we are done with the first three parts of gram negative basilai the kollang natin part one are the small pleomorphic gram-negative bacilli part two are the non-fermenters negative bacilli and then partly human and gram-negative bacilli natin so the first uh the fourth part will be the gram-negative enteric basili otherwise known as the enterobacteriaceae now guys please take note now young lecturing attend for enterobacteriaceae will be composed of three parts as well so back part a part b in part c okay so we are going to first discuss the methods of identification the different biochemical tests and then the second part will be the opportunistic members of enterobacteriaceae and the new third part will be the two entities not two entities so expect the recordings for this topic i hope now i can record it today and post it by by tomorrow siguro but at least you will be able to watch it at your most convenient time and then on monday in the moon face to face available culture media for water and so hindi dao unfortunately hindi available okay device run away or in an activity not ready nothing going on wednesday so on monday anu monetario fill up discussion monetary of whatever activity we will be performing on wednesday okay so let's start with our gram negative enteric basilic once again this will be part a or the first part for this topic okay all right so family entitled bacteria can be divided into two groups we have the coliforms qualiforms or the lactose fermenters and the non-lactose fermenters so coliforms are the normal inhabitants of the colon they are mostly abundant in the intestinal tract these organisms under this division of enterobacteriaceae while the organisms the members under the division of non-lactose fermenters can be can be further divided into invasive organisms or opportunistic organisms soon invasive organisms nathan are salmonella shigella in yavchenia while the opportunistic snowman are the potatoes providencia morganella next okay so this table shows us the seven tribes under the family enterobacterishe and within each of these tribes are the different genus and of course the species of enterobacter okay so we have esc salmonella cytobacterium okay as the seven tribes and then please again okay look at the different genus under each of these tribes okay so next okay so the following are the general characteristics of enterobacteriaceae as a whole now so please take note of this very important tito first of course obviously the organ negative bacilli okay number two they are inhabitants of the intestinal tract except for your senior pestis except for your sinja pastis now they are facultative and aerobes okay all are glucose fermenters and they utilize glucose and aerobically all are glucose fermenters and they utilize glucose and aerobically in addition oil members can be juiced nightly to nitrite except for i think among one to two species i'll be showing later on so all are glucose fermenters and can be juiced nightly to nitrite they are catalyst positive except for pleshomonas members are oxidase negative they are oxidase negative in addition with a very important malimitable baseboard examination all members are motile political all members are motile with peditricus flagella so all members are motile except for shigella except for clibb sheila and except for yersinia okay another one is that all members are non-encapsulated all members are non-except encapsulated except for two okay except for two uh enchanted kingdom except for two we have entered so these two are encapsulated and the rest are non-encapsulated okay and lastly okay some organisms may tend to grow at lower temperature okay a colder temperature okay particularly one to five degrees okay so we have two under this okay sy school year we have sevashah india okay okay so please take note guys examination okay so please take note of the exemptions okay further by offer the general characteristics and features of the different enterobacter so this is another table showing us the biochemical characteristics of the tribes of enterobacteriaceae so we have here the seven tribes okay and the result in the different biochemical tests so once again what you can do is to take a print screen a screenshot of this table so you will know at least you will have an idea of what to expect or what results to expect for these different biochemical tests among h2s in vic reaction we'll be discussing those later so biochemical characteristics of the seven tribes of enterobacter okay next is the microscopic morphology okay so as what was mentioned they are short no shortsila negative okay or filament like okay they will appear like this short gram negative threads okay most are motile okay with pediclicus platula once again except for three we have the sky in terms of culture okay please take note that we can divide the cultural media into two okay you know the left side okay this already primary isolation cultural media primary isolation cultural media okay and then to to isolate specific pathogen okay we employ selective cultural media okay so once again primary isolation media autonomous selective okay so under primary we have three aucin methylene blue or emb no emb squeeze methylene blue is emb okay so agents inhibitory agents incorporated in the media is aucin y and methylene blue so ibix being an inhibitory they will not allow the growth of other microorganisms okay only enterobac okay each indicator ao syn y and methylene blue dissolve casings and primary carbohydrate primary isolation cultural media is lactose obviously we are to look for lactose fermenters so padding red and possible okay for e coli the positive result is green metallic shin and then for