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Final Exam - Flashcards

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Class:BIO 3722 - Microbiology Laboratory
Subject:Biology
University:University of Texas - San Antonio
Term:Fall 2013
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Capsule Stain
  • differential
  • both G(+) & G(-) can produce capsules
  • environment ---> favorable
  • increases virulence
  • heat sensitive
  • Water and alcohol soluble
  • nonionic
Primary dye: congo red
Secondary: maneval
Bile Esculin Test BEA plate
  • Selective: bile salts - inhibits everything but sero group D
  • Differential: esculin & ferric citrate

Esculin --------> Esculetin + glucose
                             **reacts with Fe
Purpose: ID group D strep

black: positive
NC: negative
DNA Hydrolysis DNA plate
  • differential: methyl green - forms a complex with polymerized DNA, clearing around growth indicates DNase activity

DNA + MG -------->DNase-------> nucleotides

positive: clearing
negative: no change
Bacitracin Susceptibility BAP plate
  • Selective: Bacitracin - inhibits bacterial cell wall synthesis; effective only on bacteria with a cell wall & in the process of growing

Reagent: Taxo A disc; 0.04 units Bacitracin
Purpose: ID group A strep

positive: clearing >10mm
negative: clearing <10mm
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CAMP Test BAP plate
**testing or the presence of CAMP substrate**

CAMP factor of S. agalactiae acts synergistically with β-hemolysin of   S. aureus and forms an arrowhead shaped zone of hemolysis (positive)
Coagulase Test Coagulase Plasma Tube (pink cap)
  • rabbit plasma
  • Fibrinogen----->coagulase----->fibrin

positive: clot - bacteria form protective fibrin barriers to shield from phagocytosis

negative: no change
Triple Sugar Iron Agar TSIA: red slant with black cap
Selective:
  • 0.1% glucose
  • 1.0% lactose
  • 1.0% sucrose
Differential:
  • phenol red
  • ferric ammonium citrate (FAC)
Na thiosulfate---->thiosulfate reductase---->H2S + FAC
*Cracks in agar is positive for gas production*
Fluid Thioglycollate Medium FTM (teal cap)
Differential: Resazurin - indicator; pink when oxidized, colorless when reduced

Purpose: Determine aerotolerance category
  1. aerotolerant
  2. facultative anaerobe
  3. strict anaerobe
  4. microaerophile
  5. strict aerobe  
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Temperature
  1. Minimum: enzymatic activity ceases under min. temp
  2. Optimum: bacterial growth is most efficient
  3. Maximum: enzymes/proteins begin to denature above max temp
psychrophiles <20 °C
*mesophiles 15-45 °C
thermophiles >40 °C
  • obligate
  • facultative
  • extreme 65-110 °C
UV Radiation Bacteria that form endospores are more resistant to UV radiation
  1. monotrichous
  2. amphitrichous
  3. lophotrichous
  4. peritrichous
  • resistant to heat and staining
  • form in poor environment
  • made of keratin

Primary stain: malachite green
Secondary: safranin

  1. central
  2. terminal
  3. subterminal 
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Antibiotics
  • natural 
  • synthetic
  • broad spectrum
  • narrow spectrum
  • bacteriostatic
  • bacteriocidal
  • IV, IM, SQ, PO
Carb. Fermentation

Purpose: differentiate members of Enterobacteriaceae from other Gram(-) rods

 

Differential: phenol red, durham tube

  • glucose/sucrose/lactose/mannitol fermentation - yellow color
  • peptone deamination - pink color
  • bubbles in durham tube - gas production

 

red cap = lactose; green cap = glucose; yellow cap = sucrose

Gelatinase

Purpose: Determine organism's ability to produce gelatinase

 

Differential: Gelatin

  • liquid after 3 minutes on ice - gelatinase is present

Gelatin--> Gelatinase--> amino acids

Urea

white lid

Purpose: test for enteric bacteria which can metabolize urea

urea-->urease--> CO2 + H20 + NH3

Selective: urea

  • only rapid urease-positive organisms can alkalinize the medium; Proteus, Morganella, and Heliobacter

Differential: phenol red

  • pink: positive for rapid urease-positive organism
  • orange/yellow: negative

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Citrate

Purpose: differentiate members of Enterobacteriaceae; G(-) rods

Selective: citrate

  • only bacteria with the enzyme citrate-permease will grow

Differential: bromthymol blue

  • citrate-positive bacteria alkalinize the medium and turn it blue

 

