Lab Exam 1 Flashcards
Place one drop of water in the center of the slide, transfer small amount of bacterial inoculum from the culture, spread into the size of a nickel, Air dry, heat fix.
Preparation of bacterial smears from solid medium
Wet mount
Determine how to do a wet mount
Let slide dry, heat fix, apply stain (1 to 2 minutes), gently wash with tapwater, blot slide with bibulous paper
Simple staining Crystal violet 20 to 60 seconds
Place a drop of Negrosin at one end of the slide, place loop full of inoculum in the drop, mix, place slide against the drop at 45° angle and spread, Air dry.
Negative staining
Prepare a slide (Inoculum, water, smear, heat fix). Flood slide with crystal violet. One minute. Wash with tapwater (Still purple). Flood slide with gram’s iodine. One minute. Wash with tapwater. Decolorize with ethyl alcohol (Almost clear). Wash with tapwater. Counterstain with saffranin for 45 seconds. Wash with tapwater. Blot dry.
Gram staining
The bending power of light passing through air from glass slide to objective lens
Retractive index
When one lenses and focus, the other lenses will also have the same focal length, and can be rotated into position without further major adjustment.
Parfocal
Vibratory movement of the cells due to their bombardment by water molecules in the suspension
Brownian movement
Pair of cocci
Diplococcus
Chain of cocci
Streptococcus
Cluster of cocci
Staphylococcus
Packet of four cocci
Tetrad
Packet of eight cocci
Sarcina
Rod shaped, pair
Diplo bacillus
Rod shaped chain
Streptobacillus
Spiral bacteria, curved rods
Vibrios
Spiral, helical and Rigid
Spirilla
Spiral bacteria, helical and flexible
Spirochetes
Two Loopfulls of the cell suspension, spread suspension, allow the smear to air dry, heat fix.
Preparation of bacterial smears from broth medium
Requires the use of at least four chemical reagents that are applied sequentially to heat fixed smear
Differential staining
Imparts it’s color to all cells, Crystal violet
Primary stain
Killing agent, increases the cells affinity for a stain, Used to intensify the color of the primary stain, grams iodine
Mordant, Crystal – Violet – iodine complex CV –I
Protein dehydrating agent and lipid solvent. In gram-negative cells, the alcohol increases the porosity of the cell wall by dissolving the lipids in the outer layer. CV – I complex is washed out. Gram-positive cells retain CV – I complex do you to thicker peptidoglycan. May or may not remove the primary stain from the entire cell or for only certain cell structures.
Decolorizing agent, ethyl alcohol
Contrasting color to that primary stain. Gram-negative.
Counterstain, safranin
Divides bacterial cells into two major groups, gram-positive and gram-negative
Gram stain
Malachite green
Used to stain free spores with impervious coats
Used to isolate specific groups of bacteria. They incorporate chemical substances that inhibit the growth of one type of bacteria while permitting growth of another, thus facilitating bacterial isolation.
Selective media (Phenylethyl alcohol auger, crystal violet auger, 7.5% sodium chloride auger)
Distinguish among morphologically and biochemically related groups of organisms.
Differential/selective media (macconkey agar, mannitol salt auger (staphylococci), eosin-methylene blue auger)
7.5% sodium chloride selects for staphylococci
Mannitol salt auger (differential selective)
Crystal violet allows the isolation of gram-negative bacteria by inhibiting gram-positive organisms
Macconkey agar (differential selective)
Lactose and dyes permit differentiation between enteric lactose fermentors and non-fermenters as well as the identification colon bacillus E. coli.
Eosin-methylene blue auger (differential/selective media)
Highly nutritious Materials for fastidious organisms. Blood, serum, or yeast extract.
Enriched media
Permits the demonstration of hemolytic properties particularly streptococci
Blood agar (enriched media)
No lysis of red blood cells
Gamma hemolysis, enriched media, blood agar
Incomplete lysis of red blood cells, greenish halo around bacterial growth.
Alpha hemolysis, enriched media, blood agar
Lysis of red blood cells, clear zones surrounding the colonies.
Beta hemolysis, enriched media, blood agar
Non-antigenic oxygen stable lysin
Streptolysin S
Use both aerobic and anaerobic pathways, fermentors of carbohydrates, degradation of glucose by the way of the glycolytic pathway
Facultative anaerobes
Inverted vile for the detection of gas production in fermentation
Durham tube
Design to differentiate among different groups of Enterobacteriaceae. All gram-negative bacilli capable of fermenting glucose with the production of acid and Gram-negative intestinal bacteria. (Differences in carbohydrate fermentation patterns and hydrogen sulfide production).
Triple sugar – iron auger test
Aerobic and anaerobic
Cellular respiration
Bioxidations in which molecular oxygen can serve as the final electronic acceptor
Aerobic
Bioxidations in which inorganic ions other then oxygen such as nitrate (no3-) or sulfate (so4 2-) can serve as the final electronic acceptor
Anaerobic
A bioxidative process not requiring oxygen in which an organic substrate serves as the final electronic acceptor, produces an organic acid that may be accompanied by gases, facultative anaerobes are usually fermenters of carbohydrates
Fermentation
Nutrient broth ingredients for all organisms, specific carbohydrate, pH indicator
Carbohydrate fermentation medium
Fermentation
Net two ATP, alcohol, lactic acid (in animals and some organisms)
One glucose to two pyruvic acid to Krebs cycle
Glycolytic pathway
Lack of carbohydrate fermentation. Alternative nutrients can be used as energy sources. These reactions liberate ammonia producing an alkaline environment. Phenyl red turns deep red in it now basic medium indicates aerobic respiration.
Peptones
Alkaline slant (red) and acid butt (yellow) with or without gas production (breaks in the auger butt)
Results of triple sugar iron auger test, glucose fermentation only, peptones used in the production of alkali
Acid slant (yellow) and acid butt (yellow) with or without gas production.
Results of the triple sugar iron Auger test, lactose and or sucrose fermentation
Alkaline slant (red) and alkaline butt (red) or no change (Orange – red) butt
Results of the triple sugar iron auger test, no carbohydrate fermentation, peptones catabolized under anaerobic and or aerobic conditions create ammonia.
A substrate for hydrogen sulfide production, blackening in the butt because of the precipitation of the insoluble ferrus sulfide
Sodium thiosulfate in TSI auger, indicates the production of h2s.
Differentiation of the principal groups of Enterobacteriaceae.
Indole, methyl red, voges – proskauer and citrate utilization
Imvic
Determines the ability of microorganisms to degrade the amino acid tryptophan
tryptophanase and Indole production test
Detects indole, cherry red reagent layer
Kovacs reagent
Detect the presence of a large concentration of acid end products, ph 4, indicative of E. coli. PH 6 indicative of e. Aerogenes
Methyl red test
Differentiates further along enteric organisms such as E. coli, E. Erogenes, and K. pneumoniae. Determines the production of nonacidic or neutral end products
Voges-proskauer test
Barrett’s reagent, pink complex formed imparting rose color to the medium, indicates acetylmethylcarbinal
Voges-proskauer test
an organism can use citrate has its sole source of carbon. Indicates the presence of citrate permease, medium becomes alkaline, the carbon dioxide that is generated changes bromthemol blue indicator from green to Prussian blue
Citrate utilization test
