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BIOL 2321_Microbiology for Science Majors - 2321_10_3_23_CH04
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00:05 Like I, I, I, , let's see. How do you
00:19 the test, like eighties or 80 something? Ok. Ok.
00:34 , testing, testing. Hello. . Hello. There we are.
00:43 , folks. Um let's uh get , uh what we got. So
00:52 got um, chapter four. So this is a relatively short,
00:58 units. So three chapters, We already, uh as you can
01:02 there, it's like already time to exam two going. Oh my
01:06 Not again. Ok. But uh Friday, so I'll be aware of
01:10 and um, I know there's still small handful that are still not sure
01:18 this Casa interface, right? So not going to the old CASA
01:23 right? So, remember remember to to the CCS blah, blah casa
01:29 whatever. Ok. And I'll send usual information on, on Thursday.
01:35 , um, so just remember you're , you're not, don't go to
01:38 old process like you're going to the one. Ok. So it opens
01:42 Friday for example, two, which uh 20th 21st. And then,
01:49 , so we'll start on, viruses on Thursday, ok. And
01:56 continue on your next week and then so on. Let's see. Uh
02:03 we got uh so let's do a summary here. So we're in chapter
02:08 , let's remember a lot of the concepts you learned in metabolism like
02:13 14. Uh these certainly apply Um So let's look at uh some
02:22 these things we talked about. Um right. So we get, so
02:29 put this in here just kind of uh little bit of a road
02:33 OK. So we haven't gotten that honestly, but we've gotten through a
02:40 requirements. So the basic things we about before, right? The CHO
02:46 , OK. Essential nutrients, which can be broken down. Uh
02:50 we have essential types in terms of , macro, micronutrients, right?
02:57 micronutrients, carbon nitrogen, phosphorus, sulfur micronutrients, things like typically mineral
03:06 , we call them iron, Uh uh Well, no, that
03:12 go in macronutrients, calcium, um magnesium, those kind of things micronutrients
03:18 more like uh elements like coals or , these kinds of things. Uh
03:25 um coal factors and enzymes very Um So that's kind of where they
03:30 . So based on quantity and so lot of your micronutrients, you really
03:35 have to add those because they're usually as trace contaminants in your water.
03:41 , um anyway, so, so that, you know, we gotta
03:45 supply these things no matter what type are. All right, the nutritional
03:50 you are because all the biomolecules are on this structure, right?
03:55 carbon based life form, right? , um of course, there's variations
04:00 terms of nutritional types, right? already know that, right? You're
04:05 uh hetro autotrophs and we'll break that further today. Um And then we
04:13 about growth factors. So these are additions, um for those that kind
04:21 are deficient metabolically speaking, they can't some things they need to like amino
04:26 or whatnot. So you supply growth to them. Um vitamins, amino
04:33 , blood serum, these all fall that category. And then we looked
04:37 uh the protropin Oxytrol. So proto basically the think of it as
04:43 if you look at E coli, , there's all kinds of strains of
04:46 coli, but there's one, you , one that represents the group,
04:51 ? That has all the, you , features of a of a basic
04:54 coli, OK. Um And that call the proto troph and then those
04:59 are deficient in certain pathways very often acids or what have you, we
05:05 term them an oxo troph for for that particular nutrient. OK.
05:10 it, so it is specific, it would be an Oxytrol for a
05:15 pathway. OK. So I think pretty much what we finished,
05:20 So we're going through growth requirements. just did that then of course,
05:25 know, nutritional type, if we're to grow a particular type of,
05:30 organism microbe? Uh Is it a hetro, is it a li
05:36 Is it a photo troph? Then you've got to obviously combine the
05:42 uh physical chemical factors, light photo temperature, certainly with the optimum
05:50 optimum ph all these kinds of And then you put that collection of
05:56 chemical factors together uh into a formulation there and this is stuff we'll talk
06:03 today um that we can have, know, you want solid media,
06:08 want liquid media, those have different . Um gross media type, the
06:14 complex defined, we'll get into that quickly or shortly rather. And then
06:19 within that, within that context, you can still have media, you
06:25 make up media for different purposes. ? Maybe you wanna see,
06:29 Can this organism, does this have particular metabolic pathway? I can look
06:34 it on a this medium and I'll a color change and that will tell
06:38 , oh OK. You can, has this property, things like
06:41 OK. Um Maybe you have a from the environment and what you wanna
06:48 is in very small numbers in But maybe they have a particular nutrient
06:57 that you can simulate and then that favor their growth over the others.
07:02 ? And that's what we call enrichment . We'll get into this. So
07:06 worry if you, you know, get everything written down, don't worry
07:09 it. We're gonna cover this. general purpose. That's your, the
07:13 that you use in lab all the . Nutrient. That's general purpose and
07:17 all different types of heterotrophic heterotrophic Um High yield. This is,
07:25 know, this is manipulating growths uh formulations to, to get your
07:32 . There is high, lots of , lots of cells in a short
07:35 of time. OK. So uh go through all these things and
07:40 not finally, but uh we'll cover today as well. Cell numbers,
07:45 how you calculate cell numbers. Uh then uh at the end of the
07:52 growth curve in the different parts of . In fact, so let's uh
07:57 at uh so this is where we're today. The, the next part
08:01 this, the part two of this uh covers like in those four formation
08:06 , and biofilm formation. So we'll about that Thursday anyway. So there
08:11 be some, there, there, are two, maybe three calculations to
08:17 on exam two that involve growth. You'll, you'll be allowed to have
08:22 calculator and hill calculator. You can't a cell phone, but you can
08:26 a handheld calculator, any kind. to do those calculations, nothing
08:32 We'll go through it today. Um . So let's look at this question
08:38 . All right. So let me this thing up. Um Right.
