Biomedical subjects

D I Andersson

Publications and source records attributed to D I Andersson.

At least 19 recordsLinked to original sources

PubMed2005

Bacterial genome size reduction by experimental evolution.

Bacterial evolution toward endosymbiosis with eukaryotic cells is associated with extensive bacterial genome reduction and loss of metabolic and regulatory capabilities. Here we examined the rate and process of genome reduction in the bacterium Salmonella enterica by a serial passage experimental evolution procedure. The initial rate of DNA loss was estimated to be 0.05 bp per chromosome per generation for a WT bacterium and approximately 50-fold higher for a mutS mutant defective in methyl-directed DNA mismatch repair. The endpoints were identified for seven chromosomal deletions isolated during serial passage and in two separate genetic selections. Deletions ranged in size from 1 to 202 kb, and most of them were not associated with DNA repeats, indicating that they were formed via RecA-independent recombination events. These results suggest that extensive genome reduction can occur on a short evolutionary time scale and that RecA-dependent homologous recombination only plays a limited role in this process of jettisoning superfluous DNA.

Base Pair Mismatch↗

PubMed2001

Mutation frequency and biological cost of antibiotic resistance in Helicobacter pylori.

Among the several factors that affect the appearance and spread of acquired antibiotic resistance, the mutation frequency and the biological cost of resistance are of special importance. Measurements of the mutation frequency to rifampicin resistance in Helicobacter pylori strains isolated from dyspeptic patients showed that approximately 1/4 of the isolates had higher mutation frequencies than Enterobacteriaceae mismatch-repair defective mutants. This high mutation frequency could explain why resistance is so frequently acquired during antibiotic treatment of H. pylori infections. Inactivation of the mutS gene had no substantial effect on the mutation frequency, suggesting that MutS-dependent mismatch repair is absent in this bacterium. Furthermore, clarithromycin resistance conferred a biological cost, as measured by a decreased competitive ability of the resistant mutants in mice. In clinical isolates this cost could be reduced, indicating that compensation is a clinically relevant phenomenon that could act to stabilize resistant bacteria in a population.

Adenosine Triphosphatases↗

PubMed2001

An adenosyl-cobalamin (coenzyme-B12)-repressed translational enhancer in the cob mRNA of Salmonella typhimurium.

Expression of the cobalamin (Cbl) biosynthetic cob operon in Salmonella typhimurium is repressed by the end-product. This regulation is conferred mainly at the translational level and involves a cobalamin-induced folding of an RNA hairpin that sequesters the ribosomal binding site (RBS) of the cob mRNA and prevents translation initiation. A combined structural and mutational analysis shows that a cis-acting translational enhancer (TE) element, located 83 nucleotides upstream of the Shine-Dalgarno sequence in the 5'-untranslated region (5'-UTR) of the cob mRNA, is required to unfold the inhibitory RBS hairpin in the absence of cobalamin. The TE element, which consists of 5 nucleotides, is proposed to confer its enhancer function in the absence of cobalamin by interacting with nucleotides in the stem of the RBS hairpin. This interaction destabilizes the RNA hairpin and allows ribosome binding. In the presence of cobalamin, the enhancer function is inhibited. As a result, the RBS hairpin forms and prevents translation initiation. Several additional RNA hairpins in the 5'-UTR were also identified and are suggested to be important for repression. The above data suggest that normal cobalamin repression of the cob operon requires that the 5'-UTR has a defined secondary and tertiary structure.

5' Untranslated Regions↗

PubMed2001

Biological cost and compensatory evolution in fusidic acid-resistant Staphylococcus aureus.

Fusidic acid resistance resulting from mutations in elongation factor G (EF-G) of Staphylococcus aureus is associated with fitness costs during growth in vivo and in vitro. In both environments, these costs can be partly or fully compensated by the acquisition of secondary intragenic mutations. Among clinical isolates of S. aureus, fusidic acid-resistant strains have been identified that carry multiple mutations in EF-G at positions similar to those shown experimentally to cause resistance and fitness compensation. This observation suggests that fitness-compensatory mutations may be an important aspect of the evolution of antibiotic resistance in the clinical environment, and may contribute to a stabilization of the resistant bacteria present in a bacterial population.

Amino Acid Sequence↗

PubMed2000

Effects of environment on compensatory mutations to ameliorate costs of antibiotic resistance.

Most types of antibiotic resistance impose a biological cost on bacterial fitness. These costs can be compensated, usually without loss of resistance, by second-site mutations during the evolution of the resistant bacteria in an experimental host or in a laboratory medium. Different fitness-compensating mutations were selected depending on whether the bacteria evolved through serial passage in mice or in a laboratory medium. This difference in mutation spectra was caused by either a growth condition-specific formation or selection of the compensated mutants. These results suggest that bacterial evolution to reduce the costs of antibiotic resistance can take different trajectories within and outside a host.

