Lecture 1 Flashcards

Question

antibiotics as selective media

Answer 1

very heat sensitive * prepare aseptically must be added to media after autoclaved and cooled CNA: * has Colistin & Nalidixic acid which allows gram positive organisms to grow while preventing the growth of most gram negative NYC: * has numerous antibiotics to try and grow only N. gonorrhoea – See later slide for specific antibiotics and which organisms are killed

Answer 2

Control pH fluctuations in media due to bacterial metabolism * Peptones * Inorganic phosphates: buffer and serve as a source of phosphorous

Answer 3

Absorb toxic substances * inhibit the growth of fastidious organisms -Transport media, BCYE (Buffered Charcoal Yeast Extract agar)

Answer 4

Simple media, only barest amount of nutrients water, agar, simple proteins Fastidious organisms will not grow No differential or selective properties Trypticase soy Agar * Nutrient Agar * Brain Heart Infusion Agar * Columbia Agar Base * Mueller Hinton Agar

Answer 5

-Basic media + extra nutrients like vitamins or blood -usually a broth -Brilliant green, selenium salts (feces) or tetrathionate suppress growth of normal flora for a short period of time Selenite broth, Tetrathionate broth, Brain Heart Infusion broth, Cooked meat broth

Answer 6

Has ingredients that allow you to tell one bacteria from another when you have mixed bacteria growing.

Answer 7

A Basic media for susceptibility testing (AST) of non fastidious organisms * Has proteins in the form of extracts & enzyme hydrolysates * Has starch to absorb toxins ,helps with antibiotic diffusion & has dextrose for energy * Agar is loose for better diffusion of the antibiotics * Shows batch-to-batch reproducibility for AST testing * Is low in sulfonamide, trimethoprim, and tetracycline inhibitors like thymidine and thymine can add sheeps blood to make more selective

Answer 8

Enriched because has 5-10% Sheep blood that has been hemolysed by heating to 800C or by adding an enzyme – agar looks brown like chocolate because has 2% hemoglobin + supplements * Grows most organisms but used especially, to grow more fastidious microorganisms * Not a differential media because you can’t see whether hemolysin enzyme produced by bacteria- blood already hemolysed you cannot see homolysis on this plate because the blood is already hemolysed

Answer 9

-General-purpose, enriched, differential media -Enriched with hemin and vitamin K to grow fastidious & anaerobes. * Differential because you can see O2 requirements of what is growing * Some have redox resazurin dye to monitor levels of O2 – turns pink in O2 when growing aerobes the colonies will be at the top - highest oxygen anaerobic - at the bottom of the test tube facaltative anaerobes - everywhere but concentrated on the top

Answer 10

CNA Enriched because has 5-10 % whole sheep blood * Selective because has antibiotics Colistin and Naladixic acid that will inhibit gram neg growth * Allows gram positive organisms and yeast to grow * Differential because allows you to see which organisms can hemolyze RBC

Answer 11

selective Ingredients: * Bile salts * Crystal violet * Eliminate Gram positive & grows gram negative Differential ingredients: * Lactose (carbohydrate) * Neutral Red (pH indicator) * Lactose fermenters (LF) – bright pink colonies * Non-lactose fermenters (NLF) – colourless colonies

Answer 12

Selective differential agar for Staphylococcus aureus Selective ingredients: * A high concentration of salt 7.5% (halophilic) Differential ingredients: * pH Indicator: Phenol red (yellow in acid, pink/red in basic) * Carbohydrate: Mannitol * Differentiate between pathogenic and nonpathogenic Staphylococcus * S. aureus grow and ferment mannitol – yellow * Other Staph species grow but do not ferment mannitol – pink

Answer 13

Selective and differential media * Selective for gram negative – inhibitory to gram positives * Same ingredients as MAC except sorbitol instead of lactose * Differentiates between entero-hemorrhagic Escherichia coli (EHEC) which fail to ferment sorbitol and Escherichia coli which will ferment sorbitol and do not cause diarrhea Sorbitol Fermenters = PINK

Answer 14

Selective & differential agar for enteric (intestinal) pathogens Selective ingredients: * Bile salts + sodium citrate + brilliant green * Inhibits all gram-positive & gram-negative that are normal flora * Highly selective for enteric pathogens - that cause food poisoning Differential ingredients: * Lactose and neutral red (pH indicator) * Differentiate between LF and NLF * Sodium thiosulphate + ferric ammonium citrate * Differentiate between H2S and non-H2S producing colonies

Answer 15

▪ hydrogen sulfide reacts with ferric ions from the ferric ammonium citrate to make an insoluble heavy metal black precipitate ▪ H2S positive result is seen as a blackening of the center or entire colony LF, H2S positive Pink colonies with black centres NLF, H2S positive Clear colonies with black centres NLF H2S negative Clear colonies

Answer 16

Selective and differential agar used to isolate enteric pathogens Selective ingredients: * Bile salts inhibit gram positive organisms Differential ingredients: * sugars- lactose, sucrose and salicin * Indicator: Bromthymol blue + acid fuchsin * Na thiosulfate and ferric ammonium citrate in the medium produces a black precipitate in the presence of H2S fermenters that are H2S postive - orange/yellow colonies with black center non fermenters with h2s postive - green with black centers fermenters with NO H2S -orange/yellow non fermenter with no h2s blue green colonies

Answer 17

Selective and differential media specifically used to isolate the enteric pathogen Yersinia enterocolitica Selective ingredients * Cefsulodin, Irgasan, Novobiocin, Na desoxycholate & crystal violet all inhibit gram positive organisms * Also inhibit gram negative that are normal flora in stool Differential ingredients * Sugar = Mannitol & Neutral red pH indicator * mannitol fermenters -pink with clear edges * Look like a bulls eye