08:44 , so read this thing chart from top, top down. OK.
08:51 you're gonna start up here and go and just follow the um,
08:57 OK. So the first question here this one. So you have a
09:02 is A through G OK. ABC . OK. So which is chemo
09:10 Trophy? OK. So remember start here and B Source and then
09:27 Mhm We got this question and a more on this slide, you learned
09:38 stuff in 13 and 14, Comes right out, comes right,
09:43 right out. Yeah. Ok. . 17 16, 987 5432
10:27 Ok. What we got? So says the um yeah, that's
10:36 OK. I I'm, I'm gonna through all these again once we get
10:40 them, let's do another one All right. Uh So we're looking
10:45 the aerobic header troph. Ok. I wonder who that is. It's
10:51 aerobic hetro there mirror. OK. . 32nd time point. Ok.
11:46 and survey eight. OK. And more just for grins here. Let's
11:59 . Oops. Oh, ok. . Oh, never mind. That's
12:07 . That's, that's fine. So certainly both, both of those
12:12 select it correctly, obviously. um uh so we can obviously categorize
12:18 you see the categories in green, green boxes that you see uh in
12:23 middle here. OK. Different new metabolic types. Um obviously the chemo
12:31 , right? Autotroph is the the key part of the word.
12:34 co2 is a carbon source, um hetero tropes. So again, hetero
12:41 uses complex organic uh carbon source. And then aerobic, obviously 02 final
12:50 . OK. So I think the one on here was plant or
12:53 you don't have to click or which one is the plant or allergy
12:57 here? Uh My guess dish herbally , of course, it's f obviously
13:13 is this right? Uses H2O, ? H2O versus 21 ad P,
13:20 ? So, um OK. and this is what you have to
13:26 , obviously, if you kind of one of these things is prime plan
13:30 one is what's the carbon, Then? Oxygen, no oxygen,
13:34 , no light, et cetera. . Um And so here is kind
13:39 the same similar breakdown. So we've based on carbon energy and electrons.
13:45 ? There's gonna be overlap here. Hemo Organo chemo heteros, the same
13:53 , basically the same thing. Um chemo lipe chemo Autotroph, same
13:59 . OK. This actually spelled out chemo Lit Autotroph. OK. But
14:03 are, they're the same thing as chemo Autotroph. OK. Using inorganic
14:09 for energy and fixing CO2. Uh the um question here, can a
14:15 hetro use carbon dioxide? OK. know photo hetro use CO2. Who
14:28 yes, who says yes, anybody no, because what's the operative word
14:37 ? Right? Al always Trumps uh pun intended. Uh everything.
14:42 Hetro is the key. They, can't fix CO2. So, uh
14:46 remember the Hetro and the two for deal they get with their carbon
14:51 Um, what does that refer to ? Remember that? The two,
14:55 get a two for one deal with carbon source, you can break it
15:02 , right? You break it you can use those as building
15:04 And what else do you get? , well, you you you're on
15:11 right track so you get, you it down, you get, you
15:14 , that's you get the building blocks make your molecules. But what do
15:17 get as a result of that? metabolism, metabolism releases energy, which
15:24 you form a TPS, right? . So you get the two for
15:28 is you get your carbon that you break down to get the unit,
15:34 black units to make your molecules. the bonuses is you also get energy
15:39 that right? Oxidation metabolism use that make a tps, right? So
15:44 that's not the case for a right? They don't get that because
15:49 have a fixed CO2 because that CO2 is a building process. They're taking
15:56 and making unit, taking those and a lot more complex organic molecules that
16:02 a lot of energy, right? they have to have an energy source
16:05 support their CO2 fixation habit, so speak. OK. So the energy
16:11 come from light or it can come breaking down in organic molecules like a
16:15 troph. So they, they don't the, the benefit as a
16:22 Um and getting the both, both from one source. OK. Um
16:28 . So what kind of medium is ? OK. Fine complex selective
16:36 So where you're mulling that over any about the my head, all,
16:41 those names and things. So just that somebody else have overlap.
16:47 Yeah. OK. OK. Cut from 10. OK. There it
17:35 . OK. So complex were defined who said um complex complex? Why
17:46 it complex? So my um what what let me rephrase it. What
17:54 on here? Tell you what's Yeah, the peptone beef effect,
18:01 ? So um so this if you look above the line or let
18:05 let me draw a line, then can look above it. Here we
18:09 . Um Oh, there we My pen. I had to find
18:12 cursor. So if you go above line here. Wow, that was
18:15 weird stray mark. All right. try this again. A pen
18:19 So I, I have one on . So we're having to use this
18:24 . So hold on one more OK. Let's try it again.
18:32 . So above that, right? you look above, there you go
18:35 defined medium and you know, you basically see all the, all
18:40 elements, right? And their you could have a calculator in periodic
18:45 and go OK. I know how grams per mole of every atom is
18:47 this thing. That's a defined right? You can almost do that
18:52 you can't because peptone and beef right? So even just adding one
18:57 those automatically makes it a complex OK. So uh you know,
19:04 getting so um the peptone and beef , these are basically think of it
19:10 just um many of these things basically beef extract. Um cryptic soy.