Adaptation, Physiological↗

PubMed2000

Salmonella typhimurium mutants that downregulate phagocyte nitric oxide production.

To examine the potential and strategies of the facultative intracellular pathogen Salmonella typhimurium to increase its fitness in host cells, we applied a selection that enriches for mutants with increased bacterial growth yields in murine J774-A.1 macrophage-like cells. The selection, which was based on intracellular growth competition, rapidly yielded isolates that out-competed the wild-type strain during intracellular growth. J774-A.1 cells responded to challenge with S. typhimurium by mounting an inducible nitric oxide synthase (iNOS) mRNA and protein expression and a concomitant nitric oxide (NO) production. Inhibition of NO production with the use of the competitive inhibitor N-monomethyl-L-arginine (NMMA) resulted in a 20-fold increase in bacterial growth yield, suggesting that the NO response prevented bacterial intracellular growth. In accordance with this observation, five out of the nine growth advantage mutants isolated inhibited production of NO from J774-A.1 cells, despite an induction of iNOS mRNA and iNOS protein. Accompanying bacterial phenotypes included alterations in lipopolysaccharide structure and in the profiles of proteins secreted by invasion-competent bacteria. The results indicate that S. typhimurium has the ability to mutate in several different ways to increase its host fitness and that inhibition of iNOS activity may be a major adaptation.

Animals↗

PubMed1999

Novel Salmonella typhimurium properties in host--parasite interactions.

Inflammatory bowel disease (IBD) comprises different diseases in the gastrointestinal tract in human, of which Crohn's disease (CD) and ulcerative colitis (UC) are the most prominent. A key factor in the etiology of IBD is the chronic inflammatory process, and a large body of evidence suggests that the transcription factor nuclear factor-kappa B (NF-kappaB) is the key regulator of responses determining the clinical inflammatory condition. Recent findings using antisense oligonucleotides provide direct evidence that the p65 subunit of NF-kappaB plays a central role in chronic intestinal inflammation. It has previously been shown that the Gram negative bacteria Yersinia pseudotubercolosis targets the eukaryotic signal transduction pathway(s) that lead to NF-kappaB activation (and thus avoid an anti-bacterial inflammatory response). In this paper, growth-based selected Salmonella typhimurium clones have been used to generate a clearer picture of the molecular mechanisms involved in host-parasite interactions. From the results presented here, S. typhimurium and Y. pseudotubercolosis may use the same mechanism to block NF-kappaB activation, following host cell infection. A new adaptational feature could also be shown, where a growth-based selected bacteria avoided the normally induced translocation of NF-kappaB in host cells.

Biological Transport↗

PubMed1999

The biological cost of antibiotic resistance.

The frequency and rates of ascent and dissemination of antibiotic resistance in bacterial populations are anticipated to be directly related to the volume of antibiotic use and inversely related to the cost that resistance imposes on the fitness of bacteria. The data available from recent laboratory studies suggest that most, but not all, resistance-determining mutations and accessory elements engender some fitness cost, but those costs are likely to be ameliorated by subsequent evolution.

Adaptation, Physiological↗

PubMed1999

Novel ribosomal mutations affecting translational accuracy, antibiotic resistance and virulence of Salmonella typhimurium.

Many mutations in rpsL cause resistance to, or dependence on, streptomycin and are restrictive (hyperaccurate) in translation. Dependence on streptomycin and hyperaccuracy can each be reversed phenotypically by mutations in either rpsD or rpsE. Such compensatory mutations have been shown to have a ram phenotype (ribosomal ambiguity), increasing the level of translational errors. We have shown recently that restrictive rpsL alleles are also associated with a loss of virulence in Salmonella typhimurium. To test whether ram mutants could reverse this loss of virulence, we have isolated a set of rpsD alleles in Salmonella typhimurium. We found that the rpsD alleles restore the virulence of strains carrying restrictive rpsL alleles to a level close to that of the wild type. Unexpectedly, three out of seven mutant rpsD alleles tested have phenotypes typical of restrictive alleles of rpsL, being resistant to streptomycin and restrictive (hyperaccurate) in translation. These phenotypes have not been previously associated with the ribosomal protein S4. Furthermore, all seven rpsD alleles (four ram and three restrictive) can phenotypically reverse the hyperaccuracy associated with restrictive alleles of rpsL. This is the first demonstration that such compensations do not require that the compensating rpsD allele has a ribosomal ambiguity (ram) phenotype.

Alleles↗

PubMed1998

Evidence that gene amplification underlies adaptive mutability of the bacterial lac operon.

