Microbio LAB 3 – Flashcards
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thick slime layer of a bacteria is called what? 2 |
| capsule or glycocalyx |
| Why are Special techniques have to be used to stain a capsule? |
| b/c capsule is damaged with heat fixxing, capsules also repel most stains |
capsule staining begins with what? cells are spread in a film with what? and are not What?
what kind of dye are used for capsule staining?
what do they appear like on microscope? |
capsule stain- negative stain
on acidic film-no heat fixing
indian dye
halo
afterwards. similar neg staining protocol |
| resting stages of genera bacillus and clostridun, it's function |
| ;resting stages called endospores when they exhaust essential nutrients. |
| what function does endospre serve |
dorment stage of in bacterial help protect against enviromental conditions not suitable for growth ; --> making stains hard to penetrate keratin of cell wall
|
| terminal |
| end of cell |
| central |
| middle |
| subterminal |
| b/w terminal and middle |
| two different kinds of shapes |
| Spherical or elliptical (oval) |
| method using to stain endospores |
"futon shaver" |
stain used for endospore
stain used for vegetative portion of cell
color of each |
malachite green to stain
safranin to stain the vegetative portion
GIVES |
| function of streak plate method |
isolate mixed colonies to produce COLONIES (one type of cell) or colony-forming unit (CFU)
mixed--> isolated |
| 2 types of streak methods |
T-streak & quadrant streak
T quadrant |
| function of streak plating |
| Purpose is to reduce the number of cells being spread with each successive series of streaks thus diluting the concentration of cells. |
| wat should we not do when preparing streaks |
| Do NOT Gouge the Agar!! Try to make all streaks as close as possible |
lab 4 FLASHCARDS |
| DEFINE AEROTOLERANCE |
| ABILITY TO LIVE WITH OXY PRESENT |
| OBLIGATE AEROBES |
| REQUIRE HIGH O2 |
| AEROBES |
| REQ O2 - LOWER CONCEN |
| MICROAERIPHILES |
| REQ REDUCED O2 |
| MICROAEROPHILES |
| REQ REDUCED O2 |
| AEROTOLERANT ANAEROBES/ FACULATIVE ANAEROBES |
| DO NOT REQUIRE AND ARE NOT NEG AFFECTED BY O2 |
| AEROTOLERANT ANAEROBES |
DO NOT REQUIRE O2 BUT DOES NOT MEAN IT IS HARMFUL EITHER |
| FACULATIVIVE ANAEROBES |
| FLEXIBLE, NOT ADVERSELY AFFECTED BY O2 |
| OBLIGATE ANAEROBES |
| hates O2- DOES NOT WANT IT AT ALL |
| WHERE DOES OBLIGATE GROW (STRICT) |
| TOPWHERE OXYGEN IN ABUNDANT |
| FACULATIVE ANAEROBES |
| FLEXIBLE, GROWTH IS MORE RAPID WITH, BUT WILL DO WITHOUT |
| Aerotolerant anaerobes |
"IMMUNE" |
Microaerophiles |
- Survive only in environments containing lower than atmospheric levels of oxygen.
- DESIRE SMALL AMOUNTS |
| HABITAT OF Obligate (strict) anaerobes |
| DEEP MUD, LAKES,OCEANS AND SOILS |
WHAT FUNCTION DOES INOCULATING FLUID THIOGLUCOLLATE MEDIA DO?
what kind of agent is thio and why? |
IT TEST FOR OXYGEN REQUIREMENTS OF AN ORG.
-SUPPORTS GROWTH OF BOTH AERIOBIC AND ANAEROBIC BACTERIA
thioglco- is a reducing agent which reduces oxygen to water |
WHAT IS THE DYE IN THIOGLYCOLLATE, WHAT FUNCTION DOES IT SERVE?
