awesome you guys okay so let's go ahead with chapter 11 here we're talking about physical and chemical control of microbes how do we kill microbes and keep them at Bay there are historical ways that we have treated uh mainly food products you know to control microorganisms you know we salt Foods smoke foods pickle Foods dry Foods you know so our foods are preserved right these are ancient ways of preserving food before Refrigeration was you know invented also you could expose things to sunlight we all know that UV damages bacterium and they also dry out so the combination of drying out and UV kills the microbes burning clothing burning corpses you know the people burn their dead this helped to prevent the spread of disease storing water in Copper and silver container did you know that heavy metals are toxic to microorganisms so by keeping your water in these metal containers you actually keep it relatively germ free by the pro tip for those of you that have pets too if you put your if you if you have a metal Bowl that's better than a plastic bowl for your pets too for their water because it again that those heavy metals are antimicrobial in nature now these are terms we should know about microbial control these terms may seem similar but they mean distinct different things so for instance sterilization when I say sterilization that is a whole different that is a whole different Beast than saying disinfection right those are two completely different terms but mostly people use them interchangeably right people don't know the difference between disinfect and sterilize but if you're going into Healthcare you should definitely know the difference between disinfect and sterilize sterilize is the gold standard okay it's the gold standard sterilized means you've killed anything on that object um that could possibly cause disease right so any organism any endospore any fungal spore any any bacteria protozoa whatever sterilization is you know difficult to achieve you know you need something like literally fire you know this is why we flame our Loop right that sterilizes the loop so after you Flame the loop that's a sterile Loop um an autoclave we're going to talk about what an autoclave is but an autoclave is another way to sterilize uh something you know but that's the gold standard under sterilized the next level down is disinfect right disinfect is a less stringent requirement when you disinfect something you could kill I don't know 99.99% of all possible pathogens but you know you're never sure it's not 100% right and usually it's the endospores that you missed right you ever seen a Lysol bottle does it say kills 100% of everything on the Lysol bottle does it say 99.99% right so Lysol is an example of a disinfectant right it might kill 99.99% of germs but guess what it probably didn't kill any endospores or many endospores right does that make sense antisepsis also known as degermation antisepsis is when you basically you add a some kind of disinfect but on a living tissue on a living tissue so have you ever heard of antiseptic mouthwash right why do you think they call it antiseptic mouthwash and not disinfectant mouthwash well because it's going on your living tissue it's going in your mouth your mouth is a living uh tissue right inside of there living tissues you got your tongue you got your cheeks if you're applying a chemical agent to a tissue a living tissue it's an antiseptic okay sometimes you could use something as a disinfectant or an antiseptic so for example you know people use hydrogen peroxide you're not supposed to because it causes a lot of damage right they they say don't use hydrogen peroxide but they used to use hydrogen peroxide on the skin right and so if you're using hydrogen peroxide on your skin you're using it as a antiseptic right but if you're using hydrogen peroxide on a object then you're using it as a disinfectant does that make sense you see the difference there and then the last the lowest stringency the lowest requirement is decontamination also known as sanitation this is where you you know you can use mechanical removal of most microbes you know you wipe something off or you know you wash dishes with some regular soap you know it's not very stringent right it's just you got most of those microbes off of there right so when you do your dishes you're sanitizing you're decontaminating you're not you know that's the lowest Health standard okay and again why why do we need microbial control well because we've we learned early this semester that microbes are ubiquitous bacteria is ubiquitous that means everywhere there are fungal spores everywhere bacterial spores are pretty abundant you know there's protozoa I mean there's there's microbes these microbes might make you sick right especially in the health care setting where certain instruments might be used to probe you or to cut you open right so we need to understand how to sterilize those types of you know instruments right and here you can see different organisms have different resistances okay not all microorganisms are EAS as easily killed as let's say enveloped viruses you see enveloped viruses are the easiest to destroy they're the least resistant to antimicrobial techniques right and methods but on the other end of the scale prons look at them they are really hard to treat really hard to destroy I just saw um a story about a lab that was working with prons did you guys hear about this story there's a lab that was working with prons and two of the former lab members came down with basically Crutchfield jack up disease they came down with the prion disorder um years later so and it's still a mystery like how they caught it from the lab because they didn't eat anything one of them may have caught it through a a break in the glove like they broke the glove and they cut right through the blood the other one they still don't know how he caught it so you know we still don't fully understand prons and they're the hardest wants to treat because it's just a denatured protein what are you going to do denature a denatured protein even more it's really hard to treat right and also endospores right endospores are really hard to treat you see that endospores very difficult this why we need fire this is why we need you know steam we need that that uh autoclave and again it's not sterile unless you've destroyed all of these right every single one then it's sterile okay endospores are a huge problem remember endospores destruction of endospores is the goal of sterilization okay again does boiling water stabilize you guys no no no not at all no no the boiling water destroys many organisms but it's more disinfecting it's not sterilizing endospores laugh in the face of boiling um so you know you can boil all you want the endospores are still viable and so you cannot sterilize with boiling here we can see different types of methods different methods of of disinfecting or sterilizing and how long it takes to kill the vegetative cells and the endospores right so here you can see with moist heat this is an autoclave when they say moist heat they mean Steam and this is an autoclave an autoclave reaches temperatures of 120° C okay vegetative cells die at around 80° C endospores they are are at least one and a half times more resistant than the vegetative cell so it takes a lot longer to kill uh endospores than it does vegetative cells same thing with x-rays x-rays it takes a