Classifying mutagens as to their specificity in causing the six possible transitions and transversions: a simple analysis using the Salmonella mutagenicity assay

The standard Salmonella tester strains used to detect base substitution mutations carry the hisG428 ochre mutation (TA102 and TA104) and the hisG46 missense mutation (TA100). These mutations can be reverted by base changes at their mutant his loci or at extragenic suppressor loci. The base changes resulting in each class of revertants of these mutations have been identified, and simple phenotypic screens have been developed to distinguish among them. Revertants at extragenic suppressor loci are distinguished from those at the his loci by their sensitivity to inhibitory histidine analogs. The four ochre suppressor loci of hisG428 are distinguished by their ability to support growth of nonsense mutants of phage P22. These screens are the basis for a rapid and simple system for determining the base substitution specificity of mutagens using hisG428- and hisG46-containing tester strains. Diagnostic mutagens specific for each of the six possible base changes (transitions and transversions) have been identified. Using these diagnostic mutagens, two additional strains, each specifically reverted by a single base substitution mutation, have been developed to provide a minimum of two loci at which to detect each type of base change. The ability of this system to provide detailed information about mutational specificity in a variety of DNA repair backgrounds will allow further elucidation of the mechanisms of mutagenesis and DNA repair.     

Effects of Salmonella genotypes and testing protocols on H2O2-induced mutation

Hydrogen peroxide (H2O2) was shown to be mutagenic in a number of strains of Salmonella typhimurium. Strain SB1106p (hisC3108, hisO1242, pKM101), a newly-constructed strain carrying the histidine mutation at a UGA chain-terminating codon, was more responsive to H2O2 than TA104 or TA102, the two hisG428 strains originally developed for detecting oxidative mutagens. The largest proportional increase in revertants of strain TA104 was in the fraction of intragenic deletions. Three other strains (TA97, SB1111 and SB1106) gave unequivocal positive responses to H2O2 in both the liquid pre-incubation procedure and standard plate incorporation procedure. The response of TA100 varied among experiments, ranging from negative to a weak positive. Variations in the catalase content among the tester strains did not correlate with the relative responses obtained in the mutagenicity assays.     

Mutagenic activity in synthetic pyrethroids in Salmonella typhimurium

Four pyrethroids, allethrin, resmethrin, permethrin and fen-valerate,were tested for mutagenicity in bacterial reversion assay systemswith seven strains (TA1535, TA100, TA1538, TA98, TA1537, TA97and TA104) of Salmonella typhimurium. Our results show thatthree pyrethroids, namely resmethrin, permethrin and fenvalerate,were not found to be mutagenic in S. typhimurium in the presenceor absence of a rat liver activation system. Allethrin was foundto be mutagenic with TA100, TA104 and TA97 strains and requiredmetabolic activation (S9 mix) in order to show its activity,mainly with TA100 and TA104 strains.     

The effectiveness of Salmonella strains TA100, TA102 and TA104 for detecting mutagenicity of some aldehydes and peroxides

Several aldehydes and peroxides were tested for mutagenicityusing Salmonella typhimurium tester strains TA97a, TA100, TA102and TA104, in the presence and absence of Aroclor-induced liverS9 mix from F344 rats and B6C3F₁ mice, in either preincubationor vapour phase rotocols. Some chemicals were tested in additionalSalmonella strains. Benzaldehyde, butyraldehyde, benzoyl peroxide,4-chlorobenzaldehyde, isobutyraldehyde, propionaldehyde andveratraldehyde were non-mutagenic Acetaldehyde and dicumyl peroxidegave inconsistent results and furfural gave equivocal responsesin TA100 and TA104. Cumene hydroperoxide, formaldehyde and glutaraldehydewere mutagenic in TA100, TA102 and TA104. trans-Cinnamaldehydeexhibited a weak mutagenic response in TA100 with mouse liverS9 only. 2,4,5-Trimethoxybenzaldehyde was mutagenic only instrain TA1538 with rat liver S9. With the exception of butanoneperoxide, which was mutagenic only in TA104, all chemicals mutagenicin strains TA102 and/or TA104 were also mutagenic in TA100.The data do not, therefore, support the preferential use ofstrains TA102 and TA104 for screening aldehydes and peroxidesfor mutagenicity. For a number of these chemicals the advantagesof using TA102 or TA104 was in the increased responses comparedwith those obtained with TA100. Two of the four peroxides weremutagenic and one of these was mutagenic only with TA104. Thissuggests that strains TA102 and TA104 be used if peroxides arenot mutagemc in TA100 or TA97. ⁴Present addresses: ⁴British American Tobacco Ltd, SouthamptonSO15 8TL, UK ⁵FRAME, Nottingham NG1 4EE, UK ³To whom correspondence should be addressed. Tel: +1 919 541 4482; Fax: +1 919 541 2242; Email: zeiger{at}niehs.nih.gov      

