| 00:09 | mm hmm. Mhm. Ah Welcome folks. Um So this |
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| 00:29 | remember that we have first exam uh this week thursday and friday. Um |
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| 00:37 | questions about content? Look at the review sheet. Um Got questions uh |
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| 00:45 | office hours. Of course you can't officers can always arrange for another |
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| 00:49 | it's fine. uh chapter five So I just did that before flat |
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| 00:55 | uh weren't able to cover those on um unit exam or a quiz |
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| 01:03 | So I just put up some there's maybe six or 7 questions um After |
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| 01:11 | questions the answers were on on last , so it's not for Great or |
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| 01:18 | but just so you can have some to go through the latest chapter five |
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| 01:23 | . Um and that's in the unit one folder. Okay. Um Let's |
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| 01:31 | . So remember also that although there's a couple of problems on the exam |
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| 01:37 | are the tax relation type with the growth, you can't have a calculator |
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| 01:43 | you. Not a cellphone calculator obviously a calculator of any kind, handheld |
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| 01:48 | casa will be made aware of that that you won't be bugged because you |
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| 01:53 | a plague on a calculator or Okay. Uh what else? Smart |
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| 01:59 | . So this week there's no no quiz because we have the examiners |
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| 02:03 | Smart work. Do sunday. There the The Chapter five due on Tuesday |
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| 02:09 | nothing nothing next Sunday will transition to next unit. Um Next monday monday |
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| 02:19 | start unit too. So in that six, 13 and 14. Okay |
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| 02:27 | And that material will be up tomorrow . So if you're eager to start |
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| 02:33 | that already, you can if you to. Um So they were gonna |
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| 02:37 | up Chapter four. that's a that's mistake we're gonna talk about in those |
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| 02:43 | at the beginning, the definitive check that's all we got left and then |
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| 02:47 | for five as short as well. and so Wednesday we'll get through most |
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| 02:54 | pretty chunk of five today. It's that long but it means that Wednesday's |
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| 02:59 | they're not gonna be along. I'm going near for the whole period. |
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| 03:03 | luckily I don't know, 45 minutes something like that. Uh to finish |
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| 03:09 | the last of Unit one which is end of Chapter five. Okay. |
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| 03:14 | let's see um Any questions. yeah, yeah. Formulas. |
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| 03:25 | That formula. You need to memorize formula that that will be, there'll |
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| 03:28 | a question saying blah blah whatever. then there'll be the equation at the |
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| 03:34 | of it the here here here. . Um alright, so let's start |
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| 03:42 | the question. Okay, so this into um Plympton who may or may |
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| 03:50 | not talked about yet. Let's see you, can you answer this |
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| 03:55 | Sorry, surface and the ability to our absolute requirements for which one of |
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| 04:07 | ? Mm hmm. And do I've uploading the clicker points on blackboard. |
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| 04:14 | if you see, don't see I'm Oh is is it giving you |
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| 04:25 | on your on your and the Okay. One more time. What's |
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| 04:33 | what's it doing to options? jesus, okay. All right. |
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| 04:45 | then that yeah, I'll just give credit for it. So But what's |
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| 04:49 | the what's the answer here? that's the that's biofilm formation. So |
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| 04:55 | completely weird. Why don't you give two choices? All right. My |
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| 05:00 | just give you credit for that So, um Alright. Hopefully other |
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| 05:04 | aren't like that. Um Okay. yeah, so service. So also |
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| 05:12 | , so just we talked about this the end last time or towards the |
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| 05:16 | end of covered biofilm formation. So uh you know, it's it's |
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| 05:24 | couple of things to remember. You , obviously the surfaces, the surface |
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| 05:29 | required uh to be able to stick that surface and not receive having the |
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| 05:34 | of a pillai, we're gonna be to be able to stick to that |
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| 05:40 | uh initially um uh being reformed the . So remember the biofilm is not |
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| 05:48 | just a random event? Uh It this material that kind of sticks everybody |
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| 05:55 | . Right? The former film. it's it's a planned event if you |
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| 06:00 | it's a gene um encoded event. genes involved. Um It's it is |
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| 06:08 | but a random process. Okay. to be able to one thing when |
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| 06:11 | remember the last time I mentioned you to visualize kind of in this diagram |
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| 06:16 | that there is a source right of supplying this thing. Right? And |
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| 06:23 | um whether it's a pipe, you look what's going through your or shower |
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| 06:28 | maybe or something. You know there's there's a flow of nutrients which because |
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| 06:33 | what drives it. So it's the of nutrients it has to be there |
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| 06:38 | to create the favorable environment. And what the quorum sensing mechanism is |
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| 06:43 | Is presumably if you have a lot members there then it is a favorable |
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| 06:49 | because they aren't settling. They're they're dividing. And so that's how you |
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| 06:55 | that threshold level of these chemical signals they get to a certain threshold then |
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| 07:01 | triggers the process. And so The be well they wouldn't be gathering and |
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| 07:07 | bigger numbers and getting to that threshold it work. It's an indicator that |
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| 07:11 | a favorable environment. So that's 400 . Okay, so the chemical signals |
