| 00:01 | Welcome. This is uh chapter six we're talking about microbial growth. Uh |
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| 00:06 | gonna stretch this out over two Uh Part one focuses on um requirements |
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| 00:13 | growth, both chemical and physical um well as uh bacterial growth medium. |
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| 00:20 | . In the next module, part , we'll talk about growth dynamics, |
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| 00:25 | growth curves and um biofilms. So, uh as always, uh |
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| 00:32 | are uh uh lectures are preceded by learning objectives. So just think of |
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| 00:38 | as a checklist uh after you've gone the material, uh you should uh |
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| 00:44 | familiar with what the, what these objectives are stating. OK. So |
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| 00:49 | go into microbial growth. So uh microbes can grow very fast, |
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| 00:56 | ? Um Are are can under optimal . Uh They have very diverse metabolisms |
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| 01:01 | we saw in chapter five, utilize variety of different energy sources. Um |
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| 01:07 | so when we look at growth, , we're basically combining the various requirements |
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| 01:12 | growth into what we call a growth that we then inoculate and then observe |
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| 01:18 | . OK. And of course, we do that, uh we're all |
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| 01:22 | that we can look at growth in ways we can look at it under |
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| 01:25 | microscope, taking uh a sample of culture that's been inoculated and, and |
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| 01:31 | the uh can see the the increase cell density over time. As you |
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| 01:34 | here in liquid medium. Uh you also of course, take samples of |
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| 01:40 | liquid medium and put them on solid and see that uh as well. |
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| 01:47 | , there's a, there's an increase the number of colonies. So cells |
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| 01:51 | cells are plated out, they And of course, a a denser |
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| 01:55 | will produce more colonies on a And of course, we can monitor |
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| 01:58 | over time. So um and of , we can look at a liquid |
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| 02:03 | and we can see that the the culture there is very uh turbid uh |
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| 02:09 | cloudy. And as you um cells in liquid in a liquid uh |
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| 02:15 | um more cells cells multiply those more and that creates more turbidity. |
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| 02:20 | And we actually, we can actually that using a spectrophotometer. It from |
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| 02:25 | or devices that will allow you to substance that you're measuring will absorb |
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| 02:30 | So of course, cells can do same thing as they multiply liquid. |
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| 02:33 | can, we can moderate the absorbance light. And as as the turbidity |
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| 02:37 | , we can produce a growth curve you see here. OK. Now |
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| 02:41 | , we'll talk about this in the part, but this is how you |
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| 02:44 | these things is by formulating growth supplying the nutrients, not them with |
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| 02:50 | and then following their pattern of growth as they go through what's called the |
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| 02:56 | batch growth cycle, it's often Ok. And so um so for |
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| 03:04 | this first part, we're gonna mainly focus on what are the requirements for |
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| 03:09 | and the different types of growth media we can use. Ok. So |
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| 03:13 | course, we're talking about requirements for . Think of that as what are |
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| 03:16 | conditions that will make the cells happy want to divide at a fast |
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| 03:21 | And of course, there's gonna there's gonna be physical uh parameters, |
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| 03:25 | temperature creates so concentrations. Um and course, chemical, the the uh |
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| 03:32 | of the molecules living cells are made including us, of course, uh |
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| 03:39 | comprised of these basic elements. Uh abbreviate as CHO NPS. Uh and |
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| 03:46 | term macron nutrient refers to the fact needed in higher quantities typically because they're |
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| 03:51 | used more. OK. And of , then uh also falls in there |
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| 03:56 | trace elements which are essential but are much smaller quantities. And of |
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| 04:01 | we look at oxygen as well and um different microbes have different responses |
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| 04:06 | to oxygen. OK. So uh about physical requirements first, so we'll |
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| 04:12 | on uh temperature, osmotic pressure. uh ph so as you collectively, |
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| 04:20 | three parameters um are optimized, uh microbes will have different optima for these |
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| 04:30 | . Ok. Regardless it's all about it's temperature, whether it's ph, |
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| 04:36 | it's so uh osmotic pressure, it's about what's going to make the proteins |
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| 04:41 | a cell happy, right? Because the function of any living thing is |
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| 04:45 | to the function of proteins for the part and keeping proteins um functioning properly |
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| 04:53 | key, it's key, it's the to, to being able to |
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| 04:56 | And so proteins have particular um um at which they function. And so |
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| 05:05 | you recall protein structure, right, assume once they're synthesized, they perform |
