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A long-standing hypothesis regarding antibiotic resistance assumes that the evolution of resistant genotypes comes at a fitness cost to individuals. This predicts that resistance will disappear from populations when antibiotic stress is removed because susceptible individuals can then outcompete resistant individuals. However, recent studies have shown that low fitness costs and the evolution of fitness compensation cause very low levels of reversion to susceptibility in some situations. This study evaluated the fitness parameters associated with ampicillin resistance in Escherichia coli under optimal, temperature stressed, and nutrient limited conditions by comparing optical density values between resistant and susceptible individuals. Unexpectedly, the ampicillin resistant population grew faster than the susceptible population under optimal and limited nutrient conditions and experienced no fitness cost under temperature stress conditions. Though ampicillin is not used in clinical settings and E. coli may have different fitness parameters for other antibiotics, using ampicillin as a model suggests that reversion to susceptibility may be unlikely under both optimal growing conditions and stressful conditions. This study illustrates the need for greater understanding of each species' fitness parameters for individual antibiotics and the development of antibiotics that have higher fitness costs and low probabilities of compensation evolution.
Magnesium (Mg2 ) is the most abundant divalent cation in living cells and plays structural and biochemical roles in many cell processes. Although the importance of Mg2 as a cellular nutrient has been well established, the process by which organisms obtain Mg2 from their environment is still unclear. The Gram-positive bacterium Bacillus subtilis is used as a model system to understand Mg2 uptake and utilization in bacteria. The B. subtilis genome codes for a homolog of the MgtE family of Mg2 transporters. The protein is 34% identical to the structurally well-characterized MgtE homolog in Thermus thermophilus. This study was designed to characterize phenotypes of a B. subtilis mutant deleted for the mgtE gene. Compared to wild type, the ΔmgtE strain exhibits a Mg2 -dependent growth defect, consistent with a role of MgtE in Mg2 transport. Additionally, mutant cells are more resistant to manganese, show a decrease in sporulation efficiency, and are more sensitive to hydrogen peroxide than wild type cells.
Morphogenesis is a fundamental developmental process that encompasses myriad cellular processes, including cell shape changes that determine an organism's form and function. The RhoA signal transduction pathway is highly conserved at cellular and molecular levels and significantly influences morphogenetic processes through regulation of cell shape changes. RhoA accomplishes this, in part, by regulating contraction of nonmuscle myosin and the subsequent sliding of the apical actin cytoskeleton to change cell shape. Since these processes are so highly conserved, Drosophila melanogaster serves as an effective model organism for these studies. Drosophila requires RhoA function during embryonic and pupal morphogenesis. RhoA mutants are recessive embryonic lethals that disrupt head involution. However, later developmental stages also require RhoA. In order to study the RhoA pathway during post-embryonic phases, a heat-inducible wild-type RhoA transgene (hRh) was constructed using PCR amplification and standard subcloning techniques. The hRh transgene was inserted into the Drosophila genome by injection into pre-cellularized Drosophila embryos by Rainbow Transgenic Flies, Inc. Seven transgenic lines were generated, and insertion locations were chromosomally mapped by a series of segregation analysis matings. Four of the seven lines had single transgene inserts; one of them mapped to the X-chromosome and the other three mapped to the Second chromosome. The remaining three lines exhibited two or three insertions within the genome, mapping to the Second, Third and/or Fourth chromosomes. A series of crosses were performed to accurately map these insertions while simultaneously generating sublines from each of them that carry only a single transgene insertion. Finally, the X-linked hRh transgene insertion was successfully introduced into a RhoA loss-of-function mutant background. In the future, these stocks will be analyzed for heat-induced rescue of the RhoA embryonic lethality.
Previous data suggest that perceived foot heat and comfort may not coincide with actual foot temperature during treadmill running and that different shoe and sock materials can variously impact foot temperature. The purpose of this experiment was to determine if there were significant differences in foot temperature produced by cotton versus synthetic socks during longer runs, which more closely resemble endurance training, and if subjects were able to perceive those differences in terms of either comfort or temperature. Twelve adult males (22.4 ± 1.8 yrs, 180.6 ± 1.2 cm, 70.1 ± 1.6 kg) participated on two separate occasions one week apart. Subjects ran for 30 minutes with two temperature probes attached to the lateral dorsal aspect of the right foot in the same location: one directly on the skin and the other on the sock. All subjects wore the same shoe model. Foot temperature, heart rate, heat perception, and comfort perception were recorded. Perception was measured by using 10 cm visual analogue scales. Each subject ran once in a cotton-based sock and once in a synthetic (olefin-based) sock. Subjects perceived no significant difference in comfort or temperature between cotton and synthetic socks, and heart rate did not vary significantly between the two trials. The olefin-based sock was associated with significantly lower absolute temperatures at the sock thermometer site but not the skin thermometer site. However, changes in temperature from one time point to the next were the same between the two socks for either thermometer site. The results cannot conclusively state that one sock has an advantage over the other, but they suggest olefin-based socks may dissipate heat better than cotton-based socks under certain conditions.
