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00:27 Okay folks, um let's go ahead get started. Uh Okay, I

00:38 , put my usual announcement slide in . But so of course exam friday

00:48 this week. Uh if you haven't up, please do sign up on

00:55 website if you have questions or so I'm available. If not,

01:00 can't make regular alpha sours, just me, figure something out. So

01:06 , if you have any last minute , what have you, everything uh

01:12 and thursday is not on the so we're through with what's on the

01:17 . We finished that last thursday. uh so we start seven and we

01:22 much will finish seven today I Uh then on to chapter eight.

01:28 again this week's stuff, we're doing here is not on the exam.

01:32 um, so again the exam is . So make sure you look at

01:40 real exam review sheet. Okay. the blackboard, the blackboard, quit

01:48 quiz answer key will be available first actually at midnight tonight if you

01:54 if you want to be up late when it's open when it opens,

01:59 anyway, do look look through that if you have questions relating to

02:04 let me know. Um So like I said, I'm available,

02:10 you need, if you have any , what have you um if you

02:13 , that's fine, but just make specific question is not really broad

02:18 I have to write a book. , so um so again today's stuff

02:24 material, not an example. We're with unit one basically. Okay.

02:29 today's material is an extension of really growth because of course we went through

02:37 whole growth cycle stages of growth. batch growth. So we have a

02:43 of cells right over time. We have that typical growth response.

02:52 ? Um Black log log stationary And so we're actually what we're doing

02:56 focusing on this part here. What's on out here and basically forcing death

03:05 occur. Right, adding antiseptic or or radiation or what have you?

03:12 , so we're actually actively killing them trying to make that part of the

03:16 of course go as fast as Okay um and of course we're also

03:22 uh of course when you do a of treatment whether your radiation or chemicals

03:28 have you you even though you're targeting you of course are reducing numbers of

03:35 . Okay. Obviously. But you of interesting though of course is targeting

03:41 because that's that's what causes humans the problems. Right. So sorry about

03:48 . The so anyway so that's what focusing on today. And so there's

03:52 be some terms you're gonna you know gonna have a question here. Right

03:56 a second. Uh so just briefly actions into microbial agents, physical methods

04:05 control. So things like temperature pressure ways to control microbial growth that way

04:13 which you're familiar with? Of course ? Antiseptics. And then you know

04:19 those things work and why? Why would use some types of killing mechanisms

04:26 some you don't. Okay, so , so let's start off. So

04:30 you know that my favorite thing, of my things is the misuse of

04:36 word. Okay, that one. um so we had a question relating

04:44 before. I think back in Chapter . So let's try this one

04:48 Okay, so sterilization equals. Mhm. Remember what sterilization is?

05:13 not talking sterilization in the context of have Children. Okay, this is

05:20 this is the other the other Okay. Alright. I'm gonna set

05:40 countdown from five 32. Okay, of you are correct. Okay.

05:54 above sterilization is is to find out next slide. Is this their removal

06:05 destruction of all living microbes? You the area you sterilized or the material

06:11 ? It is zero. Nothing is there. Okay. Absolutely devoid of

06:18 spores, viruses. What happened? , nothing is in there.

06:24 Everything else here will reduce numbers. , obviously right, they will all

06:30 the levels of microbes. Okay, they don't bring it down to

06:35 OK. Um and there's different sterilization are typically will do that.

06:41 but not everything can be autoclave. . Um There are certain chemical agents

06:46 can do it. Um uh So look at some of those things.

06:51 , so uh this infection and sepsis and sanitation. Okay, so I

06:59 we're pretty much aware of this infection sepsis is right. So they both

07:04 about reducing pathogen numbers is just, the surface? You are using inanimate

07:12 , tabletop wall bench top. Or is it a skin?

07:19 Is a living tissue? So, so for that reason and disinfectants can

07:25 much more concentrated. Much have much harsher type chemicals. Okay, I

07:32 bleach. Okay, uh sepsis is to be have to be more gentle

07:36 the skin. Right? But you still be effective in certain reducing microbial

07:41 . Okay. Um uh the german more mechanical action when you wash your

07:47 . You're basically the german, Because you also have this going

07:51 right, That mechanical action also um in kind of reducing microbial levels.

07:58 , So a combination of those Okay, sanitation is more sanitary

08:05 Okay, so if you've ever worked a in a restaurant, right,

08:10 a server or you worked in the itself. Um then you're probably aware

08:16 these things um unless your restaurant was the restaurant report on friday and then

08:21 probably weren't. Okay, so, so what that is, is doing

08:28 in a way to minimize, you , in a restaurant setting, of

08:32 , minimizing foodborne contamination. Right? having a foodborne outbreaks to do bad

08:39 . Right? So wearing a hair wearing gloves, right? Keeping the

08:44 clean where you prepare food, the clean. The soda fountains can get

08:50 gunky and stuff with that sugary syrup in there. And so making sure

08:53 clean. Um These are all potential where you can introduce uh foodborne

09:00 Of course restaurant doesn't want that. . These are all hygienic practices.

09:04 so so there whereas um and sepsis are specifically targeting pathogens and sanitation is

09:13 more overall kind of a thing that you do and reduce all levels.