Answer 18

Selective enriched media for Campylobacter – is microaerophilic Enriched ingredients: * 10% sheep blood Selective ingredients: * Vancomycin inhibit gram positive organisms * Amphotericin B & Polymyxin B inhibit yeast/fungus * Trimethoprim inhibits a swarming gram negative called Proteus * Cefoperazone inhibits gram negative organisms * Na bisulfite lowers the O2 levels enhancing the recovery of microaerophilic organisms

Answer 19

Selective enriched media used specifically for the isolation of Neisseria gonorrhoea and Neisseria meningitidis Enriched ingredients: * Has hemoglobin from lysed horse blood, yeast dialysate & horse plasma Selective ingredients: * Vancomycin inhibits gram positive * Colistin inhibits gram negative * Amphotericin B inhibits yeast * Trimethoprim inhibits swarming Proteus

Answer 20

has soluble colourless molecules called chromogens * Chromogens consist of a substrate and a chromophore * If the target organism is present the specific enzyme it makes cleaves the colourless chromogenic conjugate, releasing the chromophore * When the chromophore is not in the conjugated form (unconjugated) it exhibits it’s distinct color which precipitates on the colony advantages - Provides a color based differentiation * Clearly distinguishable * Colonies of specific microorganisms can be recognized by their colour

Answer 21

Expiry date, lot numbers * Physical appearance: depth of agar (4mm), cracks, excessive bubbles, contamination * Performance: test with known ATCC strains to ensure proper growth and appearance

Answer 22

RBC are destroyed by exotoxins or enzymes produced by the bacteria as they grow on media only report on (BA, CNA) NOT choc * Causes complete breakdown or lysing of RBC cell membrane * Seen as a clear zone (media becomes see through) around and under the colony Some bacteria produce multiple toxins or enzymes and have larger zones of beta hemolysis * Some produce haemolysins that are oxygen labile-only active in decreased O2 Clostridium perfringens is a strict anaerobic** organism Called double zone beta hemolysis Theta & alpha toxin staph aurea

Answer 23

partial breakdown of RBC * Bacteria produce hydrogen peroxide which oxidizes hemoglobin to methemoglobin * Result is a green zone in the media around and under colony BUT THE COLONIES ARE GREY * Seen more intensely on the CNA plate * Streptococcus pneumoniae

Answer 24

innie”, “like a checker piece” ^-^ * Streptococcus pneumoniae * tilt plates to examine

Answer 25

“outie” Viridans streptococcus _^_

Answer 26

pinpoint, small , medium and large

Answer 27

white yellow tan grey red

Answer 28

Pseudomonas (a strict aerobe), is a NLF gram negative organism only for this organism

Answer 29

Scant Light Moderate Heavy depending on quadrant

Answer 30

based on cell wall structural differences of GN GP infections caused by a single organism = monomicrobial * by multiple organisms = polymicrobial

Answer 31

Thick, peptidoglycan layer (90%) * Many teichoic acid cross-linkages * Lower lipid content , 0-2% * No periplasmic space * Resists decolourization

Answer 32

Thin peptidoglycan layer (5-20%) * No teichoic acid cross-linkages * Higher lipid content 10-20% * Have both an outer and inner membrane * Has periplasmic space * Does not resist decolourization

Answer 33

Basic stain so it binds to the negatively charged groups in the cell with ionic bonds (po4) * Diffuses through cell wall pores & bonds with negatively charged cell groups (e.g. phosphate groups in nucleic acid) -water does not decolorize

Answer 34

Diffuse into the cell * Combines with crystal violet forming crystal violet-Iodine (CVI) complex * Traps crystal violet making it hard for it to leave the cell * CVI complex is stable in water but soluble to solvents such as alcohol & acetone TRAPPING AGENT

Answer 35

Most important step …. Timing is critical * ethanol and acetone * Extracts lipid creating larger pores in cell wall makes it more permeable * Dehydrates peptidoglycan reducing pore size & movement In GN cell wall: * high lipid, low peptidoglycan * Lipids extracted and CVI complex leaves cell In GP cell wall: *low lipid, high peptidoglycan * Peptidoglycan dehydrated and CVI complex trapped Looking under the microscope at this point: GN will look colorless and GP will appear bluish purple

Answer 36

can also use dilute Carbolfuchsin * No effect on GP as binding sites were blocked by CVI complex * Forms bonds with cell groups in GN cell wall * GP are bluish purple and GN are pink

Answer 37

Keep petri dishes closed when possible Clean and disinfect lab surfaces prior to and after use Avoiding talking/breathing on cultures Use a face shield when procedures create aerosols Wash your hands & wear gloves & lab coat Open packages in a manner where you do not touch the parts that will come in contact with your sterile media Sterilize inoculating loops and other equipment that comes into contact with cultures

Answer 38

white(staph) gold/yellow staph aur , grey - shiny on the edges - strep smaller colonies - CNA

Answer 39

coccobacilli - negative

Answer 40

always either large or small (small chinese letter paliside)

Answer 41

football or cat eye shape in gram pos only cant have gram neg

Answer 42

to determine the antibiotic type can be used to differentiate and quntitate inflammatory and epithelial cells first test in culture test

Answer 43

transparent

Answer 44

differntial stain because it uses two dyes and seperates two types of bacteria

Answer 45

decolorization time was too short gram negs appear blue

Answer 46

decolorization time too long gram pos are red can happen if the cells are too old