Isolation and characterization of the symbiotic phenotype of antibiotic-resistant mutantsof Frankia from Rhamnaceae

Induced rifampicin-resistant mutants of Frankia were isolated by treatment of spores from strain ChI1 with 2 mg/ml of nitrosoguanidine. The mutagenic treatment was followed by growth for 72 h on non-selective medium to allow the expression of the mutation, before plating on selective medium. Spontaneous rifampicin-resistant mutants were isolated from strain ReI4 and chloramphenicol-resistant mutants were derived from strains ReI6 and TtI42. The mutants grew in medium containing up to 20 μg/ml of antibiotics. They showed similar morphology and growth pattern compared with their parent strains. However, they exhibited differences in symbiotic properties, such as infectivity and nitrogenase activity, from their parent strains.     

Isolation and characterization of the symbiotic phenotype of antibiotic-resistant mutantsof Frankia from Rhamnaceae

Induced rifampicin-resistant mutants of Frankia were isolated by treatment of spores from strain ChI1 with 2 mg/ml of nitrosoguanidine. The mutagenic treatment was followed by growth for 72 h on non-selective medium to allow the expression of the mutation, before plating on selective medium. Spontaneous rifampicin-resistant mutants were isolated from strain ReI4 and chloramphenicol-resistant mutants were derived from strains ReI6 and TtI42. The mutants grew in medium containing up to 20 g/ml of antibiotics. They showed similar morphology and growth pattern compared with their parent strains. However, they exhibited differences in symbiotic properties, such as infectivity and nitrogenase activity, from their parent strains.     

变质甘蔗中毒病原菌节菱孢的分类鉴定

对163株从中毒甘蔗和变质甘蔗样品中分离的产毒节菱孢进行了分类鉴定,其中98株定名为甘蔗节菱孢(Arthrinium sacchari M.B.Ellis),占60.1％;43株定名为蔗生节菱孢(A.saccharicola Stevenson),占26.4％;22株为暗孢节菱孢[A.phaeospermum(Corda)M.B.Ellis],占13.5％。前两个种为国内新记录。并用小培养法及扫描电镜观察了分生孢子的形态及着生特点。     

Lack of Chemically Induced Mutation in Repair-Deficient Mutants of Yeast

Two genes, rad6 and rad9, that confer radiation sensitivity in the yeast Saccharomyces cerevisiae also greatly reduce the frequency of chemically-induced reversions of a tester mutant cyc1-131, which is a chain initiation mutant in the structural gene determining iso-1-cytochrome c. Mutations induced by ethyl methanesulfonate (EMS), diethyl sulfate (DES), methyl methanesulfonate (MMS), dimethyl sulfate (DMS), nitroquinoline oxide (NQO), nitrosoguanidine (NTG), nitrogen mustard (HN2), beta-propiolactone, and tritiated uridine, as well as mutations induced by ultraviolet light (UV) and ionizing radiation were greatly diminished in strains homozygous for either the rad6 or rad9 gene. Nitrous acid and nitrosoimidazolidone (NIL), on the other hand, were highly mutagenic in these repair-deficient mutants, and at low doses, these mutagens acted with about the same efficiency as in the normal RAD strain. At high doses of either nitrous acid or NIL, however, reversion frequencies were significantly reduced in the two rad mutants compared to normal strains. Although both rad mutants are immutable to about the same extent, the rad9 strains tend to be less sensitive to the lethal effect of chemical mutagens than rad6 strains. It is concluded that yeast requires a functional repair system for mutation induction by chemical agents.     

The action of nitrosoguanidine and other DNA-inactivating agents on Streptococcus pyogenes K56 strains with normal and reduced dark repair ability

Summary The response pattern for ultraviolet light, nitrogen mustard, and ethyl methane sulphonate of Hcr⁺ and Hcr^- strains ofStreptococcus pyogenes K 56 is similar to that observed for analogous strains ofE. coli, whereas repair-apt streptococcal strains are much more sensitive to nitrosoguanidine and mitomycin C thanE. coli. Theuvr gene(s) appear(s) to be without effect upon survival, prophage induction, and mutation to streptomycin resistance caused by nitrosoguanidine and only of little influence on repair of mitomycin C damage.     