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| 07:17 | as they are reached that threshold that themselves actually induce expression of. And |
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| 07:24 | jeans the first of those being the party saccharine information. So that's a |
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| 07:29 | Zachariah protein. It can be proteins there as well think of this kind |
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| 07:33 | glue holding everything together. Um and of course growth proceeds from um on |
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| 07:41 | surface and two dimensions to three dimensionally it gets of course massive with billions |
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| 07:47 | selves um the environment in the You can have cell types in the |
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| 07:55 | that may differ somewhat from those on interior. Okay. Um The ones |
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| 08:01 | the next year can be more susceptible say antibiotics if it's a it's a |
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| 08:06 | important biofilm, um a pathogen on catheter or what have you uh surface |
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| 08:14 | a heart valve maybe or something uh uh antibiotics may work better against those |
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| 08:22 | the outside of biofilm represented on the because they can't penetrate that well. |
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| 08:28 | Maybe that sets up some kind of types inside that biofilm. So uh |
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| 08:34 | you're pathogen biofilms can be quite um because that in certain trying to get |
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| 08:41 | of it, uh the dissolution process kind of our film falling apart. |
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| 08:47 | you will, if nutrients cannot be , write the nutrient flow is shut |
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| 08:53 | . Now members can't aren't surviving. it's like okay, time to go |
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| 08:58 | . And the dissolution process occurs. so remember, we have 22 cell |
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| 09:05 | or types that are kind of basically swimmers. And those that once they |
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| 09:10 | on the surface, they tend to that. Uh so plant tonic for |
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| 09:16 | cells versus uh sticker cells they call . Okay, so those are kind |
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| 09:23 | the highlights really of the biofilm formation . Any questions about that. All |
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| 09:32 | , So um So in those So next is our last thing to |
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| 09:38 | about in Chapter four. Um So is just to illustrate uh if you're |
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| 09:43 | a lab uh you'll look at these if you're in the 5 30 |
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| 09:50 | Um But or tomorrow's lab of course the uh we're actually doing a gram |
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| 09:58 | . We're not doing a sports statement because you can look at you can |
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| 10:01 | you can see sports not doing a fan and you can tell from this |
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| 10:04 | here. So um they look like tile. Well they are refract tile |
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| 10:12 | . So here's an endospore in in cell. This is probably a what's |
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| 10:18 | a free in those pores been Okay. Um This is likely just |
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| 10:23 | completely vegetarian called vegetative cell vegetative The functioning fully functioning healthy normal |
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| 10:32 | Um we'll talk about that in a . But this is this is material |
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| 10:36 | I think it's the one from the 250 million year old bacteria found in |
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| 10:42 | , Sea salt needs Carlsbad. So the one below 40 million year old |
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| 10:48 | found in the stomach of a be in Hamburg. Okay. So the |
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| 10:55 | is in those spores spores are prevalent old. The kingdom's. Okay. |
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| 11:03 | plant, What have you? Um forms are typically in the reproductive |
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| 11:10 | Okay, fungal sports for example. . Um uh There are other types |
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| 11:16 | bacterial sports that aren't like this that involved in uh reproductive aspect. Um |
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| 11:25 | protozoa ones can form not necessarily sports they form a form called a |
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| 11:31 | Okay, so it's a cyst or . These are all typically what we |
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| 11:37 | dormant dormant forms viable. Okay. but not really actively reproducing. |
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| 11:46 | So I think it was plant seed plant seed represents an embryo inside that |
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| 11:52 | . Okay. That will not begin grow until favorable conditions. Right, |
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| 11:59 | . Right. Um So these are similar that the as you can |
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| 12:05 | these things remain dormant for millions of and then revived under the right |
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| 12:13 | And so um so that's the thing the endospore is particularly unique. So |
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| 12:19 | saying sport, but when you put window in front of it that raises |
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| 12:23 | to another level of resistance. And uh frankly why we use the |
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| 12:30 | in lab to sterilize media, we the autoclave to basically kill endorsed sports |
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| 12:37 | may be present. Okay. They particularly resistant resistant to chemicals. Um |
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| 12:44 | uh extremes of ph um temperature. , radiation. So they can be |
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| 12:53 | uh um already formed. Okay. so and they particularly form under these |
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| 13:02 | of stress conditions, lack of nutrients elevated temperature. What happened? Um |
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| 13:09 | the endospore themselves? The endo spore are only in two groups of bacteria |
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| 13:16 | and bacillus. So clostridium you're familiar the is tetanus bike system. This |
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| 13:23 | caused by members of that group. anthrax was really the only dangerous one |
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| 13:30 | in the bacillus group. Most bacillus pretty much benign soil, they're both |
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| 13:36 | of soil inhabitants. Okay. Um clostridium group and they're both grand |
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| 13:46 | And what really differentiates them both. there Um we'll talk about this as |
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| 13:51 | start. Chapter five um behavior in of oxygen. Okay, so bacillus |
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| 13:58 | typically aerobic like us. Um maybe aerobic, what we call micro era |
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| 14:06 | . But Australians are obligate anaerobic, live in the presence of oxygen. |
