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Uptake and Replication of Salmonella enterica in Acanthamoeba rhysodes. Dilek Tezcan-Merdol, 2004.The ability of salmonellae to become internalized and to survive and replicate in amoebae was evaluated by using three separate serovars of Salmonella enterica and five different isolates of axenic Acanthamoeba spp . In gentamicin protection assays, Salmonella enterica serovar Dublin was internalized more efficiently than Salmonella enterica serovar Enteritidis or Salmonella enterica serovar Typhimurium in all of the amoeba isolates tested . The bacteria appeared to be most efficiently internalized by Acanthamoeba rhysodes . Variations in bacterial growth conditions affected internalization efficiency, but this effect was not altered by inactivation of hilA, a key regulator in the expression of the invasion-associated Salmonella pathogenicity island 1 . Microscopy of infected A . rhysodes revealed that S . enterica resided within vacuoles . Prolonged incubation resulted in a loss of intracellular bacteria associated with morphological changes and loss of amoebae . In part, these alterations were associated with hilA and the Salmonella virulence plasmid . The data show that Acanthamoeba spp . can differentiate between different serovars of salmonellae and that internalization is associated with cytotoxic effects mediated by defined Salmonella virulence loci . Description and Interpretation of Adaptive Evolution of Escherichia coli K-12 MG1655 by Using a Genome-Scale In Silico Metabolic Model. Stephen S. Fong, 2003.Genome-scale in silico metabolic networks of Escherichia coli have been reconstructed . By using a constraint-based in silico model of a reconstructed network, the range of phenotypes exhibited by E . coli under different growth conditions can be computed, and optimal growth phenotypes can be predicted . We hypothesized that the end point of adaptive evolution of E . coli could be accurately described a priori by our in silico model since adaptive evolution should lead to an optimal phenotype . Adaptive evolution of E . coli during prolonged exponential growth was performed with M9 minimal medium supplemented with 2 g of  -ketoglutarate per liter, 2 g of lactate per liter, or 2 g of pyruvate per liter at both 30 and 37°C, which produced seven distinct strains . The growth rates, substrate uptake rates, oxygen uptake rates, by-product secretion patterns, and growth rates on alternative substrates were measured for each strain as a function of evolutionary time . Three major conclusions were drawn from the experimental results . First, adaptive evolution leads to a phenotype characterized by maximized growth rates that may not correspond to the highest biomass yield . Second, metabolic phenotypes resulting from adaptive evolution can be described and predicted computationally . Third, adaptive evolution on a single substrate leads to changes in growth characteristics on other substrates that could signify parallel or opposing growth objectives . Together, the results show that genome-scale in silico metabolic models can describe the end point of adaptive evolution a priori and can be used to gain insight into the adaptive evolutionary process for E . coli .
TcaR, a Putative MarR-Like Regulator of sarS Expression. Nadine McCallum, 2004.TcaR, which shares sequence homology with MarR-like transcriptional regulators, has been identified as a novel Staphylococcus aureus regulator affecting the expression of the global regulatory element SarS (SarH1), as well as that of the cell surface-associated protein SasF (N315-SA2439) . Microarray analysis, confirmatory Northern blots, and genetic complementation experiments showed that TcaR upregulates sarS and thus spa transcription . In addition, it attenuates whole-length transcription of sasF, thereby producing a truncated transcript lacking the 3' terminus, which codes for the cell wall anchor motif . Hence, in strains containing an intact tcaR gene, TcaR is likely to decrease the amount of the surface-associated protein SasF and to increase that of the surface-associated protein A . The widely used laboratory strains derived from NCTC8325 were found to be natural, truncated mutants of tcaR, harboring an inactive TcaR and therefore expressing very low levels of sarS . The data presented here identified TcaR as a further activator of sarS, and a modulator of sasF expression that has to be taken into account in studies of virulence gene expression in S . aureus . Inactivation of Bacteria in Seawater by Low-Amperage Electric Current. Jong-Chul Park, 2003.Seawater used in mariculture has been suspected of being a potential source of infection . In this study, the lethal effects of low-amperage electric treatment on microorganisms were examined in natural seawater and in seawater inoculated with Vibrio parahaemolyticus . In both cases, bacteria including V . parahaemolyticus in seawater were completely eliminated in 100 ms by a 0.5-A, 12-V direct current . Electron microscopic investigation of the electrically treated bacteria revealed substantial structural damage at the cellular level . In conclusion, our results indicate that low-amperage electric treatment is effective for rapid inactivation of microorganisms in seawater . Identification of cry1I-Type Genes from Bacillus thuringiensis Strains and Characterization of a Novel cry1I-Type Gene. Fuping Song, 2003.A PCR-restriction fragment length polymorphism method for identification of cry1I-type genes from Bacillus thuringiensis was established by designing a pair of universal primers based on the conserved regions of the genes to amplify 1,548-bp cry1I-type gene fragments . Amplification products were digested with the Bsp119I and BanI enzymes, and four kinds of known cry1I-type genes were successfully identified . The results showed that cry1I-type genes appeared in 95 of 115 B . thuringiensis isolates and 7 of 13 standard strains . A novel cry1I-type gene was found in one standard strain and six isolates . The novel cry1I gene was cloned from B . thuringiensis isolate Btc007 and subcloned into vector pET-21b . Then it was overexpressed in Escherichia coli BL21(DE3) . The expressed product was shown to be toxic to the diamondback moth (Plutella xylostella), Asian corn borer (Ostrinia furnacalis), and soybean pod borer (Leguminivora glycinivorella) . However, it was not toxic to the cotton bollworm (Helicoverpa armigera), beet armyworm (Spodoptera exigua), or elm leaf beetle (Pyrrhalta aenescens) in bioassays . Subsequently, the Cry protein encoded by this novel cry gene was designated Cry1Ie1 by the B . thuringiensis  -endotoxin nomenclature committee .
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