19:19 , cryptic uh crypto is another, lot of these are trade names,
19:23 trade names, but they either boil the source, they uh enzyme
19:29 the source um other treatments to kind help break down. It's basically just
19:35 a lot, a lot of these just taken from the slaughterhouse floor,
19:39 ? Just taking the meat and the and stuff, boiling it, digesting
19:43 and that be and dehydrating it to a medium. So, you
19:47 when you see beef extract, think , right? You're eating a
19:51 right? So it's gonna be everything in there, right? You have
19:55 of preformed nutrients, right? And amino acids are gonna be in
20:00 Uh You're gonna have cho NPS, course, right? And you're gonna
20:04 them in, you know, the forms, right? You're gonna have
20:07 acids present, you're gonna have you can have fats, right?
20:12 so you get a lot of preformed . So if you're a microbe and
20:17 sitting in a complex medium, you getting lots of stuff, you can
20:22 assimilate, incorporate and grow very OK? If you're in a,
20:28 a defined medium, right? Defined . So just think of the ingredients
20:32 are above the line, you're just it, you're just sitting in a
20:37 that only has uh what's above the , right, then you pretty much
20:43 to make everything for yourself. You to take those basic elements and make
20:49 of your various molecules, right? a completely different story than being handed
20:54 already premade for you. OK? And so it typically means slower growth
21:00 it's a minimal medium. OK. let's, let's look at examples of
21:05 . OK. So the top, um the what's called, it's LBLB
21:13 short is that top one? Um um the very common medium that they
21:20 to grow E coli typically in like that do molecular biology work with E
21:25 . Um So that's nothing but complex they do have sodium chloride in
21:30 But you know, krypton yeast right? Yeast extracts are sources of
21:35 , I believe like b vitamins, rich or rich and yeast extract.
21:41 So, I mean in lb you know, you're pretty much getting
21:44 kind of preformed nutrients for the most . OK? But you don't know
21:48 they call it complex nutrients you don't the, you don't know exactly what
21:54 various elements are and their exact but you just know it has all
21:58 all in there. Ok. And a, an M nine medium in
22:03 middle, right? Um, uh, as you can see,
22:10 they, they wrote out glucose instead putting the formula, but of
22:12 that C six H 12 06 for , um, you, you know
22:17 that's in there exactly down to the Adam. OK? And the
22:22 So, uh and so they both different uses. OK. Absolutely.
22:28 you wanted to figure out the exact requirements for an organism, um you
22:34 use a defined medium because you can , you can take out right,
22:39 elements and see what happens and then them back and, and see uh
22:44 at growth that way. So you determine these kind of uh make these
22:48 with defined medium. OK? Um uh you know, maybe you have
22:54 mutant strains, OK, that you're with and you want to supply
22:58 you wanna find out what's the, the particular nutrient they need to
23:01 right? So you'd have to do this kind of in the defined
23:05 OK. If your goal is I'm get lots of growth, right?
23:11 want lots of cells in a relatively amount of time, then you're gonna
23:16 with a complex medium. But Um from a practical standpoint. What
23:23 do is actually something like we saw is this, you do have most
23:30 your ingredients that are defined, but you add a couple of complex nutrients
23:35 that's, that's the way to get growth. Ok. Because what you
23:39 in that circumstance is you find that there it is you manipulate.
23:50 Let's see if I can do Mhm. Ok. So that's,
23:55 what you manipulate the carbon, So you, you make a so
24:00 , even if you have this one nutrient in there is it's a complex
24:03 . So that's what you do, ? Uh But you have these other
24:06 in there, then you add as growing, you feed it glucose or
24:12 the carbon source is and that allows to sustain a high level of growth
24:16 you're feeding it carbon. But then pepto yeast extract are supplying those things
24:20 vitamins and amino acid and stuff stuff don't need to bother to make and
24:28 they can just grow very quickly because you having to stop and make stuff
24:32 means turning on, turning on a , turning on uh you know,
24:37 turning off certain pathways. So it's involves having to do a lot more
24:42 , right? Because remember protein synthesis doing stuff, it's making stuff,
24:47 takes energy, right? So if can minimize that and provide preform things
24:53 , then that can translate into faster , more growth, more cells.
24:58 . And so um so you kind keep, we used to call it
25:03 defined medium, but we don't do anymore to avoid confusion for people are
25:07 to learn this stuff, but semi is basically a combination. So now
25:11 just call it even if it has one complex ingredient, it's just a
25:16 done. OK. But so the the bio person out there that was
25:22 ? Uh Is he here today? you should be familiar with this.
25:25 you're gonna run bio reactors, bio , biotech measures, I'm giving you
25:30 here. OK. You know, stuff in the textbook. OK.
25:36 Anyhoo. So with any questions about is that logic makes sense if they
25:41 a carbon source doing these other things when you, when you need lots
25:45 growth, if you're commercializing a you can't deal with just what you
25:50 in a lab which is like a mier flask 100 mils. That's not
25:55 , that's not gonna work. You buckets of stuff, right? If
25:58 an enzyme, you're trying to mass , you need tons of stuff.
26:02 you need to maximize growth in those . And so that's, this is
26:06 thing, these are the things you . OK. So um all
26:12 So OK, so define, so , there's other names for that as
26:15 . So if you see synthetic you see M medium, all three
26:20 those mean the same thing. They're , those three terms are synonymous.
26:25 . Um, fastidious. Ok. that's best described by showing a gross
26:33 , right? So just hold on , for a second. Ok.