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| 05:12 | certain shape and that shape is also for the functioning of that protein because |
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| 05:17 | is typically bind other molecules, other as part of the how they carry |
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| 05:22 | their functions. And if these are proteins aren't properly folded, they won't |
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| 05:29 | . All right. And what helps that folding is optimal temperature, optimal |
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| 05:35 | optimal Sodi concentrations, all these things to keep those proteins and keep their |
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| 05:42 | and function. OK. Um So with temperature, uh too high a |
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| 05:48 | will cause the proteins to come apart we call the nature. OK. |
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| 05:52 | that's detrimental function. So there's gonna organisms that, that have different, |
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| 05:58 | for temperatures because there are different Um as you see there, um |
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| 06:03 | mom, especially among bacteria and these can expand ranges of different conditions |
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| 06:09 | that there's a term called extremophiles, ? They can live on the very |
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| 06:13 | of these conditions. Now, when look at these terms, uh hyper |
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| 06:18 | means a file thermoph hyper Theophile. and for all these parameters, temperature |
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| 06:25 | popularity, most of life is gonna in the middle range, what |
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| 06:31 | what you, what you might call normal conditions. OK? Whether it's |
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| 06:37 | most limit things are around neutrality ph . Uh for temperature, most things |
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| 06:42 | in the temperature range, you see . OK? But there are of |
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| 06:46 | , uh those that can live above below those conditions, right? |
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| 06:50 | quote normal conditions, right. So of course, you're a meso |
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| 06:57 | right? Uh pathogens that would be infecting you would be measles as |
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| 07:02 | Um But as we go above temperature , so heat loving thermophilus and there's |
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| 07:06 | degrees hyper Theophile is very hot right? We see these, there's |
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| 07:11 | environments on earth uh could be in water environments that have these uh underwater |
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| 07:17 | , basically super heating the water, we call thermal vents. And we |
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| 07:20 | hyper the files living there also in Yellowstone Park, we have geysers and |
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| 07:25 | know, firstly boiling water, these uh basically steam coming out of the |
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| 07:30 | , then you actually can have bacteria in these uh areas. Uh CYC |
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| 07:35 | are in the, so these are can, can grow, can grow |
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| 07:40 | refrigeration temperature. OK. Um uh , in different ranges below where music |
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| 07:47 | live uh one of the um important there, uh, medically important strains |
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| 07:53 | listeria, uh, that causes uh, the food borne pathogen and |
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| 07:59 | can live, you can live and in your refrigerator on certain foods. |
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| 08:02 | one of the strains we'll talk about . But, uh, the point |
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| 08:05 | is that you, you're gonna have will fit, you know, |
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| 08:08 | most uh microbes will be in the range, but you're gonna have a |
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| 08:13 | of them that will be above and that. OK. Now, one |
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| 08:16 | the concepts to understand here this is just go for temperature. Although I'm |
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| 08:22 | the example of temperature here is the of tolerance versus optimal growth, |
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| 08:27 | You can use that same, that concept applies whether it's temperature, whether |
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| 08:30 | ph uh whether it's a number of physical parameters, right? Because um |
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| 08:36 | you're, if you're, there's a if you're tolerant condition and versus that's |
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| 08:41 | condition where you optimally grow. So I made an example here uh |
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| 08:46 | at temperature versus growth rate, And three different strains. So X |
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| 08:50 | course, is a meso file and can see it has an optimal temperature |
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| 08:54 | by that, I mean, it its highest growth rate at somewhere I |
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| 08:59 | around 34. So I'm just guessing uh temp uh sea temperature where it |
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| 09:04 | where it grows optimally and it's in mesa file range. Why is still |
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| 09:09 | the mess of file range, but is able to remain viable as shown |
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| 09:15 | the dash line right extending out OK. So, so yeah, |
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| 09:23 | growing very slowly obviously, but it able to be viable right in that |
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| 09:29 | . So you can say it's tolerating condition. It's not growing optimally obviously |
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| 09:34 | there's a meso file. But in higher temperature range, it can tolerate |
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| 09:38 | temperature for a period of time. of course, you have Z that's |
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| 09:42 | thermoph file, right? It's optimal it requires for growth is in the |
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| 09:47 | file range right here about 55 Uh And that's an optimal growth, |
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| 09:52 | ? It doesn't grow at temperatures below , very well below 45 degrees. |
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| 09:58 | . So hence it's a thermoph OK. Why is thermo tolerant? |