Adaptive mutability is the apparent alteration in specificity or rate of mutability seen in bacteria during stress. A model is proposed by which gene amplification during selective growth can give the appearance of adaptive mutability without requiring any change in mutability. The model is based on two assumptions, that a mutant lac locus with residual function allows growth if its copy number is increased, and that true reversion events are made more likely by replication of chromosomes with many copies of the locus. Apparent directed mutability, its recombination requirement, and its apparent independence of cell growth are all accounted for by the model. Evidence is provided for the required residual function and gene amplification.

Adaptation, Physiological↗

PubMed1998

[Antibiotic resistance here to stay? Compensatory mutations restore virulence of resistant bacteria].

Bacterial antibiotic resistance has increased alarmingly because of overuse of antibiotics both in humans and animals. One way of reversing this development is to reduce the use of antibiotics, thus promoting the disappearance of the resistant bacteria already present in humans and the environment. This approach is based on the assumption that resistance is conferred at the cost of impaired survival fitness in the absence of antibiotics, as compared with sensitive strains. It seems to be generally true that resistant bacteria are less fit than the respective sensitive strains, which suggests that resistance may be reversible. However, a complicating factor is the frequent finding in resistant strains of various types of compensatory mutations that restore fitness without concomitant loss of resistance. Thus, second-site compensatory mutations may allow resistant strains to persist and compete successfully with sensitive strains even in an environment depleted of antibiotics. It is concluded in the article that, if compensatory mutations are as common in clinical settings as they are in the laboratory, many types of resistance will be irreversible.

Anti-Bacterial Agents↗

PubMed1998

Virulence of antibiotic-resistant Salmonella typhimurium.

We show that most Salmonella typhimurium mutants resistant to streptomycin, rifampicin, and nalidixic acid are avirulent in mice. Of seven resistant mutants examined, six were avirulent and one was similar to the wild type in competition experiments in mice. The avirulent-resistant mutants rapidly accumulated various types of compensatory mutations that restored virulence without concomitant loss of resistance. Such second-site compensatory mutations were more common then reversion to the sensitive wild type. We infer from these results that a reduction in the use of antibiotics might not result in the disappearance of the resistant bacteria already present in human and environmental reservoirs. Thus, second-site compensatory mutations could increase the fitness of resistant bacteria and allow them to persist and compete successfully with sensitive strains even in an antibiotic-free environment.

Animals↗

PubMed1997

Vitamin B12 repression of the btuB gene in Salmonella typhimurium is mediated via a translational control which requires leader and coding sequences.

Expression of the Salmonella typhimurium btuB gene, which encodes an outer membrane protein required for vitamin B12 uptake, is repressed by the presence of external vitamin B12. We have, by means of a mutational analysis, investigated which btuB sequences are required for repression. Analysis of btuB::lacZ transcriptional and translational fusions of various lengths showed that the control was exerted mainly at the translational level and required both coding and leader sequences in the btuB transcript. Regulatory mutants with a B12 non-repressible phenotype were isolated and the mutations were shown to be located at several sites within the btuB leader. Analysis of constructs carrying site-directed point mutations, which either destabilized or restabilized a putative RNA hairpin that sequesters the btuB ribosomal binding site, demonstrated that this hairpin was essential for normal repression. Comparison of the S. typhimurium btuB gene with the previously characterized S. typhimurium cbiA and Escherichia coli btuB genes reveals significant similarities as well as differences in the cis-acting sequences required for repression.

Bacterial Outer Membrane Proteins↗

PubMed1997

Carbon starvation of Salmonella typhimurium does not cause a general increase of mutation rates.

Mutation rates in bacteria can vary depending on the genetic target studied and the specific growth conditions of the cells. Here, two different methods were used to determine how rates of mutation to antibiotic resistance, auxotrophy, and prototrophy were influenced by carbon starvation on agar plates. The rate of mutation to rifampin resistance was increased by starvation as measured by fluctuation tests, similar to what has been reported previously for Escherichia coli. In contrast, the rates of mutation to various types of auxotrophy were unaffected or decreased as measured by both fluctuation tests and a repeated-streaking procedure. Similarly, the rates of reversion to prototrophy of his and lac nonsense and missense mutations were unaffected by starvation. Thus, mutation rates of different genetic targets can be affected differently by starvation and we conclude that carbon starvation is not generally mutagenic in Salmonella typhimurium.

Drug Resistance, Microbial↗

PubMed1996

Muller's ratchet decreases fitness of a DNA-based microbe.

