACT TRUNCATED AT 250 WORDS)

A defined medium for Aquaspirillum serpens VHL allows the replacement of the complex media now in use. It was developed by batch culture methods but supports growth in continuous culture. A basal salts medium supplemented with L-aspartic acid, L-alanine, and L-glutamic acid provided the best growth (turbidity), as long as ammonium chloride was omitted. Ammonium chloride caused either a lag or a reduction or a complete inhibition of the growth of A. serpens VHL on the above amino acids and other organic supplements depending on the combination used. Ammonium sulfate and ammonium hydroxide with L-glutamic acid allowed growth, but the lag period was increased in shake flask cultures. Vitamins, cysteine hydrochloride, and carbon dioxide had no effect on the growth rate. Viability (less than 50%) was inadequate to maintain continuous culture with L-glutamic acid as the sole source of carbon and nitrogen. Combinations of amino and carboxylic acids were then tested and, of these, L-glutamic acid (1 g/liter) and L-histidine (75 mg/liter) without ammonium chloride in the basal salts medium supported growth in batch and continuous culture. L-Glutamic acid was the limiting substrate for growth.

48] Pantothenase from Pseudomonas fluorescens

Nutrition and metabolism of marine bacteria. V. The inhibition of growth of a marine bacterium by amino acids and the development of resistant strains

Collecting the soil which had applied Atrazine for more then 5 years from The Xiang Fang experimental farm of HarBin and the corn field of WuChang in Heilongjiang Province. We isolated 7 microbes which have the ability of Atrazine degrading by the way of the traditional enrichment and isolation of bacteria in the laboratory, then we filtrated 2 microbes which the ability of atrazine degrading was relatively higher and named J2 and J6. They were inoculated into 20 mL the corresponding medium which the concentration of Atrazine was 100 mg/L by 5% percent of the medium, they could degradated the Atrazine completely in 5 days and 4 days respectively. According to the condition of enrichment and isolation, we known that J2 used Atrazine as the sole nitrogen source and J6 used Atrazine as the sole carbon and nitrogen source. The preliminary identifications indicated that J2 belonged to Microbacterium sp and J6 belonged to Aureobacterium.

The present investigation is concerned with l-glutamic acid production in the presence of pyrrolidone carboxylic acid and glucose in Bacillus megaterium st. 6126. This strain does not grow on dl-pyrrolidone carboxylic acid (dl-PCA)1) as the sole source of carbon and nitrogen. The optimal concentration of yeast extract required for the maximal production of l-glutamic acid was 0.005% under the conditions used. As the yeast extract concentration was increased, growth increased proportionally; but the l-glutamic acid production did not exceed the control’s to which glucose and ammonium chloride had been added. l-Glutamic acid produced by both growing cultures and resting cells was derived from glucose and ammonium salt of dl-PCA. Isotope experiments suggested that the l-glutamic acid produced was partially derived from ammonium salt of dl-PCA in the growing culture which had been supplemented with d-glucose-U-14C or dl-PCA-1-14C and that ammonium salt of dl-PCA was consumed as the source of nitrogen and carbon for l-glutamic acid.

Growth of moderately thermophilic iron-oxidizing bacterium strain TI-1 in synthetic medium

Cultures of Streptomyces lividans TK24 grown in defined media containing certain rapidly used carbon and nitrogen sources excreted high levels of organic acids. These were identified by HPLC and enzymic assays as pyruvic acid and 2-oxoglutaric acid. Acidification occurred only with glucose as the principal carbon source, and depended on the nitrogen source used. With nitrate as the sole nitrogen source, high levels of pyruvate and small amounts of 2-oxoglutarate were produced. Carbon from D[U-14C]glucose was converted into both organic acids. Combining glucose with a selection of amino acids as primary nitrogen/secondary carbon sources yielded less pyruvate and more 2-oxoglutarate. Carbon from both 14C-labelled glucose and amino acids was metabolized to both organic acids. Adding nitrate to this combination caused a reversion of the acid production pattern to that of the glucose-nitrate combination, as if the amino acids were absent. Addition of ammonium salts to any combination of carbon and nitrogen sources completely prevented organic acid formation.