26:38 , um, so you see the different media types there, right?
26:40 is two of them are defined. . Um, one of the top
26:45 complex medium. Um, and to be honest, that lb you
26:52 think, oh, it's full of nutrients. Yeah, but it,
26:55 , it's not really a gross That's great to get lots of cell
27:00 on. OK. It's great for fast growth to a certain point because
27:03 the old preform things, but you're super high in carbon, right?
27:08 why you add like something like a or, or whatever your favorite carbon
27:12 to and manipulate that in, in of quantity to get lots of
27:16 Ok. So the, the the guess you say the benefit of the
27:22 nutrient is the providing of preformed nutrients amino acids and whatnot, vitamins and
27:28 like that. OK. Um So look at this question. So
27:33 we're back to kind of to relating , nutritional type that we did at
27:36 beginning to media type. OK. putting that together so carefully, there's
27:45 lot of words here. So let just kind of um pause for a
27:49 and read all this. OK? basically a through D or four media
27:57 that are like just written out instead being like a like we just
28:01 OK. Um Pay attention to the in D right? Carbonates or non
28:08 sources of CO2. All right. you're looking for the medium that will
28:16 a hit. A OK. So , you're not a pro carry
28:22 you know. Yeah. Right. . Ok. Let's count down
28:55 You do. No. Mhm. . 0 10 9. OK.
29:41 got B and E good one. BB and E All right. Um
29:50 . Let's just go one through. a why or why is it not
29:58 Yeah, soy digest. So the digest gives that one away.
30:02 So remember even just one of those of ingredients, it's automatically complex.
30:06 . So the soy digest gives eliminates OK. Um C what eliminates that
30:14 be be extra, right? So is out and D is out because
30:23 a medium for a auto, Yeah, because so it's the
30:29 You can, you can bubble in . Um You can use that.
30:36 the um the thing is though the this part of the molecule just carbonate
30:45 thing. OK. Um uh an ha actually has to have the enzyme
30:51 can convert that to CO2 but some do not all but you could,
30:55 could use that as a substitute. the point is it's a CO2
30:59 right? So an autotroph is what on B OK. Um lipe.
31:05 B is the only one here that . OK? Because there's no,
31:08 kind of complex ingredients that's strictly all . OK. And it has,
31:13 know, glucose as a carbon OK. Um All right. Any
31:19 about that? All right. So another one. This is about
31:27 what we talked about last time, ? The oxy, OK. So
31:29 question here is would a histidine Oxytrol being amino acid? I'm sure you
31:40 , would it be able to grow that medium? Yes or no.
31:48 . Sorry. OK. I think can speed this up a little
31:57 Yeah. OK. For 12. or no. Mhm OK. All
32:27 . Here's a look at that any . Look at that. All
32:33 Um Who said yes. Who said . So why? Yes, growth
32:49 , growth factors would supply the uh peptone and beef extract are gonna be
32:54 source of amino acids, right? gonna contain those things will contain certainly
33:01 histadine in there, right? So you see peptone beef extract, that's
33:05 like meat, right? Meat's gonna these things in it. OK.
33:12 acids, vitamins, et cetera. . Or soy, the soy,
33:18 soy ones, those are plant plant . So plants can also serve as
33:22 complex nutrient as well. OK. um All right. OK. What
33:29 grow on this medium? OK. would grow on that, right?
33:38 just think about it and then a of hints here. What would grow
33:42 that? Yeah. All right. missing, here's another hint. So
34:04 would grow in that medium? Which one? Uh I Right.
34:15 what side of that triangle is, something that might grow on that?
34:22 actually, it's actually how you would the uh one of those types that's
34:27 one side of that triangle to Which side, left side? the
34:36 22. Yeah, that's how that's how you uh enrich for.
34:42 you have a soil sample, you find nitrogen fixers, you pop them
34:46 a medium that looks just like OK? That's gonna be very
34:51 right? Um There's no end there's no in source in that
34:56 OK? So if something's gonna grow , it's gonna have to basically fix
35:01 two from the air to grow. um that's, that's, that's enrichment
35:08 , right? You are providing the uh to support a particular metabolism and
35:17 will be favored to grow over OK? So if you have a
35:21 of soil very quickly, the hetero in there will overgrow everything,
35:27 And so you in order to favor numbers to get something in there that's
35:33 , so doesn't grow that way, you use a medium that will favor
35:39 growth. And so that will absolutely a lot of things. And so
35:45 have a pretty good chance of finding particular metabolism. OK. That's,
35:49 the essence of enrichment culture. Um Now there's, we'll talk about
35:54 medium. It's like subtle, subtle there. OK. Um So,
36:01 , fastidious, get back to that . So fastidious bacteria. They will
36:08 a, basically the stuff on the we've seen before, like carbon energy
36:14 various minerals, right? But then right side, right, you typically
36:18 to add a bunch of stuff because , they're very deficient in various
36:24 So you guys supply amino acids, vitamins, et cetera, et
36:29 Um, and that's something that we it. They have a lot of
36:33 requirements, right? So if you're a medium for one of these,
36:37 got a long list, you are a bunch of stuff out.
36:40 it's very annoying really. So you wanna have to deal with fastidious bacteria
36:45 like, you don't have to deal fastidious people. They're, they're very
36:48 , right. They're the ones uh, order a salad and no
36:51 , don't want this on it or on it. How they want,
36:54 , this kind of dressing is, is a chicken, you know,
36:58 range kind of go see the chicken there really is, you know,
37:01 range or they just stuff in the or, you know, a lot
37:04 requirements, high maintenance. All Think of that. High maintenance.