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| 14:12 | . Or very very low levels of . They don't use auction. Their |
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| 14:16 | anna ropes. Okay. And uh I mentioned, there are a number |
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| 14:21 | disease causing types in the group and for producing a plastic toxins. |
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| 14:27 | toxin of botulism toxin, gas, causes tissue death in contaminated balloons um |
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| 14:36 | so forth. So there we'll talk them in the context of diseases at |
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| 14:42 | end of the semester. But but , those two groups facilities and frustration |
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| 14:47 | these. And those sports which is unique, unique type of sport. |
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| 14:51 | so two former sport is not a thing. It takes several hours. |
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| 14:57 | number of genes involved. It too a is a organized process. |
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| 15:04 | coordinated process ending up with a um , proceeded by D. N. |
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| 15:11 | replication and then a what's called There's an unequal compartmentalization occurs or in |
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| 15:19 | smaller compartment. The endless performs. in the process we formed this double |
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| 15:26 | we form, we deposit a layer pepper look like. And in their |
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| 15:32 | different types of what are called coat . Uh So it becomes a very |
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| 15:37 | um resistant structure. Okay. And it used the terms mother cells, |
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| 15:43 | floor. Those are the two unequal that form um the the removal of |
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| 15:51 | . So water in a Sell you as well. My life is pretty |
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| 15:55 | 70% water. And when temperatures it's the increase in water temp that |
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| 16:02 | does the damage inside of the Okay. Um so or if it |
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| 16:08 | ice crystals form and cause damage So it's it's the just the amount |
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| 16:13 | water present in cells that can actually the thing that kills them when extreme |
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| 16:18 | of temperature may arise. And so taking water out, that actually helps |
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| 16:24 | terms of viability. Okay. And when those four formation occurs, a |
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| 16:30 | of water is taken out. So gotta go down from about 72, |
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| 16:33 | 20-30% of water. And that helps keep it viable when it's in this |
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| 16:39 | dormant state. Okay. Um The process is really just coming out of |
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| 16:45 | end of those four states into a cell. Okay. And so we'll |
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| 16:51 | over those terms here. So When you look at a endospore former |
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| 16:59 | culture, I'm just throwing it not for the purpose of getting into form |
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| 17:05 | . But you're just growing it for reason, you'll typically see three types |
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| 17:10 | selves. Uh we're looking under the . Okay. You'll see. Uh |
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| 17:15 | these are what we call the completely is a gram stain, completely bluish |
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| 17:22 | colored cells. This guy, this , this guy. These are completely |
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| 17:32 | . So let me get hold There were these, Yeah, pretty |
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| 17:41 | ahead of mark. Okay. Um again, you see on the |
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| 17:45 | right vegetative cell, there's no speculation on. It's kind of going through |
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| 17:49 | hole, normal growth phase, so divided etcetera. Um But then you'll |
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| 17:57 | cells that have this fact out That's the end of the sport for |
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| 18:01 | . Okay, so these cells are the process of and speculation. |
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| 18:07 | form the end of sport. You have so they have completed that |
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| 18:10 | will stay in the south. It's a free end of sport at that |
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| 18:15 | at that time. Okay, so the end of the process. That's |
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| 18:18 | happens. Okay, so um the of sports, I don't don't worry |
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| 18:27 | memorizing these things down here. I wanted to mention it because with these |
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| 18:33 | groups of endospore formers, you can the fact that they species will form |
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| 18:40 | how this force formed and how they . And you can use that as |
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| 18:45 | way to identify members of those And so the endospore itself can cause |
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| 18:53 | and it can kind of blow up swell the cell like you see here |
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| 18:59 | here here Or it may not it just you know not swell like |
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| 19:04 | A non sworn numbers are swollen. position of the sport, is it |
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| 19:08 | one end is a kind of toward end. So sub terminal, this |
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| 19:13 | terminal. Is it central in the ? So you can have three |
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| 19:17 | Can be swollen, hands swollen. Something maybe club shift is typically what |
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| 19:22 | call that. But anyway, the being you can identify these members based |
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| 19:28 | their sport and those four characteristics. . Um Now the cycle. |
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| 19:35 | so speculation begins with um DNA replication so the chromosome replicates but it forms |
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| 19:46 | elongates alright. Taking up the whole of the self and that's what we |
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| 19:52 | an actual filament. Okay. And it will uh copy goes into each |
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| 19:59 | . So now we get the compartmentalization unequal in size for the mother cell |
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| 20:06 | than the four sport for sport is the end of the four formation will |
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| 20:11 | occur. Okay, the mother cell uh function is really to direct the |
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| 20:21 | chelation process. There's a bunch of involved. And so the mother cell |
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| 20:26 | have proteins being expressed and those are proteins that are acting on the force |
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| 20:34 | DNA and directing the the process. . And so once the mother cell |
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| 20:41 | kind of started that process then the sport kind of just does its |
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| 20:47 | And what happens is that the mothers actually engulf that forced board to form |
