Transcript for:
Understanding Bacterial Growth and Classification

foreign good day good evening okay so it doesn't matter what time you have opened this lecture video but I'd like to say that I hope everyone is in um the best of condition for uh listening to a lecture video okay so for today we have the prelim lecture three uh bacterial growth and classification and later I'll be making another lecture video for bacterial genetics so for this particular lecture uh this will be divided into two parts now we have the classification of bacteria and the learning outcomes for this particular part of the topic would be the following so at the end the students are expected to be able to categorize the basic principles of microbial classification system and to be able to discuss the structural and biological characteristics that we use in classifying bacteria so there are three known classification systems that are widely used in microbiology and that would include bacteriology of course so we have numerical taxonomy uh phylogenetic classification and of course phenotypic classification which is the most common now for numerical taxonomy this makes use of computer well the computer would cluster different strains of organisms at selected levels of overall similarity usually on the basis of the frequency with which they share uh traits okay so you can see this in Burgess manual usually I think page 39 so if you can get a copy of that you could check that out okay so like uh when we use machines right like vitec for example so it uses computer taxonomy or numerical taxonomy so we provide bacterial organisms with a certain set of numbers to identify them and to separate them from uh the other members of the same genus okay then we have phylogenetic classification so uh the basis for this type of classification system is the uh genetic similarity you know so groups reflect genetic similarity as well as evolutionary relatedness okay so say for example uh feline family you know the cat family for example so they are grouped together because of the presence of the whiskers the presence of pointed ears okay uh the eyes that would reflect light and so on so phylogenetic classification of course the similarity of the proteins okay that are shown on uh genetic studies okay and then of course phenotypic classification so this would be based on the overall similarity so this is what we usually do in the laboratory when we uh culture and then we review those cultured organisms microscopically and check out their morphology okay and so this is an example of a machine that we use or a computer that we use to identify organisms uh numerically okay so we have the white Tech 2 okay so it's an automated instrument for identification as well as sensitivity testing so AST here is antibiotic or antimicrobial sensitivity testing okay so this one uh the colored thing here and this is actually API we use this for identification of enterobacterial so this scholar things that you see here actually gel not gelatin which includes uh particular carbohydrates and materials that are needed in order for us to identify uh the organism and so we have numbers here to indicate the intensity or the presence of the reaction that you would like to see now to identify a particular bacteria okay and then so number four this is what we see you know as the uh on the interface of the computer the lighting after the assay is done okay so we have here ID values or numbers that are used to identify specific species of a uh group of organisms right of course for phylogenetic uh classification so we base solely on this uh phylogenetic tree note that we have three domains of living organism okay so domains are the large groups into which we classify or categorize organisms so we have uh the domain bacteria which includes of course everything that we will be discussing in this course and then we have domain Archaea so the domain Archaea are organisms that are considered ancient okay so included here are square and star-shaped bacteria that we only get to see from the Red Sea specimen and then of course we have the domain eukaryotes which would include us humans no under the kingdom Animalia and then we have fungal organisms or also eukaryotes they belong to the domain eukaryotes same with the parasites so you have here the ciliates the flagellates the trichomonas okay so you have met those in your parasitology course Last Summer okay so uh of course part and parcel of classifying organisms are to provide them with names now before they are provided with names they are given the complete you know classification and they are usually there are usually seven one two three four five six seven taxa or levels when we classify organisms so their names are not just limited to two not the genus and the species actually they have a very very very long name okay so uh the levels of classification called taxon or taxa if there are a lot so in this case that's how Okay so taxon is a group or a level of classification so this are the hierarchical systems within the domains recall earlier there are three domains right so you have the domain bacteria the domain Archaea and the domain Eukarya so each of those domains are further divided into seven levels of classification or taxa so we have Kingdom after a domain we have Kingdom then phylum or division then class then order family genus and species so before we were asked to memorize a mnemonic you know so King Philip or King David came over for good spaghetti you can easily memorize the uh order of the taxa okay so Kingdom phylum or division class order family genus and species of course in the references that we get to read they usually make use only of the genus and the species because it would take so much space if you include the domain the kingdom the phylum the class the order the family okay so of course it would be good for us no for people who's going to read those books okay to limit ourselves to genius and species only okay so specie is technically the basic unit of taxonomy you know so it's that specific name you know that would identify