CLS 271 UNIT 1 EXAM STUDY GUIDE

what does a clinical microbiologist help perform regarding one of its three roles?

helps perform cultures of microorganisms from patient specimens

what does a clinical microbiology do after isolation regarding one of its three roles?

classifies and identifies the microorganism after isolation through staining and biochemical reactions

what does a clinical microbiologist do related to a microorganisms patterns regarding one of its three roles?

predicts and interprets antimicrobial susceptibility patterns through its cell structure and biochemical pathways

what is the cell structure of a eukaryote?

• more complex than the prokaryotic cell structure —> fungi, parasites, viruses

• cytoplasmic structure = has nucleus, nucleolus, ER, and golgi

• cell envelope structure = has a plasma membrane, cell wall, and organelles

more specifically, what is in the cytoplasmic structures within a eukaryotic cell? explain their roles too

• nucleus contains DNA in the discrete chromosomes and is bound by bilayered lipoprotein nuclear membrane (IMPORTANT)

  • nucleolus = site of rRNA synthesis

• endoplasmic reticulum which is rough covered with ribosome (protein synthesis) and smooth-synthesize phospholipids

• golgi apparatus = modifies and packages proteins that is sent by rough ER

what are the membrane bound organelles in a eukaryotic cell? what are their roles?

• mitochondria = energy production

• lysosomes = degrade microorganisms in cell

• peroxisomes = break down in H2O2

• chloroplasts = energy production in plants

more specifically, what are the cell envelope structures in the eukaryotic cell?

• its plasma membrane is composed of a phospholipid bilayer that is embedded with proteins and cholesterol

  • plasma membrane envelops the cytoplasm and regulates transport of chemical that go in and out of the cell

• cell wall is rigid and only fungi will have chitin

• the way it moves is through cilia and flagella

what is the cell structure of a prokaryote?

• small and has less compartments

• has no nucleus

• cell envelope structure = plasma membrane and cell walla

• has surface polymers = capsule and appendages

  • helps stick to a eukaryotic cell

more specifically, what is the cytoplasmic structure of the prokaryotic cell?

• often will see a single circular chromosome attached to the mesosome in the cell membrane

• ribosomes which are the site of protein synthesis will be free in the cytoplasm

• cytoplasmic granules which store granules like polysaccharides, lipids, or polyphosphates in the cytoplasm

• can form endospores which is the response to a harsh environment

  • dormant asexual spores that have a thick protein coat that is highly resistant to things that will kill it like chemical agents, UV, dehydration, dessication, gamma radiation, and temperature changes

  • when favorable conditions arise, it germinates

what is the principal component of a gram positive cell wall?

its wall is composed of a THICK protective peptidoglycan (murein) layer

what does the gram positive cell wall absorb?

absorbs the primary stain, crystal violet, but is not dissolved by alcohol which gives its purple color

how do antibiotics like penicillin act on a gram positive cell wall?

acts by preventing the synthesis of peptidoglycan

what are the two things unique to the gram positive cell wall?

• teichoic acid which is anchored to peptidoglycan

• lipteichoic acid which is anchored to the plasma membrane

what are the layers in a gram-negative bacteria? what is unique in the gram negative cell wall?

• has two layers —> a thin INNER peptidoglycan layer and an additional outer membrane that is unique to the gram-negative bacteria

• periplasmic space which is a gel-like area that is between outer and inner membrane and contains nutrient-binding proteins and detoxifyinng enzymes

what does the additional outer membrane of the gram-negative cell wall contain? what does it function as? what is LPS?

• it contains proteins, phospholipids, lipopolysaccharide (LPS)

• it functions as a barrier to harmful substances, attachment sites to the host cell, and acts as a sieve which allows water soluble molecules to come into the cell

• LPS Lipid A which is an endotoxin is responsible for fever and shock conditions in patients who are infected with gram-neg bacteria and is often the cause of septicemia

what are the three different shapes of bacteria?