37:08 ok. Um, let's see. right. So culture media type,
37:16 solid liquid. So, real um uh the utility of liquid is
37:23 volume, right? You can control volume there. So you can uh
37:27 certainly if you're gonna grow cells to yield and you wanna harvest um you
37:34 them in liquid, OK? You uh grow cells on a plate for
37:38 purpose. OK. If you're gonna a growth curve, you grow them
37:42 liquid, OK? Um You can the liquid and take measurements and
37:46 So if you wanna measure a particular for enzymatic activity, you typically do
37:53 analysis by growing them in liquid. . Um Plates have their utility in
38:00 have to be a part of the if you're trying to get a pure
38:04 , OK? Because you need to them on a plate to see uh
38:10 manipulate them. OK. Yes, can look at a liquid culture on
38:13 microscope and see if there's different types there, right? But you can't
38:17 anything beyond that. You have to them on a plate to then be
38:21 to, you know, isolate different and do whatever work you wanna
38:26 right? So plates have to be part of the pure culture process.
38:32 ? Um Now, selective, selective differential. OK. So is so
38:43 versus enrichment media. OK. So media, you are basically combining nutrients
38:51 and supplying the physical factors, et cetera to favor the growth of
38:57 types. OK. Selective medium is are actively adding inhibitory agents to basically
39:09 things from growing, right? Um could add an antibiotic to the
39:15 right? That's gonna be very OK. So only those resistant to
39:20 will grow, for example, So and there's other chemicals you can
39:25 uh very often um why this media selected media were developed for like wastewater
39:31 . And so in wastewater analysis very looking for indicator organisms, indicators of
39:38 contamination of water, right? And classic definition is a gram negative lactose
39:46 rod E coli for example. And so media was developed to kind
39:52 favor their growth and select against grand , for example. So this kind
39:57 media you see here hea for short in the lab, you use that
40:04 um and others. And so they're way to identify one of these fecal
40:09 in your water very easily. So media here and I'm not gonna test
40:15 on hea media on this exam. just giving you this as an
40:18 OK. Um So it's gonna a selected because there is chemicals in their
40:24 in poss, OK? It's so, differential media allows you to
40:33 to um to distinguish between different metabolic . OK. And so very often
40:42 a color change that occurs or it's presence of a clearing zone because they
40:48 an enzyme activity that kind of that up the the substrate in the medium
40:53 the clear zone, for example. with blood o you can see um
40:58 that can uh destroy red blood have an enzyme and they secrete
41:04 And in the process of destroying their cells, they create a halo uh
41:08 zone, a clear zone around, the growth that's positive. Like you
41:13 here um over here is a color . So lactose fermentation, right?
41:19 lactose fermenters, I know this is great, great contrast here. But
41:24 like for example, um e coli cola form produces a more yellowish colony
41:30 this medium. Then does a nonlactose men. It's more kind of just
41:34 . OK. Uh We have another here is the black colonies. These
41:38 production of H two S OK. another feature that distinguishes salmonella. And
41:44 point is you see color differences in of do they have the metabolism or
41:50 they not? Right? Can they lactose or not? You can see
41:53 a color reaction. There's many different of differential media that you know,
41:58 test for some kind of some kind uh enzyme activity of the organism has
42:03 doesn't have and you can differentiate, ? So in lab, we're doing
42:06 unknown project starting this week. And you'll be doing some of this in
42:10 of not on plates but in liquid . OK. Uh looking at differential
42:17 and liquid media. OK. So so you can combine, you can
42:22 selective and differential together. OK. is very often what you do.
42:26 selecting for certain types and then seeing grows. Do we have different meta
42:32 here as well? Um OK. questions about that? Mhm Right.
42:43 , all right. So I'm gonna gears here a little bit and we're
42:46 talk about growth. OK? And calculations and uh there's not, there's
42:52 that complicated in time. So on exam, you will have the formula
42:59 need. OK. Um You'll be to have a calculator in time.
43:05 so we're gonna do a couple of and uh and, and there's some
43:10 problems on uh ca uh canvas as . Um They're worked out so you
43:17 see how it's done. OK? um so this, so we're talking
43:21 growth. OK. So we've already that obviously, bacteria grow fast,
43:25 ? They have a small chromosome, have the polyribosome formation, right?
43:30 produce proteins very quick. Um They a small cell size, right?
43:35 all these things um lend them to able to grow very quickly,
43:39 So this is just kind of show in terms of this parameter called doubling
43:45 . OK. And so that equates how fast for the population to
43:51 OK, you can look at it the time for one cell to split
43:55 two right? To make a Uh These are all represent dumping
43:59 And so bacterial types all have different times. Ok. So under optimal
44:06 , E I can double every 15 or so. Ok. Others slower
44:13 that. Uh there's a range. . So this is just looking at
44:17 with 10 cells. Ok. Uh type doubles every four hours.