lot of it takes a lot more grays of X-ray radiation to kill the the endospores than it does vegetative cells it takes a lot more sterilizing gas to kill endospores than regular vegetative cells it takes a lot more glut alahh which is a steril it's a sterilant cross- leaking agent it takes it 3 hours to kill endospores as compared to 10 minutes with vegetative cells so if you notice all of these it takes a lot more fire power right to kill the endospores than it does to kill the veget of cells right okay by the way um did you guys know when you boil water what temperature does it boil at water boiling water 100° C right 100° C but now if you crank it up so the water's boiling and then you crank up your stove even more so now it's boiling even more in fact it's on maximum what's the temperature of the water 100 it's still 100° C you you can't boil water higher than 100° C it doesn't matter if it's boiling only a little it doesn't matter if it's boiling like crazy it's still 100° C you didn't go above it okay the only way to increase the temperature of water past 100° C is to do what pressure like a pressure cooker you know if you prevent the steam you know the uh Steam from escaping and you pressurize it so the steam can't escape then the temperature of the steam can reach 120 right that's what that's what a autoclave is an autoclave is basically a steamer it's a pressure cooker you know you ever cook with a pressure cooker it it fixes your food in no time like you know your food's ready in low time because in a pressure cooker the temperature went above 100° so you cooked your food faster right an autoclave is like a glorified pressure cooker an autoclave is basically this pressure cooker that gets the moist heat up to 120 and increases the pressure and the combination of the pressure and the steam kills all endospores everything right so it sterilizes again sterilization is a process that that destroys or removes all viable microorganisms including viruses including endospores everything any material that has been subjected to this process is said to be sterile so when that when you pass someone a sterile scapel that means it's been autoclaved right now if I leave that sterile scapel at out on a table for a while is it sterile anymore no the second even one particle of dust lands on that scapel that scapel is no longer sterile a't that interesting so the second you touch that scapel accidentally to the table or you touch here it's not sterile right it's only sterile if it's been autoclaved and it's inside that autoclaved packaging right when you bring it out and leave it out no longer sterile if you accidentally touch something for a second no longer sterile right so um what products need to be sterilized does your food need to be sterilized no that's not necessary right but there are certain things that we need to sterilize right so surgical equipments right so if we're going to do a surgery on you all of those tools need to be sterilized sterile syringes right syringes need to be sterile you take the needle out of its sterile packaging you need to use it and then dispose of it immediately right and then some packaged foods but these are usually for you know MREs or some kind of long longterm shelf life stuff and for canned goods too you don't want your canned goods to have endospores cuz then you could get botulism remember that what about disinfection remember disinfection is the next level down from sterilized it's not as stringent but it's sufficient for most cases right in your kitchen do you do you need to sterilize your countertops not necessarily you can disinfect your countertops just spray them down with Lysol that's plenty sufficient for something like that okay so you're killing vegetative pathogens but not necessarily many endospores when you do uh this type of disinfection okay disinfectants are used on anatom objects right like a table whereas antiseptics remember antiseptics are used on living tissue antiseptics are used on living tissue okay we learned about a setic technique right aseptic technique the sterile method of transferring microbes or dealing with microbes in a in a way that prevents contamination and prevents breaking containment some of these chemicals we work with these antimicrobial chemicals are called cidal agents like bacteriocidal or sporo cdal you know what what does that suffix mean it means kills right kills so if something is bacteriocidal it's kills bacteria right if something is sporo cdal that means it kills spores fungicides kill fungi right anytime you see side that means kills now virus side is kind of a misnomer because viruses aren't alive so you can't kill them however they're still called Verdes but just understand you're not killing the virus you're inactivating the virus right because there's the active ver thyroid the active virus then the inactive virus the denatured virus so virus sdes they don't kill the virus they inactivate the virus and a germicide or a microbios side these are just general terms for you know like Lysol would be a germicide it just kills a bunch of different microorganisms and by the way are fungal spores as hard to kill as bacterial spores no right fungal spores aren't super tough they're not super tough like endospores are fungal spores are designed to disseminate easily and spread and just for reproduction right they're more reproduction oriented and less defense oriented so it's way easier to kill a fungal spore than it is an endospore a bacterial endospore those are the really tough little buggers to kill so what are static agents the suffix static static means not moving right so static agents prevent the organism from dividing it prevents the organism from growing right so if I if I treat you with a fungi static chemical it's not necessarily going to kill the fungi it will keep the fungi from growing right or a microbiostatic agent will keep the microb from growing or bacteriostatic agent keeps bacteria from growing sometimes all you need is to keep the infection from growing and then allow your immune system to do the rest right your immune system kills the infection but at least you prevent it from spreading right that's sometimes sufficient you don't always have to kill the microorganism many agents are static and what about decontam ination or sanitation remember this is the lowest standard this is the lowest uh stringency okay this is what they do in General Health practice like at you know restaurants and stuff they're required to wash the dishes or scrape the food off and then rinse it off you know this is this is just the lowest standard to meet Health requirements if they say reduces contamination to a safe level right to a safe level okay and so when you go on to you know when you go on to health care you're going to have to ask yourself you know does this instrument need to be sterilized or can I get away with it being disinfected you know how do I treat this or can I just sanitize it you know and it all depends like is it going to is it going to you know be in proximity to the patient or is it going to be used in inside of the patient's organs you know you have to like assess the level of control you need for each you know scenario right so like does the item need sterilization like if I'm giving someone a spoon so they can eat their Jello do does it need sterilization no obviously not it just needs Sanitation right but if I'm handing the