Citrate-->citrate permease-->pyruvate

           -->                           -->acetate

SIM

Used for determination of sulfur reduction, indole production, motility

 

black: positive for sulfur reduction, H2S production

Na thiosulfate-->thiosulfate reductase-->H2S + ferric ammonium citrate


red after addition of Krovac's reagent: positive for indole production

tryptophan-->tryptophanase-->indole


radiating growth pattern: positive for motility

5 Tests for Strep organisms
  1. Bile Esculin Test
  2. DNA Hydrolysis Test
  3. Bacitracin Susceptibility Test
  4. CAMP Test
  5. Coagulase Test
  1. aerotolerant
  2. facultative anaerobe
  3. strict anaerobe
  4. microaerophile
  5. strict aerobe
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Antibiotics Mechanism of Action: Cell Wall Synthesis Inhibits cross linking of peptidoglycan
Penicillin, cephalosporins, bacitracin, & amoxicillin
β-lactam ring
  • β-lactamase
  • β-lactamase inhibitor
Antibiotics Mechanism of Action: Protein Synthesis Eu: 40s/60s subunits
Pro: 30s/50s subunits
  • tetracycline
  • streptomycin
  • chloramphenicol
Antibiotics Mechanism of Action: Nucleic Acids DNA or DNA associated enzymes
  • ciprofloxacin
  • nalidixic acid
  • topoisomerase II = cipro + NA
Antibiotics Mechanism of Action: Anti-Metabolites Folic acid synthesis is inhibited

PABA------------------------------->DHF-------------------------------->Folic acid
                   folate synthase                        DHF reductase

  1. sulfa drugs inhibit folate synthase
  2. trimethoprim inhibits DHF

1 + 2 = 5:1 ratio co-trimoxazole


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Antibiotics Mechanism of Action: Plasma Membrane Alters membrane integrity

  • polymixin & colistin

Neosporin = bacitracin + neomycin + polymixin
Antibiotic's Effects
  1. Additive - sum of individual effects of 2 or more drugs
  2. Synergistic - acting together, enhancing the effect
  3. Antagonistic - a therapeutic result less than the sum of each drug's effect
Mueller-Hinton Agar MHA **looks like TSA plate with a black line**
measure zone of inhibition (mm) & use chart to determine susceptibility or resistance

  • Te-30: tetracycline
  • P-10: penicillin
  • C-30: chloramphenicol
  • Cip-5: ciprofloxacin
  • Cro-30: ceftriaxone
  • Amc-30: amoxicillin
Blood RBC
platelets
WBC: 
  • granulocytes
  • agranulocytes
 
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Granulocytes 55-65% neutrophils

0.5-1.0% basophils

1.0-3.0% eosinophils

Agranulocytes 3.0-7.0% monocytes

25-33% lymphocytes
  • B cells
  • T cells
Endospore stain
neutrophil
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eosinophil
basophil
monocyte
lymphocyte
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Fungi Mycology
  • nonmotile eukaryotes
  • absorptive feeders
  • aflatoxin
  • statins 
Zygomycetes
  • terrestrial
  • nonseptate
  • nonmotile
  • sporangospores or zygospores
  • Ex: rhizopus
Ascomycetes
  • yeast & mold
  • sexual reproduction
  • spores: ascospores and ascus
  • Ex: penicillium & saccharomyces
Basidiomycetes
  • fleshy fungi (mushrooms)
  • basidiospores
  • septate hyphae
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Deuteromycetes
  • no sexual reproduction
  • phase has been observed
Ophiocordyceps **Zombie ant fungi**
Ringworm
  • multiple fungi
  • cause athletes foot and yellow toe nails
Helminths Part of Animalia
Hosts
  1. Definitive: the host in which the sexual reproduction of a parasite takes place 
  2. Intermediate: a host which is normally used by a parasite in the course of its life cycle and in which it may multiply asexually but not sexually 
  3. Paratenic: a host that is not necessary for the development of a parasite but maintains the life cycle
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Trematodes "fluke"
bilateral symmetry & suckers
Clonorchis singergis
Trematode
  1. undercooked fish
  2. operculum
  3. monoecious
  4. infection, liver damage, bile duct damage
  5. eggs in fecal matter
  6. mebendazole
Schistesoma mansoni Trematode
  1. skin penetration in contaminated water
  2. egg has a lateral spine
  3. dioecious
  4. fever, rash, GI symptoms
  5. eggs in fecal matter
  6. Praziquantel (disrupts calcium channels)
Cestodes "tapeworms"
flat
ribbon shaped
absorb nutrients
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Taenia solium Cestode
**has hooks**
  1. undercooked pork
  2. circular eggs
  3. monoecious
  4. asymptomatic / cystercerosis (brain & eyes)
  5. eggs in fecal matter
  6. Praziquantel or Albendazole
Taenia saginata Cestode
**no hooks**
  1. beef
  2. round eggs
  3. monoecious
  4. abdominal pain, allergic reaction
  5. eggs in fecal matter
  6. Praziquantel
Nematodes "round worms"
cylindrical
invade digestive tract
Ascaris lambricoidis Nematode
  1. fecal-oral
  2. lipid layer on eggs
  3. dioecious
  4. fever & diarrhea
  5. eggs in fecal matter
  6. mebendazole
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Enterobius vermicularis Nematode
  1. fecal-oral
  2. n/a
  3. dioecious
  4. perianal itching
  5. eggs (tape test)
  6. mebendazole
Protozoa
  • absence of cell wall
  • locomotion
  • single cell
  • heterotrophic
  • asexual
Sarcodina Pseudopods
FL: Amoeba proteus
P: Entamoeba hitolytica
  • necrosis of GI tract
  • dysentary
Mastigophora Flagella
FL: Euglena
P: Giardia lamblia