The nitrogen utilization pattern of two spe? cies of Leucostoma was examined using virulent isolates of L. persoonii and L. cincta, and two virus-infected hypovirulent isolates of L. persoonii. Virulent L. per? soonii were able to utilize all of 47 nitrogen sources tested except anthranilic acid, cysteine, homocysteine, hydroxylamine, indole, lysine, urocanic acid, methyl- ated purines and pyrimidines. Leucostoma cincta isolates were unable to use these latter compounds, acetamide and the branched side chain aliphatic amino acids: isoleucine, leucine, and valine. The ability to utilize branched aliphatic amino acids might be a species- specific characteristic. A virulent isolate of L. cincta, ATCC62910, also could not utilize nitrate and nitrite. The two hypovirulent L. persoonii isolates had a further restricted list of usable nitrogen sources. In addition to the nitrogen sources above, they were unable to grow on nitrate, nitrite and the branched aliphatic amino acids, as sole nitrogen sources. Growth of the two hypovirulent isolates on glutamic acid and aspartic acid, as well as aromatic amino acids was poor com? pared to virulent isolates. The patterns of inorganic nitrogen utilization were determined in reference to the known gene loci that control the process in fungi. Inability to utilize specific nitrogen sources is likely the result of mycovirus infection. Poor growth on sev? eral utilizable nitrogen sources might be influential in hypovirulence. trees, dieback of young branch tips greatly reduces vigor, and large perennial cankers in older trees often lead to death (Hildebrand, 1947; Rozsanyay, 1977). As fungicides have proven to be ineffective in con? trolling this disease (Biggs, 1989; Chandler, 1974; Hel- ton and Rohrbach, 1967; Royse and Reis, 1978), a biological control agent that could reduce the impact of this disease has been sought. A diseased isolate of L. persoonii (NC14.4A) has been isolated from peach in North Carolina. Isolate NC14.4A is a hypovirulent nonsporulating isolate that contains many virus like particles (VLPs) in its cytoplasm when viewed in the electron microscope (Snyder et al., 1989). The strain contains at least six segments of dsRNA. Isolate HT is derived from isolate NC14.4A by successive hyphal tipping and has lost the viral particle and four of the six bands of dsRNA. HT has wild-type growth rates and can sporulate, but retains hypovirulence (Ham-

Neptunomonas sp. BPy-1 is an epiphytic bacterium isolated from in vitro culture of the red alga Pyropia yezoensis. It uses ethanol as a sole carbon source and promotes the growth of host alga. A related bacterium, Neptunomonas sp. BZm-1, was isolated from leaves of Zostera marina found in the Yatsushiro Sea (Japan). BZm-1 showed 99% 16S rRNA sequence identity with Neptunomonas sp. BPy-1. Similar to BPy-1, BZm-1 grew in artificial seawater (ASW) medium containing ethanol or butanol. When thalli were treated with a multi-enzyme cleaner, the growth of treated thalli was retarded, but the addition of BZm-1 to the medium promoted growth. To explore the benefits of epiphytic bacteria, indoleacetic acid (IAA) production by isolated bacteria was examined under conditions of limited nutrients. Salkowski assays and GC-MS analysis revealed that both BZm-1 and BPy-1 excreted IAA during growth in ASW medium containing glucose or ethanol in the presence of tryptophan. In ASW medium containing tryptophan but lacking a carbon source, neither isolate grow, but produced IAA. ASW medium includes nitrate as the sole nitrogen source. In the absence of carbon source, different nitrogen forms in the presence of tryptophan did not affect IAA production by the two isolates. These findings indicate that IAA production by the two isolates is strictly dependent on tryptophan but less affected by carbon and nitrogen sources. Based on the different origins of BPy-1 and BZm-1, this mode of IAA production seems to be conserved among relatives of BPy-1.

The amdS gene of Aspergillus nidulans encodes an acetamidase enzyme that hydrolyzes acetamide and a limited range of other amides. This allows acetamide to be used as a sole source of nitrogen or as a source of carbon. As far as we know, the function of this gene is of no great intrinsic interest with respect to the physiology or development of A. nidulans. Deletion mutants of amdS have no detectable phenotype other than an inability to catabolize acetamide (Hynes 1979). It should also be noted that acetamide is a very poor sole source of carbon or nitrogen, and is unlikely to be a significant source of metabolites in the natural environment. However, the study of the amdS gene and its regulation has provided much information regarding mechanisms of gene regulation in fungi, as well as insights into how catabolic pathways are coordinated. Furthermore, the amdS gene has been of considerable importance in the development of fungal transformation systems (Tilburn et al. 1983; Kelly and Hynes 1985; Fincham 1989). In this chapter we discuss in detail the genetical and molecular analysis of the amdS gene and how this has contributed to the field of fungal molecular genetics.