what color does it change, what does this represent |
DYE: RESAZURIN- INDICATOR FOR OXYGEN --> BECOMES PINK
res-a-zur-in turns pink in the pressence of oxygen |
| DYE RESAZURIN BECOMES W/ COLOR IN THE PRESENCE OF OXYGEN |
| AFTER BOILING FTM BROTH AND IS PINK,(GETTING RID OF OXY) DEPENDING ON OXYGEN REQUIREMENTS WHERE WILL AEROBIC AND ANAEROBIC ORG BE LOCATED IN CONTAINER? |
AEROBIC ORG- AT THE TOP
ANAEROBIC- BOTTOM |
| FUNCTION OF ANAEROBIC JAR |
| CREATE A ANAEROBIC ENVIROMENT FOR THE CULTURE OF STRICT ANAEROBES |
| WHAT IS USED TO CONVERT H2 AND 02 --> WATER IN ANAEROBIC JAR |
| PLADDIUM CATALYST |
KEY METABOLITES IN METABLOISM
2 |
ACETYL CoA
PYRUVIC ACID |
| ARE METABLOITES LINEAR |
| NO SEVERAL DIFFERENT PRODUCTS FOR DURING REACTION |
| PYRUVIC ACID IS FORMED IN WHICH PROCESS |
| GLYCOLYSIS (GLUCOSE) |
| DEFINE FERMINTATION |
| process of deriving energy from the oxidation of organic compounds, such as carbohydrates. |
| Carbohydrate fermentation is the metabolic process by which |
an organic molecule acts as an electron donor
OXYGEN IS NOT THE FINAL E- ACEPTOR |
| PRODUCTS OF FERMENTATION |
| ACIDS AND ALCOHOL |
| MICROORG PRESENT IN FERMEN |
| ANAEROBIC BACT/ EUKARYOTES/ YEASTS |
| F OF GLUCOSE TEST |
| determine if an organism is able to ferment glucose |
| IF ORG ABLE TO FERMENT GLUCOSE W/ IS THE BYPRODUCT |
| acid will be produced and gas may be produced. |
| IF ACID IS INDEED FORMED IN GLUCOSE TEST WHAT COLOR WILL OF TUBE B AND WHY? |
YELLOW-->PHENOL RED INDICATOR positive fermentation of glucose |
IN METHY RED AND VOGES PROSKAUER TESTS HOW MANY MEDIAS ARE YOU USING
|
| 1-SAME |
| FUN OF METHYL RED TESTS |
| TEST FOR PRODUCTION OF MIXED ACIDS FROM GLUCOSE FERMENTATION |
| HOW MANY DROPS OF METHYL RED INDICATOR ARE PLACED IN OVERNIGHT CULTURE |
| 5-6 |
| RED COLOR MEANS |
| POSITVE ACID |
| ORANGE |
| INCONCLUSIVE |
| NO COLOR CHANGE |
| NEH RESULT |
| F OF METHYL RED AGAIN |
| MIXED ACID FERMENTATION PRODUCT |
| FUNCTION OF VOGUES MAGAZINE |
| to see if the organism produces 2,3 butanediol instead of acids from glucose fermentation |
| THERE IS NO WAY TO DIRECTLY TEST FOR 2,3 BUTANEDIOL SO WHAT IS THE VOGUES REALLY TESTING FOR?? |
| ACETONIN- PRECURSOR TO 2,3 BUTANEDIOL |
| ACETONIN IMPORTANCE |
| no way to test the presence of 2,3 butanediol directly. Therefore, the VP test is actually testing for the presence of acetoin, which is a precursor of 2,3 butanediol |
AFTER ADDING BARRITS REAGENT a (ALPHA NAPTHOL) And BRB + KOH(s/p 3 days)
if reagents cause acetonin to oxidize to diacetyl which further reacts with guanidine nuclei from peptone what color will be produced? |
| red or pink, indicates a positive reaction for acetoin, which tells you that the organism is a butanediol fermenter |
| what does a positive reaction of acetoin indicate( red or pink) |
| org is a butanediol fermenter (remember can't direct test for this) GOal is this! |
| recap: function of VP |
Tests for the precusor of acetoin, which is a precursor of |
| in respiration final electron acceptor is always.... |
| inorganic molecule |
| function of catalase |
converts hydrogen peroxide into oxygen and wawter
h2o2--> h2 + o2
Mainly used to distinguish between staphylococcus and streptococcus** |
| function of nitrate test |
| test for presence of enzyme nitrate reductase |
| what is enzyme nitrate reductase |
-produced by faculative bacteria to carry out anaerobic respitation -enzyme catalyzes the transfer of electrons from |
| red or pink color change reprsents what for nitrate reduction |
| represents positive for indicating that org capable of anaerobic respiration |
| function of nitrate reduction test |
| indicates org capable of anaerobic respiration |
| if nitrate red test has no color change that means it's neg--> if you suspect this to be the case add some zinc dust if it turns red what does this represent |
confirmed to be negative-
there is no color change |
define oxidation
define reduction |
|
| lab 5 flash cards |
| function of media |
| to differentiate org based on ability to grow when esstential nutriens are limited |
| function of citrate slant |
| some organisms can utilize citrate as ONLY carbon source |
| function of CITRATE PERMEASE |
| enzyme some org that only use carbon to transport citrate into the cell producing pyruvate from it |
| what is contained in medium for utilization test |
medium contains:
- sodium nitrate--C source - ammonium phosphate--N source |
| what is the indicator dye used for citrate test |
| bromthymol blue due - indicator |
| as the pH increase medium color changes to what |
| green-----> Blue |
| bacteria using citrate and survive in media convert ______ into ____ and ____ ____ causing media to become more ____ |
ammonium phsphate into ammonia and ammonium hydroxide, alkaline
bacteria using citrate and survive in media convert ammonium phosphate into ammonia and ammonium hydroxyde causing media to become more alkaline |
Blue color represents
green color change
can organism use citrate as sole source of carbon |
blue-- org uses citrate as sole Carbon source
green- NO media stay green |
| function of hydrolytic enzymes |
| perform hydrolytic reactions - using water to split compelx molecules |
| intraceulluar vs exoenzymes |
intracellular enzymes- catalyze the reaction inside the cell
exoenzymes- secreed from the cell to catalyze the reactions outside the cell |
| 3 different test for hydrolytic exoenzymes |
1. starch hydrolysis 2. casease 3. gelatinase |
| test for intracellular enzyme |
urease
urine comes from the inside |
| 2 glucose subunits |
amylose (linear) -losing streak
alpha-amylase and oligo 1,6-glucosidase |
| what are we using to test wheather an org can produce alpha-amylase and oligo 1,6 glucosidase? |
| Starch agar- simple plated medium of beef extract soluble starch, and agar |
| if test proves positive for alpha amylase/ or oligo what does this mena |
| will hydrolyze starch |
| indicator for starch |
| iodine- turns blue with positive starch |
| when starch indicator reacts with starch the media should change what color? |
| dark brownish color |
| for a positive result of exoenzymes what should petri look like? |
| clear zone around bacteria |
| negative result |
| no clear zone |
| when do you add iodine to starch hydrolysis ( testing for exoenzymes) |
| 24 hours |
| function of urease enzyme |
| hydrolyzes urea into ammonia and CO2 |
| what will be using to determine urease prod |
| urea broth |
| what is the ingrediants of urea broth |
- yease extract and urea - has phenol red as pH indicator***
|
| what is the pH indicator in urea broth |
| phenol red |
| if org grown in urea broth is urease, pH and color is effected how? |
| ph will increase, phenol red will change to yellow to red to PINK |
| how long does urease phenol red take to complete |
| 8 days |
| function of casein hydrolysis test |
| to test if bacteria requires protein as a source of amino acid. |
| function of CASEASE enzyme?? |
| breakes down large proteins into subunits that can be brough into the cell |
for casein hydrolysis test how do you interpret result??