Trypanosoma:
  • T. cruzi - Chaga's
  • T. brucei: African Sleeping Sickness (tsetse fly)
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Ciliophora Cilia
FL: Paramecium caudatum
P: Balantidium coli
  • ulceration of the GI tract
  • constipation / diarrhea
Sporozoa no mature locomotion
no free living
Plasmodium vivax
P. falciparum
  • 80% deaths
  • 90% infections
  • humans - immature host
  • Anopheles mosquito - mature host
Saccharomyces cerevisiae
Aspergillus septate hyphae
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Rhizopus nonseptate hyphae
Aspergillus conidial heads
Penicillium conidiophore
Schistosoma mansoni adults
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Dipylidium caninum scolex
Taenia solium scolex
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  Capsule Stain
  • differential
  • both G(+) & G(-) can produce capsules
  • environment ---> favorable
  • increases virulence
  • heat sensitive
  • Water and alcohol soluble
  • nonionic
Primary dye: congo red
Secondary: maneval
 Bile Esculin TestBEA plate
  • Selective: bile salts - inhibits everything but sero group D
  • Differential: esculin & ferric citrate

Esculin --------> Esculetin + glucose
                             **reacts with Fe
Purpose: ID group D strep

black: positive
NC: negative
 DNA HydrolysisDNA plate
  • differential: methyl green - forms a complex with polymerized DNA, clearing around growth indicates DNase activity

DNA + MG -------->DNase-------> nucleotides

positive: clearing
negative: no change
 Bacitracin SusceptibilityBAP plate
  • Selective: Bacitracin - inhibits bacterial cell wall synthesis; effective only on bacteria with a cell wall & in the process of growing

Reagent: Taxo A disc; 0.04 units Bacitracin
Purpose: ID group A strep

positive: clearing >10mm
negative: clearing <10mm
 CAMP TestBAP plate
**testing or the presence of CAMP substrate**

CAMP factor of S. agalactiae acts synergistically with β-hemolysin of   S. aureus and forms an arrowhead shaped zone of hemolysis (positive)
 Coagulase TestCoagulase Plasma Tube (pink cap)
  • rabbit plasma
  • Fibrinogen----->coagulase----->fibrin

positive: clot - bacteria form protective fibrin barriers to shield from phagocytosis

negative: no change
 Triple Sugar Iron AgarTSIA: red slant with black cap
Selective:
  • 0.1% glucose
  • 1.0% lactose
  • 1.0% sucrose
Differential:
  • phenol red
  • ferric ammonium citrate (FAC)
Na thiosulfate---->thiosulfate reductase---->H2S + FAC
*Cracks in agar is positive for gas production*
 Fluid Thioglycollate MediumFTM (teal cap)
Differential: Resazurin - indicator; pink when oxidized, colorless when reduced