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| 20:55 | double membrane. Right? So you the double membrane here and then that's |
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| 21:00 | people like him will be synthesized inside . And that double membrane. Okay |
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| 21:08 | you form a type of light can within there. You have some other |
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| 21:12 | being deposited there as well. What called coat proteins. Okay. Um |
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| 21:17 | unusual molecule called diabetic Olynyk acid or . P. A. Is thought |
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| 21:25 | be involved in binding to the chromosome the chromosome calcium. Mines are thought |
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| 21:32 | have some process in the water extraction of the forming spore. Um Also |
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| 21:40 | in nature. So calcium gets lines deposited in there and so were forming |
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| 21:46 | what they call ex operandi um um Spork hoped eventually is the at that |
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| 21:54 | that we're pretty much done. And so so from here up to |
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| 22:00 | from up to here throughout I'd say stage here. So all these stages |
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| 22:09 | here where we see that the like recall the gram stain the cells with |
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| 22:19 | endospore in the middle right? The cell containing an endospore. That's what |
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| 22:25 | see at these stages. That's what cell would look like. Okay. |
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| 22:30 | like real quick beep beep that would like this. Okay so it's just |
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| 22:40 | don't know by looking at the cell a microscope. We don't know which |
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| 22:45 | of these is not identifiable with light . Which which of these stages in |
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| 22:50 | . It is. But that's what would look like under the microscope. |
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| 22:54 | . And finally, the free sport be the seller vegetative cell kind of |
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| 23:00 | around it in the end those Those four. Okay. Of |
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| 23:04 | In that state uh that we've seen can stay like that for thousands of |
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| 23:13 | . Okay. Um More typical of is uh is not that long but |
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| 23:20 | know, it can be as long it takes until there's a favorable |
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| 23:24 | Okay. And they'll they'll germinate. . Um so that's in the nature |
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| 23:30 | the end of this is only with windows. Because you hear the terms |
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| 23:34 | a lot. But this this is for this for this for this |
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| 23:38 | These two groups. Okay. Um questions about that. Yeah. |
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| 23:46 | So And that concludes chapter four. . And uh Let's move on to |
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| 23:57 | . And so we're only covering a of this which is focused on what |
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| 24:02 | call zero tolerance, which is bacterial if you will in the presence of |
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| 24:09 | . Okay. Uh it's it spans spectrum of you know how the growth |
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| 24:16 | you'll see. Okay. And then talk a little bit about physical chemical |
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| 24:21 | uh control of bros. Okay, first with oxygen. Okay. |
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| 24:29 | we haven't yet. Yes, we'll this in unit two. Uh talking |
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| 24:36 | respiration and all what's involved with But um in a aerobic world that |
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| 24:44 | live in. Um number one oxygen is a very reactive molecule. |
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| 24:53 | . The molecule that likes to grab . Okay, and become reduced. |
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| 24:59 | a powerful oxidizing agent. Okay, course we use it to help us |
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| 25:05 | . Right with the old. If recall the uh auction at the end |
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| 25:10 | our electron transport chain. So you auctions the terminal except er right, |
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| 25:17 | worry about that yet. I'm just to you its role here. And |
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| 25:24 | in the in the process of we then form water. Right, |
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| 25:27 | picks up electrons are reduced to Right? And so that's how we |
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| 25:32 | that to breathe. And so um oxygen being very reactive can also interact |
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| 25:42 | molecules enzymes involved in the process. . And the results of that are |
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| 25:50 | side reactions that occur. Okay, these species you see here super |
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| 25:56 | hydroxide, radical hydrogen peroxide all reactive themselves. Okay, Especially super |
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| 26:04 | So um the uh and that can damage. So those molecules will interact |
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| 26:11 | proteins create acids uh causing that. . Uh same in our own |
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| 26:18 | Same thing happens. Okay. And too, so the cells would have |
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| 26:24 | as a result of the damage You have to have protection. |
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| 26:29 | And the protection comes in the form three different enzymes. Okay, so |
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| 26:35 | call these three reactive species not surprisingly oxygen species. Again, this process |
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| 26:44 | . We just happen to looking at cell. That is an aerobic inspired |
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| 26:49 | , same molecules Okay. Are generated the organism uses oxygen or not. |
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| 26:59 | . Because auction can still wrapped in molecules. Okay, so you don't |
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| 27:05 | this could be a a bacterium that doesn't use oxygen at all. But |
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| 27:10 | will still be affected by it and still get these species form and it |
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| 27:14 | damage the cell Again unless there's Okay, in the form of these |
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| 27:21 | ourselves have all three of these. . If you've ever poured hydrogen peroxide |
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| 27:26 | the cut, you may have had your finger or leg whatever. Well |
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| 27:30 | as a bubble. Right? The is due to the catalyst that those |
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| 27:35 | have. Okay, neutralizing the effect the hydrant profit. Um anyway, |
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| 27:42 | three of those those three molecules Okay, afford protection against damage. |
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| 27:50 | , so here is how auction can with this is a component that's in |