the doctor a SCA so they can do an operation then it needs to be sterilized so you know this this slide is basically asking you what you know you know there's there's no there's no cookie cutter way to go about things you have to assess each scenario differently you know here I need to sterilize here I don't here I need to disinfect here I can sanitize again it all has has to do with How likely is it to cause disease in the patient right so look a critical medical device a critical medical device is expected to come into contact with sterile tissues remember inside of your body inside of your tissues inside of your fluids right so let's say you're putting a pacemaker in a patient right you want to make sure that pacemaker is sterile right yeah for sure right so that that needs to be sterile right you can't just put you know this random pacemaker into a person without St without making sure ensuring that it's sterile but what about a semi critical medical device it could come into contact with mucosal membrane so you know you have like a feeding tube you know or something like that that would be a semi critical medical device you know it's not inside of your tissues but it's among your membranes you know your it goes through your nose and everything what about non-critical medical devices those that do not touch the patient and are only expected to touch the skin so you know like those little pads for monitoring your EKG or whatever you know that would be a non-critical so you have to ask yourself you know to what extent is this device going to you know interact with my patient you know obviously it needs to be sterile if it's a critical medical device right and then the goal of all of this is to kill microorganisms right the goal of control is to kill the microorganisms but it's hard to detect whether you've killed the microorganisms because you can't even see them right I don't know if there's living bacteria on this table or dead bacteria on this table I don't know I can't see it you know I'd have to do some studies so it's hard to see if you know organisms are dead or alive okay why because even if I spray it with Lysol even if I spray the table with Lysol I still can't tell if anything's Dead or Alive because it doesn't alter the overt appearance of the microbial cells even if I did take those cells and put them under the microscope even if they were dead they would still look like cells if I stained them so it it's kind of it's a pain because you just have to rely on your techniques right you know you know um you have to rely on doing things correctly so for instance do you guys know I say spray down your table and wait how long five minutes right why do I say that well because they've done studies to show that if you leave that disinfectant on for at least five minutes then you've achieved good disinfection then you've killed 99.99% of everything right but if you just spray the table and wipe it immediately they've shown that you may kill far less than 99.99% right but that's why you just have to go off of what it says so when you go into the clinic and they say you know did you know a common sterilant in the clinic is this uh sporicidal liquid glut alide right glut alide they might say you know what this patient in room this patient had uh SEI remember this the endospore forming SEI and we're concerned that it's like contaminated the room right so why don't you spray it down with glut alahh and wait three hours right wait three hours would it make sense to just be like ah whatever I'll just wait 10 minutes you know yeah if you wait 10 minutes you've killed the vegetative cells but you've really not killed the problem then you know you wipe it up you're like whatever it's it should work you know the next patient comes in those spores are still intact they touch something they touch their food they touch their mouth next thing you know the next patient has C right because you didn't wait the full three hours so again you can't see on a table if these spores are living dead what you know so you have to go off of what it says on the label of the disinfectant you're using there's a reason why Lysol says you know wait a minute and then wipe it there's a reason why you know in your lab the disinfect the disinfectant says wait five minutes and then wipe it there's a reason why glitter alahh needs to be left on for 3 hours and then wipe it because they've done all these lab experiments they've done all these tests and that's what the their data showed right so try to adhere to those things now let's talk about factors that affect the death rate of microorganisms the number of microorganisms is important if you're trying to destroy billions or trillions of microorganisms obviously that's going to be harder than a few dozen the nature of the microbes in the population are they endospore formers are they easily disrupted are they easy to kill or are they more hearty do they have let's say uh molic acids are they acid fast those are typically harder to to kill off the type of microbial growth are the these microbes in a bofilm for instance that's going to be tougher to kill off temperature and pH of the environment can play a role the concentration of the agent so the chemical you're using are you using a strong concentration a high concentration of let's say bleach or are you using diluted bleach that could play a role in the death rate the mode of action of the agent what part of the cell does that agent attack that could take uh uh account as well the presence of solvents interfering organic matter and Inhibitors all of these things can affect the death rate so you have to take them into account and by the way speaking of the death rate these microorganisms when they die they die off in a logarithmic fashion this means that when you add a uh disinfectant when you add a antimicrobial chemical not all the microbes die instantaneously they die off in a logarithmic fashion they die off over time in a logarithmic fashion so this is something to be aware of they don't instantaneously die instead they die off logarithmically now how do antimicrobial agents work typically these agents Target either the cell wall or the cell's cytoplasmic membrane or either DNA or RNA synthesis by targeting uh RNA polymerase or DNA polymerase or they can even Target protein synthesis for instance they could Target the ribosomes which synthesize the proteins these are typical targets of antimicrobial agents and this can lead to all kinds of effects if we're talking about agents on the cell wall well if you block cell wall synthesis you could have a negative or dilus effect you could cause the cell to perhaps lice um this is how penicillin Works penicillin prevents the maintenance of the cell wall of bacterium and this results in osmotic stress water forces its way through osmosis into the cell causing the cell to lice some of these agents can even digest the cell wall of poog glycan break down the surface of the cell wall any of these could be fatal for the cell and how about the cell membrane I told you about the membrane of the cell uh we talked about surfa actants which can insert themselves into the membrane of the cell and this can cause the cell to lice or break open these cause the cell to become more permeable they can lose their selective permeability this means that vital inerts could flow out of the cell you know and the cell lices the this is a damaged