Analysis of the ciprofloxacin-induced mutations in Salmonella typhimurium

The mutagenic events induced by ciprofloxacin, a potent antimicrobial agent, have been characterized. For this, a battery of His⁻ mutants of Salmonella typhimurium (hisG428, hisG46, hisC9070, and hisG1775 targets) that detects the six possible transitions and transversions [Levin and Ames (1986): Environ Mutagen 8:9–28] and two additional His⁻ strains (hisC3076 and hisD3052 targets) carrying frameshift mutations have been used. Our results indicate that GC→TA transversions are the major base-pair substitution induced by ciprofloxacin and that GC→AT transitions are also produced, but to a lesser degree. However, we cannot discard the fact that AT→TA transversions are also induced. In addition, the data indicate that the mutational specificity of ciprofloxacin depends on the location of the target. Intragenic base-pair substitutions are the most frequent mutations at the hisG428 target when it is on the chromosome, whereas 3 or 6 base-pair deletions are the major mutagenic events when this target is on the plasmid pAQ1. We have shown that ciprofloxacin also induces deletions/insertions at the hisC3076 and hisD3052 frameshift targets. Therefore, this inhibitor of DNA gyrase promotes a wide pattern of mutations including different kinds of base-pair substitutions, 3 or 6 base-pair deletions, and insertions/deletions resulting in frameshifts. All of these mutagenic events require the MucAB proteins involved in the error-prone repair, with the exception of base-pair insertions/deletions at the hisD3052 target, which are independent of the presence of plasmid pKM101. © 1996 Wiley-Liss, Inc.     

Comparative mutagenic and genotoxic effects of three antimalarial drugs, chloroquine, primaquine and amodiaquine

Comparative mutagenic and genotoxic effects of three antimalarialdrugs, chloroquine, primaquine and amodiaquine, were assessedin the Ames mutagenicity assay (in strains TA97a, TA100, TA102and TA104) and in vivo sister chromatid exchange (SCE) and chromosomeaberration (CA) assays in bone marrow cells of mice. These arethe most commonly used antimalarial drugs available at presentthroughout the world. The results of the bacterial mutagenicityassays showed a very weak mutagenic effect of all three drugsin Salmonella strains TA97a and TA100 both with and withoutS9 mix and in TA104 only with S9 mix. The results of the invivo SCE and CA assays indicate that these three drugs are genotoxicin bone marrow cells of mice. ³To whom correspodence should be addressed. Tel: +91 33 473 3491; Fax: +91 33 473 5197; Email: iichbio{at}giascl01.vsnl.net.in     

Relevance of plasmid pKM101-mediated mutagenicity in bacteria to genotoxicity in mammalian cells