Sin Nombre virus (SNV) is a hantavirus hosted almost exclusively by deer mice (Peromyscus maniculatus) that causes high (∼40%) mortality in humans. Transmission patterns of SNV in natural host populations are not well understood and, as such, represent a knowledge gap that has significant implications for human health. Since its identification in the early 1990s, the Four Corners region of the southwestern United States (i.e. geographic area encompassing southeastern UT, northeastern AZ, southwestern CO and northwestern NM) has remained a “hotspot” for SNV infections in both deer mice and humans; however, the reason for this skewed geographic distribution of SNV remains unclear. Therefore, our objective was to observe regional patterns of SNV prevalence in deer mice along a latitudinal gradient in the Four Corners region and determine the extent to which prevalence is influenced by factors including deer mouse density, small mammal diversity, vegetation cover, plant moisture content, soil pH and soil water content. Although we did not find a significant correlation between SNV prevalence and small mammal species diversity, SNV prevalence and deer mouse density were positively correlated. We found no relationship between soil pH and SNV prevalence; however, there was a strong negative correlation between SNV prevalence and soil water content. We also found strong positive correlations between SNV prevalence and total vegetation cover and between SNV prevalence and plant moisture content. Collectively, our results suggest that SNV prevalence is driven primarily by increased deer mouse densities and that deer mice are more likely to be associated with habitats that have higher plant moisture content, as well as greater total vegetation cover. Our results also indicate that, whereas soil pH is not a direct predictor of SNV prevalence, sites with dry soils at pH levels close to the optimal for SNV (6.9) that also contain high densities of deer mice may be hantavirus “hot spots”.
Escherichia coli and Salmonella enterica species are Gram-negative bacteria that have recently shown acquired resistance to multiple antimicrobials. These resistant strains, referred to as multidrug resistant (MDR), are not effectively inhibited by current antimicrobials and therefore cause a range of untreatable infections. Peptide nucleic acids (PNA) have been proposed as an alternative antimicrobial treatment. These synthetic nucleic acid mimics are effective through steric inhibition of bacterial transcription and translation. This study examines the effect of PNA on MDR strains of E. coli and S. enterica serovar Typhimurium (S. Typhimurium). Kirby-Bauer disk diffusion susceptibility and minimum inhibitory concentration determinations (MIC) were completed to show bacterial resistance levels to current antimicrobials. Using S. Typhimurium specific dnaK PNA, MIC was determined to assess the effectiveness of PNA on MDR strains of E. coli and S. Typhimurium. Results showed that while the dnaK PNA was only mildly effective at inhibiting growth of MDR and control E. coli strains, there was a more noticeable effect of PNA on S. Typhimurium. Scrambled sequence control PNA did not have an effect. These results exemplify the species specificity of PNA and its ability to inhibit pathogenic bacteria in a targeted manner. Although future studies are still needed, the findings demonstrate that anti-dnaK PNA are an effective in vitro treatment for S. enterica strains, and that they are a possible alternative antimicrobial treatment for use against multidrug resistant bacterial infections.
The MTT assay was used to quantify the viability of human osteosarcoma cells (Saos-2) in the presence of chelators administered to alleviate cadmium-mediated cytotoxicity. The chelators evaluated were potassium bis(2-hydroxyethyl)dithiocarbamate, K[bhedtc] (1) and potassium O-[2-[bis(2-hydroxyethyl)amino]ethyl]dithiocarbonate hemihydrate, K[bhexan]·0.5H2O (2). In this work, cell viability was measured as a function of chelator concentration (0-1000 μM) and administration delay post cadmium inoculation (0-16 hrs). Upon simultaneous administration, cytotoxicity mediated by the presence of 100 μM CdCl2 was best alleviated by the presence of 100 μM chelator 1. Chelator 2 was much less potent: optimal alleviation was attained at 1000 μM (1 mM) 2. Relief from cadmium toxicity was still afforded by 100-500 μM chelator 1 or 600-1000 μM chelator 2 administered up to 8 hours after 100 μM CdCl2. However, cell viability lower than that exhibited by 100 μM CdCl2 (the -control) was observed for chelator 1 when the delay was extended to 16 hours. These results indicate that chelator 1 is more potent at alleviating cadmium-mediated cytotoxicity, but its effectiveness is more sensitive to administration delay than that of chelator 2. Overall this work has demonstrated a method to evaluate the efficacy of chelators to alleviate cadmium-mediated cytotoxicity.
Transposable elements, mobile DNA sequences present in virtually all organisms, have previously been thought to be inactive in mammals. However, current studies have revealed the first evidence of recent piggyBac and hAT activity in Myotis lucifugus. The objective of this study was to further characterize the abundance and diversity of five transposable elements in various Myotis bats. The five elements were confirmed present in 10 species (M. ciliolabrum, M. californicus, M. velifer, M. yumanesis, M. lucifugus, M. thysanodes, M. volans, M. austroriparius, and M. keaysi) spanning the new world clade of Myotis using PCR. Preliminary sequence data focusing on the transposase region was collected and analyzed for two of the elements; hAT3 data set contained 1200 bp and piggyBac1 contained 942 bp. The older hAT3 element had a mean sequence diversity of 1.88%; whereas the younger piggyBac1 element had little diversity (0.01%). A small number of polymorphic sites within the sequence of a single individual suggested multiple copy types of the hAT3 element; these polymorphic sites were not evident in the piggyBac1 element. These preliminary results support the hypothesis that the piggyBac element is a relatively young transposon and a recent addition to the Myotis genome.