cell and remember again surfactants are an example of chemicals that disrupt the cell membrane opening up leaky spots that really damage uh the cell allow damaging chemicals to seep into the cell and important ions and important cellular uh components to leak out here you can see how the surfactants work again the surfactants enter into the cell membrane disrupting the cell membrane opening up these leaky spots and what about agents that affect protein or nucleic acid synthesis these are also typical targets of antimicrobial chemicals substances that inhibit ribosomes they they're going to inhibit protein synthesis and then the cell can't divide these are typically static agents static agents prevent the cell from dividing also so nucleic acid blockers these are agents that can impede transcription of DNA into mRNA or even DNA replication these typically these typically act on DNA polymerase the enzyme that copies DNA or RNA polymerase the enzyme that uh transcribes DNA into RNA either way you're not going to allow the cell to grow and remember some uh of these microbial agents they can attack the proteins themselves remember that a protein needs to be folded in order to function and so when a cell produces proteins that protein needs to fold into a native state and that that protein then functions normally but several of these chemicals including like let's say alcohol for instance can denature proteins causing the protein to misfold and become nonfunctional and that can lead to Cellular death again agents that alter protein function typically they cause proteins to misfold they cause these proteins to denature once a protein is is denatured that means the protein can no longer function and the cell usually dies moist heat can cause this denaturation of proteins also strong organic solvents such as alcohols or acids or phenolics these can have the same effect here you can see the native state or the or the properly folded protein which would be functional and then the the denatured uh protein the protein that has become unfolded once a protein becomes denatured or unfolded it can no longer function and it can no longer renature and pick up function again next let's discuss methods of physical control moving on from chemical control to physical control this would include methods like radiation filtration ultrasonic waves as well as cold now starting with heat as an agent specifically moist heat moist heat can refer to water uh it can refer to steam hot water boiling water and steam the temperature ranges that would destroy a typical uh mopile or a type of bacteria that would usually cause human disease is about between 60° C to 135° C with water or steam the temperature of steam can be regulated by adjusting its pressure in a closed container uh bo water boils at 100° C and no matter how much you turn up the temperature the water will not rise above 100°c it will simply become steam faster and faster so if you want the temperature of water to increase if you want moist heat to be warmer than 100° C what you need to do is apply pressure like a pressure cooker and essentially that's what an autoclave is an autoclave is a pressurized chamber that's allows water to form Steam and that steam can reach 120 to 135° C and that's sufficient moist heat to sterilize ize now the reason you sterilize is because that hot temperature that really warm temperature uh causes coagulation and denaturation of proteins now what about dry heat dry heat is when either air with low moisture content or a flame or electric heating coil is applied to the sample this is a dry heat so when when we flame our Loops in the lab or we use incinerators or Flames to incinerate our Loops this is an example of dry heat it dehydrates the cell it oxidizes cellular components and it burns them oh there's Wicket behind me I think you guys saw him walking behind me what that that's Gizmo Gizmo come say hi Gizmo's back back there adventuring so again let's compare moist heat to dry heat moist heat remember to reach temperatures of 121 to 134° with moist heat with water you you're talking about a pressurized container you're talking about an autoclave and this can sterilize within you know just minutes at 12 121° in an autoclave sterilization uh is achieved in 15 minutes um and this means that even endospores are destroyed in 15 minutes and the warmer you go the less you can uh the the less time is required for sterilization with dry heat however at the same exact temperatures dry heat takes longer okay dry heat takes longer to sterilize but it is possible to sterilize with the same heat the main point of sterilization is to kill even endospores endospores have the greatest resistance to disinfection and to heating because of their you know dried out seed like nature they are very tough structures and the point of sterilization is to destroy endospores vegetative cells are far easier to destroy and to uh kill off vegetative cells can vary in their sensitivity to heat but it's typically nowhere near as robust as an endospore now both temperature and length of exposure should be considered for adequate sterilization we need to have the right temperature for the right amount of time to kill off the microbes at higher temperatures this would allow shorter exposure times lower temperatures conversely require longer exposure times each organism has two distinct times the thermal death time and the thermal death point I want to explain the difference here the thermal death time of an organism is the shortest length of time required to kill all test microbes at a specified temperature while the thermal death point is the lowest temperature required to kill all microbes in in a sample in 10 minutes so let me explain the difference um for instance let me let's say we're trying to kill off eoli and I put eoli at I don't know 55° C and then we notice that it takes uh an hour to kill all of the eoli that would mean that at 55 deges C the thermal death time for eoli is 1 hour does that make sense but we have not reached the thermal death point the thermal death point is the temperature where all of the ecoli would die in 10 minutes so let's say we did the experiment again at 100° C and then we saw that all of the eoli were dead before 10 minutes we could call that the thermal death Point does that make sense so the thermal death time is how long does it take for all of the microbes to die at a particular temperature and the thermal death point is the lowest temperature that kills all of them in 10 minutes and remember we said that an example of moist heat is boiling water I want you to know that boiling water is good for disinfection but not for sterilization boiling water is not a sufficient means for sterilization uh this is the problem 100° C will not kill all resistant cells nor will it kill most endospores so why do we boil water it is useful in the home for disinfecting water killing most pathogens but remember you are not sterilizing anything by boiling it you are simply reducing the bacterial load but not sterilizing pasteurization is an even lower standard with pasteurization we want to heat a particular food or beverage to retain its flavor this means that we cannot heat the food past a particular temperature we all know what happens when you boil milk for instance right if you boil milk then it tastes off it does not taste right so with pasteurization which was developed by Louis