one particular bacteria okay uh from a group of similar organisms okay so this would be equivalent to our given names right so there would be many tago pero there would only be one tiny okay so uh so that would be species okay so it would represent a specific recognized type of organism and uh the identity we usually identify them by comparing you know the uh organisms that we were able to grow in the lab with known pipe strains okay so how do we know that uh the organism that we were able to grow on our culture media in our laboratory would be that particular organism of course there would be uh peer cultures or type strains okay so the most widely used uh type strains or pure cultures uh here okay is the atcc so we usually buy this so we can grow them and keep them in our laboratory so anytime that we get confused we get to look at this we can grow samples from this set no the the atcc and so we can observe their growth and compare whatever it is that we have grown from the sample from the patient sample comparatory or the American type culture collection of organisms so you can buy this in uh Dost they have this you know so uh come your research for example you'll be needing bacterial specimen uh bacterial species you can buy them from dosd or from uplb okay but that would be much more expensive okay strain okay so strain is a population of microbe that has the descended from a pure culture okay so if you have grown your peer culture in the laboratory and regrown them okay so say for example from the initial culture you have subculture them into another uh plate or another uh media okay so the succeeding growth would be cold strains okay so different strains represent genetic variability with the species so usually uh the longer we culture them or the farther they get away from their pure culture sometimes uh they change no morphology they lose some of their virulence some bacteria may lose some structures in the process okay so we have different strains because of that okay and then uh streets can be divided into different types of we have bio varieties or simply biovars morphovirus and then serovars okay so biovars uh this may mean they are biologically variable from their pure culture okay so one example is europlasma urea lithicum okay so we have the biovirus parvo and biovar t960. a for more for VAR so morphologically they are different no uh in this case for Quran the bacterium diphtheria they are uh morphologically different in terms of their colonies okay so their colonies are morphologically different okay those that are small are called metis okay those that are intermediate or medium-sized we call them intermediaries and then we have dravis okay so it is intermediaries and gravis are the more for varieties of Quran bacterium diphtheria okay so complete name would be for example bacteria morphovar metis okay so we insert the terms more for VAR biovar and server in between okay and then we have for Server variety they are serologically VAR uh varied from the period culture so we have here salmonella enterica serovar TV okay now in writing the scientific name always remember they should be in italics no so italicized or underlined so sometimes when you're writing on your yellow paper say for example for a quiz it could be difficult to determine whether or not you have used italics okay so to be sure when we have our quizzes our um face to face or on-site quizzes remember to write the scientific names and underline them underline okay so remember to underline scientific names okay most scientific names are latinized okay so they're Latina it's like eskerikia Kali you know so esquerica is actually the name of the discoveries esketic okay so uh we simply add a um letters at the end to make it sound Latin then sometimes they are descriptive you know like uh safilococcus Oreos okay so or use is descriptive primarily because the colonies are yellow colored okay so from aurum which is the Latin term for gold right so descriptive okay so the names can be descriptive or it could be as uh to honor the scientists to discover them okay so scientific in the team okay so species are never ever abbreviated so what we abbreviate is the genus like but we only abbreviate if we were able to uh introduce the complete name in the first hand okay so do not introduce the abbreviated form so first introduce the name of the organism in full okay so A genus name may be used alone to indicate a group so you can simply say sapulco's group or the safilopokai that you cannot say orios [Music] okay but you cannot use okay so these are the examples as mentioned earlier so staphylococcus aureus is actually a descriptive no it's a description of what the organism looks like microscopically okay so safilo means clustered and kokai refers to their spherical structure or use is because of their golden I golden yellow colonies okay they may not look golden microscopically because they will be stained with a different color altogether but when we grow them on culture media on gelatin they're actually yellow okay and then for escarakia coli yes it honors the Discover Theodore aesthetic okay so escarity nice and then Koli refers to the colon where we usually see or isolate the organism okay so again once you have introduced the complete name no scientific names may be abbreviated but never start with an abbreviation okay so always introduce the complete name first and then you can abbreviate thereafter okay so other examples we have here salmonella species so you can use that no salmonella species okay so say salmonella and terica subspecy enterica okay uh one common cell variety is the Dublin okay so on complete name Dublin would be uh salmonella and terica subspecial completely okay so serovar is um capitalized not the Dublin okay the the first letter of the sarovar is capitalized but it is not italicized okay so if you'd like to abbreviate it you can only abbreviate the first letter of the genus so s dot enterica subspecialvar Dublin but no uh this shortened version is also acceptable okay so salmonella Dublin would also mean that it is salmonella