Purpose: Determine aerotolerance category
  1. aerotolerant
  2. facultative anaerobe
  3. strict anaerobe
  4. microaerophile
  5. strict aerobe  
 Temperature
  1. Minimum: enzymatic activity ceases under min. temp
  2. Optimum: bacterial growth is most efficient
  3. Maximum: enzymes/proteins begin to denature above max temp
psychrophiles <20 °C
*mesophiles 15-45 °C
thermophiles >40 °C
  • obligate
  • facultative
  • extreme 65-110 °C
 UV RadiationBacteria that form endospores are more resistant to UV radiation
  
  1. monotrichous
  2. amphitrichous
  3. lophotrichous
  4. peritrichous
  
  • resistant to heat and staining
  • form in poor environment
  • made of keratin

Primary stain: malachite green
Secondary: safranin

  1. central
  2. terminal
  3. subterminal 
 Antibiotics
  • natural 
  • synthetic
  • broad spectrum
  • narrow spectrum
  • bacteriostatic
  • bacteriocidal
  • IV, IM, SQ, PO
 Carb. Fermentation

Purpose: differentiate members of Enterobacteriaceae from other Gram(-) rods

 

Differential: phenol red, durham tube

  • glucose/sucrose/lactose/mannitol fermentation - yellow color
  • peptone deamination - pink color
  • bubbles in durham tube - gas production

 

red cap = lactose; green cap = glucose; yellow cap = sucrose

 Gelatinase

Purpose: Determine organism's ability to produce gelatinase

 

Differential: Gelatin

  • liquid after 3 minutes on ice - gelatinase is present

Gelatin--> Gelatinase--> amino acids

 Urea

white lid

Purpose: test for enteric bacteria which can metabolize urea

urea-->urease--> CO2 + H20 + NH3

Selective: urea

  • only rapid urease-positive organisms can alkalinize the medium; Proteus, Morganella, and Heliobacter

Differential: phenol red

  • pink: positive for rapid urease-positive organism
  • orange/yellow: negative

 Citrate

Purpose: differentiate members of Enterobacteriaceae; G(-) rods

Selective: citrate

  • only bacteria with the enzyme citrate-permease will grow

Differential: bromthymol blue

  • citrate-positive bacteria alkalinize the medium and turn it blue

 

Citrate-->citrate permease-->pyruvate

           -->                           -->acetate

 SIM

Used for determination of sulfur reduction, indole production, motility

 

black: positive for sulfur reduction, H2S production

Na thiosulfate-->thiosulfate reductase-->H2S + ferric ammonium citrate


red after addition of Krovac's reagent: positive for indole production

tryptophan-->tryptophanase-->indole


radiating growth pattern: positive for motility

 5 Tests for Strep organisms
  1. Bile Esculin Test
  2. DNA Hydrolysis Test
  3. Bacitracin Susceptibility Test
  4. CAMP Test
  5. Coagulase Test
  
  1. aerotolerant
  2. facultative anaerobe
  3. strict anaerobe
  4. microaerophile
  5. strict aerobe
 Antibiotics Mechanism of Action: Cell Wall SynthesisInhibits cross linking of peptidoglycan
Penicillin, cephalosporins, bacitracin, & amoxicillin
β-lactam ring
  • β-lactamase
  • β-lactamase inhibitor
 Antibiotics Mechanism of Action: Protein SynthesisEu: 40s/60s subunits
Pro: 30s/50s subunits
  • tetracycline
  • streptomycin
  • chloramphenicol
 Antibiotics Mechanism of Action: Nucleic AcidsDNA or DNA associated enzymes
  • ciprofloxacin
  • nalidixic acid
  • topoisomerase II = cipro + NA
 Antibiotics Mechanism of Action: Anti-MetabolitesFolic acid synthesis is inhibited

PABA------------------------------->DHF-------------------------------->Folic acid
                   folate synthase                        DHF reductase

  1. sulfa drugs inhibit folate synthase
  2. trimethoprim inhibits DHF

1 + 2 = 5:1 ratio co-trimoxazole


 Antibiotics Mechanism of Action: Plasma MembraneAlters membrane integrity

  • polymixin & colistin

Neosporin = bacitracin + neomycin + polymixin
 Antibiotic's Effects
  1. Additive - sum of individual effects of 2 or more drugs
  2. Synergistic - acting together, enhancing the effect
  3. Antagonistic - a therapeutic result less than the sum of each drug's effect
 Mueller-Hinton AgarMHA **looks like TSA plate with a black line**
measure zone of inhibition (mm) & use chart to determine susceptibility or resistance

