16 research outputs found
Research of White Sucker Cell lines and Associated Viruses
Catostomus commersonii, also known as the White Sucker, is a vital part of most aquatic ecosystems in the U.S. Specifically looking at the Upper Midwest, these fish are widely distributed around the states of Minnesota and Wisconsin. The importance of these organisms is due to their species reputation, they are indicator species that are very useful in informing whether the environment around them is healthy or if contamination has occurred. To better understand the organism, we needed to start at the molecular level. Understanding the nature of White Sucker cells through cell cultures. To get the individual fins of the White Sucker to flasks with growth media and begin to have the cells grow from the tissue. Through process of cell culturing, the cells differentiated from one another through each passage and adapted to different media to create contrasting growth rates. The differentiating cells were then characterized through DNA barcoding and examined for susceptibility to a variety of viruses to obtain better information about the multiple cell types from the fins of the White Suckerhttps://openriver.winona.edu/urc2018/1045/thumbnail.jp
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Divergent evolution of the activity and regulation of the glutamate decarboxylase systems in Listeria monocytogenes EGD-e and 10403S : roles in virulence and acid tolerance
The glutamate decarboxylase (GAD) system has been shown to be important for the survival of Listeria monocytogenes in low pH environments. The bacterium can use this faculty to maintain pH homeostasis under acidic conditions. The accepted model for the GAD system proposes that the antiport of glutamate into the bacterial cell in exchange for γ-aminobutyric acid (GABA) is coupled to an intracellular decarboxylation reaction of glutamate into GABA that consumes protons and therefore facilitates pH homeostasis. Most strains of L. monocytogenes possess three decarboxylase genes (gadD1, D2 & D3) and two antiporter genes (gadT1 & gadT2). Here, we confirm that the gadD3 encodes a glutamate decarboxylase dedicated to the intracellular GAD system (GADi), which produces GABA from cytoplasmic glutamate in the absence of antiport activity. We also compare the functionality of the GAD system between two commonly studied reference strains, EGD-e and 10403S with differences in terms of acid resistance. Through functional genomics we show that EGD-e is unable to export GABA and relies exclusively in the GADi system, which is driven primarily by GadD3 in this strain. In contrast 10403S relies upon GadD2 to maintain both an intracellular and extracellular GAD system (GADi/GADe). Through experiments with a murinised variant of EGD-e (EGDm) in mice, we found that the GAD system plays a significant role in the overall virulence of this strain. Double mutants lacking either gadD1D3 or gadD2D3 of the GAD system displayed reduced acid tolerance and were significantly affected in their ability to cause infection following oral inoculation. Since EGDm exploits GADi but not GADe the results indicate that the GADi system makes a contribution to virulence within the mouse. Furthermore, we also provide evidence that there might be a separate line of evolution in the GAD system between two commonly used reference strains
Divergent evolution of the activity and regulation of the glutamate decarboxylase systems in listeria monocytogenes egd-e and 10403s: roles in virulence and acid tolerance
The glutamate decarboxylase (GAD) system has been shown to be important for the survival of Listeria monocytogenes in low pH environments. The bacterium can use this faculty to maintain pH homeostasis under acidic conditions. The accepted model for the GAD system proposes that the antiport of glutamate into the bacterial cell in exchange for c-aminobutyric acid (GABA) is coupled to an intracellular decarboxylation reaction of glutamate into GABA that consumes protons and therefore facilitates pH homeostasis. Most strains of L. monocytogenes possess three decarboxylase genes (gadD1, D2 & D3) and two antiporter genes (gadT1 & gadT2). Here, we confirm that the gadD3 encodes a glutamate decarboxylase dedicated to the intracellular GAD system (GAD(i)), which produces GABA from cytoplasmic glutamate in the absence of antiport activity. We also compare the functionality of the GAD system between two commonly studied reference strains, EGD-e and 10403S with differences in terms of acid resistance. Through functional genomics we show that EGD-e is unable to export GABA and relies exclusively in the GADi system, which is driven primarily by GadD3 in this strain. In contrast 10403S relies upon GadD2 to maintain both an intracellular and extracellular GAD system (GAD(i)/GAD(e)). Through experiments with a murinised variant of EGD-e (EGDm) in mice, we found that the GAD system plays a significant role in the overall virulence of this strain. Double mutants lacking either gadD1D3 or gadD2D3 of the GAD system displayed reduced acid tolerance and were significantly affected in their ability to cause infection following oral inoculation. Since EGDm exploits GAD(i) but not GAD(e) the results indicate that the GAD(i) system makes a contribution to virulence within the mouse. Furthermore, we also provide evidence that there might be a separate line of evolution in the GAD system between two commonly used reference strains
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Interfacial Solar Evaporation by a 3D Graphene Oxide Stalk for Highly Concentrated Brine Treatment.