Gibberella fujikuroi mutants obtained with UV radiation and N-methyl-N'-nitro-N-nitrosoguanidine

N-methyl-N'-nitro-N-nitrosoguanidine (nitrosoguanidine) and to a lesser extent UV radiation are very mutagenic for Gibberella microconidia. The recommended nitrosoguanidine doses lead to much higher frequencies of mutants than are found in other microorganisms. The frequency of mutants among the survivors increases linearly with the nitrosoguanidine dose (molar concentration X time); the absolute number of viable mutants in a given population reaches a maximum for a dose of ca. 0.7 M X s. The microconidia are uninucleate. The onset of germination brings about increased lethality of nitrosoguanidine, but it does not modify the action of UV radiation. Mycelia are more resistant than spores to both agents. Visible illumination effectively prevents lethality when given immediately after UV radiation. Auxotrophs and color mutants are very easily obtained. Pink adenine auxotrophs and several classes of color mutants are affected in the biosynthesis of the carotenoid pigment, neurosporaxanthin.     

Comparison of spontaneous, UV-induced, and nitrosoguanidine-induced mutability to drug resistance in myxobacteria.

The UV survival curves of different strains of myxobacteria exhibited shoulders; in the case of Polyangium luteum, an unusual double shoulder appeared. Repair inhibitors like acriflavine, caffeine, and coumarin reduced the survival of UV-irradiated cells if the drugs were incorporated in the post-irradiation plating medium. The shoulders were reduced, but the final inactivation slopes were not affected by the repair inhibitors. Those strains that were resistant to UV were also more resistant to being killed by nitrosoguanidine. A variety of drug-resistant mutants occurred. The spontaneous mutation frequencies to drug resistance varied with the drug and the strain used. Drug-resistant mutants were inducible by UV irradiation and nitrosoguanidine. The UV mutability of Myxococcus xanthus was high compared to Cystobacter sp. However, the nitrosoguanidine mutability of M. xanthus was low compared to the other strains.     

Mutation and DNA replication in Escherichia coli treated with low concentrations of N-methyl-N′-nitro-N-nitrosoguanidine

N-Methyl-N′-nitro-N-nitrosoguanidine (nitrosoguanidine) causes an unexpectedly high frequency of closely linked double mutants because of its specificity for chromosome regions in replication. Low nitrosoguanidine concentrations (1 μg/ml) in liquid cultures allow replication at the normal rate and are mutagenic. It was expected that mutations would be spread over the chromosome as it replicated, but a high frequency of closely linked double mutants was found.If a thymine auxotroph is grown in the presence of 5-bromodeoxyuridine (BUdR) and nitrosoguanidine, then exposed to 313-nm radiation (which destroys BUdR-substituted DNA), the mutation frequency is much higher among survivors than among non-irradiated cells. It is concluded that nitrosoguanidine inhibits DNA replication in a small fraction of the population and that mutations are induced in that same fraction.Nitrosoguanidine treatment leads to a high frequency of closely linked double mutants under all known conditions.     

Isolation of Mutants of Euglena gracilis With Impaired Photosynthesis

Four mutant strains of Euglena gracilis have been isolated after treatment of wild type cells with ultraviolet light or the chemical mutagen nitrosoguanidine. None of the mutants is capable of autotrophic growth or photosynthetic carbon dioxide fixation.The mutant strains contain normal amounts of the enzymes of the reductive pentose phosphate cycle and are qualitatively similar to the wild type in pigment composition, but are unable to carry out the Hill reaction (light induced reduction of 2,6-dichlorophenol indophenol). Isolated mutant plastids cannot photoreduce NADP with water as the electron donor but can carry out this reaction when the electron donating system is ascorbate and 2,6-dichlorophenol indophenol. Whole cells of the mutants show the light induced oxidation of cytochrome f by light reaction I but are unable to bring about cytochrome f reduction by light reaction II. The mutants appear to be blocked at or near light reaction II in the photosynthetic electron transport chain.The mutants may represent alterations of the chloroplast genome since the mutation isolation was carried out under conditions where chloroplast viability was severely impaired, but cell viability was unaffected.     

Mutagenesis byN-methyl-N-nitroso-N-nitroguanidine in synchronized cultures ofMycobacterium phlei

Conditions of maximum induction of back mutations byN-methyl-N-nitroso-N′-nitroguanidine (“nitrosoguanidine”) were studied in auxotrophic mutants ofMycobacterium phlei. In asynchronous cultures the effects of pH, buffer molarity and concentration and exposure time to nitrosoguanidine were studied. It was shown that between 6 and 10, pH does not affect the induction of back mutations but that with increasing pH up to 9 the lethal effect of nitrosoguanidine on cells is increased. Protracted treatment with nitrosoguanidine or buffer molarity did not affect the induction of back mutations. It was found with several strains ofMycobacterium phlei that it is most efficient to treat a culture with 0.5 mg or 1 mg nitrosoguanidine/ml for 20 min at pH 6. On the basis of these findings a method of induction of back mutations by nitrosoguanidine was developed for populations with synchronous cell division.     

