Microbiology Labs 1,2,3 Flashcards
What does aseptic mean?
Free from contamination
What is the aseptic slant technique?
You stick an inoculating spear into the sterilizing heat source. Then you stick it into your vial of microbes. Then you get a “slant” which is a vial with diagonal gel. Stick the spear all the way to the bottom of the slant, pull it out, and draw a squiggle on the top of the smear, slowing coming up towards the lid of the vial.
What is the aseptic streaking technique?
You get an inoculating loop and stick it into the sterilizing heat source. Then take your loop and swirl it around in your vial. Rake your loop gently over about 1/4th of your agar gel. Re-sterilize the loop. Rake gently across the next quadrant, initially overlapping to pull up some bacteria from the previous quadrant, but eventually diluting by no longer crossing. Cross very few times on the last quadrant. The goal is to dilute the bacteria so that individual colonies are grown from a single source cell.
In which cases do you need to crack the lid on a slant vial?
If the cells or the reaction requires oxygen, you need to have a slight crack in the lid. Without a crack you could have a sealed environment and oxygen will quickly be depleted. This can ruin your results, but you might not be able to tell that it’d happened, so then you’d make the wrong conclusions.
What is the catalase test?
Catalase is an enzyme (found in our peroxisomes, which are used to digest long-chain fatty acids and detoxify alcohol). Catalase reacts with hydrogen peroxide, turning it into O2 and H2O. If you have catalsae in your cell you will turn any available H2O2 into O2 gas and H2O. The O2 gas is what causes the characteristic bubbles of catalase tests. The test is used to distinguish between staphylococci (catalase-positive) and streptococci (catalase-negative).
What is the coagulase test?
Coagulase is an enzyme that turns fibrinogen (a zymogen in the blood used in clotting) into fibrin. Coagulase presence in a pathogen means it will often be coated in fibrin and thus more difficult to attack with antibiotics. The fibrin barrier proteins a pathogen from phagocytosis (getting eaten by macrophages/monocytes) and antimicrobial agents. The test is used to distinguish between staph aureus and staph epidermis.
What is the oxidase test?
Oxidase is used to find any cells that produce the enzyme cytochrome Oxidase C. This enzyme is present in any cell that uses oxygen as an electron acceptor in generating energy. A positive test will show a purple stain. It is used to confirm the presence of moraxella catarrhalis. Other compounds it can be used for include pseudomonas species and neisseria species
What is the sheep blood agar - hemolysis test?
It’s an agar gel coated with RBCs (I assume from sheep). The test is used to see if plated cells are partially, fully, or not hemolytic. A hemolytic pathogen is going to eat up/kill the RBCs, and you’ll see a clearing around the bacteria. You should be able to see clearly through the plate near the bacteria if the pathogen is fully hemolytic. There are 3 classes of hemolytic ability: alpha (partially hemolytic, the environment around the bacteria will seem sort of green), beta (fully hemolytic, the environment near the bacteria will be bright yellow or totally cleared of RBCs), and gamma (no hemolytic activity, no clearing or change in agar coloring). Be sure to check by individual colonies, since sometimes large bodies of bacteria generate waste/junk that might look like the wrong type of activity. This is a commonly used differentiating media.
Common alpha: streptococcus pneumonia, streptococcus viridans
Common beta: staphylococcus aureus, streptococcus pyogenes, non-group A beta-hemolytic streptococci
Common gamma: enterococcus faecalis, moraxella catarrhalis, staphylococcus epidermis
What is the optochin sensitivity test?
Also know as “P” disc. It’s a disc diffusion test that looks at bacterial resistance to optochin. streptococcus pneumonia is optochin sensitive (will have clearance) and other alpha-hemolytic streptococcus bacteria are resistant (the only other alpha hemolytic streptococcus we’ve learned about so far is streptococcus viridans).
What is the bacitracin sensitivity test?
Also known as “A” disc. It’s a disc diffusion test to see if bacteria are resistant or sensitive to bacitracin. Bacitracin is a selective test because it’s being used to inhibit growth of certain cell types. Can distinguish between Group A B-hemolytic streptococci (streptococci pyogenes) and any other type of b-hemolytic streptococci (non group a b-hemolytic streptoccoci)
What is the bile esculin test?
It’s used to distinguish between enterococcus faecalis and any other type of gamma-hemolytic bacteria (often streptococci). Esculin can be digested by enterococcus feacalis but not other gamma-hemolytic bacteria. Esculin breakdown creates iron-binding agents that make iron insoluble. The iron in the bile becomes insoluble and turns black. This is what stains the vial black. So if you get black in the gel it means you have a positive result and have enterococcus faecalis.
What are common gram positive pathogens?
All strep bacteria are gram positive
All staph bacteria are gram positvie
Most eneterococci bacteria are gram positive (eneterococcus faecalis, enterococcus faecium)
All clostridium bacteria are gram positive.
What are common gram negative pathogens?
All neisseria bacteria are gram negative.
Salmonella, shigella, proteus, providencia, serratia are gram negative
E coli is gram negative
What is a selective test media?
A selective test media is one that keeps some bacteria from growing while allowing others to grow. This is often used as first-pass testing, distinguishing between families of pathogens.
What is a differential test media?
A differential test media is one that visually distinguishes between two types of bacteria that both grew. Usually differential medias are for distinguishing between closely related species.