and terica subspecial emperica sarovar Dublin okay so pedrine as words [Music] okay and then they do have common name search reveal names no descriptive names like see mycobacterium tuberculosis we know here it rather we know it also as tubercle bacillus and the Nigeria meningitis is also meningo Cocos and the group a Streptococcus which is specifically we simply know it as or refer to it as gas or gas okay so uh shortened group a Streptococcus right so the Burgess Manual of systematic bacteriologists actually a very important tool in identification of unknowns so for newly identified organism this serves as a guide okay so it's the main source resource for determining the identity of bacterial species utilizing every characterizing aspect no so there uh the three types of classification is actually present in the Burgess Manual of uh determinative bacteriology okay so here they use the key features now we have the dichotomous key to narrow down identification so I need so this is a process really that would help us identify the organism present in the patient specimen so gram reaction is it gram negative or gram-positive if it is if it is gram-positive no purple colored check the morphology are they rods or spherical organisms if they are rods they could be this organism if they are kokai they could be this organism okay now what if they are gram negative no red and color okay check their ability to ferment glucose can they ferment glucose and produce acids they could be this organism or if they could produce acids and gas no they could ferment glucose and produce acids and gas if so are they motil can they move okay if yes this could be the organism if no check if they're able to hydrolyze urea if they could it could be this organism so check could they utilize citrate if so this could be the organism now so this is what we use this we call this dichotomous key because reactions can go either way you know so there are two ways by which a reaction can go so that would serve as a guide to us next if we come up with this particular result okay so we call that dicotine they call the mosquito and that can be found in the vergee's manual of classification okay and moving on to part two of this discussion we have the physical and nutritional growth requirements of bacterias because we work with them in the laboratory it's important for us to know how to grow them right so what are the nutrients that organisms would need in order for them to grow artificially in an artificial environment rather okay so the learning outcomes for this part of the topic would be you know for you to be able to identify the growth requirements of bacteria and for you to be able to appreciate the importance of physiological and traditional requirements for bacterial growth okay so like human beings microbes also need several elements for growth okay so they would need for major elements they need carbon of course oxygen or nitrogen okay so they need all this major elements in order for them to grow but they also need this trees elements you know so for crease elements they only need this for minimum uh at the minimum but they should be present of course no so this elements would act as cofactors for enzymatic reactions our bacterial organisms will not be able to perform in the needed metabolic functions if they do not have this major and Trace elements available okay so they only need a small amount okay and it does not need to be added to the culture media because it's there now it's included in the culture media however sometimes there are some elements that may not be found in the best culture media for a particular organism so you get to add them okay or there are some organisms that would need more of a particular element so we need to provide them in order to achieve you know uh the amount of growth that we want for the organisms that we have to grow in the laboratory okay So based on their nutritional requirement or physiologic requirements you can also classify organisms okay so an organism is called a phototroph if they would get their energy from light okay so some organisms need uh light not necessarily sunlight no artificial light inside the oven or the incubator would be uh added or would be necessary in order for them to grow okay some organisms would need different types of chemicals in order for them to gather energy and so we call them chemotrophs okay so phototrophs are organisms that gather energy from light chemotrophs or organisms that gather energy from chemicals now chemotrophs are further divided into the types of chemicals that they would need now organisms that need inorganic chemical for energy are called chemolitotrophs those that would need organic chemical or energy source are called chemoorganotrops okay now organisms that would require carbon Source okay and use carbon dioxide as the source are called autotrophs okay or we call them also as capnophiles now Capital files are organisms that would need more carbon dioxide than normal okay so all organisms would need some organisms would need carbon dioxide but those that would need a lot or a lot additional rather additional uh percentage or carbon dioxide and normal are called capnophiles okay so again you know to avoid confusion those that would need carbon dioxide are called autotrophs and those that would uh those that would require a larger okay amount of carbon dioxide are called capnophiles still they're autotrophs right and those that would need organic compounds as their carbon Source are called heterotrophs so for growth factors some organisms would require additional growth factors apart from those major increase elements okay so growth factors are essential substances that the organism is unable to synthesize and so we add them okay so they are required in small amounts also okay so they could be purines or pyrimidines no so that they could gather nucleic acids if needed okay uh to to create rather nucleic acids uh some organisms would require amino acids okay so that they could synthesize their own proteins and some would require a variety of vitamins you