non-lactose fermenter of course colorless the second primary isolation culture media is mcconkie agar okay or mac mcconkie agar the inhibitory agents includes crystal violet and bile salts once again carbohydrate source is lactose ph indicator is neutral red positive result for lactose fermenter is pink or red and non-lactose are colorless and then we have the oxycolate citrate agar pedinan dc aditon acronym deoxycolith citrate agar so this contains sodium deoxycolate sodium citrate and fetic ammonium citrate once again lactose and carbohydrate sourcing ph indicator is neutral the positive result for lactose fermenters is spin colonies while negative or colorless so once again this is our primary isolation cultural media so ibxabihin our specimen will be first isolated or cultured or inoculated in this cultural media okay and then if we want to further study them and to isolate specific pathogen so we are going to employ our selective cultural media okay metaphor so we have hectoring entity agar orga okay cylo slicing the oxycolate angle or ssa okay so this three contains bile salts as their inhibitory agent however is a carbohydrate source lactose okay except for salmonella shigella agar but for heya my silos and sucrose for xldinam and my salis my salicin and sucrose page indicator for helia is spinal red xld blonde thymol blue while ssa is neutral lactose fermenters for hey and xlv are yellow colonies okay while ssanam and pink or red colonies and the non-lactose fermenters haya green or blue-green is red and ssa is colorless okay colorless so please take note of this so in the laboratory later on we will try to seguro selective um so the following are the biochemical tests we perform and employ to differentiate the different species of enterobacteriaceae so the first one is oxidase test as we all know oxidase test is a test to detect the presence of cytochrome c oxidase enzyme however for the members of enterobacteria shape please take note they are oxidized negative so what result are we expecting okay from an oxidase test for enterobacteriaceae no color change the bomb positive the sultan oxidized test is blue or purple color within 10 seconds after the application of the organism or of the sample okay so lithium for intel back oxidized test negative number two is nitrate or nitrite reduction so please take note that all species are positive for this test except for pantoeya agglomerants and members of armenia okay so you median again the gamete nut n4 um for nitrate reduction test is incorporated with 0.1 percent knot and the positive result for this is either azodi noted as diet next is carbohydrate utilization test we have two types of carbohydrate utilization tests unai untsi a iron agar or kia or kia no we have already discussed tsia last time okay i hope you still remember the principle of how and why we perform tsia okay so triple sugar epic sabine it contains three types of sugar okay so these three sugars are glucose lactose and sucrose don't forget also know the amount or the ratio among these three so glucose zero point one percent plan yen lactose sensors are both one percent in ten [Music] so we need to look at the slant portion in the back portion of psia there is glucose utilization meaning to say the end product is an acid and how do we deport a yellow slant we report it as a okay how about for example color then a color that young slant this means that the organism is not not a glucose fermenter okay but rather sorry the organism okay may not be a lactose or a sucralose fermenter or not a glucose experimenter is operating negative fermentation however it can be a protein utilizer okay so gradient slant it is a glucose fermenter and glucose fermenter so once again yellow acid and product we deport it as a what if naman for example vegeta like this this means that the organism is a non carbohydrate fermenter it will not use carbohydrate or utilize carbohydrate okay so what will be the reporting for this yellow slant in yellow but this will be reported as a a over psia also has the ability to detect the presence of h2s and of course gas gas formation okay the other type of carbohydrate utilization letter b is gligler iron agar or kia so like psia it also contains sugar however edito dalvolan is 0.1 percent of glucose and one percent of lactose with a ratio of 1 is to 10 pepo basically reporting parampara slang density sia also you will also look for the red color of the slant or the butt this will mean an alkalinity and then yellow color means acidity okay so what is the importance once again of psia so please take note now our our entire bacteria can be can be differentiated by their result in psia so we can group them into four okay so the first group are the k over a h2 is positive index [Music] construction can be grouped into four based on the result in tsia so first group are those with this result so k over e yellow okay h2 is positive the second group are the a over a so yellow yellow young now my black but h2s positive the third is k so red over a yellow and then the fourth group are the a over a okay yellow yellow while i'm blocking because h2s negative okay so four groups okay certainly over the member our non-entities or non-fermenters the bokeh over quesilla so members for each of this tsia grouping you know it indito tsi grouping are the following human organisms under this okay so later on we will look at the different or the other tests biochemical tests that will further differentiate each members of this tsia grouping [Music] so there is no other way to to study this but so trying to make in munich so that is so after identifying their tsia grouping or tsia the soul the next tests are and my invic pad