Three structurally related compounds, 4-acetoxy-3-acetoxy-methyl-acetophenone(AAMAP), 1-[4'- hydroxy-3'-hydroxy-methylphenyl]-2-[benzyl-t-butylamino]ethanone hydrochloride (HHBEH) and 1-[4'-hydroxy-3'-hydroxymethyl-phenyl]-2-[benzyl-t-butylamino]ethanol (HHBE), gave positive dose-related mutagenic responsesin the Ames test when Salmonella typhimurium strain TA100 wasused as the test organism. Strain TA100 carries the hisG46 allele,which is revertable by base changes, together with plasmid pKM101,which encodes mucAB genes that are analogous to umuDC, the chromosomalSOS-repair genes of Escherichia coli K-12. None of the compoundswas mutagenic in Ames strain TA1535, which is the plasmid-freederivative of strain TA100. Only AAMAP, and that at only thehighest concentration tested, was mutagenic in strain TA98,which detects frameshift mutations and carries plasmid pKM101.No compound was significantly mutagenic in strain TA1538, whichis the plasmid-free derivative of strain TA98. When the threecompounds were tested for the induction of sister-chromatidexchanges (SCEs) in Chinese hamster cells, the two more potentmutagens, AAMAP and HHBEH were found to increase SCEs, whereasHHBE did not give a significant response at any concentrationtested. Ames test data showing plasmid pKM101-dependent mutagenesisare therefore, at least for these compounds, relevant indicatorsof eukaryotic genotoxicity. Parts of this paper were communicated to the Science Group atthe 123rd British Pharmaceutical Conference, Jersey, 1986.      

Induction of SOS genes in Escherichia coli and mutagenesis in Salmonella typhimurium by fluoroquinolones

The induction of several SOS genes of Escherichia coli by fluoroquinolones has been studied. Three different SOS gene fusions (recA::lacZ, umuC::lacZ and sulA::lacZ) have been introduced into the E.coli MC1061 strain to study the induction of these SOS genes in the same genetic background. Data on the basal level of expression of these fusions, as well as their induction by mitomycin C and N-methyl-N'-nitro-N-nitrosoguanidine are presented. Using these strains, we have found that, like nalidixic acid, ofloxacin, enoxacin and ciprofloxacin are strong inducers of the SOS genes tested, umuC gene expression being the highest. Furthermore, fluoroquinolones produced a significant increase in the reversion of the base substitution hisG428 mutation in the TA102 Salmonella tester strain, while no effect was found in strains TA98, TA100, TA1537 and TA1535. These data indicate that the error-prone repair pathway can participate in mutagenesis induced by fluoroquinolones and also that the damage produced by these chemicals may be similar to that produced by nalidixic acid.     

Mutagenicity spectra in Salmonella typhimurium strains of glutathione, L-cysteine and active oxygen species

Glutathione and L-cysteine, in the presence of rat kidney post-mitochondrialsupernatant (S9) fraction, and various forms of active oxygenwere investigated for mutagenicity in seven his^– strainsof Salmonella typhimurium. Glutathione and L-cysteine showedqualitatively and quantitatively virtually identical mutagenicactivities. The number of mutants induced in strain TA97 was3–4 times higher than in TA100, the strain in which themutagenicity was originally detected. Mutagenic effects werealso observed in strains TA92, TA102 and TA104, but not in TA1535and TA1537. Hydrogen peroxide, superoxide and glucose/glucoseoxidase in the presence and absence of kidney S9 fraction showedpronounced mutagenic effects in strains TA104 and TA102. Additionally,weak mutagenic effects were observed in TA100, while the remainingstrains, including TA97, were not responsive. These mutagenicityspectra suggest that the mutagenic species formed from glutathioneand L-cysteine are similar, if not identical, and are differentfrom hydrogen peroxide, superoxide and other oxygen speciesderived from them. Further support for this notion was givenwhen it was observed that catalase did not affect the mutagenicityof glutathione and that superoxide dismutase showed a significanteffect only when used in milligram quantities. This study showsthat mutagenicity spectra may be useful in the elucidation ofactivation pathways. Furthermore, it is interesting to notethat all the compounds and preparations showing a positive responsein the Ames test in the present study occur endogenously inorganisms: glutathione, L-cysteine, hydrogen peroxide, superoxide,glucose, glucose oxidase and kidney S9 fraction (which was mutagenicin several strains).     