In this work, we demonstrate a 3-dimensional graphene oxide (3D GO) stalk that operates near the capillary wicking limit to achieve an evaporation flux of 34.7 kg m-2 h-1 under 1 sun conditions (1 kW/m2). This flux represents nearly a 100 times enhancement over a conventional solar evaporation pond. Interfacial solar evaporation traditionally uses 2D evaporators to vaporize water using sunlight, but their low evaporative water flux limits their practical applicability for desalination. Some recent studies using 3D evaporators demonstrate potential for more efficient water transfer, but the flux improvement has been marginal because of a low evaporation area index (EAI), which is defined as the ratio of the total evaporative surface area to the projected ground area. By using a 3D GO stalk with an ultrahigh EAI of 70, we achieved nearly a 20-fold enhancement over a 2D GO evaporator. The 3D GO stalk also exhibited additional advantages including omnidirectional sunlight utilization, a high evaporation flux under dark conditions from more efficient utilization of ambient heating, a dramatic increase of the evaporation rate by introducing wind, and scaling resistance in evaporating brines with a salt content of up to 17.5 wt %. This performance makes the 3D GO stalk well suited for the development of a low-cost, reduced footprint technology for zero liquid discharge in brine management applications
The standard model for the action of the GAD system.
<p>(<b>a</b>) A membrane bound antiporter carries glutamate into the cell in exchange for GABA. A cytosolic decarboxylase enzyme converts glutamate to GABA, with a consumption of H<sup>+</sup>. (<b>b</b>) The genomic structure of the genes encoding the GAD system in <i>L. monocytogenes</i> EGD-e.</p
GABA production from <i>L. monocytogenes gad</i> mutants.
<p>(<b>a</b>) Production of GABA<sub>e</sub> by EGDm and 10403S gad mutants with (grey) or without (black) 1 h exposure to acid at pH 4.0 (EGDm) or pH 3.5 (10403S). (<b>B</b>) Production of GABA<sub>i</sub> by EGDm and 10403S <i>gadD</i> mutants with (grey) or without (black) 1 h exposure to acid at pH 4.0 (EGDm) or pH 3.5 (10403S). Dashed horizontal lines indicate the detection limits for GABA in each experiment. Error bars represent the standard deviation from the mean of three individual biological repeats for each sample. An asterix represents signifcant difference of less than 0.05 between a given mutant and respective wild-type as determined by a student’s <i>t-</i>test.</p
Acid survival of <i>L. monocytogenes gad</i> mutants.
<p>Stationary phase EGDm (<b>a</b>) and 10403S (<b>b</b>) Δ<i>gad</i> mutants were challenged at pH 2.5. Cell counts were taken every 20 min. Values are the means of data from three individual cultures, with the cell counts for each culture being the means of counts from three platings. Relative transcript levels of EGDm <b>(c)</b> or 10403S (<b>d</b>) <i>gadD1</i> (dark grey fill), <i>gadD2</i> (hatched) and <i>gadD3</i> (grey) genes to 16S gene prior to acid exposure in each mutant strain. Error bars represent the standard error from the mean value of three individual biological repeats. The numbers over the bar charts (<b>c & d</b>) indicate the <i>p</i>-value for the difference between each gene expression compared to wild-type levels as determined by student’s <i>t</i>-test.</p
Acid survival and GABA<sub>i</sub> production of <i>L. monocytogenes</i> EGDm double GAD system mutants indicates a key role for <i>gadD3</i>.
<p>(<b>a</b>) Stationary phase EGDm <i>gadD</i> mutants were acidified to pH 2.5 with 3 M HCl in BHI broth. Cell counts were taken every 20 min. Values are the means of data from three individual cultures, with the cell counts for each culture being the means of counts from three platings. Error bars represent the standard deviation from the mean value for each time-point. (<b>b</b>) Stationary phase EGDm gad mutants were acidified (grey) or not acidified (black) with 3 M HCl to pH 4.0 and GABA<sub>i</sub> accumulation was quantified. Error bars represent the standard deviation from the mean of three independent biological replicates.</p