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Microbiology test 1

Quiz yourself by thinking what should be in each of the black spaces below before clicking on it to display the answer.

Communicable disease   when an infected host can transmit the infectious agent to another host and establish infection in that host  
contagious.   Highly communicable diseases  
Non-communicable infectious diseases   do not arise through transmission from host to host.;occurs primarily when a compromised person is invaded by his or her own normal microflora;Can occur through direct contact with organism in natural, non-living reservoir  
how many of nosocomial infections could be prevented.   More than 1/3rd; 2 to 4 million cases/year in U.S. with approximately 90,000 deaths Most commonly involve urinary tract, respiratory tract, and surgical incisions Most common organisms involved: Gram-negative intestinal flora;E. coli, Pseudomonas, Staph  
Hand sanitizer   is a non-water-based hand hygiene agent. Active ingredient is usually isopropanol or ethanol Needs to contain at least 60% alcohol to be effective Added benefit of viral inactivation  
Antibacterial soap   most Triclosan-based (antibacterial compound) based: Many Triclosan resistant bacteria can be found in the environment Triclosan has no effect upon viruses Antimicrobial effects are concentration dependent High – biocide – multiple cytoplasmic and mem  
emulsification.   allows water to remove normally-insoluble matter  
Soaps Anionic surfactants   have both a hydrophilic end, which dissolves in water, as well as a hydrophobic end, which is able to dissolve nonpolar molecules like grease; emulsification  
Joseph Lister   father of antisepsis; cleaned using carbolic acid solution; stiches; sterile instruments could be left in patients; got idea from Pasteur,  
Ignaz Semmelweis   handwashing in Vienna General in 1846; puerperal fever in babies;  
Normal microbiotia or normal flora   More or less permanent residents of your body Actual numbers and types may vary over time Different people will have different distributions  
Transient microbiota or transient flora   Present only for days or weeks in/on the body Some of these are pathogens; i.e. bacteria that could colonize and cause an infection – resulting in disease  
Oceans   1 billion of microbial cells per liter of sea water  
Soil   There are more microbes in a gram of soil than there are humans alive today. There are 10,000 different species found in one gram of soil  
Humans   There are 100 trillion (1014) microbial cells in the gut. Ten times the number of human cells in an adult body Over 1,000 different species; prevalence and type vary from individual to individual  
Robert Hooke   reported that living things were composed of little boxes or cells  
Antonie van Leeuwenhoek   construction of microscopes which could magnify up to 30 - 100X Described microorganisms that he observed in teeth scrapings & rain water “animalcules”  
Germ Theory of disease   The theory that proposes that microorganisms are the cause of many diseases.; Pasteur and Koch  
Louis Pasteur   Disproved theory of spontaneous generation  
Robert Koch   Developed postulates that validated the germ theory of disease  
Spallanzani – 1700s   Experiment effectively disproved spontaneous generation Previous experiments performed in this manner were refuted in that “the boiling kills the life force” How did Spallanzani disprove this? Critics then argued “air” was required for spontaneous gen  
Why was Koch successful?   Developed techniques for culturing microorganisms  
Agar   Not degradable by most microorganisms Liquefies at 100°C Solidifies at temps under 40°C Once solidified, can be incubated at temps up to 100°C  
Streak plate   Most common Determines the “what” is in the culture  
Serial dilution and plating   Determines the “how much” is in a culture Used in conjunction with spread plating Expressed in terms of cfu/ml Colony forming units per milliliter of culture  
Pipetman   Dual stop: should go down to 1st stop to take up sample and should go to 2nd stop to expel sample P-1000 100 – 1000 µl sample transfer P – 200 20-200 µl sample transfer P- 20 2-20 µL sample transfer  
cfu/ml   Colony counts are expressed from serial dilutions;Colony forming units/ml Each colony represents outgrowth from a single cell, but an individual colony is made up of millions of bacterial cells!  
Spot plating: TNTC   too numerous to count More than 30 colonies per spot  
Spot plating: TFTC   too few to count Less than 5 colonies per spot  