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| 27:57 | respiratory chain over here. Okay, it's also present in other other cells |
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| 28:02 | don't use auction. It's it's it's an uncommon molecule. And so the |
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| 28:09 | is the formation of super oxide Yeah, now that's one step. |
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| 28:14 | , so how do we neutralize Well, S. O. |
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| 28:17 | Okay. Super oxide. Disney taste S. O. D. For |
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| 28:21 | . Okay, that will combine with to form hydrogen peroxide still reactive but |
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| 28:29 | peroxide is not as bad as super but still bad. Okay, but |
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| 28:33 | the catalysts and paroxysms can take care that. Okay, the end result |
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| 28:39 | basically to form water. Right? that's not harmful obviously. So so |
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| 28:45 | catalyst and proxies do their job to the um reduction of these reactive |
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| 28:53 | Okay, so um so when we at how bacteria behave in the presence |
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| 29:02 | air I. E. Oxygen. . They will have various responses. |
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| 29:09 | basically five types of responses. We'll . Okay, there's those it's all |
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| 29:15 | correlates to the protection they have or have for level of protection. So |
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| 29:27 | may have some protection. Maybe have or more of the enzymes. Maybe |
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| 29:32 | have all three but they don't have at maximum levels. Right. So |
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| 29:36 | gonna be variation. Okay, but behavior they have in terms of growth |
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| 29:42 | in the presence of their directly relates what do they got in terms of |
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| 29:48 | . Do they have all of Some of them not not the maximum |
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| 29:52 | or do they lack it completely because do not have any of it. |
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| 29:58 | , and that's gonna tell you what patterns will will be. Okay, |
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| 30:03 | let's look at Okay, this Well, not yet questions coming up |
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| 30:08 | this. Like Okay. How do detect these patterns? Okay, so |
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| 30:14 | have a medium that will has a of options hide the top or the |
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| 30:21 | . Okay, so we create that this medium formulation called fluid dialogue |
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| 30:30 | Okay, so it contains a dye there that turns pinkish when oxygen is |
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| 30:37 | , less pink when there is less present. So so you see how |
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| 30:41 | tube is dark red, dark red pinkish at the top, and less |
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| 30:44 | you go down because the levels of are going down from top to |
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| 30:49 | Um It's a it's a um it's jelly like matrix. It's not completely |
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| 30:57 | and it's not completely solved. It's between, it's kind of a jelly |
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| 31:00 | matrix. Um There's chemicals in there can bind the pockets. Um When |
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| 31:07 | make this you um of course boil put in the autoclave that will drive |
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| 31:13 | any gas in there. Okay? then as it cools down and air |
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| 31:18 | to diffuse back in it. Um because it's a jelly like matrix air |
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| 31:27 | and slowly. Right, so go it sets up a grading because those |
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| 31:31 | chemicals kind of bind up oxygen. the net effect is uh to create |
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| 31:37 | grading. And so um the um when you annihilate this medium, |
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| 31:46 | you're gonna take a wire loop. ? And you will cede, you'll |
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| 31:52 | take some binoculars and you'll just go the whole length of the tooth with |
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| 31:57 | . Okay, so what you wanna is basically seed that entire tube with |
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| 32:04 | through the whole length. Okay, what you're doing. So then what |
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| 32:09 | wanna do is incubate, allow themselves grow. Okay so then what you're |
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| 32:13 | is asking is which in which area the tube? One of the |
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| 32:20 | I see it begin to replicate because at the they're at the part of |
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| 32:24 | tube they like to be at from best growth. Okay. So is |
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| 32:29 | going to be here? Is that only place we'll see growth? It |
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| 32:34 | be there, we'll be there or it be throughout the whole too? |
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| 32:39 | right. All the pimps, Goes back to what the protection it's |
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| 32:44 | . All right. Um That will where it will grow in that |
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| 32:48 | Okay. Um because even though you the whole the whole thing top to |
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| 32:55 | itself, we're only depending on what what its capabilities are. The only |
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| 33:03 | cells that you see that that will begin to replicate will be where it's |
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| 33:07 | for. Okay so uh so with in mind now we can get the |
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| 33:14 | . Okay so um let me reset . Okay that does it give you |
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| 33:23 | answers again, jesus christ Hold on me see if I can fill it |
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| 33:28 | that here, just real quick mm . I I don't I can't I |
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| 33:41 | do it from here to go back the other software. So alright you |
|
| 33:45 | there's another three questions to try. . Alright so this is rude |
|
| 33:50 | five strains are grown medium uh to the growth pattern. So based on |
|
| 33:59 | results, which strain has S. . D. Catalysts and proxies but |
|
| 34:06 | reduced levels of these or maybe missing or two of the enzymes. |
|
| 34:13 | Um So mm hmm. What would your guess on that one? What |
|
| 34:22 | be your guess which strain? Based these patterns? Which strain A. |
|
| 34:27 | . C. D. Or Has these protective enzymes but but not |
|
| 34:34 | maximal levels or maybe missing one or . Okay. What respond? Which |
|
| 34:41 | can you eliminate? Start there. ones can you say? It's not |
|
| 34:46 | at all, can't we? Okay. Um See see because she |
|
| 34:55 | what complete an arrow? Right. . Stick at the bottom of the |
|
| 35:03 | . But there's no oxygen. that's the only place that grows. |
|
| 35:07 | . Remember we actually we seeded this tube. Yeah. Right. But |
|
| 35:17 | but only the ones down here are ones that actually continued to grow. |
|
| 35:22 | . So yeah, I wouldn't be it. Um You wouldn't think it |
|