Muller proposed that an asexual organism will inevitably accumulate deleterious mutations, resulting in an increase of the mutational load and an inexorable, ratchet-like, loss of the least mutated class [Muller, H.J. (1964) Mutat. Res. 1, 2-9]. The operation of Muller's ratchet on real populations has been experimentally demonstrated only in RNA viruses. However, these cases are exceptional in that the mutation rates of the RNA viruses are extremely high. We have examined whether Muller's ratchet operates in Salmonella typhimurium, a DNA-based organism with a more typical genomic mutation rate. Cells were grown asexually under conditions expected to result in high genetic drift, and the increase in mutational load was determined. S. typhimurium accumulated mutations under these conditions such that after 1700 generations, 1% of the 444 lineages tested had suffered an obvious loss of fitness, as determined by decreased growth rate. These results suggest that in the absence of sex and with high genetic drift, genetic mechanisms, such as back or compensatory mutations, cannot compensate for the accumulation of deleterious mutations. In addition, we measured the appearance of auxotrophs, which allowed us to calculate an average spontaneous mutation rate of approximately 0.3-1.5 x 10(-9) mutations per base pair per generation. This rate is measured for the largest genetic target studied so far, a collection of about 200 genes.

Adenine↗

PubMed1994

Vitamin B12 repression of the cob operon in Salmonella typhimurium: translational control of the cbiA gene.

Expression of the cob operon is repressed by B12 via a post-transcriptional control mechanism which requires sequence elements within the leader region of the mRNA and the first gene of the operon, the cbiA gene. Here we show that B12 repression of cbiA gene expression occurs at the level of translation initiation through sequestration of the ribosomal binding site (rbs) in an RNA hairpin. Analysis of mutations that destabilize or restabilize the secondary structure demonstrates that folding of the hairpin is essential for repression. The existence of the hairpin was confirmed by a secondary structure analysis of RNA from the wild type and three mutants. Deletions that remove the upstream part of the leader confer a drastic reduction in translation efficiency. This low-level translation is caused by the hairpin, as indicated by the finding that suppressor mutations that destabilize the hairpin restore efficient translation. Thus, the native upstream RNA functions as a translation enhancer and acts to relieve the hairpin's inhibitory effect on translation initiation. The inhibitory effect of the hairpin was confirmed by a ribosomal toeprinting analysis. We propose that the translational control of the cbiA gene mediates repression of the entire cob operon.

Base Sequence↗

PubMed1994

The control region of the pdu/cob regulon in Salmonella typhimurium.

The pdu operon encodes proteins for the catabolism of 1,2-propanediol; the nearby cob operon encodes enzymes for the biosynthesis of adenosyl-cobalamin (vitamin B12), a cofactor required for the use of propanediol. These operons are transcribed divergently from distinct promoters separated by several kilobases. The regulation of the two operons is tightly integrated in that both require the positive activator protein PocR and both are subject to global control by the Crp and ArcA proteins. We have determined the DNA nucleotide sequences of the promoter-proximal portion of the pdu operon and the region between the pdu and cob operons. Four open reading frames have been identified, pduB, pduA, pduF, and pocR. The pduA and pduB genes are the first two genes of the pdu operon (transcribed clockwise). The pduA gene encodes a hydrophobic protein with 56% amino acid identity to a 10.9-kDa protein which serves as a component of the carboxysomes of several photosynthetic bacteria. The pduF gene encodes a hydrophobic protein with a strong similarity to the GlpF protein of Escherichia coli, which facilitates the diffusion of glycerol. The N-terminal end of the PduF protein includes a motif for a membrane lipoprotein-lipid attachment site as well as a motif characteristic of the MIP (major intrinsic protein) family of transmembrane channel proteins. We presume that the PduF protein facilitates the diffusion of propanediol. The pocR gene encodes the positive regulatory protein of the cob and pdu operons and shares the helix-turn-helix DNA binding motif of the AraC family of regulatory proteins. The mutations cobR4 and cobR58 cause constitutive, pocR-independent expression of the cob operon under both aerobic and anaerobic conditions. Evidence that each mutation is a deletion creating a new promoter near the normal promoter site of the cob operon is presented.

Amino Acid Sequence↗

PubMed1992

Involvement of the Arc system in redox regulation of the Cob operon in Salmonella typhimurium.

Transcription of the cobalamin biosynthetic (cob) genes in Salmonella typhimurium is induced during anaerobic growth. Two mutants defective in induction of the Cob operon were isolated. The mutants showed decreased anaerobic induction, both during conditions of fermentation and anaerobic respiration, relative to the wild type. The mutant genes were localized by Hfr and transduction mapping to 100 minutes and 69 minutes on the chromosome. The mutants were sensitive to the dye toluidine blue. The mapping and the dye sensitivity suggest that these mutants are defective in the arcA and arcB genes, not previously identified in Salmonella typhimurium. Thus the regulatory function of the Arc system is expanded to include anaerobic activation of the Cob operon.

Anaerobiosis↗