44:24 Will be the population size after 20 in the same time frame. What
44:29 they had a doubling time of 15 ? OK. OK. Four hours
44:33 15 minutes, there'll be a But can it be that much of
44:36 difference? Well, we'll see. . This is the kind of I'm
44:40 the gun a little bit. This the formula we use. You
44:43 you could use, we're gonna, gonna manipulate this to get a,
44:47 equation. That's, there's something we do more with. There's limitations to
44:52 simple equation. But um so NT a time N zero, I'm
44:58 N zero is your start time. . And NT is a uh time
45:05 in the future and then in this , number of generations that have passed
45:10 to the end. OK. So starting with the same number of
45:15 same time frame of 20 hours. many generations, right? So uh
45:20 a doubling time, that's one generation four hours. OK. 20 hour
45:25 frame that gives us five generations. we just plug it in right?
45:30 times two to the 5th, 320 OK. Who you do? All
45:37 , for a 15 minute doubling that's one generation every quarter hour,
45:42 ? 0.25 hours and 15 minutes times it gives you 80 you can already
45:48 the difference here, right? 2 80 10 to 25th cell versus 320
45:53 , right? Humongous difference, right? And so the doping
45:57 uh it's, it's, of it's a exponential, right? Uh
46:02 to 4 to 8 very quickly, ? Produces a steep curve.
46:07 So under optimal conditions, like I before, I think um you collect
46:12 form 20 generations in about 6 to hours, it takes humans 400 years
46:17 do that, right? So you the idea how fast they can
46:21 OK. And so that's what kind what we're focusing on a little bit
46:25 in this section is um um you , if you combine the proper nutrient
46:33 that, that it needs physical temperature ph whatever uh that you can
46:38 these kinds of uh rates of OK. And so, uh of
46:44 , this uh may recall from intro , right? I think you talk
46:48 some of these basic principles in right? So the J shaped
46:55 right cell cell number over time, growth is J shaped, right?
47:01 we know that this isn't, doesn't in that way forever, right?
47:06 it will eventually flatten out. And so generation times is what we
47:11 in terms of um monitoring uh microbial . And so, um again,
47:18 for population to double is usually more way to do this. OK.
47:22 you can measure so growth very easily a a spectral photometer, right.
47:27 you get an absorbance measurement and you , well, how long does it
47:31 for that absorbance measurement to double? that's gives you uh an of the
47:37 time. OK. And so here showing you the, the rapid growth
47:43 it can occur. So when of , when we're doing, when you're
47:46 um large changes in numbers over you try to compress that into a
47:53 scale. OK. Same as you with Ph, right? Um And
47:59 that gives us a way to better these patterns. So you see what
48:03 hear very typical of what you would . You, you would see a
48:07 of growth in this period here where the the absorbance is doubling. So
48:12 doubling time and generation time, you it as a very steep curve
48:16 OK. And so every organism has intrinsic doubling time, right? An
48:23 coli on, on nutrient broth versus sub list on nutrient broth, they're
48:32 gonna have differing uh generation times. . So this is, this is
48:38 specific number um but it can change on what it's growing on uh
48:43 et cetera. And obviously, microbes growing in lab are quite different from
48:48 you're doing monitoring on nature, Because they're competing with so many of
48:52 types, right? So nutrients are be limiting in nature. So only
48:57 , as we, we learned right, in terms of the,
49:00 reification process, right, that can to spurts of growth in nature when
49:05 provided excess of nutrients, right? for the most part, things are
49:09 much kind of imbalance, right? um uh except for those times when
49:13 do get influxes of nutrients, Um So in, in taking this
49:22 , right? So we have this one here here um that uh so
49:28 have to be able to, you , count the number of generations,
49:33 ? The end, the number that get kind of cumbersome if you're,
49:37 trying to get a a lots of trying to figure that out,
49:40 So we want to simplify things to it easier. So we're gonna take
49:43 equation and basically just gonna solve for , right? So we just,
49:48 remember this is just, you need base 10 is what we're using here
49:52 we're just, and you don't have worry about deriving this for the
49:55 And I'm just showing you how we're to the, the, the final
49:59 . So we just basically multiply through log to base 10, right?
50:02 remember you can express this in this , right? Log to base 10
50:08 two to the end is the same in times the base 10 of
50:12 right? And then you can solve that and becomes 0.301. OK?
50:17 so now we just solve for right? And so that gives us
50:21 that, all right. And then is your equation. OK. And
50:27 thinking that 0.301 is kind of that of two if you will.
50:32 And so from here now we can any uh two uh cell numbers,
50:39 ? Because we can, we we can figure out cell numbers pretty
50:43 , right? We can measure to , we can do an actual,
50:47 an actual sample and get a cell , right? So getting in the
50:51 number is pretty easy. OK? so in doing that, then we
50:55 figure out the number of generations and are things we can do with that
50:59 as we'll see. OK. And very uh and certainly we, we
51:06 , we can slightly modify that to time in a time factor in
51:11 Um a rate or gross rate. . So just putting T in
51:15 Um and so K we call K rate constant. So growth rate
51:20 that number is what is specific for , right? Microbe X will have
51:25 certain growth rate constant versus microbe It may be very close, but
51:30 one kind of has its own distinctive . OK? The, the max
51:34 which they will grow at under optimal . OK. Um Like I
51:39 for most bacterial types, procaryotes, between 15 minutes to two hours is
51:46 the average. OK. There's some are longer. Uh but that's kind
51:50 the average. And so the generation is basically taking the inverse of that
51:58 . OK. So time per generation typically it's, it's, it's expressed
52:04 minutes. OK. So minutes per say, oh, this, this
52:08 has a generation time of 15 So that means uh one generation uh
52:14 15 minutes, one generation. And so we're gonna use this
52:20 this equation here, a couple of and uh in a couple of different
52:26 . And so the bottom line is trying to get the end number and
52:29 use that uh to get some OK. So again, on the
52:35 , you'll have this equation will be there in the problem. So you
52:40 to worry about memorizing. OK. So here's the question. So take
52:45 shot at this. OK? And go through it. So this bacterium
52:58 a generation time of 40 minutes. . So we start with five cells
53:05 log phase, we'll go over log and all that stuff a little
53:09 Uh How many minutes does it take produce about 10,000 cells? OK.