09:18 of course pathogen levels as well. So what numbers you have to get

09:25 to? That's kind of up to local health department. Okay. They

09:30 codes and things for for these the you have to do and and the

09:36 you should be at. These kind things. So that's your county or

09:42 health department. Um So any questions these terms. Alright, so obviously

09:51 of you, most of you going health care and acceptance and disinfection in

09:56 Germany will be what you'll be familiar sterilization. Well you have to sterilize

10:02 sterilize surgical instruments. So you use that um prepackaged things, You would

10:10 catheters and things like that will be sterilized and those things are typically I

10:15 radiation is how they sterilize those The manufacturers do that at least.

10:21 But anyway, so this is just example of, I'm sure we've all

10:25 this on your favorite cleaner. Whether Lysol disinfectant rather lifestyle. What have

10:31 ? Uh they all have a number the front kills 99.9 or 99.99.

10:36 have you? Right. They always some value like that. Uh They'll

10:41 list um a different types of viruses bacteria that are medically important.

10:49 Right all manufacturers do this. They'll a list and generally it's fairly consistent

10:55 they all test. Okay so here see avian flu virus, this is

10:59 somewhat dated. I'm sure now the all have covid on it I would

11:04 . Um So different types of viral . Salmonella of course. Foodborne

11:10 E coli klebsiella, respiratory asian and on. Right so there's the

11:16 Right? That's the chipotle E coli um responsible for various food foodborne

11:24 But anyway so typical for any kind disinfectant. Okay um and it has

11:30 other things on here. Uh This hundreds of services in your home.

11:34 that's kind of a marketing thing But anyway you can see this for

11:40 . So sanitize the soft surfaces. it as a as a disinfectant is

11:47 it's more of a disinfectant on hard . Right? Because you can really

11:51 soak it well and rub it in well the soft surface like maybe a

11:55 or something is what I'm guessing out that you could spray it lightly And

11:59 can have some a little bit of , reducing overall numbers effect.

12:04 But that's not really the application for thing, an application for these of

12:08 is countertop toilet bowls, things like . Right. Anyway, so uh

12:14 kind of let's go back to this here, 99.9% killed. Okay.

12:20 if we do that, this is to a parameter that's used commonly used

12:26 measure death. Death of microbes by method. Okay, here, we're

12:32 looking at disinfection. Ok, by , this Lysol stuff. Okay.

12:37 uh so in terms of numbers we're talking about Big numbers. Of

12:41 . So here's a million cells. ? And so if he killed

12:45 you know, basically getting rid of . That's a lot. Right.

12:52 it's not everything right, because um still gonna have viable cells left.

12:58 . 10,000 viable cells. So we've down 10 to 6 to 10 to

13:02 fourth. Alright, that's two Okay. And logs of death is

13:07 have a lot of manufacturers measure Right? Measure their whatever their product

13:13 , and it's an antimicrobial agent. how many logs of death do we

13:17 ? That's kind of the parameter they to test it. And so of

13:22 it's getting lots of logs in a time period, that's optimal.

13:27 and so if we go from that million to 99.9. Which is what

13:33 claims it can do. Well then are going down 2000 bible cells.

13:38 three logs of death if we started this number. Right? These are

13:43 done. Um We have something like pretty good. Like a template typically

13:48 a two by two template square template is of course open in the middle

13:53 they put on the surface and they will swab to see what's actually

13:59 right? And then take the same area and then spray it with their

14:03 or apply whatever the chemical is. it sit there, follow the instructions

14:08 then tested again to see how many cells are there. And then you

14:12 up with this is this is the of death were getting. So that's

14:16 uncommon to do it that way. there's a bazillion ways to do to

14:21 these things. This is probably one the more popular science fair things kids

14:27 like in junior high and whatnot or younger is um I have some chemicals

14:33 the house and how can they be . I saw one that was

14:37 They went to the spice rack and up different spices and use that as

14:42 as the agent to reduce from a numbers. And so so very

14:47 There's lots of visual things. You just do. You have to always

14:50 numbers. You can get a visual see if there's an effect going on

14:53 we'll see a couple of these Um So okay so terminology it's a

15:00 static bacteria seidel and bacterial link terms used to assess how an agent is

15:09 on is having its effect. And so um again we're trying to

15:17 a negative rate very fast killing right . Okay. So but bacterial static

15:25 from the other two in that it kill it inhibits growth. Okay So

15:33 this and so the graphic we're looking right here is two things right?

15:39 have two lines. One is a line. Okay. And that's basically

15:44 taking samples. We're gonna take samples we're going to measure the total number

15:49 living cells in the flask. And um at the arrow. Right

15:56 arrow, that's where you see the . That's when the treatment was given

16:03 the culture. Okay. And so see you know what's the effect after

16:08 . Okay. And so again we the solid line which is viable

16:13 How many living cells per per volume in that culture. Okay. Um

16:21 there's a way you can do All right. The dash line is

16:26 what you're seeing under the microscope to the samples over time. You look

16:31 the microscope looking at the cells Or the cell numbers visually. Cell

16:36 increasing over time or what are they ? Okay. So of course prior

16:41 adding anything to it. The culture happily growing along. Alright so both

16:45 are going up. Okay then you the agent and all of a sudden

16:49 flattens out. Okay, so the to bacterial static effect is that the

16:57 viable cells count is not dropping It's raining steady. Okay. So

17:05 basically means you're not actively killing cells kind of inhibiting the roof. You're

17:08 able to grow anymore. Okay, the the bacterial seidel, bacterial

17:16 both are killing agents. Right? have that same effect. Okay.