  • Te-30: tetracycline
  • P-10: penicillin
  • C-30: chloramphenicol
  • Cip-5: ciprofloxacin
  • Cro-30: ceftriaxone
  • Amc-30: amoxicillin
 BloodRBC
platelets
WBC: 
  • granulocytes
  • agranulocytes
 
 Granulocytes55-65% neutrophils

0.5-1.0% basophils

1.0-3.0% eosinophils

 Agranulocytes3.0-7.0% monocytes

25-33% lymphocytes
  • B cells
  • T cells
  Endospore stain
  neutrophil
  eosinophil
  basophil
  monocyte
  lymphocyte
 FungiMycology
  • nonmotile eukaryotes
  • absorptive feeders
  • aflatoxin
  • statins 
 Zygomycetes
  • terrestrial
  • nonseptate
  • nonmotile
  • sporangospores or zygospores
  • Ex: rhizopus
 Ascomycetes
  • yeast & mold
  • sexual reproduction
  • spores: ascospores and ascus
  • Ex: penicillium & saccharomyces
 Basidiomycetes
  • fleshy fungi (mushrooms)
  • basidiospores
  • septate hyphae
 Deuteromycetes
  • no sexual reproduction
  • phase has been observed
 Ophiocordyceps**Zombie ant fungi**
 Ringworm
  • multiple fungi
  • cause athletes foot and yellow toe nails
 HelminthsPart of Animalia
Hosts
  1. Definitive: the host in which the sexual reproduction of a parasite takes place 
  2. Intermediate: a host which is normally used by a parasite in the course of its life cycle and in which it may multiply asexually but not sexually 
  3. Paratenic: a host that is not necessary for the development of a parasite but maintains the life cycle
 Trematodes"fluke"
bilateral symmetry & suckers
 Clonorchis singergis
Trematode
  1. undercooked fish
  2. operculum
  3. monoecious
  4. infection, liver damage, bile duct damage
  5. eggs in fecal matter
  6. mebendazole
 Schistesoma mansoniTrematode
  1. skin penetration in contaminated water
  2. egg has a lateral spine
  3. dioecious
  4. fever, rash, GI symptoms
  5. eggs in fecal matter
  6. Praziquantel (disrupts calcium channels)
 Cestodes"tapeworms"
flat
ribbon shaped
absorb nutrients
 Taenia soliumCestode
**has hooks**
  1. undercooked pork
  2. circular eggs
  3. monoecious
  4. asymptomatic / cystercerosis (brain & eyes)
  5. eggs in fecal matter
  6. Praziquantel or Albendazole
 Taenia saginataCestode
**no hooks**
  1. beef
  2. round eggs
  3. monoecious
  4. abdominal pain, allergic reaction
  5. eggs in fecal matter
  6. Praziquantel
 Nematodes"round worms"
cylindrical
invade digestive tract
 Ascaris lambricoidisNematode
  1. fecal-oral
  2. lipid layer on eggs
  3. dioecious
  4. fever & diarrhea
  5. eggs in fecal matter
  6. mebendazole
 Enterobius vermicularisNematode
  1. fecal-oral
  2. n/a
  3. dioecious
  4. perianal itching
  5. eggs (tape test)
  6. mebendazole
 Protozoa
  • absence of cell wall
  • locomotion
  • single cell
  • heterotrophic
  • asexual
 SarcodinaPseudopods
FL: Amoeba proteus
P: Entamoeba hitolytica
  • necrosis of GI tract
  • dysentary
 MastigophoraFlagella
FL: Euglena
P: Giardia lamblia

Trypanosoma:
  • T. cruzi - Chaga's
  • T. brucei: African Sleeping Sickness (tsetse fly)
 CiliophoraCilia
FL: Paramecium caudatum
P: Balantidium coli
  • ulceration of the GI tract
  • constipation / diarrhea
 Sporozoano mature locomotion
no free living
Plasmodium vivax
P. falciparum
  • 80% deaths
  • 90% infections
  • humans - immature host
  • Anopheles mosquito - mature host
  Saccharomyces cerevisiae
  Aspergillus septate hyphae
  Rhizopus nonseptate hyphae
  Aspergillus conidial heads
  Penicillium conidiophore
  Schistosoma mansoni adults
  Dipylidium caninum scolex
  Taenia solium scolex
36, "/var/app/current/tmp/"