| 35:27 | be a right because that's up there the highest levels of oxygen. The |
|
| 35:32 | levels of oxygen are at the top the tube. Okay. So that |
|
| 35:36 | has the full complement of enzymes and normal amount. Okay, so um |
|
| 35:46 | B. And C. Okay. two our um grows throughout. |
|
| 35:55 | And but there is a they may the same but is there a difference |
|
| 36:03 | B. And D. Yes. the difference at? That's right. |
|
| 36:10 | is more even distributed. Be tends have more cells up here. All |
|
| 36:15 | . It grows throughout like need does there's more cells at the top. |
|
| 36:18 | the more cells at the top because why or auction it means more because |
|
| 36:26 | means more different. Well, because you're on the right track. |
|
| 36:31 | more opportunities more growth and there's more because not more nutrients. But it's |
|
| 36:41 | has to do with what we haven't about it yet but it has to |
|
| 36:43 | with um an oxygen metabolism. so an oxygen using the tablets and |
|
| 36:49 | his 90 PS more https generally always more. Okay. So that's why |
|
| 36:57 | indeed differ. You distinguish those two looking at the top because B is |
|
| 37:04 | we call a faculty native animals. . Like an E coli just like |
|
| 37:10 | . Um If there's you know in where there's oxygen present, okay. |
|
| 37:15 | will grow uh and it will grow a little bit more because it's just |
|
| 37:20 | nature of an oxygen using the tablet you more a little more https. |
|
| 37:25 | De is what we call a and is on the next slide anyway. |
|
| 37:31 | d is an aero tolerant and a . zero tolerance being the operative |
|
| 37:37 | Okay. You can tolerate oxygen because has a protection but it doesn't grow |
|
| 37:44 | at the top because it does not oxygen at all. They're typically fermented |
|
| 37:51 | . Maybe they're anaerobic or aspire ear's , but they don't use oxygen. |
|
| 37:56 | there's really no difference for it at top compared to a faculty of |
|
| 38:00 | which can use oxygen. Okay. B Is the micro era file? |
|
| 38:10 | . And um I would say that probably what the answer would be to |
|
| 38:16 | one. Okay. Um It has it uses oxygen. Okay? Um |
|
| 38:24 | because it has reduced protection. It's going to be sensitive to high |
|
| 38:30 | of oxygen. So like In the it's 20% 02 atmosphere conditions, |
|
| 38:37 | Um It likes 10%, Something like . Okay. And so uh so |
|
| 38:46 | me just flip to the slide so can break these down. You |
|
| 38:52 | I break them down kind of this . A robe and a robe, |
|
| 38:55 | , faculties can go either way. . Um So Arab types, you're |
|
| 39:02 | . Arabs like us. That's what are. Um And the micro era |
|
| 39:08 | . Okay. So the key thing the era of is they use oxygen |
|
| 39:14 | respond. That's the only way they Okay, Must have it. It's |
|
| 39:20 | that one type is a little more to the atmospheric concentrations. The micro |
|
| 39:26 | . Okay. And to be in a precarious world, most things |
|
| 39:32 | really micro aero files or anna Rome's caves, sheer numbers. That's many |
|
| 39:41 | that's kind of the categories they fall . Okay. Um You're gonna robes |
|
| 39:49 | not. So in contrast to the do not use auction. All |
|
| 39:55 | Be the ferment and or inspire and , which we'll talk about later, |
|
| 40:01 | they don't use auction. Okay. so now the difference between these two |
|
| 40:07 | the protection, right? Obligation. don't have any protection against those |
|
| 40:11 | So there that's toxic to them. , So they're the ones that grow |
|
| 40:15 | the bottom of the tube. Uh paws treaty is in that |
|
| 40:20 | Okay. The aero tolerant and Arabs not use auction, but it's not |
|
| 40:26 | they have the protection against it. have the S. O. |
|
| 40:29 | The catalyst etcetera. Okay. So can go throughout the tube. |
|
| 40:36 | Um but they just don't grow more the top because they can't use |
|
| 40:40 | Okay. Your faculties of types can can use 02. If it's |
|
| 40:45 | you don't have to use it. was not available. They can grow |
|
| 40:49 | way. Okay. E coli is faculty in general. It can grow |
|
| 40:55 | with oxygen. It can grow, can ferment, no auction can and |
|
| 41:00 | can also respond to an honorable thing do all three of those things. |
|
| 41:04 | know what it does, how it , depends on what's available to it |
|
| 41:07 | what the conditions are. Okay. um so so of course it has |
|
| 41:13 | protection has a complete protection against the effects of options? Okay. Um |
|
| 41:22 | questions. Yeah, but it's Do you still want to take |
|
| 41:30 | No, because I'm gonna get free you only had two choices on your |
|
| 41:33 | . So it's not working right? got if you're here you got you |
|
| 41:40 | golden. No problem. It's it's to me to do that. Figure |
|
| 41:45 | what's going on with this. But not penalized for that, so, |
|
| 41:50 | hmm. Yeah. Okay. So you have so if it has the |
|
| 42:04 | the serotonin and a robe and effective a robe, you would guess would |
|
| 42:10 | the full complement of protection because they throughout the whole two top to |
|
| 42:14 | Right? Highest levels of 02 to . Oh, too. Right. |
|
| 42:19 | uh the Albuquerque and a robe doesn't any protection. Right? The arab |
|
| 42:27 | the top. So you're gonna think gonna have it should have the max |
|
| 42:30 | , right? All three enzymes at the high levels. Right? So |
|
| 42:37 | what's that leave us with the micro have protection, but not the maximum |
|
| 42:44 | . So that tells me, well, it can use oxygen or |
|
| 42:48 | lower levels of oxygen the time. it wouldn't be at the top. |
|
| 42:53 | . Wouldn't be throughout the tube. ? You would think would be somewhere |
|
| 42:57 | the middle. That's kind of the process. Yeah. Makes sense. |
|
| 43:05 | . Um Okay, so we just about that. All right. Um |
|
| 43:15 | , so now talk a little bit get into physical chemical control methods to |
|
| 43:24 | microbial growth. And so we talked So number one we we might be |
|
| 43:30 | whole batch growth curve? So the here we're talking about now we're focused |
|
| 43:38 | the death death rate, death the phase or stage of the curve. |
|
| 43:45 | this is all about what kind of can we use to control microbial |
|
| 43:50 | Um The and of course there's different for that. Obviously whether it's not |
|
| 43:57 | setting, hospital etcetera, four million restaurants, right? Or other services |
|
| 44:04 | you know um can be a situation people can get infected. Okay. |
|
| 44:13 | you always obviously when you're testing different and seeing how they control growth, |
|
| 44:21 | target pathogens but of course everything is to be reduced and leveled. |
|
| 44:26 | But you specifically you're trying to target for the most part. Okay so |
|
| 44:31 | you're testing your treatments, whether it's disinfectant and accepted will have you you |
|
| 44:38 | have a set of target pathogens, ? And they're pretty much common among |
|
| 44:44 | the manufacturers of these things. This just one example where you see the |
|
| 44:49 | virus uh cold virus, um salmonella . Coli. So and you always |
|
| 44:57 | test among bacteria, gram negative gram because they will you you'll see differences |
|
| 45:02 | the nature of the cell envelope. , so um but what they all |
|
| 45:08 | do as well our state as well the house effective. Right percent killing |
|
| 45:16 | they produce? Okay 99% 99.99 9.99 happened? Okay. So what what |
|
| 45:26 | that really mean when we talk about one of the parameters they're used to |
|
| 45:37 | two evaluate or two uses the metric cell death. That's what we call |
|
| 45:45 | logs of death. Okay So when look at for example an area that |
|
| 45:51 | um a million cells. Right? when they do these things they typically |
|
| 45:56 | like a little especially with the surface . Ah They'll have like a little |
|
| 46:02 | that's like two inch by two and open in the middle right put on |
|
| 46:07 | surface and they'll take a sample and like a cell count and then take |
|
| 46:13 | area and then maybe spray with disinfectant then let it sit there and then |
|
| 46:18 | another cell count kind of how to these things. And so if we |
|
| 46:22 | like a million cells and then we disinfectant for example. And that disinfectant |
|
| 46:28 | well 99.9% killed. And then we're off 990,000 cells in that area. |
|
| 46:36 | so certainly uh significant. Okay so we're down to 1000 viable cells. |
|
| 46:45 | ? Um and we can keep going in that instance we've we've done three |
|