reaction is composed of four tests okay namely we have indole methyl red okay vogue's postcard or vp test and citrate utilization test reaction is a test to detect if an organism can secrete the enzyme lipophage okay and if the organism can secrete the prophecies it can convert tryptophan into indul okay and this indole product will be detected using erlix reagent or covax regen okay why because this indole product will now react with aldehyde from these reagents to form the end positive result which is caller then so on gagovinatindito is of course inoculate our culture media by the way and cultural media ponatin for indole my so we can use strip to find agar okay we can use sim okay i believe you are familiar now with sim see a sulfide indulge motility agar and we can also use motility in dol or anything agar okay but among these three uncommonly nagina gamete is sim sim so if we inoculate our cultural media semi-solid so anchor up arduino inoculation is by stabbing the culture media we incubate it and then after 24 or 18 to 24 hours we place drops now drops off our earlix reagent or covax reagent and then wait for a color development once again the positive result will be a red link on top of the cultural media like this that means the organism is indole positive a negative number an absence of the red ring media okay that's indole in the second the action is methyl red test okay metal dead is used to detect the ability of an organism to produce and maintain stable acid end products not from glucose fermentation now please take note the sum that some bacteria produce large amounts of acids from glucose fermentation that they overcome okay the buffeting action of the system in short methylated test detects acid production via mixed acid fermentation pathway now mixed acid fermentation pathway okay a media point again a gamete not in detail it's the same media that we use in vogue's past tower so the mrvp media and the positive result no positively sold after after the addition of additional mrd agent or methylated agent they will show a red or pink color on top of the culture media so red or pink color the counter part of methylated test is vogus plus scour okay so while the metal bed test is useful in detecting mixed acid producers the vp test number detects butylene glycol producers okay butylene glycol producers okay so the positive test for vogue's plus tower okay once again is added or pink color on top of the cultural media and the last part of the in vic reaction is citrate utilization test so this detects the ability of the organism to utilize citrate as the sole source of carbon and energy okay the ph in ph indicator is utilization is the blue color edition of the cultural media or of the sitelit or siemens site re-tagger this is a chart not showing us the result of the invic reaction for the commonly isolated enterobacter shape so indole mrvpn cy3 so you can see godot take a screenshot of this [Music] uh you need to memorize it [Music] they are very good in memorizing biochemical test results of bacteria okay i just do not [Music] right before the internship my first same second sim for me internship and thank you later on before you you go for your internship try to review i'll try to scan your notes and your materials section or at least okay next next is uda's test so you these tests determines whether a microorganism can hydrolysiodia okay so the hydrolysis of urea will release a sufficient amount of ammonia that will now change the color or the ph indicator from an original orange color okay like this one being negative to a positive and positive uh pink red or magenta color okay by the way on cultural media nagina gamete is christian sense peace [Music] foreign the next is sugar fermentation and so this determines the ability of an organism to ferment a specific carbohydrate and the ph indicator for this is pinhole red no final dead and the media that we use is peptone peptone media incorporated with a carbohydrate so please take note once again all enterobacteriaceae can ferment glucose and that's why for sugar fermentation for members of enterobacter we are to expect it is salt like this yellow it was a non-carbohydrate fermenter another test is lysine the carboxylation so this detects the ability of bacteria to cleave under the liberation of carbon dioxide the end result or n product of this test is an alkaline alkaline product which will now turn the original color of the culture media color orange to a positive result of purple by the way the ph indicator of lysine the carboxylation test is brom block the sole purple okay number eight is phenylalanine the amines test or pad so this differentiates among the ud-positive gram-negative bacilli on the basis of the ability of the organism to produce penile pyruvic acid by oxidative the ammunition okay by oxidative the ammunition okay so detail after after inoculating after incubation my additional the agent in the gamete pina patagona indonesia a positive result will show a progression of light to dark green color or purple to black color okay within one to five minutes after the application of ten percent fed the chloride agent okay so this will now suggest no phenylalanine in the ammunition reaction okay with the addition of pericle died so if for example phenylalanine in the amines is present or is secreted by the organism so it will now first convert l-phenylalanine okay within one to five minutes after the addition of pedicle into color green okay ibxabihen pasitiv4 phenylalanine the amines okay positive results [Music] okay so once again this is a chart now is somebody showing us the tsia a test lysine iron agar a growth and result for the commonly isolated gram negative