17:21 so we see that in both. , Because the solid line, the

17:26 of living cells in that liquid have down following addition here with the arrows

17:34 Okay, so solid line solid line are going down. So living cells

17:41 of those are going down. That's the key parameter. Okay. And

17:46 basically the cells are no longer They're dying as prolonged exposure to the

17:54 . So, I think that's easy to understand now. It may be

17:57 of the oddball here is the dash . Okay, so remember that's what's

18:03 on under the looking at with your eyeballs under the microscope. Right,

18:08 what are the cells doing over Okay, so we've got in bacterial

18:14 It looks very much like bacterial static that respect, in terms of what

18:19 seeing under the microscope. Right? flat. Okay. Whereas bacterial lyric

18:23 going down. Okay, So what's on there? Well, it just

18:29 the type of killing going on, ? You can looking under the

18:36 Right. So wrote a right And Right here is where the arrow

18:41 Right, So, we're taking Post edition. Right? So post

18:45 of Chemical Agent, we see there's number of cells are remaining the

18:50 Right? So, this could be . Whether it's bacterial static or bacterial

18:55 . All right, Because you can without blowing up the cell. That's

19:01 what it is. You don't have you can kill the cells without blowing

19:05 up. Right? So, they're still be intact and you'll see them

19:08 every time point following exposure. So, agents like these can get

19:14 the cell. They can interfere with synthesis that would cause the cell to

19:19 , but it wouldn't be something that necessarily cause it to lice.

19:23 But you can have agents that are logic that can maybe do something like

19:31 the membranes, Right. Agents that dissolve membranes that certainly will pull the

19:37 apart and cause it to lice. , so, that would be a

19:40 lyric agent. So, you can that after the addition you have cells

19:45 have fewer fewer and there's only Okay, so, that's classic bacterial

19:50 agent. Okay, um from the of you know, you're killing whether

19:58 bacterial seidel materialistic, you're killing So, what's the concern?

20:04 well, a bacteriological agent. If was something if it were an

20:12 Okay, Might that have an impact it were bacterial lyric versus bacterial

20:18 Had to think back to what's the that gram negatives have that? Grand

20:23 don't? Well the the begins with nope en en endo toxin. No

20:39 effect right? Almost got them. And the toxin effect is um that

20:47 be problematic. So for a gram infection. Okay and you had a

20:52 lyric agent antibiotic? Well then we that. I think I had that

20:57 during class that scenario where the patient was given that and so it was

21:03 gram negative right? The gram negative up. Right? And so in

21:06 toxins being released. Okay and so have to come in with this other

21:11 that would then bind up the endo . But anyway that's literally just came

21:16 me as I was looking at this . But you know we're talking about

21:20 disinfectant that you're putting on a bench doesn't matter. You know how you're

21:25 him? As long as it goes . Okay um Is any question about

21:34 ? Alright so let's see here. so that that is log arrhythmic.

21:41 so you're not gonna get a profile it's everybody dying at once.

21:48 That will rarely happen if ever Okay the population of microbes um especially if

21:59 different species you don't know what you're the surface for example there's gonna be

22:03 there you don't know won't know what is necessarily in terms of genius and

22:08 . Okay. So certainly in that they're gonna be even if it's just

22:13 population of something, right? There's to be very slight variations from cell

22:17 cell, Right? Um There could be affected to be affected by

22:22 What it was really crowded with it might have some more on the

22:27 that are more susceptible than those down the in the in the middle or

22:30 bottom. Right? So that two create variations and how they're affected with

22:36 agent. So the point is killing always at some kind of a

22:40 Okay, so accumulation of damage Okay. Get enough hits.

22:46 So the chemical comes in and attacks a little bit of the membrane attacks

22:51 the proteins. Then you get enough damage in the cell dies and that's

22:54 to happen from cell to cell is different. Okay. So it's gonna

22:59 at some rate it could be very . Okay. But the point is

23:03 that some right? Don't expect it be all at once. Okay.

23:09 so the so we talked in the example was about logs of death.

23:16 . So that's where the value fits . Right? So the value is

23:20 one log one log of death how um to get there. So we

23:27 it D value. So this data for a heating 200 degrees.

23:32 Okay. I don't know what the is but um but you're gonna get

23:37 profile like this right? It's always be if you're typically going to be

23:41 kind of downward negative slope, Because we're killing cells. So it's

23:47 . How fast are we doing And so what you just look for

23:50 two points That are one log Right? 10-4 then the fifth.

23:57 ? You just extrapolate because you know that's going to be uh the

24:02 Value right? Time to kill Right so approximately a minute.

24:08 It's you know depending on the treatment and what you're doing it will of

24:11 vary. But you're just looking for points that are that are along

24:15 Okay now this devalue It's kind of different manufacturers of these products typically can

24:24 can have different parameters. They don't not necessarily consistent. Some some used

24:30 the parameter of 12 logs of How long to get 12 logs of

24:34 ? Okay. Others have you one log it depends. Okay um

24:40 so the factors that influence. All . So it could be certainly the

24:45 of microbes present. Okay. You a lot versus a little right.