Pastor here we go to you know either 71.6 Dees C for 15 seconds in a b in a Flash method or in a batch method to 63 to 66° for 30 minutes you see this is well below boiling but it's sufficient temperature for denaturing proteins uh of of bacterium so it can inactivate most viruses and destroy the vegetative stages of 97 to 99% of bacteria and fungi however it does not kill endospore and it does not kill heat resistant microbes but remember the main point here is not to sterilize the beverage or the food here the main point is to retain the flavor and reduce the microbial loads and so it's uh nice uh for milk you know when we uh pass Iz milk wine beer and other beverages we reduce the microbial load and retain Flavor now if you do want to sterilize we want to use steam Under Pressure which can achieve St sterilization and we do this with a pressure cooker called an autoclave with an autoclave this is a device that can subject pure steam to pressures greater than one atmosphere we're going to 15 psi uh of pressure for to for 20 minutes and this achieves a temperature of around 121° C the steam is very warm and this is sufficient to sterilize glasswar cloth medical instruments liquids Etc anything that's heat resistant um we can put in an autoclave for sterilization and yes this means that endospores are neutralized here is an image of the inner workings of a standing autoclave essentially it's a pressure cooker you place your samples in this chamber there's steam steam at high pressure reaching a temperature of 121° C for 15 psi for 20 minutes 15 to 20 minutes that's sufficient to sterilize and so in the clinic uh in the dentist's office even at uh even at tattoo artists uh tattoo parlor you will find autoclaves where instruments are sterilized now remember for dry heat methods incineration here we're using uh for instance uh a flame we can use a bunson burner it a bunson burner reaches 1,870 de C at its hottest point and this is more than enough to incinerate to destroy all living organisms or all viable cells including endospores from your instruments including your inoculation Loop your inoculation needle Etc this is common in the lab you can also use a hot air oven a low mo low moisture uh uh environment an oven to sterilize but remember with dry heat it takes quite a bit of time hours to sterilize now what about cold one thing you should know about cold is that cold does not kill the microorganisms instead cold merely slows down the microbes by placing microbes let's say in a freezer at minus 20° C you're not actually killing microorganisms you're making them slow to the point of suspended animation they stop moving um that's essentially all that's happening so we're not using cold to kill microbes cold is lower you know the freezer usually is lower than the minimum temperature required for growth of microbes and so these microbes simply stop growing but they're not dead what about desiccation what physical control method is this what does this refer to desiccation refers to dehydr ation a des a desicant is an a substance that removes moisture from the air so desiccation is drying of the cells remember that all cells require water for life and most of the volume of a cell is water so by drying the cell by drying the environment these pathogens can't grow desiccation can also preserve Foods so if we make beef jerky if we make dried fruits like dried bananas or dried apricots these foods are resistant to microbial growth because they do not possess enough water for the microbes to grow on next what about radiation this is another form of physical control radiation radiation refers to energy emitted from atomic activities and dispersed at High Velocity through matter or space there are three types of radiation that we can use for microbial control gamma rays xrays and ultraviolet radiation irradation refers to Cellular bombardment with radiation so if I use radiation to bombard cells and kill cells I'm irradiating those cells ionizing radiation uh this type of radiation causes catastrophic mutations in DNA it causes double strand breaks in the DNA and xrays and gamma rays are capable of ionizing radiation conversely nonionizing radiation nonionizing radiation causes uh these thyine thyine diers to form or cytosine cytosine diers to form in the DNA this is known as abnormal bonding mutations and UV radiation is known for causing this type of nonionizing damage now ionizing radiation can sterilize without heating this is also known as cold sterilization but don't don't uh don't uh confuse cold sterilization with sterilizing with cold the reason it's called cold sterilization is because gamma rays X-rays and UV rays they don't require a heat and so it's known as cold sterilization so it's that's why it's called cold sterilization so there are devices that emit ionizing Rays gamma ray machines x-ray machines cathode ray machines these machines means emit ionizing rays which can cause double strand breaks in the DNA causing cellular death and sterilization again ionizing radiation includes gamma rays and x-rays this causes double strand brakes to form because of the uh stripping of electrons from DNA and RNA nonionizing radiation this includes UV rays this leads to abnormal bonds within the DNA such as thyine thyine diers or cytosine cytosine diers together those are known as perimidine permiting diers because thyine and cytosine are known as peridin UV rays are a form of nonionizing radiation these rays are absorbed by DNA and cause thyine thyine diers or cytosine cytosine diers these are not as penetrating as ionizing radiation so with UV did you know with UV the bacteria have to be directly struck by UV they have to be on a like a smooth clean surface and the UV rays have to directly uh directly penetrate the cell if the cells are underneath plastic or underwater then UV will not have an effect on those cells the UV have has to directly touch the cell and the cell will develop these thyine thyine diers or cytosine cytosine diers and that will lead in cellular death so here you can see exactly what I've been talking about with UV here you can see two adjacent thines in the DNA molecule thyine thyine and here you can see the effect of UV the the effect of UV is forming a dimer between these two thyine these two residues UV radiation can also disrupt cells by generating free radicals free radicals are these toxic photochemical products that damage the cell they interfere with essential cell processes by binding to DNA RNA and proteins this can lead to cell death and lastly let's talk about filtration we can use the physical control method of filtration in order to remove microbes from air or liquids here you can see a the filter with pores and this liquid is being filtered through a membrane filtered through um this filter and you can see the bacteria are stuck on this side of the of the filter because the pores are fine the pores are small and they allow for the water to be filtered now keep in mind with filtering if it's tiny tiny viruses the size of the pores those tiny viruses may get through so often times filtering is not necessarily A sterilizing technique here you can see that filters can be ultra fine down to 0.02 micromet however even these filters May let the smallest viruses across so filtering isn't normally a uh form of true sterilization even smaller pore diameters permit true sterilization by removing viruses even large