53:36 let me pause. So you can some calculating there to remember, remember
53:56 minutes to hours conversion. OK. you need that. Mhm.
54:45 Ok. Let's uh, slowly count here. 54, right? All
55:03 . 32, one. Ok. let's see if that's right. Let's
55:12 through it here. So I'm gonna you probably more steps than what you
55:18 , but this goes to a piece here. Ok. So basically setting
55:22 , right? So we're trying to out um, um this,
55:30 So there's our equation is for you start with five cells to
55:35 So how long does it take? . Uh 40 minutes per generation.
55:41 we have all the knowns written right? To them, calculate number
55:46 generations and going from 5 to 10,000 , right? So if we uh
55:53 we have a 40 minute generation OK. If we calculate the number
55:57 generations then cancels out and we'll get long that is? OK.
56:03 So that to that. All 10,000 is your nt five is your
56:09 zero? All right, you do there. Did we get that?
56:14 is about 11 generations? OK. . A little over seven hours,
56:28 420 minutes to seven hours. So hours and 20 minutes. Ok.
56:34 So let's look at it in a different way or different question being
56:39 This is asking for how long does take to get to that number?
56:44 one, I think as for an generation time, right? So let's
56:48 here. Ok. Um calculate generation if 900 cells growing 15 hours produced
57:01 3 million cells. Ok. It's little bit, a little bit different
57:13 . Oh, sorry. There we . Mhm. You know me.
58:19 , let's count down from seven uh was ok. Ok, let's
58:41 All right. So here are, the knows, right? I can
58:45 15 hours, 10 minutes. Then are N zero and NT right?
58:53 a number of generations then just generation is minutes over generations, right?
59:02 that's gonna be what you said, minutes. OK. So, um
59:12 you might do the same experiment with same strain and maybe expose it to
59:17 disinfectant or something and see if you a difference in generation time or what
59:22 you. There's different ways you can this data. OK? Um
59:29 I have, there was like a of including these and an extra three
59:34 . It's, it's all worked If you want to go through
59:37 they're on canvas. So I don't you're gonna have any problems with
59:42 But if you do, after looking the all the problems on canvas,
59:47 you do, certainly let me know can go through it. OK?
59:51 there any anything right immediately right now you know, OK. So,
59:59 all right. And again, these will be posted as well after
60:03 So uh you can go through these OK. So let's look at this
60:09 . All right. Uh Now we're to growth curve. Yeah. So
60:19 so this term batch growth curve. . So what that refers to is
60:29 just this is my flask. I'm making a medium growth medium uh
60:37 . And then the only thing I'm with this is to just take samples
60:41 and measure growth. OK. That's I'm doing right. And I'm just
60:46 follow it throughout until it dies. , that's, that's a batch growth
60:53 . OK. There you can change up as we'll see. But
60:57 that's what batch growth is, You have a batch, a medium
61:03 , you just follow it until it growing until it dies. And then
61:07 do you get? What kind of ? Right. So um and that's
61:12 you get obviously some kind of curve that, right? But what can
61:20 is how long the different phases how fast or slow they are.
61:27 what changes, but the basic curve OK. Right. OK. Let's
61:47 . E OK. Changes in cell . That's true. B is true
61:57 is true and D is true. . So remember um penicillin is going
62:05 act most effective when the culture is growing. Like that's what's making most
62:11 the cell wall is being synthesized, ? Very active growth. So,
62:14 , none of these are false. . So let's go through these phases
62:19 stages. OK. So um So we inoculate, we have a
62:27 medium, we inoculate. OK. So imagine you're the cell, you're
62:34 cell and that inoculum. OK. you're plop down into this pond,
62:40 ? Of fresh medium? OK. what can happen? Well, what's
62:46 , what's gonna influence how fast you to grow? Well, no
62:51 Well, one, what kind of am I in? Am I,
62:54 was I growing in? And what I in now? Ok. So
62:58 can have an effect, right? Am I was I in a,
63:04 a uh defined medium? And now in a complex medium, um exactly
63:14 or increase lag phase. Anybody I'm a defined medium. I'm in plopped
63:21 a rich medium, complex. Begin grow more quickly or will it stretch
63:35 any gifts wild gift? Not? , I guess you got two
63:40 You're gonna be longer or shorter. said shorter says sure, I'm in
63:46 fine medium. I'm going into a medium. What do you say?
