April 07, 2016 at 01:59AM

Tody I Learned: 1) Today I have some super cool facts about B. subtilis, a commonly-studied sporulating bacteria. B. subtilis (or B. sub, as it is affectionately known) grows into colonies on surfaces, just like most studied bacteria. The colony displays an interesting growth pattern. It grows quickly at first, but slows down as it becomes too big for nutrients to diffuse easily into the middle of the colony. Then, at somewhere around 50 or 100 microns, the colony starts growing in regular pulses. Apparently what happens is that bacteria near the center of the colony become nutrient limited by gluatamate, which is the primary nitrogen source for B. sub (and therefore more or less absolutely required for growth). When the colony gets big enough that glutamate can't diffuse easily to the center past the mass of growing bacteria on the perimeter, the whole colony will periodically stop growing for a bit to let glutamate through. This lets the colony keep growing while keeping the cells in the interior alive. Now, how does a population of bacteria communicate their pulse across a colony that could be hundreds of microns across? The answer's a doozy -- they use propagating electrochemical signals that are functionally reminiscent of (and possibly the ancestral basis for) neuronal communication. Here's how it works. B. sub, like pretty much all cells, keeps a ton of (positively charged) potassium ions in its interior. This potassium, among other things, maintains an electrical potential of about -150 mV across the cell membrane. B. sub has a membrane channel that, when activated, lets out potassium ions, depolarizing the cell and causing the electrical potential to go away, and spewing tons of potassium into the surrounding environment. The channel is thought to be held closed by glutamate. When intracellular glutamate is low, the channels open. B. sub also has membrane channels that actively pump glutamate into the cell, BUT these channels rely on the membrane potential to function. So when a neighboring cell fires, the efflux of potassium ions depolarizes a B. sub's membrane, turning off its glutamate channels. The cell pretty quickly runs out of glutamate, which triggers its potassium channels, spewing out more potassium to trigger the next cell to fire. Using this active firing mechanism, B. sub can communicate across a hundreds-of-micron-diameter colony in a few minutes, which is *way* faster than it could communicate by diffusion. These spikes also look uncannily like neuronal firing, and honestly, the mechanism is rather similar. 2) When bacterial biofilms reach a certain size, they undergo complex patterns of (programmed?) cell death. This causes the biofilm to buckle*, forming ridge-like "veins", like you can see in this colony of Pseudomonas aeruginosa: http://ift.tt/1RQqWdV. Apparently scientists have recently observed fluid movement through these channels, suggesting that they *may* serve to move nutrients around the colony, which at this point has a lot of difficulty getting nutrients to its center. * This is because the whole surface of the colony is under a lot of physical strain -- it's full of bacteria that are actively growing and pushing on their neighbors. When a hole opens up, the whole thing expands into it, which causes things like buckling. 3) ...how to calculate Ct in qPCR (the cycle at which an amplified sample crosses a threshold, which is used to back-calculate the starting concentration of the sample). In particular, I learned that the threshold value is basically arbitrary, as long as it isn't in the noise range or the plateau. The rest is just linear interpolation.