Micro lab final

• Sulfur reduction
• Indole production from tryptophan
• Motility

• It is used to identify bacteria that are capable of producing indole, using the enzyme tryptophanase.
• It is used to differentiate sulfur-reducing members of enterobactericaeae, especially membrs of the genera salmonella, francisella, and proteus from the negative morganella morganii and providencia rettgeri.
• Motility plays a big part because it is an important differential characteristic of Enterobacteriaceae.

The triple sugar iron agar (TSIA) is a rich medium designed to differentiate bacteria on the basis of glucose fermentation, lactose fermentation, sucrose fermentation, and sulfur reduction.

• Glucose and lactose fermentation with acid accumulation in slant and butt (if experiment was done in a kliger iron agar), or glucose, lactose and/or sucrose fermentation with acid accumulation in slant and butt (if experiment was done in a triple sugar iron agar).
• Result: Yellow slant/yellow butt

• Glucose fermentation with acid production. Proteins catabolized aerobically (in the slant) with alkaline products (reversion).
• Result: Red slant/yellow butt

• No fermentation. Peptone catabolized aerobically and anaerobically with alkaline products. Not from enterobacteriaceae.
• Result: Red slant/red butt

• No fermentation. Peptone catabolized aerobically with alkaline products. Not from enterobacteriaceae.
• Result: Red slant/no change in butt

• Organism is growing slowly or not at all. Not from Enterobacteriaceae.
• Result: No change in slant/no change in butt

• Sulfur reduction. (An acid condition, from fermentation of glucose or lactose existsin the butt even if the yellow color is obscured by the black precipitate.)
• Result: black precipitate in the agar

Cracks or lifting in the agar indicates gas production.

• Includes 5% sheep blood in a tryptic soy agar base
• Several species of Gram-positive cocci produce exotoxins called hemolysisn which are able to destroy RBCs and hemoglobin.
• Blood agar allows differentiation of bacteria based on their ability to hemolyze RBCs

Three major types of hemolysis are: B-hemolysis, a-hemolysis, and y-hemolysis.-

Application: Blood agar is used for isolation and cultivation of many types of fastidious bacteria. It is also used to differentiate bacteria based on their hemolytic characteristics, especially within the genera streptococcus, enterococcus, and aerococcus.

AKA Kirby Bauer Test and Disk Diffusion Test

• A clear zone that appears around the disk where bacterial growth has been inhibited. The paper disks were “impregnated” with antimicrobic agent. The antimicrobic drug establishes a concentration gradient around the paper rings as it moves away from the rings, and that is why bacterial growth can’t take place close to the paper, b/c of the antimicrobic drug, hence the “clear zone.”
• The size of the zone of inhibition depends on the sensitivity of the bacteria to the specific antimicrobial agent and the point at which the chemical’s minimum inhibitory concentration (MIC) is reached.

• In this test, measures whether or not an organism/bacteria is “susceptible” or effected by the antimicrobial.
• Antimicrobial susceptibility testing is a standardized method that is used to measure the effectiveness of antibiotics and other chemotherapeutic agents on pathogenic microorganisms. It is also sometimes valuable in prescribing treatment.

• Resistance mechanisms can be broken down into three main categories:
• (a) altered target such that the antibiotic no longer can interact with the cellular process,
• (b) an alteration in how the drug is taken into the cell and
• (c) enzymatic destruction of the drug.

• Prolonged exposure to UV rays can be lethal to cells because when DNA absorbs UV radiation at 254 nm, the energy is used to form new covalent bonds between adjacent pyrimidines=
• cytosine-cytosine, cytosine-thymine, or thymine-thymine.
• These are known as pyrimidine dimers, with thymine dimers being the most common. These dimers distort the DNA molecule and interfere with DNA replication and transcription.

• E. Coli have the repair mechanisms:
• Light repair or photoreactivation
• Excision repair or dark repair

E.Coli performs light repair or photoreactivation, in which the repair enzyme, DNA photolyase, is activated by visible light 9240-400 nm) and simply monomerizes the dimer by reversing the original reaction.

• Excision repair or dark repair involves a number of enzymes.(look at fig 10-12 in lab manual.)
• The thymine dimer distorts the sugar phosphate back bone of the strand.
• This is detected by an endonuclease that breaks two bonds- eight nucletides, in the 5’ direction from the dimer, and the other four nuletides in the 3’ direction.
• A helicase removes the 12-nucletoide fragment (including the dimer), leaving single-stranded DNA. DA polymerase I inserts the appropriate complementary nucleotides in a 5’ to 3’ direction to make the molecule double-stranded again.
• Finally, DNA ligase closes the gap between the last nucleotide of the new segment and the first nucleotide of the old DNA, and the repair is complete.