Bacterial genotoxicity of nitrosated famotidine

Famotidine, a histamine H2-receptor antagonist, was devoid ofmutagenic activity in seven his^- Salmonella typhimurium strains(TA1535, TA1537, TA1538, TA97, TA98, TA100 and TA102) and wasequitoxic in repair-proficient (WP2) and repair-deficient (WP2uvr,WP67, CM561, CM571, WP100 and CM871) Escherichia coli strains,both in the presence and in the absence of S9 mix containingliver S9 fractions from Aroclor-treated rats. However, aftera short pre-incubation step with nitrite in an acidic environment,the drug increased, by a direct mechanism, the number of his⁺revertants in Salmonella strains TA100, TA102 and TA97 (a decreaseof mutagenicity being conversely observed in TA1535) and oftrp+ revertants in E. coli strains WPluvrA and WP67. Moreover,it enhanced the induction of non-reparable DNA damage in E.coli strains simultaneously lacking the uvrA-dependent excisionrepair and the lexA post-replication repair pathways. The mutagenicityof acidified nitrite-famotidine mixtures was related to dosesof both precursors, with a maximum production of mutagenic derivativesin a slight molar excess of nitrite. The optimal pH of the nitrosationreaction (2.0) was intermediate between the one required forcimetidine (1.5) and ranitidine (2.5). Potency of famotidineas a precursor of mutagenic derivatives was considerably lowerthan the one of the other two H₂ blockers. The nitrosation productsof all three drugs mainly induced base-pair substitutions inSalmonella DNA, to a greater extent at sites containing G-Cbase pairs (strain TA100) in the case of famotidine and cimetidine,and at sites containing AT base pairs (TA102) in the case ofranitidine. Although these experimental findings may suggestpossible toxicological consequences in ulcer patients receivinganti-secretory drugs, various considerations tend to minimizetheir practical in vivo relevance, especially for risk-benefitevaluations. Additionally, as in the case of cimetidine andranitidine, formation of mutagenic nitrosated famotidine wasefficiently prevented by equimolar ascorbic acid.     

mutation spectra of Glu-P-1 in Salmonella: Induction of hotspot frameshifts and site-specific base substitutions

We used colony probe hybridization and PCR/ DNA sequence analysis to determine the mutations in —1,640 revertants of the -1 frameshift allele hisD3052 and -260 revertants of the base substitution allele hisG46 of Salmonella typhimurium induced by the heterocyclic amine cooked food mutagen 2-amino-6-methyldipy-rido[1,2-a:3′,2′-d]imidazole (Glu-P-1). All of the mutations were at sites containing guanine, which is the base at which Glu-P-1 forms DNA adducts. A hotspot mutation involving the deletion of a CG or GC within the sequence CGCGCGCG accounted for 100% of the Glu-P-1-induced mutations at the frameshift allele in strains TA1978 (uvr⁺) and TA1538 (uvrB) and 99% in TA98 (uvrB, pKM101). To explain the induction of these hotspot mutations by Glu-P-1, we describe here a more detailed version of our recently proposed correct incorporation/ slippage model [Genetics:136:731, 1994]. We propose that after cytosine is incorporated correctly opposite a Glu-P-1-adducted guanine, various slipped intermediates may form (a total of 18), depending on which guanine is adducted and whether it remains within the helix or becomes extrahelical. This variety of mutational pathways may account for the high mutability of the hotspot sequence by Glu-P-1. Although the pKM101 plasmid does not influence the mutagenic potency or mutational spectrum of Glu-P-1 at the frameshift allele, it is required by Glu-P-1 to revert the base substitution allele, where Glu-P-1 induces G-C-→T-A transversions (75%) and G-C→T.A transitions (25%) exclusively at a single site (the second position of the CCC codon of the hisG46 allele). The limited (20–30 times less) base substitution mutagenic potency of Glu-P-1 relative to its frameshift mutagenic potency as well as the extreme site specificity exhibited by Glu-P-1 for base substitutions may have bearing on the lack of base substitutions identified in ras genes in Glu-P-1-induced rat colon tumors. © 1994 Wiley-Liss, Inc.     