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| 10:04 | . It it's a meso file. is a meso fi but it's thermo |
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| 10:07 | ? OK. So just make sure understand the difference between those two that |
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| 10:12 | that differs, right? That OK. Um osmotic pressure. So |
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| 10:18 | talked about this previously in uh chapter on the Procter cell functions. Uh |
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| 10:24 | just to remind you, you the the uh osmotic pressure relates to |
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| 10:29 | availability, right? So high solute outside of a cell will draw water |
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| 10:35 | of that cell and which is which is not a good thing. |
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| 10:39 | mean, remember that all life, know, water is the the solvent |
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| 10:42 | living things right. And uh most life is comprised mostly of water. |
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| 10:48 | so it's gonna be very important to able to have a good supply of |
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| 10:51 | . And so that's where the cells kind of manipulate their sou concentrations |
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| 10:57 | And that will allow soyuz to kind flow in. So that's kind of |
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| 11:00 | is what this relates to. So remember that um the terms hypertonic |
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| 11:06 | always uh are relative to each other whatever, if it's hypertonic on one |
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| 11:12 | , it's hypotonic on the other side vice versa. OK. So these |
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| 11:17 | uh plasmosis, right? So that's a cell in hypertonic solution will lose |
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| 11:23 | , right? Uh And um uh you see the uh cell in the |
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| 11:31 | , right, in a 10% sodium concentration, water flows out of the |
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| 11:35 | and that uh plastic membrane shrinks, ? That's what we call plasmosis. |
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| 11:42 | hypertonic solution of course is on the . And that's where water flows |
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| 11:47 | So the cell is slightly hypertonic and what bacteria tend to do is to |
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| 11:50 | of keep your cell slightly hyper Hypnic the inside. So that water flows |
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| 11:56 | . OK. And that helps to to create the uh turgid, which |
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| 12:01 | kind of a swoll a little But bacteria, many bacteria have a |
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| 12:04 | wall so that so that incoming water of helps us serve, keep shape |
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| 12:09 | it. OK. Um Now, course, your halo files right |
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| 12:13 | Files are types that um work uh in excess of, of uh |
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| 12:24 | OK. 5% solute concentration in sodium . OK. Or higher, |
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| 12:33 | there's uh 5, 10, I'd say 5 to 2020%. That's |
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| 12:39 | of the range in terms of levels sodium chloride. Very extreme. And |
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| 12:45 | are, there are environments on earth actually have this um great salt |
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| 12:50 | Uh The Red Sea are very, very high in salinity, high in |
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| 12:55 | , but there are bacteria that can this condition. OK. And for |
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| 12:59 | that, you know whether it's temperature pressure or ph as we'll talk about |
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| 13:02 | a second, the ones that live the extremes, they obviously have adaptations |
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| 13:07 | enable them to live in those conditions you, you, you'll have to |
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| 13:11 | that um in order to, to , you have to have certain adaptations |
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| 13:15 | allow you to live in those conditions , and they indeed do. |
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| 13:21 | Uh A halo phyle is constantly fighting loss of water to the surroundings because |
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| 13:26 | lives in these high salt conditions. it has mechanisms to kind of retain |
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| 13:30 | . Um uh But regardless, so something to keep in mind the uh |
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| 13:37 | requirements of growth. OK. uh again, most are gonna be |
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| 13:43 | the middle, most, most are quote, the normal conditions of, |
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| 13:46 | neutrality around P seven plus or Uh And again, it's all about |
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| 13:53 | protein function, whichever ph is That's, that's what it's about. |
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| 13:58 | so your pseudo phys are acid They grow below ph six alpha files |
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| 14:04 | basic uh above eight. Just one in, in terms of cultivating uh |
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| 14:12 | and fungi, uh these actually are, are more acid loving |
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| 14:18 | than bacteria are. And it's classic um when you cultivate, if you |
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| 14:23 | to cultivate fungi from environmental samples, in order to kind of minimize the |
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| 14:29 | types around, you make the medium slightly acidic. And that, that |
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| 14:35 | to, to um allow for the of fungi uh regardless um culture medium |
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| 14:42 | ph, so we we'll be talking growth medium and of course, you |
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| 14:46 | that to be at optimal. Um There are cells can have buffers |
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| 14:52 | which will help neutralize the um um of uh too much acid or too |
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| 14:59 | base. Um The natural buffers present cells are, are typically things like |
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| 15:05 | acids. They basically have properties of acids and bases and they can help |
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| 15:09 | these types of uh uh this level of the city or, or or |