pos neg |
positive- clear zone negative result- no clear zone |
| gelainase test function |
| test to see if organism has enzyme that allows some organisms to hydrolyze gelatin |
| gelatin |
| Gelatin is a protein derived from collagen – a component of vertebrate connective tissue. |
| what is a important step in gelatinase test |
| Chill media for (4°C) ~30 mins before looking at results. |
| positve or neg result of gelatinase test |
positive- gel liquid negative- solid gel |
| ? Starch Hydrolysis: ? Polysaccharide catabolism, exoenzyme catalyzed ? Casein Hydrolysis: ? Polypeptide catabolism, exoenzyme catalyzed ? Urea Hydrolysis: ? Protein catabolism, intracellular enzyme catalyzed ? Gelatin Hydrolysis: ? Polysaccharide catabolism, exoenzyme catalyzed |
| function of combination differential media |
- combination differential media combine compoenents of several compatible tests into one med
- include core test to differentiate members of specific bacterial group
- can be used as a follow up to selective media |
| function of SIM, what does it stand for |
sem-solid media
tests: sulfur reduction/ indole prod from tryptophan/ motility |
| sulfur reaction: function of cysteine desulfurase/ thisulfate reductase |
| bacteria can reduce sulfur to hydrogen sulfide using either of 2 enzymes both produce hydrogen sulfide gas |
| If the organism can reduce sulfur___will be seen in the tube. |
| , a black color |
indole production:
function of tryptophanase
(4) |
| bacteria that produces enzyme hydrolyze trypophan to pyruvate, ammonia, and indole |
| how to tet for indole production |
| kovak's reagent is used on an inoculated, incubated culture |
| kovac's reagent will react with indole to produce what? |
| red color |
| what indicates a positive result for enzyme tryptophanase? |
red color
brown means neg |
| motility 3rd test function |
| stabbing technique is used, if cloudiness is seen in the media away from stab line, motility is indicated |
| you want to maintain the needle how during sim motility test |
| straight |
| what does the SIM motility resemble |
| gelain media |
| how far in do you want to in, what position do you want to keep the |
~2/3, upward |
TSIA - tripple sugar iron agar 3 tests it provides |
sugar utilization gas production hydrogen sulfide production |
| 3 sugars agar contains |
| lactose, sucrose, glucose small amt |
| what is the indicator for TSIA |
| phenol red |
| what indicates ferrous sulfate |
hydrogen sulfide
no color change- no sugars fermented |
| page 25 |
| chapter 1 |
Fastidious heterotrophs
|
Fastidious heterotrophs – rely heavily on the environment to supply ready-made organic compounds
Nonfastidious heterotrophs – do not rely as heavily on |
| Never use what with oil immersion?? |
| Never use the coarse adjustment with the oil immersion lens! |
| total magnification = |
| Total magnification = Magnification of Objective X Magnification Of Ocular |
| LAB 2 |
| [image]pellicle coat |
• some float on top of medium and produce a type of surface membrane |
| sediment |
• sink to bottom |
| Uniform fine turbidity (UFT) |
| ?? look up |
| flocculent |
| clumping growth |
| describe bacteria |
Bacteria are:
Simple
Have circular DNA
Cell surrounded by peptidoglycan |
| Spirillum (spirilli |
| a rigid curved or spiral shaped cylinder |
| vibrio |
Gently curved rods
part of spirilli |
spirochetes
part of spirilli |
| Slender flexible spirals are called spirochete (spirochetes) which is greek for coil of long hair |
| gram positive |
Gram positive (+) cells have a thicker peptidoglycan
retain the primary stain of crystal
purple pos dark counter-intuitive |
crystal violet water iodine ethanol safranin |