Specificity of base substitution mutations induced by the dietary carcinogens 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) in Salmonella

The base pair substitution mutational profiles induced by the heterocyclic amine cooked food mutagens PhIP and IQ in Salmonella typhimurium strains TA100 and TA1535 were determined by colony hybridization analysis. Both PhIP and IQ induced predominantly G→TA transversions in strain TA100 (rfa,ΔuvrB/pKM101) with a pronounced preference for the second codon position (CC→CAC; 72% of total). PhIP also reverted strain TA1535 (rfa, ΔuvrB) efficiently at concentrations similar to those required for strain TA100. In contrast to the PhIP-induced mutational profile observed in strain TA100, in strain TA1535 PhIP induced exclusively G→AT transitions at the second codon position (CC→CTC; 96–99% of total). Base substitution mutagenesis induced by heterocyclic amines related to PhIP is generally SOS-dependent, requiring the presence of plasmid pKM101 in Salmonella hisG46 strains. Thus, the SOS dependent reversion of S. typhimurium strain TA100 probably reflects error-prone lesion bypass at the major PhIP- guanosine adduct at the C-8 position. The G→AT transition mutations induced by PhIP in strain TA1535 appear to be SOS-independent, however, suggesting that these mutations may arise from the formation of PhIP-DNA adducts other than the replication-blocking C8-dG lesion. Environ. Mol. Mutagen. 31:327–332, 1998 © 1998 Wiley-Liss, Inc.     

Comparison of Salmonella typhimurium TA102 with Escherichia coli WP2 tester strains

In 1982, Levin et al. published a paper describing a new Salmonella typhimurium strain, TA102, for detecting mutagenic agents that react preferentially with AT base pairs. This strain has an AT base pair at the critical mutation site within the hisG gene, which is located on a multicopy plasmid, pAQ1; the chromosomal copy of the hisG gene has been deleted. It also has an intact excision repair system, thus facilitating the detection of cross-linking agents, and carries the mutator plasmid, pKM101. Although TA102 has been shown to be reverted by certain mutagenic agents that are not detected in the usual battery of strains (TA1535, TA1537, TA1538, TA98 and TA100), there has been a general reluctance within the field to include TA102 as one of the standard screening strains. This may in part result from the difficulties which have been experienced in many laboratories in maintaining the strain, and in obtaining reproducible spontaneous and induced revertant counts. At Glaxo we routinely include certain Escherichia coli strains in our microbial test battery, and were aware that some of the genetic features offered by TA102 were already being covered by these strains. For example, E.coli WP2 (pKM101) has an AT base pair at the critical mutation site within the trpE gene, is excision proficient (and thus will detect cross-linking agents) and carries the pKM101 plasmid to enhance error-prone repair. From the published literature it was apparent that a number of the 'TA102 specific' mutagens could be detected in E.coli e.g. neocarzinostatin, UV and 8-MOP plus UV.(ABSTRACT TRUNCATED AT 250 WORDS)     

Studies of error-prone DNA repair in Escherichia coli K-12 and Ames Salmonella typhimurium strains using a model alkylating agent

4-Acetoxy-3-acetoxymethyl acetophenone (AAMAP) is muta-genicin Ames Salmonella typhimurium tester strains TA100 and TA98,which carry plasmid pKM101, but not in the isogenic plasmid-lessstrains TA1535 and TA1538. Similarly, no AAMAP-induced reversionof the his-4 allele is detectable in Escherichia coli K-12 umuCstrains in the absence of the plasmid, even when the strainsare treated with ethylene-diaminetetraacetate to increase permeability,or when the uvrB allele is introduced to increase error-proneDNA repair. AAMAP is, however, mutagenic in umuC⁺ strains orin umuC strains in which plasmid pKM101 has been introduced,suggesting that the plasmid-encoded MucAB or the chromosomallydetermined UmuDC proteins are required for mutagenesis. Mutationfrequencies are higher in E. coli umuC (pKM101) strains, whichresemble Ames tester strains of S.typhimurium, than in E.coliumuC⁺ or even umuC⁺ (pKM101) strains. Therefore, providing thatthe recommended pKM101-containing tester strains are used, theapparent absence of Umu-like protein activity in S. typhimuriummay actually increase the sensitivity of the Ames test for thedetection of mutagens that require error-prone DNA repair foractivity. ^* Parts of this paper were communicated to the Fourth InternationalConference on Environmental Mutagens, Stockholm, 1985. ²To whom correspondence should be addressed      