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| 15:16 | uh being basic. Um now where a or PH rather can't come in |
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| 15:25 | a control of uh of growth because have to also think in terms of |
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| 15:32 | we can manipulate these conditions of PH . Um and uh and uh om |
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| 15:42 | to help control growth, right. that's kind of chapter seven is about |
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| 15:46 | u using growth grammars to control or levels of microbes such as pathogens. |
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| 15:52 | certainly with temperature, we can do , you know, preventing spoilage |
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| 15:56 | or at least uh minimizing spoilage of or refrigerate, right? Because salt |
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| 16:01 | grow very slowly in, in cold most except for cys but um uh |
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| 16:09 | can use high salt. Uh uh have, have packed food and salt |
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| 16:14 | centuries as a way to preserve it the water is high salt inhibits growth |
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| 16:19 | occurring because the the water is constantly out of cells. So that is |
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| 16:23 | uh a way to inhibit growth and uh reduce spoilage of foods. Similarly |
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| 16:29 | ph right, there are certain foods are actually very acidic and that in |
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| 16:34 | acts to inhibit growth. And so of those uh one way you can |
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| 16:38 | it as well is through what are uh certain weak acids, small weak |
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| 16:44 | . So remember, acids um give protons, right. So weak acids |
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| 16:50 | partially do that, right. So solution, you have a weak acid |
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| 16:56 | the weak acid would be a small uh in its in, in, |
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| 17:00 | that form, you see circled, would be neutral and those are would |
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| 17:03 | small enough to kind of diffuse through membrane. And when it gets inside |
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| 17:07 | cell, then it'll dissociate and then the protons uh are, are formed |
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| 17:13 | they uh or are um associated from weak acid and, and then the |
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| 17:18 | ph inside of the cell begins to , it becomes acidic. And so |
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| 17:22 | , this is actually an inhibitory effect the cells. So there's various foods |
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| 17:28 | use this property. So they, have things like citric acid. Uh |
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| 17:32 | another one that's very common in food para aminobenzoic acid pab A. So |
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| 17:37 | are compounds you'll see, you see things like bread and, and others |
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| 17:42 | uh they're there to help preserve the , prevent spoilage. And they do |
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| 17:45 | through this kind of a mechanism by growth by affecting internal ph of, |
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| 17:51 | the microbes trying to grow on that . OK. So, uh |
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| 17:56 | So let's talk now about these are requirements for growth. Let's talk about |
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| 18:02 | requirements. So, uh here, as mentioned at the beginning of most |
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| 18:12 | or rather growth depends on the availability these elements because you're gonna put together |
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| 18:18 | elements to make the four major building of life, right? Your, |
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| 18:23 | DNA or RN A proteins collectively DNA A of course, is nucleic |
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| 18:29 | uh proteins, lipids, and right? The four classes of biomolecules |
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| 18:35 | you put those almost together, you , to, to make those. |
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| 18:38 | so of course, they're gonna be critical and in terms of, of |
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| 18:42 | a, on a proportionally uh what call a percent of dry cell |
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| 18:49 | Um carbon is gonna have the biggest , right? 50% of of a |
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| 18:54 | trice weight is is is carbon, , which is not surprising because all |
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| 18:59 | , all the molecules in those four you see there are carbon is the |
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| 19:04 | atom in the, in all of . OK. And thereafter, things |
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| 19:07 | oxygen, nitrogen, hydrogen, et . OK. So uh supplying CHO |
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| 19:14 | is is key to the to to growth medium. Of course, essential |
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| 19:20 | are those that can't be synthesized and be supplied like carbon source typically. |
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| 19:27 | the role of carbon, right? these are all what we call macronutrients |
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| 19:30 | they're needed in higher qualities because they're used in in higher amounts. |
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| 19:36 | ? And so carbon is mentioned, probably gonna be the most influential in |
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| 19:40 | of determining how much cell mass you get in a in a culture, |
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| 19:44 | many cells will grow it based on level of level of carbon present? |
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| 19:49 | so um so what's the the carbon for? Well, carbon skeleton remember |
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| 19:55 | is carb we are carbon based life on earth, right? So carbon |
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| 19:59 | the central molecule around which we can these other biomolecules. So I often |
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| 20:04 | the term carbon skeletons, right? as we break uh organic food sources |
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| 20:09 | , we use those as a building to make all the basic my |
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| 20:13 | right? So we can take a skeleton and add phosphorus or nitrogen or |