proteins you need to have the finest the finest filters in order to achieve sterilization with filtration um unless you use these Ultra ultra fine uh filters that can even filter out proteins and viruses the fluid is not considered uh sterile uh this has the same effect when you have a lot of sugar a lot of solute in the environment it makes it really difficult for microbes to grow they become plasm they become dehydrated the water leaves the cell okay okay so now we're going to move from physical control to chemical control methods of chemical control and when we're looking at desirable antimicrobial chemicals what properties are we looking for we're looking for Rapid action low concentration solubility in water or alcohol good shelf life we don't want our our chemical to degrade in just a week or two we want it to have a good long shelf life broad spectrum that means the ability to kill a broad range of microorganisms it should penetrate and animate surfaces to get where the microbes are it should be resistant to becoming you know in activated it should be non-corrosive and nonstaining that makes sense it should have sanitizing and deodorizing properties it should be affordable and readily available all of these make it a better chemical to use for control so these are the desired qualities of a good antimicrobial chemical to use the high level chemicals the highlevel germicides they can actually kill endospores right endospores not everything can kill endospores right like even Lysol can't kill endospores but there are some chemicals that can kill endospores like glut alahh you know hospitals use 2% glut alide because after 3 hours remember that after 3 hours it does kill endos spores so some chemicals are called highl germicides these chemicals can kill endospores and why would we want to use these sterilants we could use them on items that can't be put in the autoclave right we could use them on catheters heart lung equipment implants things that could get destroyed in the autoclave okay what about intermediate level germicides these would kill fungal but not bacterial spores these would be good against some microorganisms but not all and we could use these on semi critical items such as respiratory equipment and thermometers and then obviously there's lowlevel germicides this is probably what you have around your house as in the form of Lysol and other things like that these eliminate only the vegetative mother cells vegetative fungal cells some of the viruses especially the enveloped viruses which are more susceptible to degradation they're used in the clinic on non-critical material like the electrodes that are placed on your skin straps or furniture that touches the skin okay and of course what are some factors that can affect microbial microbi microbicidal activity well what's the nature of the microorganism is the microorganism more resistant is it more susceptible does it have like for instance does it have a capsule does it have molic acids is it acid fast is it gram positive is it gram negative all of these things can affect the susceptibility of the microb to the chemical right the nature of the material being treated right are you spraying a nice clean smooth surface are you spraying a nice table or are you trying to get them out of this cloth material right that's going to be a little bit more challenging to get microbes out of this cloth material than it is off a table that you could just spray easily right the degree of contamination you know that makes sense how badly is it contaminated the time of exposure remember I we talked about this last time you can't just spray and then wipe it immediately you know did you allowed the proper exposure time for the chemical and the microbes and then the strength of the germicide you know did you use a high strength germicide you know or did you use a diluted or watered down or low low strength germicide and again we talked about this last time but what are some targets of these antimicrobial chemicals the targets are either the proteins of the cell the nucleic acids of the cell that would mean the RNA or the DNA the cell wall you know the peptidoglycan cell wall or the cell membrane these are the these are the most typical targets of antimicrobial chemicals so let's talk about some antimicrobial chemicals let's start with the halogens halogens are elements including Florine bromine chlorine iodine all of these can be used be because of their antimicrobial uh properties you know they put uh chlorine for instance in swimming pools you know that helps to destroy microorganisms that get in the swimming pool chlorine is also a central component in Bleach right bleach has chlorine in it iodine we all know about iodine you know uh it's very standard to use iodine as a antiseptic remember antiseptic means treating the living tissue right antiseptic but Florine and bromine also have antimicrobial properties as well these are known as the halogens chlorine is a very powerful antimicrobial agent it can even kill endospores like I said chlorine is found in um bleach and bleach is a sterilant it can sterilize at high enough concentrations and the way it works you don't need to memorize this for an exam or anything but the way it works is it denatures enzymes it denatures the proteins of those microbes and this is why you don't want to get bleach on your skin because it will also start to denature your own proteins and enzymes right so you got to wear gloves if you're going to work with concentrated amounts of of chlorine or of bleach and in their swimming pool obviously they dilute it so it's effective against the microbes but not against you but it's still you know it's it's pretty toxic stuff okay what about iodine remember iodine it penetrates the cells interferes with their metabolic function it can cause disulfide or interferes with the disulfide bonds of proteins so it basically disrupts proteins and again you don't need to know each of these different chemicals and what it disrupts but I'm just mentioning it you know just so you know okay but we all know that you can use iodine to help disinfect wounds things like that as an antiseptic now if I use iodine on a Surface remember I'm using it as a disinfectant if I'm using I on my living skin then I'm using it as an antiseptic right you see the difference if something is used on a Surface it's a disinfectant if I'm using that substance on the skin I'm using it as an antiseptic exactly right we don't need to go into all these fine details here about iodine what about phenol let's move on to phenol phenolics are these antimicrobial chemicals that have a phenol ring phenol was one of the earliest chemicals used to disinfect do you remember when I told you about liser and how he treated his surgical instruments before he did surgery and he treated the surgical site he used phenol phenol is a potent antimicrobial but it's pretty toxic so it's not great uh it's pretty toxic it makes them dangerous to use as antiseptics you know uh so it's kind of like hydrogen peroxide right if you guys remember hydrogen peroxide was commonly used as an antiseptic but now dermatologists pretty much agree that you shouldn't use hydrogen peroxide on your skin it causes a lot of damage right because it's oxidative right and phenols like that so liser would have used it on your skin to help prep for the surgery but nowadays we know that's not worth it there's