Genetic differences between the standard Ames tester strains TA100 and TA98

The standard Ames tester strains of Salmonella typhimurium areseparated by many steps in their pedigree, some involving mutagentreatments, and contain independently isolated uvrB-bio-galdeletions and rfa mutations. In this work the araD531 mutationwas introduced into the Ames tester strains TA100 and TA98.The responsiveness of the resulting strains (BA15 and BA14)to a number of chemical mutagens was then assessed by monitoringthe induction of forward mutations to L-arabinose resistance(Ara test). Here we have shown that these two strains of theAmes test differ greatly in their responses to mutagens, inways that are not associated with the mutagenic specificitiesof the original his mutations. In general, the genetic backgroundof strain TA100 appears to be more sensitive to the killingeffects of chemicals than that of TA98. The greatest differenceswere found with nifurtimox (NFX) and its analogue, compound1K. The Ara test responded to the mutagenic effects of thesetwo nitrofurans when carried out in the genetic background ofstrain TA98 but not in that of TA100. A higher sensitivity tothe lethal effects of NFX and 1K together with the greater nitroreductioncapability of strain TA100 as compared with TA98 might explainthe differences. In conclusion, our results indicate that thestandard Ames S. typhimurium tester strains are not isogenicand that genetic differences at loci other than his might besignificant for mutagenicity testing. To this respect the routineuse of the isogenic set of S. typhimurium strains constructedby Popkin et al. (Mut. Res., 224, 453–464, 1989) and derivedfrom strain hisD3052 (as the standard TA98) seems advisable.     

SOS induction in Escherichia coli and Salmonella mutagenicity: a comparison using 330 compounds

To examine the concordance of two microbial genotoxicity short-termassays, 330 experimental results for the SOS chromotest usingtester strain Escherichia coli PQ37 were compared with the resultsof the Salmonella/mammalian microsome mutagenicity assay withSalmonella typhimurium TA97, TA98, TA100, TA102, TA104, TA1535,TA1537 and/or TA1538. With respect to qualitative features,the concordance between SOS chromotest and Salmonella mutagenicitytest results was 86.4% (sensitivity, 78.6%; specificity, 100%;     

Testing human faecal extracts for genotoxic activity with the SOS Chromotest: the importance of controlling for faecal enzyme activity

Two aqueous extracts of human faeces were prepared from a healthymale donor and assayed in the SOS Chromotest. Both extractswere positive in microtitre fluctuation tests in Salmonellatyphimurium TA100 and Escherichia coli WP2uvrA(pKM101). Differenceswere observed in the induction factors of these samples whenp-nitrophenyl-/5–D-galacto-pyranoskle (p-NPG) and o-nhTophenyl–3-D-galactopyranoside(o-NPG) were used as substrates for the /3-galactosidase assayin the SOS Chromotest. With one sample, a positive inductionfactor was reproducibly obtained using p-NPG but not o-NPG.When the bacterial cells were washed with fresh LB broth beforeenzyme assay, the positive induction factor obtained with p-NPGwas reduced to an insignificant level. During the 2-h treatmentperiod, both faecal samples enhanced bacterial growth abovethat of the zero-dose control. When SOS Chromotest assays wereperformed with no bacteria or with 5. typhimurium TA100 or hisG46(non-lactose fermenting organisms) in place of E. coli PQ37,it was found that the extracts contained significant levelsof endogenous 3–galactosidase and alkaline phosphatase,which, due to their carry-over in the bacterial pellet (aftercentrifugation to remove the coloured extract) gave rise tothe positive induction factor obtained with p-NPG. The resultsobtained in these experiments indicate that where the SOS Chromotestis applied to biological samples, care should be taken in theinterpretation of the data and that a washing step should beincluded to prevent possible errors occurring due to exogenousenzymes in the sample.     