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| 20:17 | cetera and make and make the all the basic biomolecules. OK. But |
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| 20:23 | , you know, remember from, uh chapter five, the form of |
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| 20:27 | that must be that an organism can , right? It can be organic |
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| 20:30 | of carbon or it maybe has to CO2. OK. So, and |
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| 20:35 | comes into play here is where the autotroph uh differences are right. Heteros |
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| 20:40 | can release CO2 as part of their respiratory metabolism. And of course, |
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| 20:45 | fix that um other elements, So carbon is number one, in |
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| 20:52 | of biggest influence in in determining nitrogen is gonna be second. Uh |
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| 20:58 | of course nitrogen for to formation of , amino acids, nucleic nucleic |
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| 21:04 | et cetera, right? And the forms of nitrogen are are typically |
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| 21:10 | nitrate, nit trite are are the that are simulated by living things. |
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| 21:15 | in particular prefer uh nitrate, nit , uh phosphorus, of course, |
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| 21:20 | again, from the fake acids, ? A tp uh fossil lipids, |
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| 21:25 | ? So, phosphate is the form assimilated and sulfate is the assimilable form |
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| 21:31 | sulfur, right? We need that make these two amino acids that contain |
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| 21:37 | mest as well as there are some proteins as well that were molecules as |
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| 21:40 | that need sulfur. But certainly uh two amino acids. And then beyond |
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| 21:47 | , there's there's cat ions, So things like potassium magnesium, calcium |
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| 21:51 | are major uh uh cat ions and have different roles. Uh some act |
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| 21:57 | part of enzyme reactions. Uh Other of like signaling fact, uh signaling |
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| 22:02 | , things like that. OK. uh now, among those constituents that |
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| 22:08 | needed in much smaller quantities, micronutrients things like this. Uh Cobalt, |
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| 22:14 | , manganese, mole, nickel, , even tungsten in some cases. |
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| 22:18 | these often serve as as uh parts um different enzymes. Uh In some |
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| 22:25 | , they're parts of uh redox uh , things like that. Uh But |
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| 22:31 | , and need them very, very quantities. OK? And then uh |
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| 22:35 | , we have organic growth factors, ? So these are constituents, uh |
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| 22:40 | , amino acids, blood serum, typical they, they, they |
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| 22:45 | we often add these uh often fast pathogens. They often need blood or |
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| 22:49 | as an additive um for blood or . They contain lots of different elements |
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| 22:54 | there in, in both blood and . We may not necessarily know exactly |
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| 22:58 | it is in there that's helping But we know if we had blood |
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| 23:01 | serum it, it, it, promotes better growth. Uh So vitamins |
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| 23:06 | acids, there may be uh uh organism that you're trying to cultivate, |
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| 23:11 | may be deficient in being able to certain vitamins or amino. And so |
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| 23:15 | supply these uh as growth factors. . So, OK. So |
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| 23:24 | we're gonna take a look at OK? And it's roll and not |
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| 23:31 | , but actually, it's a uh different microorganisms respond to oxygen. |
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| 23:37 | A term we call aero tolerance. Now, uh we learned previously, |
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| 23:44 | course, that the aerobic uh uh the restoration, of course, you |
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| 23:49 | , that utilizes oxygen as a terminal uh in the uh breakdown of um |
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| 23:56 | molecules. And so, oxidation, should say nutrient molecules. Um but |
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| 24:02 | there are enzymes particularly that are part the respiratory chain that will interact with |
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| 24:09 | as a as a side reaction and form these reactive molecules called superoxide |
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| 24:15 | hydroxyl radicals which that you see which are very reactive and uh they |
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| 24:22 | interact with um proteins and DNA and A and damage them. OK? |
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| 24:27 | basically like to scavenge electrons and in so damage the molecules they interact |
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| 24:32 | OK. So this is a problem both those that utilize oxygen or that |
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| 24:38 | utilize oxygen. OK? Because the exposure to oxygen itself can actually get |
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| 24:42 | the cell and damage other molecules even they may not even be using oxygen |
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| 24:46 | all. OK. So the bottom here is uh for bacteria ikea living |
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| 24:53 | , in the presence of oxygen or they use it or not, they |
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| 24:58 | likely they're gonna have to have some of protective mechanism to prevent damage from |
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| 25:03 | from these radicals. OK. And that's where detective enzymes come in that |
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| 25:10 | shown here, right? Superoxide mitta and Pera is another one. |
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| 25:16 | Ok. What you see there? so these reactions, these enzymes help |
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| 25:22 | these toxic products into more benign products aren't damaging. So, uh sod |
