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A Multicomponent System Is Required for Tetracycline-Induced Excision of Tn4555. Anita C. Parker, 2004.Bacteroides spp . are the predominant organisms in the intestinal tract, and they also are important opportunistic pathogens . Antibiotic therapy of Bacteroides infections often is complicated by the prevalence of drug-resistant organisms which acquire resistance genes from a variety of mobile genetic elements including conjugative transposons (CTns) and mobilizable transposons (MTns) . Tn4555 is an MTn that encodes ß-lactam resistance, and it is efficiently mobilized by the Bacteroides CTns via a tetracycline (TET)-inducible mechanism . In this study a model system with CTn341 and a Tn4555 minielement was used to examine Tn4555 excision from the chromosome . Using PCR and mobilization assays it was established that excision was stimulated by TET in the presence of CTn341 . In order to determine which Tn4555 genes were required for excision, int, tnpA, tnpC, xis, and mobA mutants were examined . The results indicated that int plus two additional genes, tnpC and xis, were required for optimal excision . In addition, there was no requirement for the mobA gene, as had been shown for another MTn, NBU1 . The Xis protein sequence is related to a family of plasmid excisionases, but the TnpC gene product did not match anything in the sequence databases . Evidence also was obtained that suggested that Xis is involved in the control of TET-induced excision and in control of mobilization by CTn341 . Overall, these results indicate that excision of MTns is a complex process that requires multiple gene products .
Evolution of Resistance to Sulfadoxine-Pyrimethamine in Plasmodium falciparum. Michelle L. Gatton, 2004.The development of resistance to sulfadoxine-pyrimethamine by Plasmodium parasites is a major problem for the effective treatment of malaria, especially P . falciparum malaria . Although the molecular basis for parasite resistance is known, the factors promoting the development and transmission of these resistant parasites are less clear . This paper reports the results of a quantitative comparison of factors previously hypothesized as important for the development of drug resistance, drug dosage, time of treatment, and drug elimination half-life, with an in-host dynamics model of P . falciparum malaria in a malaria-naïve host . The results indicate that the development of drug resistance can be categorized into three stages . The first is the selection of existing parasites with genetic mutations in the dihydrofolate reductase or dihydropteroate synthetase gene . This selection is driven by the long half-life of the sulfadoxine-pyrimethamine combination . The second stage involves the selection of parasites with allelic types of higher resistance within the host during an infection . The timing of treatment relative to initiation of a specific anti-P . falciparum EMP1 immune response is an important factor during this stage, as is the treatment dosage . During the third stage, clinical treatment failure becomes prevalent as the parasites develop sufficient resistance mutations to survive therapeutic doses of the drug combination . Therefore, the model output reaffirms the importance of correct treatment of confirmed malaria cases in slowing the development of parasite resistance to sulfadoxine-pyrimethamine . Activation of the glnA, glnK, and nac Promoters as Escherichia coli Undergoes the Transition from Nitrogen Excess Growth to Nitrogen Starvation. Mariette R. Atkinson, 2002.The nitrogen-regulated genes and operons of the Ntr regulon of Escherichia coli are activated by the enhancer-binding transcriptional activator NRI  P (NtrCP) . Here, we examined the activation of the glnA, glnK, and nac promoters as cells undergo the transition from growth on ammonia to nitrogen starvation and examined the amplification of NRI during this transition . The results indicate that the concentration of NRI is increased as cells become starved for ammonia, concurrent with the activation of Ntr genes that have less- efficient enhancers than does glnA . A diauxic growth pattern was obtained when E . coli was grown on a low concentration of ammonia in combination with arginine as a nitrogen source, consistent with the hypothesis that Ntr genes other than glnA become activated only upon amplification of the NRI concentration .
Comparing the Dehalogenase Gene Pool in Cultivated  -Halocarboxylic Acid-Degrading Bacteria with the Environmental Metagene Pool.Julian R. Marchesi, 2003.Culture-dependent and culture-independent approaches were used to determine the relationship between the dehalogenase gene pool in bacteria enriched and isolated on 2,2-dichloropropionic acid (22DCPA) and the environmental metagene pool (the collective gene pool of both the culturable and uncultured microbes) from which they were isolated . The dehalogenases in the pure-cultures isolates, which were able to degrade 22DCPA, were similar to previously described group I and II dehalogenases . Significantly, the majority of the dehalogenases isolated from activated sludge by degenerate PCR with primers specific for -halocarboxylic acid dehalogenases were not closely related to the dehalogenases in any isolate . Furthermore, the dehalogenases found in the pure cultures predominated in the enrichments but were a minor component of the community used to inoculate the batch cultures . Phylogenetic analysis of the dehalogenase sequences isolated by degenerate PCR showed that the diversity of the group II deh gene was greater than that of the group I deh gene . Direct plating of the activated sludge onto minimal media supplemented with 22DCPA resulted in biomass and DNA from which dehalogenases were amplified . Analysis of the sequences revealed that they were much more closely related to the sequences found in the community used to start the enrichments . However, no pure cultures were obtained with this isolation method, and thus no pure cultures were available for identification . In this study we examined the link between genes found in pure cultures with the metagene pool from which they were isolated . The results show that there is a large bias introduced by culturing, not just in the bacteria isolated but also the degradative genes that they contain . Moreover, our findings serve as a caveat for studies involving the culturing of pure cultures of bacteria and conclusions which are drawn from analysis of these organisms .
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