24:49 can affect um It could be the . If it's an endospore former then

24:55 might be you know they're gonna be more resistant if they're mostly in those

25:00 that can certainly be more resistant. that can be consideration. Uh I

25:05 probably part of the of the most or practice from a practical standpoint,

25:11 really organic load. I would say two and exposure time, those organic

25:16 refers to you'll often see. But you look back at that can of

25:20 and the instructions, it likely said clean your surface with just a general

25:28 cleaner. Okay to get because of service is really dirty. Then that

25:35 interfere with the agent getting to the themselves. That's what organic loading organic

25:40 is kind of a level of how is the surface, is it full

25:42 organic material? Just dirt and grime what have you. So then you

25:47 then your agent won't work that well then it's the other extraneous material that's

25:54 the chemical and not getting to the themselves. So it's always good,

25:59 the surface is very dirty just to it with something else. First,

26:03 come in with your disinfectant, exposure time dose. Um Yeah,

26:11 . So that can relate to the itself. Um can have different volatilities

26:19 think of ethanol or isopropyl alcohol, disinfectant and antiseptic. Uh to something

26:29 um something that's not so volatile alcohol go into from liquid to gas rather

26:37 . Right vaporized. And so if spray it well then its lasting power

26:42 not gonna be that long compared to like like maybe detergent type of disinfectant

26:49 will stick on the surface longer. so so that can be a

26:54 Um And how long how long are leaving it there with the concentration of

26:58 kind of things? Okay ph and can be a concern um You know

27:06 temperature, it's hot right then. volatile. It's not gonna kind of

27:11 quicker. Right? Ph may affect how the chemical works. So that

27:16 be a consideration. Uh So again factors play into this. Okay of

27:24 when you're testing these things you try keep everything obviously everything constant and minimize

27:29 effects. Um But it's a you from a practical standpoint, you know

27:33 things things things to think about. the okay so here's the question.

27:39 is just testing you testing on the . Value, right? This this

27:46 require a calculator I don't think. , fingers were enough. So uh

27:53 is added to a culture containing 10 the six soc F. You so

27:58 call a colony forming unit. So think about that cells per mill is

28:03 . Uh And the the value of disinfectant is two minutes. So how

28:08 viable cells are left after eight So there's the devalue definition. Okay

28:16 value definition. Okay. All right down 1098. Okay there we

29:20 Okay so we get consensus says d what we got. So here is

29:27 we start with 10 and six. by definition then one leg of death

29:32 two minutes. Alright so So we two minutes we got 10-5. Uh

29:40 minutes 10 in the fourth eight minutes and third I'm sorry six minutes into

29:44 third. Eight minutes 10 to the . So yeah it's d okay um

29:52 questions about that. Okay. Alright. Um Alright so here is

30:01 couple of methods. A couple of that are commonly used to evaluate these

30:07 of antimicrobial agents. So the first these was called the use dilution

30:14 Okay you do a variation of this lab. Um We use glass beads

30:22 of metal rings but the purpose is it's a surface for the bacteria to

30:27 onto. Okay so you basically put rings and a culture of bacteria and

30:32 move the liquid and they kind of to the rings. Okay And then

30:37 add those two disinfectant solution incubate and um transferred to bras without disinfectant and

30:49 check for growth. Okay. To did they grow do they not

30:53 And that gives you kind of indication maybe it's bacteria static or bacteria

30:59 Um strictly it's strictly a quant Just visual you're looking for growth plus

31:06 minus. Okay. Are you seeing effect? It's kind of like a

31:11 pass you have a chemical you have favorite chemical or something, you want

31:14 check it out and then you know results say, okay maybe maybe then

31:19 can get more quantitative on it, ? It's kind of think of it

31:22 a screening screening tool to see how it works initially. Okay. And

31:29 gonna look at a problem involving this in a second. Um The second

31:33 uh a also a qualitative uh method what we call this confusion. You're

31:44 familiar with this I guess. Um we have a plate. Okay.

31:51 so what you do is um you the one you're not going to

31:58 You take a culture. Okay. of course it's very common to take

32:04 it's grand positive and grand mega. they're gonna you'll always see you typically

32:09 uh somebody they may behave differently but very common to see differences between the

32:14 because of the nature of that cell . Right? The gram negative the

32:18 it is versus the gram positive. does lead to differences and how they

32:22 to different types of chemicals. And in any case, so you start

32:27 a clean plate, un inoculated Okay then you have a culture.

32:32 what you do is you you lay growth. So you take a cotton

32:38 basically and you like a mop on floor. You just go all over

32:41 plate like this, right? And that does is you're not going for

32:47 pure culture. I speak plate isolation of a thing where you get individual

32:52 , you don't want you want the that you want a thick mat of

32:56 . Okay, So you do that then then you lay down these

33:03 So these disks have been soaked in concentration of in this example of these

33:10 chemicals. Okay. And so then can get disk and soak them anything

33:15 want really to test. And uh then take the disks and you place

33:21 on. You have to kind of get the excess chemicals off. But

33:26 you do that, then you plop them on top of the plate.

33:30 ? And so then of course you , right? Because that that long

33:35 bacteria you lay down will then grow like a big mat. Okay?

33:41 then if there's if the chemicals inhibiting , then you're gonna see that in

33:48 form of these clear zones around the . So, these discs there on

33:52 plate, okay, chemical in them diffuse out okay, into the surrounding

34:02 . Okay. And so you now to putting the discs on the

34:08 right? You put a big swab growth over everything. Right? So

34:13 question is, if these are, the dots are the cells you've put

34:21 as part of your making your man growth, Right? So the question

34:29 , will these grow or not? , Well they grow and if they

34:39 grow then you're gonna see something like . Texas chloramine, Right? There's

34:43 all over the place. Right? chemicals doing nothing to it,

34:47 It's able to unaffected by it. . But something like chlorine.