Effect of the raw extracts of Arthrinium strains (Hyphomycetes, Dematiaceae) on the growth of some deleterious fungi in poultry feed

In previous work the authors have shown that some species of the Arthrinium genus are characterized by being able to produce secondary metabolites with antibiotic activity. The aim of this study was to evaluate the effect of raw extracts of the growth of three different Arthrinium strains against Aspergillus flavus, Aspergillus nidulans, Fusarium moniliforme and Penicillium purpurogenum when they were present in poultry feed. The results showed that the extracts reduced the growth of Aspergillus flavus and Fusarium moniliforme but could not inhibit the development of Aspergillus nidulans. Only the raw extract of A. aureum inhibited the growth of Penicillium purpurogenum.     

Repressor Mutant Forms of the Azospirillum brasilense NtrC Protein

The Azospirillum brasilense mutant strains FP8 and FP9, after treatment with nitrosoguanidine, showed a null Nif phenotype and were unable to use nitrate as their sole nitrogen source. Sequencing of the ntrC genes revealed single nucleotide mutations in the NtrC nucleotide-binding site. The phenotypes of these strains are discussed in relation to their genotypes.     

Stimulation of mutations suppressing the loss of replication control by small alcohols

Transient exposure of lysogenic Escherichia coli cells to small alcohols stimulated the frequency of mutations suppressing the lethal loss of replication control from a prophage fragment of bacteriophage lambda. The stimulation in mutation frequency paralleled the effect of mutagenic agents, and in this sense the alcohols behaved as mutagens. 10-min treatments above distinct threshold concentrations at 23%, 18%, 10% and 4% (v/v) were required in order for methanol, ethanol, isopropanol and propanol to evoke mutagenic effects. The selected mutant cells were, in general, equally or more sensitive to ethanol than the starting cells. The mutagenicity of methanol and ethanol was detected only with E. coli strains with lambda fragments that included the site-specific and general recombination genes found within the phage int-kil gene interval; whereas, stimulation of the frequency of phenotypically identical mutations by nitrosoguanidine or ionizing radiation did not require that the lambda fragment encode these genes. Treatments of lysogenic cells with mutagenic concentrations of ethanol did not trigger prophage induction and were concluded not to induce a cellular SOS response nor to denature the prophage repressor, or to disrupt repressor-operator binding. The toxicity of ethanol was pH-dependent. Cellular sensitivity to ethanol toxicity was unaffected by the integrated lambda fragment(s) or by an intact lambda prophage; but, it was increased by deletions of the E. coli chromosome extending rightward from bio into uvrB, and rightward from chlA.     

Investigations on mutagenicity and genotoxicity of pentamidine and some related trypanocidal diamidines

Pentamidine, DAPI and some related compounds (DAI, 6-Br-AI, DPTN, DIPI, 3-Am-DAI, DiaPBF) were investigated in 2 different screening test systems for their potential mutagenic and cytotoxic effects, in the light of their binding to DNA. In the Ames test using Salmonella typhimurium strains TA98 and TA100 with and without metabolic activation no mutagenic effects could be observed. All diamidines tested, except DAI, were toxic at concentrations of 0.5 and 1.0 mumole/plate. In the sister-chromatid exchange (SCE) assay with human peripheral lymphocytes all compounds tested were growth-retarding particularly in the G0 phase. A significant induction of SCEs could only be seen after treatment with the monoamidino compound 6-Br-AI at a concentration of 100 mumole/l. It is concluded from the data obtained that pentamidine and related diamidines in the 2 assays tested show no mutagenic or genotoxic effects, in spite of their tight binding to DNA.     

An Escherichia coli mutant refractory to nitrosoguanidine mutagenesis

Summary A newly-isolated Escherichia coli mutant suffers only about 10% as many mutations as normal strains on exposure to nitrosoguanidine¹. The responsible mutation, inm-1, maps at approximately minute 79 in the current E. coli genetic map. The mutant is normal for overall growth, nitrosoguanidine lethality, spontaneous mutagenesis, ultraviolet light lethality and mutagenesis, ethyl methanesulfonate lethality and mutagenesis, and the adaptive repair induced by alkylating agents. The existence of this mutation proves that nitrosoguanidine mutagenesis is not merely the result of reactions between the chemical and DNA, but requires specific cellular function(s), and underscores the peculiarity of nitrosoguanidine as a mutagen.     