1. cocci → spherical

2. bacilli → rod-shaped

3. spirochetes → spiral

how can cocci be arranged?

• singles

• pairs

• chains

• clusters

how can bacilli be arranged?

• fusiform which has taper, pointed ends

• curved

• single

• chains

• palisading which is aligned side by side

what does pleomorphic mean?

variation in size and shape which could range from 0.4 um to 2 um

what is the prokaryotic surface polymer, capsule?

it is an organized polysaccharide or polypeptide structure that prevents phagocytosis and can adhere to surfaces

why does the capsule need to be removed during serologic typing? give an example

• it needs to be removed to be able to detect the antigen underneath the capsule

ex. salmonella typhi’s capsule needs to be removed to see agglutination with salmonella somatic antisera

what does the capsule look like with india ink?

capsule shows itself as a clear halo-like structure

what is the prokaryotic surface polymers, slime layers?

• similar to capsules but has more spread-out layers

• has polysaccharides which inhibit phagocytosis and help with adhering to host tissues or synthetic implants like the hip

what is the order for gram staining?

1. crystal violet which is the primary stain for one minute

2. iodine for one minute

   • a mordant which fixes the iodine

3. alcohol or alcohol-acetone which is quickly added and then rinsed

   • a decolorizer that washes out the crystal violet out of a gram negative cell and retains the gram positive stain

4. safranin for 30 seconds which is a pink counter stain

what is an acid fast stain? what do you have to do to the cell wall?

• stains bacteria with high lipid and wax content in the cell wall like mycobacteria, nocardia, and rhodococcus

• has to treat the cell wall to allow the penetration of dye

  • heat like the ziehl-neelson

  • detergent like kinyoun

what is the order to do the acid fast stain?

1. primary stain → carbolfuchsin which is red

2. acidified alcohol which is a decolorizer

3. methylene blue which is a counterstain

the acid fast bacteria retains the red primary stain

what is flurochrome? what do you need to see it?

• a fluorescent stain that is selective for the acid fast cell wall and stains it yellow or orangeacid-fast

• need a fluorescent microscope

what is the growth media, minimal medium?

simple and completely defined

what is the growth media, nutrient medium?

• more complex and uses meat or soy extracts in its media

• is a nutrient broth, trypticase soy broth

what is enriched media? what are its growth factors?

• it is blood agar and chocolate agar

• growth factors are blood, vitamins, and yeast extract

what is selective media? give an example

• allows some microorganisms and some not

• ex. macconkey (pink) for gram negative, PEA for gram positive

what is differential media? give an example

• allows the metabolic differences between different groups or species of bacteria to be visualized

• ex: macconkey lactose fermenters (pink)

what is transport media? give examples

• prevents growth of microorganisms and preserves viability for transport to a lab

• ex: cary blair, amies, stuart broth

what is obligate aerobe?

MUST require O2

what is obligate anaerobe?

cannot grow in O2

what is facultative anaerobe?

grow either with or without O2

what is aerotolerant anaerobe? give an example

• survives in O2 but doesnt use O2 for metabolism

• ex. some Clostridium

what is capnophilic? give an example

• an increased concentration of CO2 is required (5—10% CO2)

• ex. neisseria

what is microaerophilic?

• a reduced concentration of oxygen required (5-6%)

• ex. campylobacter

what does bacterial metabolism consist of?

consists of reactions that bacteria uses to break down organic compounds and to synthesize new components

how is bacteria biochemistry and metabolism regulated?

its regulated by the presence/production and or the activity/inhibition of specific bacteria enzymes

what differs in bacteria’s biochemistry and metabolism?

differs in their ability to use various compounds as metabolic substrates and the end products that are generated

what are the three things that diagnostic schemes analyze unknown microorganisms for?