34:53 they didn't go anywhere near the Right? Because you have this

34:58 What you measure the zone of right? Measured right across the

35:04 That's an area of no growth Right? Just because the chemical that's

35:08 in the auger inhibited, they couldn't anything, right? They couldn't grow

35:13 . So that's basically how you assess you know, how big is the

35:17 area? Right? That gives you of a qualitative measure of how

35:22 Right. So you can see this is this is not a super

35:27 plate. A good plate has lots uh, what we call confluence,

35:33 growth is kind of growth altogether. looks the same, Right? Very

35:38 . Right? And so you should a nice crisp circle like you see

35:44 and not just a raggedy edge. . But nevertheless, the chlorine is

35:50 on both. Really? Okay. , it's kind of ugly over

35:53 but you see a big area Um and of course difference between gram

35:58 , gram positive be? No Right? Did have an effect here

36:02 the grand positive. That's what I . You always see differences between these

36:06 for certain chemicals. Okay. In case, just gives you a qualitative

36:12 see, okay, quick visual, work does not you can always get

36:15 technical with it from this point forward it's a good first pass kind of

36:20 thing. Okay um any questions about ? Yeah so let's look at this

36:28 . Alright this relates to the use problem. So we've got a growing

36:34 of grandpas it. Okay it's growing to mid log, okay middle of

36:40 and we're gonna add disinfectant. Okay it go for four hours and then

36:46 put it into fresh medium. So a visual of what's going on.

36:50 mid log add this in fact didn't my grammar and disinfectant. Okay and

37:01 four hours. Okay and so that's we're calling our initial. Okay then

37:10 going to take a sample of that put it into fresh media without

37:16 Okay and that next to we're calling subculture. Okay so no disinfectant subculture

37:26 here. Okay um then we see data. Right? So we have

37:32 you always do this with different delusions . Okay um to see if you

37:37 the effect the same effect with lesser of chemical. Right? Save money

37:41 way. Alright so looking for So that's why you always see a

37:45 of delusions here. Okay so we our initial right here. No

37:51 No growth no growth growth subculture growth growth growth. Okay again it can

37:58 just a visual looking for cloudiness or the case of initial increased cloudiness or

38:06 can you can take a spectrum tata measurement. Right? O.

38:10 Um And so this is the results see. Okay so uh what the

38:16 concentration this is disinfectant is. So that you can guess you know it

38:20 be something above this result somewhere one these three. Okay so would this

38:26 bacteria static bacterial sidle Or it has effect now we can all agree it

38:35 no effect that 1-1 28. Okay disregard that. Just answer it based

38:43 what you do see the effect. this is a variation kind of the

39:00 we talked about before. Like I there's lots of ways to do

39:58 Okay let's count down from 432 Okay. Right So 51 59 say

40:14 static. Um So who answered bacteria . Okay so why basically.

40:34 So basically this the subculture is assessing the state of the cells. So

40:39 was an effect. All right because didn't see that there was um we

40:45 that there was no growth further growth adding. So basically what's going on

40:50 right here they're all right so uh our experiment So we add um disinfectant

40:59 and then of course there's no growth right in our initial tube here except

41:05 the 1 to 1 28. So can imagine something like this may have

41:10 . Right if we continue the right? If there's no effect at

41:15 then it will continue to this mid and you just continue on until you

41:18 , completed its growth phase. But uh but we are sorry.

41:24 did see an effect though with these . Okay. Something like that.

41:28 then this is where you would take cells and put them into the

41:35 That's the second part of the Okay. At that point. So

41:40 what you're assessing is what's happened to cells, Are they alive or their

41:45 ? That's all you're trying to figure live or dead. Okay. And

41:49 way to do that is to say let's take them away from the

41:54 Put them in just a fresh medium any disinfectant and see if there actually

41:59 viable. So of course we found that's the case. Okay. They

42:04 . So that's that's classic bacterial static . Okay um if these had been

42:14 negative, okay then you could argue they're they're killed they weren't able they

42:21 killed in that treatment. Right? were not able to do anything once

42:25 put them in fresh media and disinfected grow because they died. Right.

42:31 that would be an indication then bacterial effect. Could you just based on

42:38 alone and nothing else Could you if did give you this result.

42:44 If you didn't didn't indeed see this the subculture could you tell if it's

42:50 , cyanobacteria lyric, just based on nothing else other than this anybody?

43:01 . Right. Because um without doing else. Right? You could look

43:07 the microscope but typically you you may see any difference between material. So

43:12 just know that they died, But you may not necessarily know how

43:16 died, but you know, in cases it doesn't matter. You kill

43:21 . That's what you wanted to Okay, so um so again,

43:26 can this can be a period you know, like all these little

43:32 we mentioned, right, whether this or the youth solution, this can

43:36 be these are kind of think of as kind of preliminary screenings. You

43:41 always get more involved, more quantitative you know, for people that work

43:47 this stuff, who are always looking different types of antimicrobial agents, it

43:53 to just have kind of a quick to see what's working and what's

43:56 And then work further with those that promising kind of a thing.

44:02 any questions about that? Okay, , so, uh actions of antimicrobial

44:10 . Okay, so you might you probably figure this out for yourself.