• Neutrophils
• Basophils
• Eosinophils

• Monocytes
• Lymphocytes

55%-65%

25%-33%

3%-7%

1%-3%

0.5%-1%

• Most abundant WBCs in blood.
• They leave the blood nad enter tissues to phagocytize foreign material.
• An increase in neutrophils in the blood is indicative of a systemic bacterial infection.
• Mature neutrophils sometimes are referred to as segs because their nucleus usually is semented into two to five lobes.
• Because of the variation in nuclear appearance, they also are called polymorphonuclear neutrophils (PMNs). Immature neutrophils lack this segmentation and are referred to as bands.

• Basophils are the least abundant WBC in normal blood.
• They are structurally similar to tissue mast cells and produce some of the same chemicals (histamine and heparin) but are derived from different stem cells in bone marrow.
• They are 12-15 um in diameter.
• The nucleus usually is obscured by the dark-staining cytoplasmic granules, but it either has 2 lobes or is unlobed.

• Eosinophils are phagocytic, and their numbers increase during allergic reactions and parasitic infections.
• They are 12-15 um in diameter (about twice the size of a RBC) and generally have two lobes in their nucleus. Their cytoplasmic granules stain red in typical preparations.

• Monocytes are the blood form of macrophage.
• They are the largest leukocytes, being two to three times the size of RBCs at 12-20 um.
• Their nucleus is horseshoe-shaped, and the cyptoplasm lacks prominent granules, but may appear finely granular.

• Lymphocytes are cells of the immune system.
• Two functional types of lymphocytes are the T-cell, involved in cell-mediated immunity, and the B-cell which converts to a plasma cell when activated and produces antibodies.
• The nucleus usually is spherical and takes up most of the cell.
• Lymphocytes are approximately the same size as RBCs or up to twice their size.
• The larger ones form a third functional group of lymphocytes, the null cell, many of which are natural killer (NK) cells that kill foreign or infected cells without antigen-antibody interaction.

Dimorphic fungi have both mold and yeast life-cycle stages

• A polysaccharide that makes up the cell wall of members of the kingdom fungi.
• A tough, protective, semitransparent substance, primarily a nitrogen-containing polysaccharide, forming the principal component of arthropod exoskeletons and the cell walls of certain fungi.

Heterotrophs that decompose dead organic matter

Individual fungal filaments, tube-like extensions of the cytoplasm

Hyphae in which walls separate adjacent cells (septums)

Walls are absent in hyphae

The two typical life-cycle stages of protazoans include a vegetative trophozoite stage and a resting cyst stage. Some have additional stages, making their life cycles more complex.

• Disease: Amoebic dysentery (amebiasis) a disease most common in areas with poor sanitation.
• Transmission: Infection occurs when a uman host ingests cysts, either through fecal-oral contact or, more typically, contaminated food or water. Cysts are able to withstand the acidic environment of the stomach. Upon entering the less acidid small intestine, the cysts undergo excystation. Mitosis produces eight small trophozoites from each cyst.
• Presence/absence of flagella: absent

• Disease: Malaria
• Transmission: Vector, the female anopheles mosquito
• Presence/absence of flagella: absent

• Disease: Giardiasis
• Transmission: typically involves fecally contaminated water or food, but direct fecal-oral contact transmission is also possible.
• Presence/absence of flagella: Present. Most commonly has four pairs of flagella and a sucking disk.

Causes the disease clonorchiasis, a liver disease, common in Japan, Korea, Vietnam, cina, and Taian, and becoming more common in the US with the influx of SE Asian immigrants.

Common parasite of dogs and cats, human infection usually occurs in children, The adult worms reside in dog/cat intestines and releases proglottids ontaining egg packets that migrate out of the anus and look like rice grains. Diagnosis made by ID of the egg packets. (didn’t list a specific name of the disease it causes, so maybe it’s just worms)

• Echinococcus granulosus: Leads to the development of a hydatid cyst in the lung, liver, or other organ, this process may take many years.
• Causes Hyatid Disease

Biofilms occur in natural environment, but are also formed in industrial and medical settings, which is a big deal. Indwelling devices like needles and catheters, are common locations for biofilm development. Reduced susceptibility of the biofilm community to antimicrobics (by a factor of 100-1000 times compared ot their planktonic counterparts) with natural detachment make these biofilms problematic in the production of nosocomial infections.

Staph aureus and S. epidermidis are notorious for forming biofilms on invasive medical devices resulting in nosocomial infections.

• Urine culture is a common method of detecting and quantifying urinary tract infections.
• It frequently is combined with selective media for specific identification of members of Enterobacteriaceae or Streptococcus.

calibrated to hold 0.001 mL or 0.01 mL of sample

• - OCD= CFU/loop vol.
• - OCD= origninal cell density
• - CFU= colony forming unit

• purpose= measures susceptibility to dental caries (tooth decay), caused by lactobacilli and oral streptococci
• glucose is the ferment-able carbohydrate

• yellow at 24 hours= high susceptibility to dental caries;
• yellow at 48 hours= moderate susceptibility to dental caries;
• yellow at 72 hours= slight susceptibility to dental caries;
• yellow at >72 hours is negative