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| 25:30 | by uh uh acting with uh superoxide forming hydrogen peroxide, which itself is |
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| 25:37 | peroxide still is, is uh not as much as superoxide, but |
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| 25:41 | , it needs further treatment to, reduce it down to water and that's |
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| 25:45 | catalase and or peroxidase come in. ? Um And so they will take |
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| 25:51 | hydrogen peroxide and, and form OK? If you ever use hydrogen |
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| 25:56 | in the cut, you'll notice that bubbles up and that's because your white |
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| 26:00 | cells also have catalase in them and they react with the uh hydrogen |
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| 26:05 | to form water and oxygen which bubbles . Ok? So when we look |
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| 26:11 | the, the tolerance levels of bacteria oxygen, it's all related to the |
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| 26:20 | enzymes that they possess or don't OK. And that they can uh |
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| 26:26 | different levels of these enzymes. They be lacking in one of these enzymes |
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| 26:30 | that all relates to how they um in the presence of oxygen as we'll |
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| 26:34 | . OK. And so we test the effects on a particular type of |
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| 26:39 | that will actually create an oxygen which I'm so let's take a closer |
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| 26:45 | at that here. And so what is this medium is called fluid thyle |
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| 26:55 | . So, it has uh a ingredients in there. One is um |
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| 27:00 | , a chemical that will help to up oxygen as well. It has |
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| 27:06 | color uh reagent and it, that it red in the presence of |
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| 27:13 | Of course, there's less red if less oxygen there and not red at |
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| 27:16 | where there's no oxygen. So as see there, it's intense right at |
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| 27:19 | top, different, then fades away we go to the bottom where there's |
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| 27:24 | red color indicating no oxygen. So medium also the medium is also a |
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| 27:29 | that's not completely liquid, but more a gel like consistency. And so |
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| 27:34 | you prepare this medium and then you it, auto Clavin boils all the |
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| 27:37 | out of it and then as air back in, it diffuses slowly because |
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| 27:43 | a gel like matrix and that plus action of the chemicals in there help |
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| 27:48 | create a ingredient of oxygen where it's at the top and zero at the |
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| 27:55 | . And so what you do is take a medium, right? You |
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| 27:57 | a uh a culture, right? typically a solid media plate culture takes |
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| 28:03 | monocular and then you will basically use inoculating loop as a straight wire and |
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| 28:10 | uh allow it to what we call stab medium straight down right in that |
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| 28:19 | and then straight out. OK. so what you're doing is you're seeing |
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| 28:22 | media all along the path through which put that needle containing your sample? |
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| 28:29 | . So then what you're looking for then, then you incubate and you're |
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| 28:34 | a question because the cells are seed throughout the medium. Uh But |
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| 28:40 | but remember the medium will have different concentrations throughout each level. OK. |
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| 28:46 | then what you're basically asking is which these cells will grow? Will it |
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| 28:50 | only those at the bottom that continue grow as you incubate? Will it |
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| 28:54 | those somewhere in the middle that are only ones that begin to grow? |
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| 28:58 | it be those at the top? . Or it'll be throughout the whole |
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| 29:02 | , right? And so the fact it's kind of a, a very |
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| 29:06 | matrix will confine the cells of, course, they'll, they'll grow in |
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| 29:10 | certain area because that's where their oxygen , optimal oxygen level is or maybe |
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| 29:14 | no oxygen, that's what they So that's kind of how we can |
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| 29:18 | a pattern based on. What part tube are they growing in? |
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| 29:23 | So that's what we see here. here's five different results looking at um |
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| 29:33 | five different uh patterns you would expect get. OK. Depending again, |
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| 29:38 | on the bacterial type. OK. obviate aerobe facultative aerobe, obviate aro |
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| 29:45 | tolerant aerobe and micro A file. . So I have divided them up |
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| 29:49 | aerobe aerobe and facultative facultative means you go both ways. OK. So |
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| 29:55 | that are Aeros that must have oxygen two types. OK. Obligate or |
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| 30:03 | file. What differentiates those two? number one, the overriding factor is |
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| 30:08 | both require oxygen. They're aerobic res . What differentiates them is the level |
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| 30:13 | oxygen. So, obligate Aeros cannot , can use, of course oxygen |
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| 30:18 | atmospheric levels, right? But micro files do not, they have to |
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| 30:22 | something much less than that. It 15 10% range, something like |
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| 30:26 | So that is where they'll grow in middle as you see on the far |
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| 30:30 | . So the obligate aerobe on the left, micro a file on the |