44:13 just thought about it for a second you know, chemical comes in contact

44:18 a cell. Right. What's it attack? Well, obviously, initially

44:22 attack the membrane, it can attack components inside the cell or the main

44:30 , proteins, nucleic acids. So the different ways they can affect

44:35 components. So there are what we detergent detergents have a chemical structure very

44:43 to fossil lipids that make up a so that enables them to dissolve

44:49 Uh The of course that leads to you break up and cell contents leak

44:55 . Sell license is killed affecting Remember proteins very specific what we call

45:03 structure. The chain itself folds in particular way that folding is important for

45:11 function. Okay. And if we it, so to speak, then

45:16 going to lose function likely be lethal itself. Okay. And there's different

45:22 to do that. You can do with acids, You can do it

45:25 other types of chemicals that interfere with interactions? Okay. Others can basically

45:32 completely break it down. Right. types of harsh acids and things.

45:36 nucleic acids as well can be radiation can break nucleic acids. So

45:44 all these are susceptible. Um So you look at the first of these

45:49 physical agents. So temperature um the uh mechanism to reduce microbial numbers in

45:59 types of chemical components self with So one thing to remember is that

46:07 depends on what you're trying to reduce microbial numbers in. What's the nature

46:13 your your um are you dealing with liquid? Are you dealing with a

46:19 ? Are you dealing with plastic Are you dealing with a liquid that's

46:24 a glass? Are you dealing with food product? So all all those

46:29 going to influence, you know what do? And does the treatment itself

46:34 ? Right. You may have a pipette tip box box, Full pipette

46:40 are using lab. How do you sterilize that? Right. And so

46:45 different ways to do it. Some better than others, Right? Liquids

46:48 particular. Right? Liquids aren't good terms of disinfecting them with radiation?

46:57 ? Um UV light for example, no good. Right? But temperature

47:01 really good. You can boil You can autoclave. So the point

47:05 it depends on what you're trying to . Okay. Which method works best

47:08 you. Okay. And so temperature very common. So the autoclave.

47:14 . Is the of course for lab rely on that to sterilize media.

47:20 so when you have steam under that's the key here. And to

47:26 honest, it's it's the auto play used to kill. And those

47:29 Right? So very resistant. But they are susceptible to uh that moist

47:36 that comes from steam and steam heated pressurized and heated to these extreme

47:41 So 121 is a temperature that that can't withstand. Okay, even a

47:50 thermal file. Okay. But generally don't need to worry about hypothermia?

47:55 because the things you claim you're not not trying to sterilize something from a

48:00 that will contain a hyper thermal Right? So it makes no

48:04 So the stuff you sterilize or things live at moderate temperature so it certainly

48:09 kill them. Okay. And the has that kind of penetration into a

48:14 effect. Um Now you can if don't have an autoclave, a lot

48:20 this, you know, if your to do with what's available to you

48:23 maybe you don't have an autoclave, what can you do? Well,

48:25 can use an oven. Okay. so incineration, that's if you're a

48:31 and your you have your loop, ? And you put it in the

48:34 burner, right? You're waiting for to glow, right? You're incinerating

48:39 right? You're you're sterilizing it as . Okay. Um so incineration,

48:44 air, right? So basically an , right convection of okay, is

48:50 reach these uh conditions but of course not gonna be the same as an

48:55 autoclave. These are standard conditions. per square inch will give you 100

49:01 C seven grade uh the equivalent and have to I wouldn't know this.

49:10 , but don't worry about these other . Okay. But there is a

49:16 can put things in the oven for uh temperature for a longer time and

49:22 can reach sterilization. Okay, um patronization. Right. So we're all

49:29 with that in the context of dairy products. Right. And so

49:37 there's been a number of so long time long temp. High temp

49:44 time. The low temp long time been around for a long time.

49:48 pun intended. Um That was one the first pasteurization methods developed and it

49:53 used for milk. I think now more common is I want to say

49:58 more common is certainly going to be temp short time. But ultra high

50:04 is not gaining more popularity. Um called ultra pasteurization um That 1 30

50:13 for 1 to 2 seconds. Um has really been beneficial uh for countries

50:20 refrigeration can be spotty. Okay, underdeveloped countries that may not have consistent

50:28 all over the place uh milk that treated that way can last for six

50:35 . Unrefrigerated. Okay, so that's that also requires you put the liquid

50:43 a sterilized container. Right? So it's but it's proven effective. It's

50:49 good. And of course with all methods that's about um preserving the

50:57 So you don't want to that that back to you can't autoclave everything.

51:01 ? So especially beverages for human you don't want to put milk in

51:06 autoclave. For example. Trust me will not come out good.

51:09 But so you have to use these methods to preserve the flavor, the

51:14 , the look, texture maybe of product. Um but you know,

51:20 it safe. Right, reduce microbial . So from milk pasteurization. This

51:26 here cox E ela is among the heat resistant. They don't form in

51:33 pores but they are among the more resistant types. So that's often used

51:39 a test organism forest for a milk process. Um Now physical agents.