63:51 course, because remember complex medium, of preformed stuff don't have to do
63:56 lot of work, right? So can probably begin to grow pretty
64:00 Ok. Even if that's the there's still gonna be a delay,
64:04 ? Because you're in a fresh likely ph like slight ph difference from
64:10 you were in slight um temperature maybe um slight. So uh uh
64:18 um a similarity, right? So kind of things, you know,
64:23 be slightly different. So it's not be a burst and start growing right
64:27 . Ok? You may have to acclimate, right? You likely will
64:34 to uh turn on and off different , right? Particularly if it's two
64:38 . You're in one type of Now you're a completely, completely,
64:41 different one, right? That will , oh, I gotta turn this
64:45 on this one on or this one . Um So yeah, there's gonna
64:50 um you have maybe you have to stuff, maybe you're growing on glucose
64:53 now you're, you're plopped in the where it's uh lactose or something,
64:58 ? You have to change, It all depends on what, what
65:01 are, what the the media types different, these kind of different factors
65:05 into this, right? Of you're not uh you're not gonna
65:10 right? Am I putting in one or I'm putting in 1000 cells,
65:13 ? That's gonna this gonna be uh , get out of lag phase with
65:17 cells and fewer cells faster. Uh these are kind of things that,
65:23 , that can affect how, how or short lag phase is.
65:28 Um Practical standpoint, if you are this on a commercial scale and you
65:34 to get lots of cells quickly, would basically be using the same media
65:38 app. So you, you, you have them already kind of
65:42 So to speak on a medium So you know that if you
65:46 there's not gonna be that much of lag face, right? So,
65:49 a practical standpoint, it's kind of you do, but there may be
65:51 when you don't. So, but that's, that is where you keep
65:55 kind of consistent, right? ok. Now, blond face.
66:03 if you are, um, the you're growing are something you want to
66:10 measuring and measure an enzyme activity with , they produce some kind of a
66:14 and you're, you wanna measure that , it's very often gonna occur in
66:19 phase. The cells will be most during that period. Um They haven't
66:24 become limited for nutrients, right? , you know, they're gonna be
66:27 well functioning high rate. So you're you will get your best measurements during
66:33 time. And that phase is they typically distinguish between early middle and
66:40 log, particularly mid to late. , um let me see if I
66:45 my, there's my pen cursor, find that thing. There we
66:50 OK. So mid log is something year that spray Marcus nuts.
66:57 Mid log. And then here, log. All right. So the
67:05 here in this block here is kind where it's gonna be most active.
67:08 . Most, most uh um where want to kind of do your measurements
67:12 whatnot. Um If you want to the culture, uh you might do
67:17 before you get the late log. be satisfied because we're running out of
67:36 , right. So then you're gonna into stationary phase. But while we're
67:39 log phase in terms of cell so of course, it's represent three
67:45 growth, right? Exponential growth and size. Remember they're gonna be
67:48 in these elongated cells that are ready divide. So the proportion of cells
67:54 in that state. So very kind they get, they kind of get
67:58 little bit bigger than they divide, ? So they're gonna have lots of
68:00 in that state, which is why trade represent cells and kind of their
68:05 size. Ok. Um So, you get the late log nutrients are
68:13 limiting, right? Growth slows right? So you enter stationary
68:17 OK? And um you know, flat. So basically growth rate and
68:24 rate are about the same. Um that's kind of a misnomer because you
68:31 have cells that are viable, They're alive, but they're not
68:38 right? So you might consider those , but they're not, right?
68:41 just, you know, you can those, those are certainly part of
68:44 mixture. OK? But um certainly not really growing at all.
68:51 So uh stationary phase and so cell decreasing. So in stationary phase,
68:57 cells are now in survival mode, ? Running out of food,
69:02 Um So what do I do to myself um survive this period of we're
69:10 out of food. Ok. you make yourself smaller, is one
69:14 the things smaller, smaller cell volume less to keep up with.
69:18 And so you can um uh prolong your life by doing that.
69:24 Um Of course, stress response. there's all kinds of um proteins are
69:30 and things to kind of uh um energy, right? To not only
69:37 the proteins that are absolutely necessary, up other processes off, right.
69:42 it's kind of what's going on in period. Again, if, if
69:45 imagine the bacterium having a mind, ? The goal is let's conserve and
69:52 the goal that food is gonna come , right? So let's keep in
69:56 mode for, for this period. they can also what happens is that
70:01 die, begin to die. they, that's food, right?
70:05 that they can actually subsist on, that as food that sells rice.
70:09 kind of cannibalizing if you will. . Uh But eventually, so
70:20 through stationary phase, there is some level nutrients are there. That's why
70:27 can, so they can kind of viable. OK. But um eventually
70:33 does get to zero then and that's very quickly when that happens here goes
70:39 too is exponential, right? That's death is exponential with no nutrients at
70:43 and very quickly they die. And so it's um and so what
70:50 like um treatments like anise antiseptics and and whatever your favorite way to slope
71:04 down. OK. How big can make that negative slope? That's
71:08 you know, different types of anti agents are all about when you study
71:13 you're fairly focused on that part of curve. OK. So, um
71:19 as mentioned, kind of in the I say kind of death cells and
71:23 phase that you might call dead in quotes may actually just be these types
71:29 we call persists. Uh They're just of hanging out, they're viable,
71:34 they're not really growing, they're not . OK. And so many times
71:39 so if you maintain right, the potential, right, that proton motive
71:44 , right? Just have enough to that, that'll give you energy to
71:48 viable, but not enough for you begin actively dividing. So, but
71:52 how you can remain kind of OK. And not growing. And
71:59 it's, I guess in the context antibiotic resistance, it's um types who
72:05 , can do that right? In presence of antibiotic. They kind of
72:08 , OK, I'm just not gonna because the antibiotic works best if I'm
72:12 . So it just kind of hangs , that's a form of really
72:17 which I, so um OK, think any questions about that.
72:25 So we'll um pick it up next with bios. OK. Uh See
72:32 Thursday. Yeah. So um I gonna come
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