| 46:49 | of death. Right? And so talked about this thing called the |
|
| 46:53 | Value shortly and the D. Value how how long to get one log |
|
| 47:00 | death? Okay. So of course want to have a time element in |
|
| 47:04 | to evaluate as well because you might how long it takes? Three days |
|
| 47:08 | to get three logs of that to five minutes to take. Three |
|
| 47:13 | Okay. So time is always gonna ported Hellman here obviously. Okay. |
|
| 47:20 | And this can vary. Other manufacturers a metric called 12 and then we'll |
|
| 47:26 | how long it takes. 12 blocks death. Okay, so, um |
|
| 47:31 | , so let's look at this Well, let's throw out an answer |
|
| 47:36 | me because obviously the quicker you only two choices here. So don't |
|
| 47:42 | this goes back to I think I this before about my pet peeves is |
|
| 47:47 | bad use of the word sterilization. is sterilization pasteurization? Yes. Yes |
|
| 47:56 | no. No, of course Is sterilization is disinfection? No, |
|
| 48:03 | is an exception. Sterilization results in final reduction of the microbial population by |
|
| 48:10 | 0.9%. No, it would be . Right? 2 0. So |
|
| 48:18 | means destruction of all cells viruses, sports spores. Okay, what |
|
| 48:27 | No detectable signs of life unless you something. Okay. Yes, there |
|
| 48:34 | um an autoclave can certainly do But you can't autoclave. Everything. |
|
| 48:40 | and there are what we call sterility . Mhm oxide is one. We'll |
|
| 48:47 | about gas. Uh certainly gamma or rays can be a sterilizing agent. |
|
| 48:53 | there are certain agents, Even liquid that can be a sterile and |
|
| 48:59 | mm hmm. Pretty damaging. But still it's still considered disinfectant. |
|
| 49:07 | Uh it's there are things that are to bleach. I mean, it's |
|
| 49:12 | . Certainly good. But it's a it's not a sterilizing agent. Um |
|
| 49:18 | see. Okay, so this we already have a disinfection exceptions. |
|
| 49:23 | the difference, terrorists. Inanimate disinfectant, walls, countertops uh and |
|
| 49:31 | this tissue. Living tissue. Um for that reason, of |
|
| 49:38 | disinfectants can be um higher concentration that be more harsh if you will. |
|
| 49:45 | . Um and accept antiseptics. Of not because you're playing to living |
|
| 49:50 | You don't want to damage the Um, sanitation is a little bit |
|
| 49:55 | . So the disinfection and accepts Um of course target pathogens. |
|
| 50:02 | Um, sanitation is more overall reduction microbial numbers through sanitary hygienic practices. |
|
| 50:11 | , example. That example I give uh Mhm. Fast food place or |
|
| 50:18 | restaurants, whether you're a server or worked in the kitchen especially grew up |
|
| 50:23 | the kitchen. You know, you , what do you do? What |
|
| 50:25 | have to have, sharing it right? You have gloves on. |
|
| 50:31 | you do practices to minimize transmission of pathogens, right? You wash your |
|
| 50:43 | when you go to the bathroom right , we all know there's a sign |
|
| 50:47 | the door before you get out. said if you work here, wash |
|
| 50:49 | hands okay. Before um uh in the kitchen leaving uh storing food at |
|
| 50:56 | temperatures. They leave it out at temp for a prolonged period of |
|
| 51:00 | Um Free cleanliness, you know, the floor clean and countertops clean things |
|
| 51:05 | that. These are all hygienic practices can do himself reduces overall numbers. |
|
| 51:11 | you do use disinfectants can be a of that process when you're cleaning off |
|
| 51:16 | , um washing your dishes right? go throughout high temp kind of process |
|
| 51:24 | sanitize. Sanitize. Okay, so for the purpose of minimizing transmission of |
|
| 51:31 | food borne agents. Okay. And what sanitation is. Okay. And |
|
| 51:38 | yeah, it says reduced numbers to to what level. And those are |
|
| 51:43 | that are are specified by. Typically county health Department will specify the level |
|
| 51:51 | then they'll audit we all know I'm sure you probably heard the friday |
|
| 51:56 | reports on on the local networks and that's what they're doing. They're checking |
|
| 52:01 | into restaurants, checking to see that are following proper protocols, etcetera. |
|
| 52:07 | . Um All right. Alright. a little bit about terminology as it |
|
| 52:13 | to killing of microbes bacteria, static bacterial politic. Okay, so these |
|
| 52:24 | are kill they obviously they killed kill actively kill cells viable cap. So |
|
| 52:36 | we don't go into the technique a account. But what you're doing is |
|
| 52:43 | we take an environmental sample. You to know what the bacterial county |
|
| 52:47 | Okay. Let's say we have and worry about writing all this down because |
|
| 52:52 | just for explanatory purposes only. So we have a sample, |
|
| 53:00 | Bacterial cells. What you do is take this and you dilute right? |
|
| 53:07 | may do it several times. And you put them in little test |
|
| 53:14 | . Okay? And then you could them out far enough, okay? |
|
| 53:21 | you take a sample and you'll get on plate. You're trying to get |
|
| 53:29 | between 30-300 on the plate. What call a countable number of copies. |
|
| 53:35 | been done for decades. And so worked out the math that statistically to |
|
| 53:39 | accurate to be in that range. . And so um And what you |
|
| 53:46 | is you count the colonies. So you have 42 climbs on the |
|
| 53:52 | . Well then, you know that know the number of delusions you took |
|
| 53:55 | get to that 42 cars, Gives you the what's called a viable |
|
| 54:04 | viable because what you're counting on the living sounds right. And each the |
|
| 54:11 | is that each colony Arises from one . Okay. So in that |
|
| 54:18 | you're counting 42 cells. Originally there just 42 cells and they made each |
|
| 54:22 | column. Okay. That's why you to divert these out so much because |
|
| 54:26 | cells are super tiny. Right? if I just took a sample from |
|
| 54:31 | very concentrated broth like say right there no delusion and I put them on |
|
| 54:37 | plate it's just gonna be a a because we call it a wand. |
|
| 54:42 | there's no it's just a lot of right? You can get a bible |
|
| 54:47 | from that so you have to keep looting amounts. So the cells are |
|
| 54:51 | apart far enough that they form individual and that takes you know doing them |
|
| 54:58 | pretty far. Okay But once you that you get a plate with accountable |
|
| 55:03 | and you can do the back calculation you can estimate what's called C a |
|
| 55:10 | . Okay per mail see if you calling the forming unit. Right? |
|
| 55:14 | none of this stuff I'm gonna You can do this and laugh and |
|
| 55:17 | later just to show you how do get a viable can't. Okay and |
|
| 55:23 | what you do is um so the is when you've added okay Added in |
|
| 55:31 | three treatments here we've got a growing to which we've added a some kind |
|
| 55:39 | antimicrobial. Okay at the arrow. let me see. Okay what's happened |
|
| 55:43 | we've done that? Okay so the solid line is a viable camp so |
|
| 55:51 | can see that with bacteria seidel materialistic counts drop. Um But you're a |
|
| 56:03 | agent is one where it's inhibiting Alright it's not actively killing cells but |
|
| 56:10 | not growing Okay once you see the . Okay now the dash line. |
|
| 56:17 | so the dash line is total Okay? So in this protocol, |
|
| 56:24 | you can do that by looking under microscope. Okay. Is there any |
|
| 56:30 | ? And you'll see sets obviously. if you're looking at looking at samples |
|
| 56:36 | the microscope from out here. All , up here, you see that |
|
| 56:46 | and dead cells look the same, ? For And you can't distinguish. |
|
| 56:53 | , under the microscope. Under the . And maybe the library that sells |
|
| 56:57 | the same they're dead. But you count them as a as part of |
|
| 57:02 | total can't. Right. So that's here in total headcount, it's |
|
| 57:08 | Right? They're not they're not growing ? The dead cells and live cells |
|
| 57:12 | basically pretty much people. Okay. no debt increase in the cells. |
|
| 57:17 | in here. Back to the Okay. Number one electrolytic bachelor recital |
|
| 57:26 | counts going down. That's a common . Right? But then the difference |
|
| 57:32 | is a total count flat total Okay, so again, it's about |
|
| 57:40 | in dead cells. Okay. It a bacterial lyric agent. What's about |
|
| 57:46 | catalytic agent Doing license is the operative there. It's blowing up to |
|
| 57:53 | Okay, so a live cell you distinguish. So the only cells that |
|