51:50 continuing with temperatures so cold temp. ? So cold temp does not

51:55 okay, not kill it inhibits Of course. We refrigerate our food

52:00 in the refrigerator obviously to next to know, prolong maintain it. It

52:07 eventually spoil at some point typically. But it does prolong that the refrigeration

52:13 for that slowing growth. We use the lab to preserve cultures,

52:17 We can grow a culture up. can add What you might call antifreeze

52:23 it, a little bit of a of drops of glycerol and they can

52:27 at -80 and be viable for Okay. Um the uh the uh

52:37 problem with this one problematic bacterium is guy listeria. We'll talk about that

52:43 the end of the semester in the . But this is what actually grows

52:47 refrigeration temperature. So um if you not one to necessarily pay attention to

52:55 dates like me, I'll eat food has to has to still smell.

53:02 . Alright. I do have my . But uh you've likely have ingested

53:08 find it like processed meats like hot and salami and deli meats you know

53:15 those kind of things. Uh and they can actively grow at for

53:23 , not fast, but they you you'll see logs of growth over a

53:27 or four week period. Um but also when it's a pathogen of course

53:33 it can uh most people that have immune systems aren't affected by it,

53:38 it can pregnant mothers need to be of it to not really eat these

53:44 of processed foods during the pregnancy because can affect the baby. Okay,

53:50 the unborn fetus. And so uh like I said, we'll talk about

53:54 later, but it is, it a no wonder the bluebell, there

53:58 a uh blue bell ice cream factory affected by a Listeria outbreak. A

54:04 fatalities occurred. This was probably eight ago, I think. And you

54:09 , google ice cream, right? refrigeration temperatures to make it. And

54:13 you're not careful, listeria can grow those conditions and it did and it

54:17 traced back to not cleaning their equipment . Um so filtration again, these

54:25 for liquids so that this term um label. Okay, this refers to

54:32 sensitive. They're sensitive to heat the them too much, basically fall

54:37 Okay, um so filtration is Um you hear the term filter

54:45 Okay, technically it's not sterilization because can get through, but honestly viruses

54:54 on a problem typically in this Um you can have air filtration for

55:00 . Have filters can remove microbes from air. Um The uh and there

55:06 certain things that just aren't amenable to ng or high heat. And we

55:11 um there's a media we have in lab that we can't use on it

55:16 all. So we have to filter Yuria broth. Um Others have to

55:21 can't be article you have to be boil gently so it just depends on

55:24 nature of the of the liquid. Other ones I'm not gonna go into

55:29 no high pressure has been used desiccation out, drying it freeze drying osmotic

55:37 . Uh So there are osmotic pressure that's probably been used for centuries as

55:44 preservation method that's packing foods and salt been used for centuries packing meat,

55:52 and salt salt itself of course creates hyper tonic environment that's gonna be

55:59 Um Of course we're moving water Because life needs water and move

56:03 That can certainly be inhibitory. So radiation. So this can be effective

56:16 Remember with radiation it's about energy. ? So high energy radiation is gonna

56:23 more effective at killing than lower Okay and so the two that we're

56:29 with here is UV light what we non ionizing radiation. And so that

56:36 so you b that's good for surface shining UV lamp on an area can

56:43 the surface. Um It works by bases in D. N.

56:49 Creating mutations that can be ultimately be to the microbe but it's easily blocked

56:56 a shirt can block it. Often we used to in lab have an

57:02 where we bombarded the culture with UV . If you forgot to take the

57:06 without the Petri dish, that would the UV light from hitting the

57:09 So it doesn't take a lot to UV light. Right? So you

57:13 want to use it certainly for liquids for surface it can be useful but

57:19 high energy are is UV light and rays. So look at the wavelength

57:26 100 nanometers for you be like for radiation gets um point less than 1000.0.1

57:34 . It's very small. Super high . And uh radiation is used for

57:41 these on meat, frozen, frozen . It's been your created use it

57:46 that. Um And other uses on like plastics have been used to being

57:54 with radiation. So it has its and it is it can be a

57:59 method for sure. Right? So breaks nucleic acids. You're doing

58:04 You're certainly going to kill the Okay. Um the and it is

58:10 more penetrating. So it's not easily by a shirt or plastic lid.

58:16 ? It can penetrate and you know because you have to wear a lead

58:21 if you're getting a Dell X Okay. Or other type of X

58:26 . Um Now chemicals so things like sanitizers um final IX. Often these

58:41 sanitizers, the phone Alex tend to types of things that um uh stick

58:50 to the surfaces. They are not easily vaporized. Okay. Have a

58:54 better staying power and they have a of effects. They basically break down

58:59 . Break down the clinic. Gasses halogen are things like iodine base of

59:05 doctor's office. You may have seen of this, this yellowish liquid beta

59:11 iodine is iodine based bleach is in category. So these basically again,

59:17 lot of these things overlap in terms what they affect can affect protein

59:22 Uh Bleach is an oxidizing agent. down chemicals in the cell alcohols,

59:29 groups that we use this in lab disinfectant. Um these work better.

59:37 add water to it. Right? we use 70% ethanol, okay,

59:45 works better than like say 95 or 100%. Right, the extra water

59:52 there helps it penetrate the membrane is and let's not get in there and

59:57 . Okay, a little bit, little bit of water helps actually.