better antiseptics to use for surgery we're not going to put phenol on your skin but it is a potent antimicrobial okay again phenol is a particular chemical but phenolic are other other types of ringed molecules phenols are ring structures molecular ring structures and these phenolics are known to be antimicrobial they destroy bacteria fungi viruses and you know like I said they're useful because they're antimicrobial but they tend to be too toxic too toxic to use as an actual antiseptic on your skin now one type of phenolic that we use on the skin is triclosan triclosan and in fact up until recently it was very very common to find trlo sand the phenolic in soap you know hand soap anytime you pick up hand soap and it's said antimicrobial hand soap most likely the antimicrobial agent in the hand soap was triclosan but unfortunately look what happened here widespread use of triclosan led to development of triclosan resistant microorganisms so this is terrible um nowadays you know most most clinicians would agree that you don't necessarily need antimicrobial hand soap you just need soap and water and that gets you pretty clean you know unless you're doing something like actually working with microorganisms or something in which case you might want some antimicrobial hand soap but just to wash your hands at home you know soap and water is fine you know you don't need antimicrobial hand soap because that causes a problem right now the microbes get more resistant to those antimicrobials is that really worth it you know not really so alcohol you guys saw alcohol in action remember when we did our Kirby bow test in the lab you took the little tweezers and I said just put the little tweezers the forceps put them in alcohol and then how long did I say to flame it yeah just for a second you just want to burn the alcohol off you know why because the alcohol itself kills the microbes alcohol itself is antimicrobial so the only reason we flamed wasn't to sterilized the forp it was to burn off the excess alcohol right alcohol has antimicrobial properties but at high concentrations the concentration that's usually potent enough to kill microbes is around the 70% alcohol range that would be 140 Proof so at least 50% but usually 70% and if you notice a lot of these hand sanitizers like this this right here says this is a hand sanitizer active ingredient alcohol 76% volume 76% that's basically the only reason this is antiseptic is because it has 76% alcohol in it that's it so instead of buying this expensive thing you could just mix alcohol and water but make sure it's it's uh alcohol that's you know meant for use on the skin cuz sometimes alcohols they have like Benzene mixed in with them and stuff so just be careful okay and again alcohol at high enough concentrations like this 91% isopropyl alcohol you probably have around your house you know people use that as a skin deter determining agent right as an antiseptic right so anytime I get a cut or I get you know something a break in my skin or a splinter I always treat it afterwards I wash my hands and I treat it with some alcohol you know like a little cotton swab of alcohol then place the bandage on top right so you might make sure you wipe off the skin and kill the microbes in that vicinity and you guys know alcohols are a colorless hydrocarbons you got the hydroxy groups on there okay we don't need to know much else about this but the way the way alcohols work is they basically attack the membrane of the cell they attack the proteins of the cell okay and as I just showed you most alcohol-based hand sanitizers contain about 60 70% ethyl alcohol and are more resistant to evaporation so they you know whatever they mix in there with the alcohol makes keeps it from evaporating because if you've ever worked with like rubbing alcohol or pure alcohol it evaporates pretty quickly so you don't want that if it's a hand sanitizer okay isopropyl alol so ethyl alcohol is ethanol isopropyl alcohol is more microbial cyto so it's more deadly to the microbes and less expensive than ethanol evaporation occurs more readily so it's you know you have to use it more quickly all right and then there are oxidizing agents let's move on to oxidizing agents remember oxidizing means to steel electrons right do you remember Leo says I always say Leo says loss of electrons means oxidized gain of electrons means reduction reduction reduced right so oxidizing agents they cause things to lose electrons they steal electrons oxidizing agents steal electrons they C cause they Cause something to lose electrons okay so for example hydrogen peroxide hydrogen peroxide is a good antimicrobial agent because it's oxidative it steals electrons from things and when when that happens that causes damage to the cell that causes damage to the DNA that causes damage to proteins it can cause cell death so again remember hydrogen peroxide used to be used as an antiseptic but nowadays we know that it's not worth it because it's better to use something like alcohol you know because hydrogen peroxide not only does it oxidize the bacteria but it oxidizes your own cells and that causes a lot of tissue damage and it can slow the healing process right when was that discovered because I used to always pretty recently yeah yeah yeah yeah it pretty recently but um but nowadays the yeah I think I think what they did was they they tested you know different ways of curing you know wounds you know curing wounds and they they they showed that you know with hydrogen peroxide takes longer to heal you know and so the problem is when you added that hydrogen peroxide you cause some more damage you know and when you cause more damage it's going to take longer to heal is not worth it so alcohol causes less damage to your own cells it's better to use as a antimicrobial uh antiseptic I should say yeah as an antiseptic 3% hydrogen peroxide was used for skin and wound cleansing mouth washing bed sore care you know but nowadays it's becoming more and more you know uh faux paw to use you know the the the the benefits don't seem to outweigh the drawbacks you know but in a pinch if I were if I were in the zombie apocalypse like Zombie Land or something or Walking Dead and you know I had a wound and the only thing around was hydrogen I definitely use hydrogen peroxide but if I have a choice I wouldn't use hydrogen peroxide yeah at High Enough concentrations we're not talking 3% but at high enough concentrations it can be not just bacterio cdal viricidal fungicidal sporicidal it can kill everything at high enough concentrations okay ozone ozone is O3 it's a gas it has similar effects to hydrogen peroxide it's it's an oxidizer and we can use ozone to disinfect the air water and in industrial air conditioners and cooling towers detergents detergents are good because detergents have part polar and part non-polar ends that's what detergents have part polar part non-polar and that means that they can embed into the membrane of the cell and disrupt the membrane right so they often act as surfactants remember surfactants they insert themselves into the membrane the plasma membrane and then cause the membrane to disrupt to open up there are some limitations to detergents ineffective against tuberculosis bacterium hepatitis virus pomonis and endospores so it's not great you know you can't use detergents to sterilize but detergents are mainly used to sanitized do you remember sanitized like