Evaluation of the genotoxicity of stevioside and steviol using six in vitro and one in vivo mutagenicity assays

Stevioside, a constituent of Stevia rebaudiana, is commonlyused as a non-caloric sugar substitute in Japan. The genetictoxicities of stevioside and its aglycone, steviol, were examinedwith seven mutagenicity tests using bacteria (reverse mutationassay, forward mutation assay, umu test and rec assay), culturedmammalian cells (chromosomal aberration test and gene mutationassay) and mice (micronucleus test). Stevioside was not mutagenicin any of the assays examined. The aglycone, steviol, however,produced dose-related positive responses in some mutagenicitytests, i.e. the forward mutation assay using Salmonella typhimuriumTM677, the chromosomal aberration test using Chinese hamsterlung fibroblast cell line (CHL) and the gene mutation assayusing CHL. Metabolic activation systems containing 9000 g supernatantfraction (S9) of liver homogenates prepared from polychlorinatedbiphenyl or phenobarbital plus 5, 6-benzoflavone-pretreatedrats were required for mutagenesis and clastogenesis. Steviolwas weakly positive in the umu test using S.typhimurium TA1535/pSK1002either with or without the metabolic activation system. Steviol,even in the presence of the S9 activation system, was negativein other assays, i.e. the reverse mutation assays using S.typhimuriumTA97, TA98, TA100, TA102, TA104, TA1535, TA1537 and Escherichiacoli WP2 uvrA/pKM101 and the rec-assay using Bacillus subtilis.Steviol was negative in the mouse micronucleus test The genotoxicrisk of steviol to humans is discussed. ⁹To whom correspondence should be addressed     

Formaldehyde is a bacterial mutagen in a range of Salmonella and Escherichia indicator strains

Formaldehyde was examined for bacterial mutagenicity using Escherichiacoli WP2(pKM101) and WP2uvrA(pKM101), and Salmonella typhimuriumTA1535, TA1537, TA1538, TA98, TA100 and TA102, in the absenceof any exogenous source of metabolic activation. Using pre-incubationexposure, clear mutagenicity was seen for TA98, TA100 and TA102,and both E.coli strains. In standard plate-incorporation assays,consistent mutagenicity was seen only for TA100 and WP2uvrA(pKM101).No evidence of mutagenicity was seen for TA1535, TA1537 or TA1538using either method of exposure. These data confirm the enhancedability of the pre-incubation method to detect the mutagenicityof formaldehyde both quantitatively, as expressed by numbersof revertant colonies, and qualitatively, in terms of the rangeof indicator strains reverted. The relatively greater sensitivityof the pre-incubation assay is probably due to better containmentof a volatile agent and/or lack of interaction with agar duringthe initial period of exposure. The findings are consistentwith the suggestion that formaldehyde induces lesions in bacteriawhich are, at least to some extent, excision-repairable, andindicate that the presence of the R-factor plasmid may be requiredfor the expression of its mutagenicity in excision repair-deficientSalmonella.     

Target sequences for mutagenesis in Salmonella histidine-requiring mutants

Nucleotide target sequences involved in reversion to the wild type phenotype are diagrammed for Salmonella frameshift histidine-requiring mutants hisD3052, hisD3018, hisD6610, and hisD6580 and for base-substitution mutants hisG46 and hisG428. Frameshift strain hisC3076 probably reverts by nucleotide changes similar to those that occur during reversion of hisD3018 and hisD6610. Multiple modes of reversion characterize each strain. Each strain also has a particularly diagnostic mutagen-susceptible sequence. These highly mutagen-susceptible stretches are the hisD3052 GCGCGCGC sequence, the hisD6610 CCCCCC sequence, the hisD6580 AAAAA sequence, and the A/T containing codon of hisG428 and G/C containing codon of hisG46, respectively. Between them, hisG46 and hisG428 are reverted by all of the six possible base substitution transition and transversion mutations.