60:02 , and so these of course dissolve dissolve membranes. Okay, heavy metals

60:10 you often see this in um uh you have an aquarium, you often

60:17 these agents to prevent algae growth in aquarium and they often contain one of

60:23 metals to prevent growth of allergy for example. Um But the thing with

60:29 metals is a little bit goes a way. You can overdo it rather

60:34 with with these because they'll be quite . So um usage and concentration is

60:41 important with these things. Too much be certainly be a hazard um surface

60:48 . This this is what I call the detergent type molecules. And they

60:53 very similar to you can see how very very similar to a phosphor

60:58 Okay. And so because of that very good at penetrating membrane and disrupting

61:05 dissolving it in time. Um Chemical . So organic gasses. So this

61:12 are things you find in very commonly different foods like bread. Um And

61:20 kind of based food cereals, things that. Um Also in cosmetics contain

61:27 . So and they kind of fit category here where you see sabic acid

61:34 , if you look on a cereal label or other food level, you

61:38 citrate or citric acid is in there have the same effect. And the

61:42 they have is this so let me get this out of the way.

61:48 So here for example is this is Zoellick acid I think. Um So

61:53 happens is in solution solution. It forms an acid all right.

62:03 so but unlike hydrochloric acid. hcl of course goes to and this

62:14 what we call a strong acid. other words, it all goes to

62:21 flooring and protons, right? There's of this left in solution. That's

62:26 nature of a strong acid, All goes to. That's why it's

62:31 . It all goes to these protons chloride ions. Okay, this

62:36 these things, organic acids are weak . Okay, so what that means

62:44 it doesn't all go to product. , you'll see this, this and

62:54 in solution. Okay, so, what, so what is this

62:59 Okay. The fact that you still some of that reactant here.

63:05 in that's neutral there's no charge associated it relatively small so we can fit

63:12 a membrane and get inside the Okay, So inside the cell this

63:18 and then this reaction occurs. And it acidified eyes the inside of the

63:24 as it comes in and that's inhibitory . That's the preservative action of the

63:31 , of the of the chemicals in food or in the cosmetic. It

63:35 that action in the cell and uh inhibits growth. That's that's why it

63:41 as a preservative. Okay. But the fact that it is a weak

63:46 . Only a little bit of this this way. And so this is

63:50 neutral compound that can squeeze in again the cell. Okay, very common

63:56 and lots of foods. You'd be . Um gas is sterling's.

64:01 so these are typically used for really different types of products. Alright,

64:10 really lab lots of lab products. things like your pipette tip boxes,

64:16 other plastics, Petri dishes not Uh and so gas a gas can

64:23 fairly well. You wouldn't want to a gas to sterilize liquid because that's

64:26 gonna mix very well. But for things like this it can it can

64:32 and it has this kind of effect it can be sterilizing. So it

64:37 this kind of structure here in the of acid or base, it kind

64:42 unfolds and then becomes very reactive. reacts with proteins and nucleic acids in

64:49 cell basically destroying their functions. And but it's kind of a chain reaction

64:54 very quickly and quickly forms these reactive products that basically destroy the components in

65:00 cell rather quickly. Okay, so , for heat sensitive material.

65:06 plastics very common. So um the . So you might think,

65:16 we're certainly aware of antibiotic resistance and talk about that later in the

65:22 but you might go, okay, about disinfectants? Antiseptics? Can microbes

65:27 resistant to those? Yes and Generally, no. Okay. Because

65:35 the varied targets that these chemicals So think about, you know,

65:40 chemical chemical like I saw right, contact the cell can likely have an

65:46 on the membrane and get inside and various proteins and nucleic acids and

65:50 So in order for a cell to resistant, it's gonna have to have

65:57 way to counter act each and every ? Okay. That would mean equate

66:03 having lots of different mutations to counteract effect. That effect this and this

66:09 this. The probability of that is unlikely. Okay. Um that's why

66:18 antibiotics, antibiotics have only like a target. Right? So comparatively

66:24 it's a little easier to become resistant you're only having to change one thing

66:29 become resistant whereas with a disinfectant antiseptic things have to change. Okay.

66:35 it's not likely unless right. Unless don't use the chemical at the proper

66:42 if you want to save money or your reasoning is and you severely reduce

66:49 concentration you use then then the effect this is less. The number of

66:56 is less. Okay. And then do have the likelihood of becoming resistant

67:02 the disinfectant or antiseptic. Okay. generally no. Okay. Or not

67:09 likely. Okay. Um Now in of uh as a function of microbe

67:18 . Okay. In resistance. So you see that Priam's was in

67:23 sport is not surprising are very resistant the effects because of that thick

67:29 Right. But prion surprising are resistant well. But then you go down

67:34 to you know, gram negative versus positive. Okay. You see the

67:40 negative more resistant that that outer membrane providing a protective covering if you will

67:48 terms of um action against disinfectants and . So, you know, it

67:54 . Okay. What uh what Certainly in a healthcare setting, what

68:01 um makes it more difficult to become ? Is the fact that when you

68:07 hospitals obviously are clean. Yeah. cleaning rotation of using different disinfectants on

68:15 rotating basis. So maybe for one they'll use an alcohol. Then the

68:22 time they clean in two weeks or , they use a detergent type.

68:27 they switch to a different you're constantly different types of disinfectants. That too

68:32 increases the probability that you're not gonna resistant. So it's uh so but

68:39 with antibiotics which we'll talk about later they they do and obviously we're aware

68:45 resistant types. Okay. Um Many . Okay. All right, okay

68:54 , that's all. Any questions writing test or that material let me

69:19 Thank you. What do you mean you say etcetera? Where?

69:35 who discovered them? They would hook etcetera. Those names?

69:45 Right. Who discovered them? Where they come? Who what?

69:52 ? When where kind of thing? .

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