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| 30:33 | , right. So you see the of the micro air files is kind |
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| 30:36 | somewhere above where there's high oxygen and somewhere below where there's the highest levels |
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| 30:43 | oxygen and and somewhere above where there's oxygen. So it's gonna be occupying |
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| 30:47 | area in there whereas the AO is the top, that's where you're gonna |
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| 30:51 | the the air liquid interface is the level of oxygen. OK? And |
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| 30:55 | where they grow. Um And so all relates again to these protective |
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| 31:01 | So micro Ayles uh either are lacking of these protective enzymes or or don't |
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| 31:07 | the same levels of the concentration levels others do, which is why they |
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| 31:12 | that middle range. Your ana of , do not use oxygen, |
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| 31:17 | They either ferment or they uh respire OK. And so the two distinctions |
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| 31:23 | are the big aero oxygen is So, obviously, they lack these |
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| 31:29 | enzymes and, and the effect of superoxide radicals, et cetera kills them |
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| 31:34 | they can't protect against it. Your tolerant aros. On the other |
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| 31:39 | have some of those um some of uh protective enzymes. So, um |
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| 31:47 | thus oxygen is not toxic. So , the aero toler arrow does not |
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| 31:51 | oxygen. It's typically ferment, but they're not killed by the presence of |
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| 31:57 | because they have those protective enzymes. they'll, they'll grow throughout the |
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| 32:02 | Ok. Um The aero but can grow at the bottom where there's no |
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| 32:08 | . That's the only place it can . OK. The a a rope |
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| 32:13 | can use oxygen, cannot use Uh E coli is a perfect example |
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| 32:18 | that. It can actually ferment, can use oxygen or it can respire |
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| 32:23 | actually. So it has actually three . Uh which one it does depends |
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| 32:27 | what's available to it. Ok. they certainly have fract of ana certainly |
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| 32:32 | protective enzymes to protect against the effects , of these oxygen byproducts. |
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| 32:39 | So, um so then the question how do you differentiate facultative aerobe? |
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| 32:44 | aero tolerant aerobe? Ok. there you have to focus on, |
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| 32:48 | the top layer of the tube. . So remember the top layer of |
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| 32:52 | tube is where you have the most presence. OK. And um remember |
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| 33:00 | metabolism, right? Chapter five, aerobic um for respiration generates the highest |
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| 33:07 | of A TPS. OK. And lead uh high energy production, high |
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| 33:14 | TP yield leads to higher better OK. So more energy produced equates |
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| 33:21 | better growth. And that's what you . The fact that the mero grows |
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| 33:25 | where there's oxygen because that provides more which provides more cells, which provides |
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| 33:30 | growth. Ok? The Ater Aro not using oxygen, right? So |
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| 33:35 | can't take advantage of that feature. either fermenting, maybe it's respiring |
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| 33:40 | But, but the point is it's growing at the where the, it's |
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| 33:45 | using the oxygen. So it can't advantage of that very high A TP |
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| 33:49 | from an aerobic respiration. So it of grows more or less the same |
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| 33:54 | the tube. But the fact that aro will have more growth at the |
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| 33:57 | for that reason. Ok. So social responses to oxygen and then getting |
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| 34:05 | to kind of the growth medium. , so we combine these elements, |
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| 34:10 | cho NPS in, in usable whether it's uh you know, an |
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| 34:14 | carbon source or a and A and uh uh ammonium phosphate to supply nitrogen |
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| 34:21 | phosphorus and et cetera. We put together, we form a medium, |
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| 34:23 | course, right? And so we have two forms, liquid and solid |
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| 34:28 | typical. Uh There can be variations this but it's, it's, it's |
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| 34:32 | either one of these two groups. so there are purposes for these, |
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| 34:35 | certainly for um isolation of, of bacterial types. Uh pure culture work |
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| 34:44 | with the plate. You have to a plate uh culture to do |
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| 34:48 | which will allow you to cultivate, visualize and work with cells on a |
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| 34:54 | , right? So the cells on plate will grow to form a |
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| 34:58 | Then you can take that colony and use that as an inoculum into liquid |
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| 35:03 | where you can grow lots of So, but so each has its |
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| 35:07 | plates again, are for um uh uh certainly for pure culture work um |
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| 35:15 | into uh uh to um isolate bacterial on the plates. So, plates |
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| 35:22 | essential for that purpose. Liquid is about uh harvesting uh to grow lots |
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| 35:27 | cells and harvest them for different Yeah, you certainly need liquid medium |
