E.coli DH5α cell response to a sudden change in microfluidic chemical environment
05 Nov 2015-Vol. 2015, pp 3213-3216
TL;DR: The studies revealed that when E.coli DH5α cells were exposed to 0.1 mM sorbitol, they showed faster chemotaxis towards the attractant and achieved steady state by 60 min, which shows that the bacterial cells respond to change in local chemical environment is within few minutes.
Abstract: Motile bacteria respond to changing chemical environment by moving towards or away from a particular location. Bacterial migration under chemical gradient is one of the most studied areas in biomedical field. In this work we looked into how bacterial cells respond to sudden change in the microfluidic chemical environment. E.coli DH5α cells were subjected to an attractant gradient (0.1 mM sorbitol - attractant to E.coli cells) and after 120 min the same cells were exposed to an inhibitor (0.1 mM NiSO 4 ) gradient in the same microfluidic device. Our studies revealed that when the E.coli DH5α cells were exposed to 0.1 mM sorbitol, they showed faster chemotaxis towards the attractant (0.1 mM sorbitol) and achieved steady state by 60 min. When we replaced 0.1 mM sorbitol with 0.1 mM NiSO 4 in the device we found that that the E.coli DH5α cells started responding to change in chemical environment within 10 min and achieved steady state at the end of 60 min. This shows that the bacterial cells respond to change in local chemical environment is within few minutes.
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TL;DR: Pilot experiments show that vectors only persist in indigenous populations when under selection pressure, disappearing when this carbon source is removed, which could prime indigenous bacteria for degrading pollutants while providing minimal ecosystem disturbance.
Abstract: Engineering bacteria to clean-up oil spills is rapidly advancing but faces regulatory hurdles and environmental concerns. Here, we develop a new technology to harness indigenous soil microbial communities for bioremediation by flooding local populations with catabolic genes for petroleum hydrocarbon degradation. Overexpressing three enzymes (almA, xylE, p450cam) in Escherichia coli led to degradation of 60–99% of target hydrocarbon substrates. Mating experiments, fluorescence microscopy and TEM revealed indigenous bacteria could obtain these vectors from E. coli through several mechanisms of horizontal gene transfer (HGT), including conjugation and cytoplasmic exchange through nanotubes. Inoculating petroleum-polluted sediments with E. coli carrying the vector pSF-OXB15-p450camfusion showed that the E. coli cells died after five days but a variety of bacteria received and carried the vector for over 60 days after inoculation. Within 60 days, the total petroleum hydrocarbon content of the polluted soil was reduced by 46%. Pilot experiments show that vectors only persist in indigenous populations when under selection pressure, disappearing when this carbon source is removed. This approach to remediation could prime indigenous bacteria for degrading pollutants while providing minimal ecosystem disturbance.
29 citations
TL;DR: The results show that the spherical AuNPs intervenes in the themotaxis of E. coli DH5α cells and inhibits the cell migration, which may be due to decreased F-type ATP synthase activity and collapse of membrane potential by Au NPs, which, in turn, leads to decreased ATP levels.
Abstract: Bacteria responds to changing chemical and thermal environment by moving towards or away from a particular location. In this report, we looked into thermal gradient generation and response of E. coli DH5α cells to thermal gradient in the presence and in the absence of spherical gold nanoparticles (size: 15 to 22 nm) in a static microfluidic environment using a polydimethylsiloxane (PDMS) made microfluidic device. A PDMS-agarose based microfluidic device for generating thermal gradient has been developed and the thermal gradient generation in the device has been validated with the numerical simulation. Our studies revealed that the presence of gold nanoparticles, AuNPs (0.649 μg/mL) has no effect on the thermal gradient generation. The E. coli DH5α cells have been treated with AuNPs of two different concentrations (0.649 μg/mL and 0.008 μg/mL). The thermotaxis behavior of cells in the presence of AuNPs has been studied and compared to the thermotaxis of E.coli DH5α cells in the absence of AuNPs. In case of thermotaxis, in the absence of the AuNPs, the E. coli DH5α cells showed better thermotaxis towards lower temperature range, whereas in the presence of AuNPs (0.649 μg/mL and 0.008 μg/mL) thermotaxis of the E. coli DH5α cells has been inhibited. The results show that the spherical AuNPs intervenes in the themotaxis of E. coli DH5α cells and inhibits the cell migration. The reason for the failure in thermotaxis response mechanism may be due to decreased F-type ATP synthase activity and collapse of membrane potential by AuNPs, which, in turn, leads to decreased ATP levels. This has been hypothesized since both thermotaxis and chemotaxis follows the same response mechanism for migration in which ATP plays critical role.
7 citations
TL;DR: In this article, the authors developed a new technology to harness indigenous soil microbial communities for bioremediation by flooding local populations with catabolic genes for petroleum hydrocarbon degradation, which could prime indigenous bacteria for degrading pollutants.
Abstract: Engineering bacteria to clean-up oil spills is rapidly advancing but faces regulatory hurdles and environmental concerns. Here, we develop a new technology to harness indigenous soil microbial communities for bioremediation by flooding local populations with catabolic genes for petroleum hydrocarbon degradation. Overexpressing three enzymes (almA, xylE, p450cam) in E.coli led to degradation rates of 60-99% of target hydrocarbon substrates. Mating experiments, fluorescence microscopy and TEM revealed indigenous bacteria could obtain these vectors from E.coli through conjugation. Inoculating petroleum-polluted sediments from a former refinery with engineered E.coli showed that the E.coli die after five days but a variety of bacteria received and carried the vector for over 120 days after inoculation. This approach could prime indigenous bacteria for degrading pollutants while providing minimal ecosystem disturbance.
2 citations
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TL;DR: A microfluidic thermal gradient system (muTGS) system is proposed which can create and maintain a thermal gradient throughout a cell-seeded gel matrix using the hot and cold water supply integrated in the system in the form of a countercurrent heat exchanger.
Abstract: Studies on the effects of variations in temperature and mild temperature gradients on cells, gels, and scaffolds are important from the viewpoint of biological function. Small differences in temperature are known to elicit significant variations in cell behavior and individual protein reactivity. For the study of thermal effects and gradients in vitro, it is important to develop microfluidic platforms which are capable of controlling temperature gradients in an environment which mimics the range of physiological conditions. In the present paper, such a microfluidic thermal gradient system (muTGS) system is proposed which can create and maintain a thermal gradient throughout a cell-seeded gel matrix using the hot and cold water supply integrated in the system in the form of a countercurrent heat exchanger. It is found that a uniform temperature gradient can be created and maintained in the device even inside a high temperature and high humidity environment of an incubator. With the help of a hot and cold circuit controlled from outside the incubator the temperature gradient can be regulated. A numerical simulation of the device demonstrates the thermal feature of the chip. Cell viability and activity under a thermal gradient are examined by placing human breast cancer cells in the device.
26 citations
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TL;DR: Most organisms, from bacteria to humans, can follow temperature gradients to an optimal temperature, but experience influences eukaryotes' preferred temperature, and bacteria also adjust their temperature preference depending on growth conditions.
Abstract: Most organisms, from bacteria to humans, can follow temperature gradients to an optimal temperature. Experience influences eukaryotes' preferred temperature, and, as it turns out, bacteria also adjust their temperature preference depending on growth conditions.
6 citations