when you wash the dishes you use a detergent right that's sanit sanitized or de germ remember that's the lowest level of you know microbial control is this you know just sanitizing yes so you had like a tower something that GRE like mold and washed it detergent it would not be not sterile so the mold would still be there maybe Well it Well there's also agitation if you put it in your D in your in your uh laundry machine there's agitation there's hot water there is the detergent so most likely the combination of the detergent and the agitation and the constant rinsing and you know you're probably going to get most of it off of there yeah yeah but it's not because just the detergent it's the detergent plus the agitation plus the hot water yeah yeah good question what's the agitation agitation so you know the yeah unless you don't have a agitator you know newer newer uh machines don't have the agitator in the they have just like the spin thing which is I have the spin yeah it's not as it's not as effective surfactant so remember surfactants are a type of detergent surfactants have a polar end a non-polar end they insert themselves in the membrane remember this slide uh I hesitate to go back this far but there was a slide here I just want to show you way way back here see these surfactant molecules remember from an earlier slide slide 25 these surfactants insert themselves into the membrane of the cell and then they cause the membrane to disrupt this is what detergents do this is how detergents work they insert themselves in the membrane the membrane gets fced quary emo compounds you know we're not going to touch on these much but they are a type of detergent as well they're mainly you know like soaps basically they're for they're mildly microbio cyal we did the handwashing activity right yeah we did the handwashing activity so with a germicidal soap you do end up killing more microbes um than a non- germicidal soap but remember usually for most applications you could just use regular non- germicidal soap around the house right but if you're going to I don't know cut raw chicken or handle raw chicken or something you know then you might want to consider using some germicidal soap to kill the salmonella for instance heavy metals we kind of touched on this last time too heavy metals are by Nature antimicrobial because they they they tend to compete for co-actors in the cell and that can cause the cell to you know not function correctly so heavy metals what are heavy metals Mercury right silver other metals just Metals in for instance if you took a old school Copper Penny when it was still made out of copper right let's say you take a Agger plate if you took an Agger plate and did a spread plate with ecoli like you just spread ecoli all over the plate and then you put a a real coin in the and by real coin I mean one with a heavy metal in it like a silver coin Co or a copper coin and you put that coin in the center of the plate and then incubate and then came back you would see a zone of inhibition isn't that neat just like the Kirby Bower test you came back and you saw a little zone of inhibition if you put a metal or even a chunk of metal if you took like a little chunk of silver or a little chunk of gold and you placed it in the center then the microbes would avoid it and this is why you know it's nice to have water bowls for your pets you know it's preferable to have water bowls that are metal for your pet than water bowls that are plastic or something like that or even glass and they also say this is why they put the copper plate on the bathroom door you know on the bathroom stall you got the copper plate cuz that if that's a real copper plate it's going to have some natural antimicrobial properties to it now if the previous person just splattered a bunch of stuff on that that's not going to help you much but you know it it it is naturally antimicrobial right see this is what I was talking about look at this this is a actual plate with silver and gold and look what's around it zone of inhibition right a zone of inhibition the microbes want nothing to do with that heavy metal okay let's move on to alahh alahh are very potent antimicrobials in fact you could call them sterilants that means they're strong enough to sterilize especially glut alahh what do we use glut alahh for remember in the hospital they use a 2% glut alahh solution to spray down inanimate objects for 3 hours and that's sporicidal what about form alahh another type of alahh is form alahh what do we use form alahh for do you know pres preserving things yeah if you want to you know if you ever been to a science lab and you see the frog in the jar you know it's a preservative form alahh is a preservative glut alahh form alahh these are types of alahh what they do is they cross-link molecules they Crosslink proteins together and that can kill microorganisms right that that can kill microorganisms and again like I said glut alahh we use this in the clinic it's broadspectrum you can use it to sterilize isn't that a powerful word sterilize that should tell you something you know that means it's the the gold standard you can use it as the gold standard it's much better than Lysol Lysol can disinfect good or alahh can sterilize well if you leave it on the requisite amount of time right three hours you know you have to leave it on for three hours but if you do you can sterilize equipment like Scopes kidney dialysis machines Dental instruments you know all kinds of things in the clinic so it's just like a spray yeah you spray it's a bottle and usually you just spray it or a lot of times they'll have a tub of it and they'll place the instruments in that tub so again glut alide form alide these are potent antimicrobials but form alahh tends to be used as embalming fluid or preserving preserving tissue samples preserving um you know organisms for labs skip that okay gaseous sterilant and disinfectants so you can use gas you can use gases to sterilize as well ethylene oxide is an example it's a o a colorless substance that exists as a gas at room temperature it disrupts DNA and proteins so it's a strong alkala agent okay so why would we want to use a gas so again remember this is a type of cold sterilization because if I put something in a gas chamber and then gas it to sterilize it did I have to heat it up no so that's another way to do cold sterilization right I don't need to put it in the autoclave I don't need to heat it up I can put it in the gas chamber right and then expose it to this gas it didn't have to get hot right so it's another way of sterilizing things that are sensitive to heat right medical devices let's skip through acids and alkaly obviously we know that acids can kill microorganisms strong bases alkalized are strong bases you know they can disrupt the cells as well these cause denaturation all right skipping ahead oh I'm GNA skip that one and that's it we did it so we've reached the end of chapter 11 uh yeah pretty straightforward chapter but an important chapter if you're going to go if you're going to go to the clinic if you're going to be in the field of nursing you know or dental hygiene you know you need to know these terms and not mix up these terms that's one way to be called out as a newbie is not know the difference between disinfect or sterilize and these terms awesome so let's call it a day let's head over to lab and remember today's the important very