| 57:58 | visible are live cells under the Okay, well in bacteriological agents are |
|
| 58:05 | down because bacteriological agent is blowing them . Okay. So you don't see |
|
| 58:12 | . Right? Don't don't see anything everything's going down. Right. Numbers |
|
| 58:17 | actively going down because the dead cells actually blowing because you can kill the |
|
| 58:22 | and leave it intact but you can kill a cell and blow it |
|
| 58:26 | Yeah there are eight. There are that work on the membrane, |
|
| 58:32 | resolve the membrane cell falls apart. agents kind of work inside the cell |
|
| 58:38 | be blocked from the synthesis and Children that way but they don't blow it |
|
| 58:42 | . Okay so back to the silo lyric agents. Right? The difference |
|
| 58:47 | those two um the you know with bacterial agents and I'm not sure that |
|
| 58:54 | are even used uh in a medical because well certainly if it's a gram |
|
| 59:01 | infection, remember the gram negative Nintendo ? If you give it back to |
|
| 59:06 | agent then you're blowing up and then in the toxins being released. Okay |
|
| 59:12 | part of something you wouldn't want to the gram negative infection. Certainly depending |
|
| 59:16 | severity of infection. Um but you really the difference in metro side |
|
| 59:23 | they both kill one just killed by themselves. One killed by not doing |
|
| 59:27 | . Okay, the material side of , they're not lowering the viable count |
|
| 59:35 | they are inhibiting growth. Okay. and so just just like, nope |
|
| 59:43 | here, this here is just meant reiterate that difference in how they look |
|
| 59:51 | the scope. Okay so here you and wrote A versus robi. |
|
| 60:01 | The total accounts. This will be total capital received many of these |
|
| 60:04 | Okay. Mhm. No change in head. Um It's kind of flat |
|
| 60:13 | then robi you see self disappearing as look at them under the scope which |
|
| 60:20 | indicates a bacteriological kind of effect right . Okay. Effect his sons are |
|
| 60:36 | blowing up so to speak back to . Um uh Any questions about |
|
| 60:44 | Okay. Yeah. Really? Yeah is again he's back to definition killing |
|
| 60:56 | . You know sell spores, A sepsis is um is a reduction |
|
| 61:03 | is a reduction in in microbe Especially pathogens. So yeah a substances |
|
| 61:09 | the is still in the same categories antiseptic sis Okay, A sepsis means |
|
| 61:16 | reduction. Yeah. Um aseptic technique kind of similar terms. It's meant |
|
| 61:23 | do things in a way that minimize . Any other portion. Yeah. |
|
| 61:36 | . Um with the back through um can be depending on the agent and |
|
| 61:43 | are you going actually down to It could be yeah but but not |
|
| 61:48 | not always it depends on the term means you're killing themselves by licensing |
|
| 61:55 | It could lead to a sterilization process sure. Yeah. Or I mean |
|
| 62:14 | terms of how they're being killed it again it depends on the agent that's |
|
| 62:18 | students sterilization so some may completely obliterate that way or maybe not. It |
|
| 62:24 | depends. Is it another question. . Yeah criterias you have to get |
|
| 62:32 | viable count on that because we're just at it just basically think considering you're |
|
| 62:37 | under the microscope that's your view of . So yeah you have to get |
|
| 62:40 | viable count to say is that bacteria that bacteria static? Right. |
|
| 62:46 | Any questions? Yeah. All Mhm. I I'm not gonna say |
|
| 62:55 | don't I'm going to say that if me I gotta I have to assume |
|
| 63:00 | if it was a gram negative infection you have to be aware of that |
|
| 63:05 | and and so it's you know how is the infection is a septic blah |
|
| 63:09 | blah blah. So yeah I'm sure do but just you have to have |
|
| 63:13 | back your mind. Yeah another Okay. Alright so let's look at |
|
| 63:22 | okay so this is a D. here. I mentioned that. So |
|
| 63:27 | all you're doing here is again focused the you know killing him fast. |
|
| 63:33 | ? How fast can we kill Obviously we're looking at the line that's |
|
| 63:36 | negative flow. So we've added a to Uh culture that started out at |
|
| 63:42 | to the 8th here. Okay. um uh and steadily dropping. Okay |
|
| 63:50 | what do you do is you pick couple of points? So for her |
|
| 63:53 | value is is it's one log one difference. Okay one log drop. |
|
| 63:59 | so we picked two points arbitrarily 10 to the 5th and 10 to |
|
| 64:04 | . Okay then just extrapolate down And so remember that that one log |
|
| 64:10 | death, you know, was 90 . Okay. And so and so |
|
| 64:19 | this is about a minute you said new value here is one minute. |
|
| 64:23 | mean that's how long it took to one log of death with this, |
|
| 64:26 | the treatment was. Okay. And one of the things that can |
|
| 64:30 | you know, how well treatment may . Um one can be. How |
|
| 64:36 | how many microbes are you dealing Okay. Um Organic loads. So |
|
| 64:42 | this refers to think of that as of the the dirtiness of the surface |
|
| 64:49 | say you're trying to clean okay if you want to disinfect the surface and |
|
| 64:54 | surface is heavily soiled okay with dirt grime or whatever. It's best to |
|
| 65:02 | do a regular cleaning first before you disinfectant. Okay. Because maybe that's |
|
| 65:08 | that organic material. That dirt if will grime is the thing that's absorbing |
|
| 65:13 | disinfectant. And it's not really even to the microbes. So uh it's |
|
| 65:19 | common to clean the surface first and disinfect uh the the types of microbes |
|
| 65:25 | dealing with. And you don't believe gonna know that specifically. But um |
|
| 65:31 | try to pick maybe uh an agent will be worked well on both. |
|
| 65:36 | negative gram positives, et cetera, kind of thing? Um The agent |
|
| 65:42 | , you know, is it very to the surface? Like are you |
|
| 65:46 | on stainless steel or something or are putting it on another type of uh |
|
| 65:51 | with a composition that's totally different. you know the stability of the of |
|
| 65:55 | compound. Is it uh light sensitive that may trigger something? Is it |
|
| 66:01 | How volatile is it? Alright if spray it on the surface and it |
|
| 66:07 | away evaporates right away, is that or not? So good? So |
|
| 66:11 | kind of all these factors play a and ultimately how effective it will |
|
| 66:18 | Um And so this log arrhythmic Right? So remember that a couple |
|
| 66:22 | the batch growth curve right? We lagged log stationary death. Right? |
|
| 66:28 | so the changes in numbers in log death are are exponential. Okay. |
|
| 66:34 | are log rhythmic. Okay. But never a scenario where you'll see something |
|
| 66:40 | that, okay. Where everything dies all at once. Okay. There's |
|
| 66:47 | that some rate whether it's a very rate but it's at some rate and |
|
| 66:52 | because it's all about members of the . My well very slightly. |
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| 67:00 | And they will accumulate damage at a rate. Okay. And once it's |
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| 67:06 | the threshold and cell dies that's gonna somewhat from cell to cell. Especially |
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| 67:12 | it's densely packed. You know, in the periphery may be more susceptible |
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| 67:15 | ones in the middle in the Okay so there's always gonna be some |
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| 67:20 | of a rate at which killing You won't, again, we're not |
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| 67:25 | see that. Okay. I'll fall the cliff at one time. |
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| 67:32 | Um, okay. So let's, , I, I don't, I |
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| 67:40 | want to do another question where you use your clickers again. So, |
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| 67:45 | , what's that for this? Let just check. Yeah, I was |
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| 67:47 | stop there anyway, so that's anybody any questions, So, okay, |
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| 67:53 | , thanks. We'll finish up. don't have a whole lot more to |
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| 67:56 | . So we'll finish it up on . Okay. So I'll fix this |
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| 68:04 | , whatever this is, hopefully. hmm. |
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