Spread plating   more sensitive than spot plating; more expensive as you need more plates  
Spread plates: TNTC   more than 250  
Spread: TFTC   less than 25  
Spread plate   Formula: #colonies x reciprocal of dilution x (1000/amt plated)This accounts for your µL to mL conversion  
Spot Plate   Formula: #colonies (average of the 3 spots)x reciprocal of dilution x (1000/amt plated)This accounts for your µL to mL conversion  
UTIs   only considered an active infection if plated cfu/mls exceed 105  
Cfu/ml numbers can be used in the following:   Estimate the amount of protein and or DNA based upon viable counts in a population for research purposes  
most compound light microscopes   have 3 or 4 objectives mounted on the nosepiece directly over the stage  
longer the lens...   greater the mag  
Magnification is   limited by resolution  
Resolution   Ability to see two objects as separate and distinct  
resolution depends upon   refraction (scattering) of light  
Refraction can be limited by:   Light wavelength Shorter wavelengths = better resolution most scopes employ a blue filter (short wavelenght) Numerical aperture (NA) of the lens The ability of the objective to capture scattered light Related to curvature of the lens  
Glass and air   different refractive indexes;- Some light is “lost” before it makes it to the objective  
Glass and oil   almost identical refractive indexes;-More light makes it to the objective = less refraction, therefore better resolution Use of oil immersion increases magnification approximately 2.5-fold  
oil immersion lens   only fine adjustment; should be able to see seal  
As you switch from low to high power   the field of view becomes darker.   To deal with this the iris diaphragm needs to be opened to allow more light into the condenser.  
As you switch from low to high power   the field of view becomes smaller.  
Parfocal   in focus with one lens – in focus with all (in theory) – higher magnifications may require slight adjustments  
Brightfield microscopy   Objects are dark and the “field” is light  
Stains   Are chromophores – charged ions Acidic (-) or basic (+)  
Direct/simple   stains organism; uses a basic stain  
Negative/indirect   stains field or background; uses an acidic stain Based upon the inherent properties of the negatively charged bacterial membrane and chromosome  
What do we stain?   smear  
what do we need?   clean slide  
Why do you fix organism to slide?   to increase adherence; to increase stain uptake; kill bacteria  
Negative Staining   Stains background not bacteria. Ionic Repulsion – like charges repel  
Pros of Negative Staining:   Bacteria less distorted- does not require heat fixation Can display cell morphology and size more clearly than other staining.  
Primary stain   Imparts color to all cells  
Decolorizing agent   May or may not remove the primary stain from the entire cell or certain cell structures  
Counterstain   Provides color contrast to primary stain  
Gram positive   peptidoglycan 90% of cell wall;  
Gram negative   peptidoglycan in 10% of cell wall; cells have outer lipolysaccharide layer (LPS)  
Primary stain   Crystal Violet (1˚) dissociates into CV+ and Cl- ions  
Iodine   (mordant) makes CV-I complexes  
. Alcohol (decolorizing agent)   washes away outer layer of Gram- along with CV-I. CV-I in gram+ cells stays in layer of peptidoglycan so stays PURPLE.  
Safranin (2˚)   gives gram- its PINK color.  
Gram Positive characteristics   thick layer cell wall peptidoglycan; low lipid (1-4%); quite susceptible to penicillin; complex for many species (pathogens); more resistant to physical disruption  
Gram Negative characteristcs   Think layer cell wall peptidoglycan; high lipid (11-22%); less susceptible to penicillin; simple; less resistant to physical disruption  
Problems with gram staining   Some bacteria will not Gram stain (Mycobacterium, Mycoplasma, Nocardia spp. to name a few examples) Accurate staining is often dependent upon culture age (18-24 hr cultures give best results) Some bacteria give variable results – (Gram – v) Stressed ba  
some bacteria contain what which makes them impermeable to staining procedures   mycolic acids  
Acid fast bacteria   Resist decolorization with acid alcohol Carbol fuschin contains phenol which solubilizes traditional cell walls Methylene blue serves as a counter stain  
Steam heat   is used to facilitate the uptake of the carbol fuchsin by the acid fast bacteria  
go over slides in the powerpoints   slides 24-28  


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