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| 35:32 | do growth uh studies with and get curves as we'll see in part two |
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| 35:37 | this uh topic. Um and to different kind of biochemical tests. Uh |
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| 35:42 | a cell is producing some kind of , you can grow in liquid and |
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| 35:46 | take samples of the liquid and then for that activity. So, uh |
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| 35:51 | , both have their roles and and both are important. OK. And |
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| 35:55 | all depends on what you want to , which form you you need at |
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| 35:59 | given time. So, in terms growth medium. Again, we combine |
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| 36:04 | macron nutrients to form growth medium. ? And the types of nutrients that |
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| 36:08 | used to find a kind of It is right. So we look |
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| 36:12 | the complex medium, this is uh want to have many preformed uh |
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| 36:22 | OK. And uh these are uh in the graphics. OK. So |
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| 36:35 | the complex, meaning you can see in the uh Maria bean which is |
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| 36:41 | LBA a complex medium or a below complex. You see these nutrients called |
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| 36:48 | peptone, beef extract, yeast These are all what we call complex |
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| 36:53 | . OK. In contrast to something sodium chloride, right? Uh son |
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| 36:59 | both medium uh with sodium chloride, can obviously see the elements present sodium |
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| 37:05 | , right? Making up that Uh you don't see that for things |
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| 37:09 | these complex nutrients. OK. So you know, so these are typically |
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| 37:15 | rich sources. So beef casings, , protein, soy, protein, |
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| 37:20 | protein. So we know that these being protein will have carbon hydrogen |
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| 37:27 | So they'll have cho NPS in OK? But we can't necessarily be |
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| 37:33 | up, we can't be given a periodic chart and a calculator and |
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| 37:37 | to come up with the exact numbers moles per liter of each atom, |
|
| 37:43 | ? So look at krypton, do see it? Do you see that |
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| 37:45 | in Krypton? You don't see? we know it's there. OK? |
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| 37:49 | it's a protein source and it's also so it provides its, it has |
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| 37:54 | nutrients like amino acids, et OK. For that reason, typically |
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| 37:59 | with complex sources like that is much . The cells do not have to |
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| 38:06 | amino acids. For example, because the the the um complex source is |
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| 38:12 | that to them, right? And energy is may be devoted to |
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| 38:16 | they can be devoted to growing OK. Uh So we look at |
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| 38:20 | middle medium, it has a composition completely known. You can see all |
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| 38:27 | the atoms right present. OK. could have a periodic chart and calculate |
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| 38:32 | grams for mold for each of those , right? Glucose, of course |
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| 38:35 | C six H 12 06. Uh That's, that's known. |
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| 38:41 | So you could again, it's, what defines a defined medium, a |
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| 38:47 | or minimal medium. All that all mean the same thing is that |
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| 38:51 | know exactly the amounts of each atom it. OK? We don't necessarily |
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| 38:55 | that with complex media, but we that it supplies the needed nutrients. |
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| 39:01 | . Uh Defined me is typically is used for um doing growth studies. |
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| 39:05 | you want, if you wanna see nutrients are required for a certain |
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| 39:09 | you'll use defined medium. Uh complex is used really for, for getting |
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| 39:13 | lots of cells for, for getting samples with lots of uh growing cultures |
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| 39:18 | you want lots of cells and so media will, will allow that to |
|
| 39:22 | . Ok. The, um, here's an example of a type of |
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| 39:29 | that's, uh, that is, , um, uh, a, |
|
| 39:36 | type of defined medium, although it have some, uh, well, |
|
| 39:43 | vitamins are actually defined. So this actually a defined medium. So everything |
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| 39:46 | here, you could, you you, you'll know all the amps |
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| 39:48 | we know all the chemicals formulas for amino acids. You see here, |
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| 39:51 | know the chemical formulas for pines and and for these vitamins. So these |
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| 39:56 | all known. So this is definitely defined medium uh that you could uh |
|
| 40:01 | one that's a very long list. this organism apparently cannot do a lot |
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| 40:05 | synthesize a lot of things for This requires having a lot of |
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| 40:08 | So this is kind of an extreme in terms of the, the numbers |
|
| 40:12 | things that need to be added to . OK. But it is a |
|
| 40:15 | medium. OK. So, uh in, in this section, then |
|
| 40:21 | kind of just uh gone over uh chemical requirements for growth, um putting |
|
| 40:26 | a medium uh looking at complex defined . Uh So next time we look |
|
| 40:32 | uh uh growing and growth dynamics uh , of uh me material culture and |
|
| 40:41 | enclose on biofilm and biofilm formation, is a type of uh uh triggered |
|
| 40:48 | uh environmental conditions where there's lots of and, and these will form, |
|
| 40:52 | be conditions where biofilms will form. . So we'll talk about that next |
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| 40:56 | . So, uh thanks folks and see you in the next |
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