know like coenzymes and enzymes in order for them to perform metabolic processes okay now the organisms that would require growth factors are technically called fastidious okay in other words growth factors and nutrients for uh growth Okay so a lot of our disease causing or pathogenic organisms are fastidious okay like Nigeria gonorrhea for example is considered a fastidious organism mycobacterium tuberculosis is also a fastidious organism it would require a multitude of growth factors and other elements in order for them to grow luxuriantly in an artificial environment okay so moving on apart from the major and phase elements and additional growth factors the usual requirement for growth are the following so number one we have oxygen of course living organisms would require oxygen however uh they have some of these microorganisms require uh a lot some require only a small amount and some would not require it at all okay so those that we really need oxygen for growth are called obligate aerobes Ayan no so these tubes will show you their growth okay the small gray circles that you see inside those yellow uh colored broths are actually colonies or bacterial growth so if an organism is an obligate Arrow meaning they really need oxygen for growth they would be seen somewhere near the surface where oxygen is easily available okay then we have facultative on aerobes so facultative and aerobes are technically aerobic organisms but they could survive even in the presence of little or low oxygen okay so you can see their growth they're dispersed right so are seen on this surface some are saying uh in the middle and some at the bottom and then we have here obligate and aerobes these are the organisms that do not need oxygen for growth otherwise they die if there's oxygen present therefore you can see them growing at the bottom of the tube right down there okay and then we have Aero tolerant or uh aerotolerant and aerobes okay so this are unaerobs that can survive or tolerate a small amount of oxygen so you can see them similarly this first no in the solution but they grow best okay again these are an aerobes but they could survive even in the presence of oxygen okay this one the facultative and aerobes this are aerobic organisms but they could survive even in the presence of little oxygen okay so facultative and aerobes are aerobic organisms Aero tolerant and aerobes are anaerobic organisms they are also known by the way you know aerotolerant and aerobs are also known as facultative aerome okay so uh they are on aerobes that could survive in the presence of oxygen facultative now micro aerophylls are aerobic organisms but they only need a very very small amount of oxygen and so they grow best at the Middle where oxygen is at its thinnest okay so dito micro aerofields counting oxygen lung behind okay so we can also uh group organisms according to their ability to uh produce enzymes okay so our obligate aerobes okay and most of the facultative and aerobes are able to produce uh superoxide okay superoxide dismitase and catalase but they cannot produce peroxidase okay most Aero tolerant and aerobs like streptococci can produce superoxide this methase and peroxidase but they cannot produce catalase that's why they are negative for catalase tests right so see obligate an arrows Naman right uh they are unable to they lack superoxide this methane is catalase and peroxidase okay so they undergo lethal oxidation by various oxygen radicals when they're exposed to oxygen okay so paracetylene suicide oxygen obligate an aerobes expose even in little oxygen so examples of obligate and aerobes are clostridia and the bacteroidis group okay now apart from oxygen heat is also very important for survival that's why we incubate them after introducing them onto gelatin media right so like humans we need heat okay so thermal requirement is also an identifying Factor so those that are able to survive at uh 20 to 40 degrees celsius you know so Optimum growth is at body temperature okay we call them mesophiles so most of the disease-causing organisms belong to this group mesophiles okay now those that can survive in temperatures above 45 degrees Celsius are called thermophiles and those who can survive Beyond 45 Beyond uh 80 degrees Celsius those that we can see in very very hot surfaces are called hyperthermophiles okay now those that do not need heat okay there are some we call them cyclophiles they survive best in cold temperatures you know so you call them cyclotrophs or facultative cyclophiles those that can survive uh zero or uh zero degrees are called Cyclery they can endure they can endure uh they can endure cold temperature freezer temperatures okay so again a note that those organisms are able to uh produce infections or diseases are mostly mesophiles you know those who can survive at 20 to 45 degrees Celsius and their Optimum growth is usually between uh 35 to 37 degrees Celsius okay together so thermophiles again uh 45 degrees to 70 degrees uh those that can survive higher than 70 degrees okay are hyperthermophiles okay so usually those are the archaeons so the old bacteria and then of course water and salinity the most indispensable requirement for growth is water okay so living organisms would require water those that would need a lot of water or a lot of moisture in order for them to grow are called humidifiers and those that live no those that can survive in dry environments like the desert are called serophiles okay seraphiles okay so salt is uh very important uh solute no of course because it is the solute present in most of the solutions inside the body okay and then um there are certain differences as to the requirements of organisms when it comes to Salt okay those that would require an environment or a place to grow with a concentration of salt that's higher than normal are called halophiles okay so salt is important everybody would need it because it's part of the solutions that we have okay however there are still some organisms that love to strive in uh environments with a slightly higher okay um salt concentration but still they