Genetic Studies of Leucine Biosynthesis in Bacillus subtilis

The mutations in a series of leucine auxotrophs isolated after treatment with nitrosoguanidine, ultraviolet light, and ICR-191 have been mapped between ilvC and pheA on the Bacillus subtilis chromosome. A fine structure map of the region was constructed by transformation. Analysis of several strains by assaying levels of their leucine bioysnthetic enzymes has shown that the region encodes three enzymes. The order of the genes with respect to the biosynthetic steps catalyzed by the gene products is 1–3–2.     

Glycogen in Methanolobus and Methanococcus

Abstract Ultraviolet light and nitrosoguanidine were used to mutagenize a red pigmented culture of Serratia marcescens , strain EB415, which produced chitinase. After mutagenesis, a stable, non-pigmented mutant designated BL40 was isolated which produced larger colonies and zones of clearing on solid medium containing colloidal chitin.
In liquid medium with colloidal chitin as the sole carbon source both strains grew similarly but BL40 produced 160 units/ml of chitinase compared with 60 units/ml for EB 415, an increase of 167%. When chitin concentration was increased in the medium, chitinase production also increased. Chitinase appeared to be extracellular, since the supernatant from washed, sonicated cells for both strains showed no detectable amount of chitinolytic activity.     

The combined action of N-methyl-N'-nitro-N-nitrosoguanidine and UV light on Bacillus subtilis

The mutagenic interaction between ultraviolet irradiation and the alkylating agent N-methyl-N'-nitro-N-nitrosoguanidine was studied in repair-competent and excision-deficient strains of Bacillus subtilis. Pre-exposure to low doses of MNNG with following treatment by low and intermediate doses of UV light increase the resistance of Bac. subtilis to UV radiation (antagonistic effect). Probably pre-exposition with MNNG leads to induction of enzymes reparation, UV damages being controlled with adaptive response genes.     

Mutagenicity of coolwhite fluorescent light for Salmonella

The most common fluorescent lamps in use today in homes and businesses in the United States, 'coolwhite' fluorescent lamps, emit light that is mutagenic for Salmonella. Strains that carry both a uvrB mutation and plasmid pKM101 are extremely susceptible to this light-induced mutation. Both base substitution and frameshift mutations can be induced without substantial lethal effects on the bacteria. Induced mutations accumulate essentially as a linear function of the time bacteria are exposed to illumination. Of Salmonella histidine-requiring strains with known nucleotide target sequences (Hartman et al., 1986; Cebula and Koch, 1989, 1990), strains either carrying one of the base substitution mutations, hisG428 and hisG46, or one of the frameshifts, hisC3076 and hisD6610, are most highly mutagenized whereas frameshift strains with hisD6580 and hisD3052 exhibit lower rates of mutagenesis. Mutagenicity does not appear to require the presence of oxygen. A filter blocking wavelengths below 370 nm eliminates mutagenesis. Polystyrene, cellulose acetate and, especially, mylar and glass filters reduce mutagenesis, indicating that at least some of the mutagenic effects can be attributed to leakage of radiations below 290 nm (far-ultraviolet light) from 'coolwhite' lamps. The more recently introduced fluorescent 'softwhite' lamps are roughly 10-fold less mutagenic at approximately equal light intensity. Incandescent light bulbs are much less mutagenic than are these fluorescent lamps. Our mutational data correlate closely with previous results in eukaryotic cells (Jacobson and Krell, 1982). A uvrB recA Salmonella double mutant is hypersensitive to the lethal effects of coolwhite fluorescent light, even when illuminated through the lids of glass Petri dishes. Thus, appropriate Salmonella strains would appear to be simple and useful screens for both the mutagenic and the lethal activities of fluorescent lamps. These systems are amenable to classroom laboratory use as relatively safe and effective means of demonstrating environmental mutagenesis.     

Effects of chemical and physical mutagens on the frequency of a large genetic duplication in Salmonella typhimurium. II. Stimulation of duplication-loss from merodiploids.

Strains of Salmonella typhimurium which contain a duplication of approximately 30% of the genome may be obtained by a simple selective procedure. These strains are highly unstable, losing the duplication when grown on non-selective medium. In this paper we report that treatment of merodiploid bacteria with mutagenic agents stimulates the rate at which haploid segregants are obtained from merodiploid strains. The mutagens which have been tested for this effect are X-rays, ultraviolet light (UV), ethyl methanesulfonate (EMS), and the azaacridine half-mustard ICR-372.