vented bacteria so slant and butt in psia whether or not they produce gas in h2s okay whether or not they are positive or negative in ud is inside the utilization or cycling test and they're the and then the invic reaction a sorry united [Music] view material you get 5.321.54321 next uh chart okay so uno once again psia visual for this organism so we have and four groups okay after tsia the ammunition test or the amines test so a positive punatin among these four is p now to further differentiate these three the an exponent is lysine the carboxylase test in the carboxylase test now to further differentiate these two nalysin a decarboxylase positive pad negative okay we can perform indole test sulfide indole motility positive proper sim is itarda negative is salmonella okay salmonella species so for example this is negative this is positive for indole lysine the carboxyl is positive negative for phenyl phenylalanine the amines test with k over a positive for h2s in psi is edward okay next autonomous shorter okay those with a over a h2s positive under tsia so we have three and what they use a cytobacter and then salmonella psychobacter from the only joint now there are some strains of cytobacter flundi that can produce a k over a result in the sia there are some a over a okay okay now to further differentiate this so we employ once again pad phenylalanine the amines test positive punatindito is p vulgaris negative this is positive for pad okay with an a over a h to s positive in the sia insago natin is so those with k over a h2s negative so mejia madame sila okay the next is to perform pad phenylalanine the amines okay so to further differentiate this now we can employ citrate utilization test okay so positive one at it for this is providencia once again negative one at informativity are shigella india the basket non-motile for example this is a a non-motile okay negative in pad with a k over e h to s negative result in tsi a and possible puna sagot not engine is either so we have those that has an a over a h to s negative in psi once again because some strains will now produce this result in psia some and will produce this result okay so next after tsia is invic reaction a very important in vic reaction because it will specifically differentiate e coli from those that belongs to the same group a in weekly action for e coli once again is pas pass okay the others are neg capacity for indole positive for mr negative for vp negative for sight rate utilization ayasha pass pass and vice versa for negneg pass is motility at 37 degrees once again negative potion non-motile clip and to further differentiate these two we employ the nh test remember our dna is test now from the first grading so positive one attend for dnac negative enterobacter okay so there's no other way not to to the view this but to at least memorize no um create some demonics [Music] presidency go to nothing going in the future as part of your destination so please do the view you can take sigurdo is a print screen or screenshot of this parameter material okay under biochemical characteristics are the enzyme tests so for this group of organisms so we employ dna test lipase and gelatinous test so please take note only sadasha only serasha are the only organism positive to all these three enzymes next okay the following are the antigenic marker no for our entire back that are detected using cytologic testing okay so number one we have the o antigen or otherwise known as somatic antigen a somatic antigen this is a type of endotoxin now it is a heat stable lipopolysaccharide okay and exists not in proteus species so we have o or somatic antigen the second okay is the h or flagellar antigen okay in contrast to o antigen this is hyplabyl now heat labyl this is found in this is i guess this is found only in motile organisms and are absent in non-motile organism such as the sky okay flagellarian so obviously this is found in members with capsule or are encapsulated okay like libcella and salmonella and some strains of e coli encapsulated then okay so these are the antigenic determinants once again tested and using several logic testing methods and lastly okay lastly are the different vivolence factors okay this video lens factors are the aid in the ability of the bacteria to produce disease or to cause infection so number one we have endotoxin a otherwise known as lipid a okay please take note that all members of the entire bacteria shape produce endotoxin now they produce endotoxin so this elicits the release of pyogenes or solid pyrogens from macrophage inducing fever okay the second is capsule okay for our encapsulated entire box okay number three exotoxin okay we have two types no enterotoxin and shiga toxin and enterotoxin and shiga toxin so these toxins acts on the small intestines causing accumulation of fluids and electrolytes in the lumen okay so basically number four is invasiveness okay invasiveness involves organisms or intestinal pathogens that are non-lactose fermenters now they are non-lactose fermenters like salmonella shigella and the arsenia and lastly type the secretion system so this is known as the major video lens a type desecration system please take note provides mechanism for the epic and the list of different metabolites and compounds from or by the bacterial cell okay that's why it is considered as the major vigilance factor so once again this vigilance factor aids the ability of the bacteria to produce disease or cause infection a cause infection okay i think that's the last slide okay for this part so the next part will be the opportunistic entire bacteria okay and then your next one will be the true enterics not true in this so any questions or clarifications