can survive even without a sodium chloride we call them halophiles okay so mild halophiles can grow at environments uh with one to six percent salt moderate halophiles at six to fifteen percent salt concentration and uh extreme halophiles at 15 to 30 percent salt concentration okay uh those that can uh tolerate okay salt but grows best no at point nine the normal concentration of Body Solution is at 0.85 or rounded off that's one percent okay but there are those that grow best at that temperature at that concentration but can still survive even if slightly higher on concentration we call them okay so salt and water actually is um would change you know the concentration of salt and water would change because of uh diffusion no processes that occur inside the body okay so simple diffusion is the movement of a solute from higher concentration to a lesser concentration okay so salt will diffuse okay or uh yeah facilitated diffusion on the other hand would require uh Gates no or channels and this would need it energy or ATP so this is what happens usually in our cell membrane so gated Okay small solute okay they would undergo simple diffusion they would simply just pass through the channels in our uh cell membrane energy however some solute would require energy in order for them to enter into the membrane and so they would require facilitated diffusion now uh water water moves across membranes in a process called osmosis okay and chambray the movement of water is similar to the movement of solute also so water moves from an area of Greater concentration one area of lesser concentration [Music] inside the body osmosis and diffusion osmotic pressure is the pressure required to stop water movement across the membrane equilibrium movement equilibrium so the concentration of solute inside the cell and outside the cell should be balanced paranormal and growth and uh processes and so no no when we talk about water and salt so the types of solution would also come to mind so you have isotonic solution hypotonic solution and hypertonic solution so isotonic solution is uh the type of solution that is Crescent in the body under normal circumstances okay so here okay uh the concentration inside and outside the cell is balanced in equilibrium so those there is no net movement of water if there's 0.9 of solute inside that would mean there's also a concentration of about 0.9 percent outside so that's isotonic solution this is the best solution for bacterial growth okay now sometimes the concentration is the concentration of uh solution is higher inside the cell or higher yeah higher inside the cell okay and then compared to outside okay so hypotonic means so what happens is water from outside would enter into the cell to balance you know the concentration okay so again concentration Solutions the normal solution would be at 0.9 or one percent the most okay so if the concentration inside the cell is three percent that's so high so water from outside will enter into the cell water moves into the cell and it may cause the cell to burst okay so if the cell wall is weak or damaged it would undergo plasmoptysis okay so it's a phenomenon undergone by bacterial cell where the cell swells and eventually it would burst okay uh young surface tensions okay then we have here hypertonic and solution so what happens what would happen okay as much as it can no the cell would try to balance the solution or the environment so it would contribute its water content to the environment no water would come out from the cell and would move into the environment in the hokna uh equilibrium is achieved okay however concentration so what happens is that right in the process of contributing water dehydrating adding cell however the unusual thing is that the cytoplasmic membrane would shrink away from the cell wall okay but the cell wall is uh it's not flexible it would not shrink it can only swell but it cannot shrink and so cell membrane from the cell wall shrink and cell membrane okay so this is what we call plasmolysis okay so plasma this is is when water from outside enters inside the cell causing the cell to swell and eventually burst that's plasma optices yeah we can see that if the environment is hypotonic okay now in hypertonic salabas compared saloom cell so what happens the cell would try to uh balance uh it would flush out its water into the environment so in the process it would cause dehydration causing the cell membrane to shrink away separate and Shrink away from the cell wall okay so there would be no communication there would be no growth membrane and C cell wall so there's no communication so the cell is uh neither growing nor undergoing processes right so this phenomenon is called plasmolysis and it's observed when the solution when the cell is in a hypertonic solution okay so I think the best type of solution to grow our organisms in would be isotonic okay so it is that would kill the organism or it would cause the organism to be in a state of a suspended animation okay so anion we have oxygen light carbon dioxide okay for energy right so energy oxygen uh uh trace and major elements water and salt and of course the pH okay so we need to be able to provide the right pH in order for organisms to grow in the laboratory okay so when we say pH or hydrogen ion concentration okay uh we're talking about the acidity or the alkalinity of a solution in which you have placed our organisms now most bacteria can survive a slightly acidic to neutral pH no so about 6.5 to 7.5 in some references it's uh more uh alkaline okay 7.2 to 7.6 okay um either way okay take note of this ranges okay so some organisms survive best at 6.5 okay until 7.6 right molds and yeast prefer acidic environment okay so five H five to six okay so those that prefer the acid environment are called acidophils a and then those that would require the neutral are called neutrophils no and then Alkali files alkaline environment okay and on to our last part for this topic we have part three bacterial growth okay so the learning outcomes for this part would be for you to be able to describe the different phases of the bacterial growth curve and for you to be able to synthesize the importance of the processes involved in culturing bacteria okay so uh bacteria divides uh through binary fission okay so Mother cell divides into two Doctor Cell so when we mean growth there's an orderly increase in the quantity of the cell constituents causing this the cell to divide into two uh equal you know and similar cells okay so binary fission simply occurs you know as the illustration shows you know so first step the cell would uh elongate and the DNA is replicated genetic material okay and then it starts to uh elongate you know it would stretch out to accommodate the size of the genetic material that's being replicated okay and then the start the cell wall and the plasma membrane would start to invaginate it would start to show the early beginnings of a division and then eventually cross walls will be formed you know and it would be uh uh it would be able to divide right the cell completely into two daughter cells simply language faces division non-human cells okay so DNA is replicated cell elongates uh invagination shows eventually uh a clear cross wall will be formed in between the two copies of the genetic material and two daughter cells will be formed now this is the bacterial growth curve it shows us the four phases of the bacterial growth first now we have the lag face lag followed by the log or the logarithmic phase or exponential or growth phase okay and then we have the stationary phase and then the death or logarithmic phase so the when the bacteria are grown in a closed system or in an environment that's artificial like in the laboratory or like in a test tube right so the population of cells almost always exhibits this growth Dynamics so cells initially adjust to the new medium you know that's the log phase so there's still no gross okay they do not yet divide as soon as they touch down on the gel okay or in your body perhaps okay so they are still adjusting you know during the long phase okay and then when they are familiar with the environment and they have assessed not the nutrients present they can start dividing regularly by the process of binary fission and so we have here the next phase the logarithmic phase no so when they start dividing you'll see here an increase no in the number of uh cells present in your culture media okay now when their growth becomes Limited the cells would stop dividing and so they enter the stationary phase here the number of organisms dividing is at a level or equivalent to the number of organisms okay eventually they show loss of viability on the gelatin media we observe this as a flat or dry colonies already glistening when they appear dry and flat that usually indicates organism so they've entered their deaths or uh logarithmic decline phase okay so again log face it's the stage where they're starting to get to know their environment so they are they haven't started dividing yet eventually when they are familiar with the area and have been acquainted with the nutrients present they'd start to grow via uh uh they start to multiply by by a binary session and so they enter the logarithmic or exponential growth phase and then they'd reach a point where the nutrients is almost depleted waste materials are almost you know um encroaching their growing space here we they enter their stationary phase so the number of organisms dividing is already equivalent to the norm the number of organisms dying eventually they would use up their nutrients and start to die okay and so they enter the death or the client phase curves now then we now uh binary fish on it occurs in a process called generation okay so each generation for organisms right so generation time is a Time taken for an organism to double its number or simply doubling time right so time required for a cell to divide uh from uh replication of the genetic material elongation of cell okay and then invagination then cross-all formation and eventually uh division of into two daughter cell that's one generation time so for example a single bacterium reproduces every 20 minutes in an hour so we know that in an hour there would be 60 minutes so tatlong generation time right okay so after one hour or three generation times is okay so first generation time meron dalawa okay and then the second generation time will also divide producing two uh producing four right four bacteria okay at on the third generation time u4 will be divided into eight microbes generation Computing number of organisms depending on their generation okay nice to know okay now what causes exponential growth to stop why is there the death or decline phase right so there are actually for reasons one is exhaustion of nutrients the nutrients are necessary for bacterial growth right secondly okay of course when they undergo metabolic processes and the use of nutrients there would be waste production as well and when waste accumulate within the environment where they are found that would be toxic to the organisms as well so it would contribute to the death of the organisms third there would be toxin production okay so toxin production may be due to an interaction of the chemicals now within the uh um environment okay or it could be produced by the organisms themselves or it could be because of the waste products present right and then all of these would lead to harmful change in PH okay the presence of the waste materials as well as the toxins can cause changes in the pH which would be harmful to the organism as well okay so again the reasons why there's a drop you know in the bacterial growth curve is because of exhaustion of the nutrients waste product accumulation toxin production and a change of the pH in the environment okay that ends our discussion on bacterial growth and classification you can use the following references to further your understanding of this topic of course again you're free to use any other references that you have aside from Mahon and Balian Scots now you can use the lost or jowitz okay and any other microbiology Books available thank you for listening