1. utilization of substrate as a carbon source

2. production of certain end products from multiple different substrates

3. production of acid or alkaline pH in the test medium which produces a color change

what is the EMP glycolytic pathway?

• it is a major pathway that converts glucose to pyruvate

• generates reducing power in the form of NADH2

• generates energy in the form of ATP

• aerobic

what is the pentose phosphate pathway?

• not much energy is produced and is a detour off the EMP pathway

• generally functions to generate NADPH for biosynthesis and provide pentoses for nucleotide synthesis

• can generate ATP but is less than the EMP pathway

• some bacteria have a shortage on the enzymes needed for glycolysis so uses PPP instead

what is the entner-doudoroff pathway?

• converts glucose-6-phosphate to pyruvic acid

• generates NADPH and is an aerobic process that is used by some bacteria that lack the enzymes for glycolysis

what can organisms that ferment lactose do?

ferment glucose

what is mRNA?

carries the genetic information that is needed to make proteins

what is tRNA and rRNA?

tRNA carries amino acids to the ribosome and rRNA is the site of protein synthesis

what is symbiosis?

two organisms living together

what is commensalism?

• when the microorganism benefits and the host is not harmed

• ex. proteus mirabillis in the GI tract

what is mutualism?

• both the microorganism and the host benefit

• ex. lactobacillus in urogenital area

what is parasitism?

• the microorganism benefits but the host is harmed

• ex. entamoeba histolytica

why is the stomach usually sterile? what are the exceptions?

acidic pH destroys microorganisms; however, some exceptions would be endospores (C. diff), parasitic cysts, and H. pylori which can live in the acidic environment

how would other pathogens entire the GI tract and alter the normal microbial flora of the GI tract?

pathogens would enter through food particles and escapes the stomach and enters the intestine then colonizes the small and large intestines (obligate and facultative anaerobes)

how can antibiotics disrupt the usual microbial flora of the GI tract?

eradicates or suppresses certain populations of the flora which causes other indigenous flora to multiply

what are opportunistic infections?

occurs because of a weakened immune system and cause disease when habitat is changed (reason why C. diff affects those who are on antibiotics)

what is the two purposes of normal microbial flora?

1. primes the immune system

2. changes the microenvironment to block/forbid colonization of pathogens

what is pathogenicity?

ability of an organism to produce disease

what are opportunistic pathogens?

pathogens that usually do not cause infection unless under special cirucumstances

what are true pathogens?

organisms that cause disease in healthy hosts with normal immune systems

what are iatrogenic infections?

happen due to medical treatment or procedures

how does the host resistance factor, physical barrier effective in preventing pathogens?

it is a mechanical barrier like how skin is effective against most pathogens

what are the three cleansing mechanisms that help hosts be resistance?

1. desquamation/shedding of skin

2. movement of liquids like through tears, urine, mucus secretion (blowing nose)

3. cilia due to it clearing debris by moving to one place to another

how does low pH make a host resistance?

prevents bacteria from colonizing like in the stomach and vagina

what does indigenous microbial flora do relating to host resistance?

prevents pathogen colonization

what are the four antimicrobial substances that make hosts resistant?

1. fatty acids on skin (prevents colonization)

2. HCl in the stomach

3. lysozymes (degrades microbes and other foreign invaders)

4. immune proteins

what are the four immune proteins?

1. IgA

2. low molecular weight cationic proteins like beta-lysins

3. complement which synergize to increase the effectiveness of killing

4. interferon

how are primary infections surveilled?

infection is related to a SINGLE, specific site

how are blood stream infections (BSI) surveilled?

infection was found in the bloodstream

how are central line-associated bloodstream infections surveilled?

infection is due to the presence of an intravascular device like a central venous catheter

how are surgical site infections surveilled? how is risk formed?

• infection is at the site where the surgery happened

• risk is often formed due to the length of surgery, site of the infection, and the degree of the anticipated contamination

how are UTI’s surveilled?

infection due to the urinary tract which is frequently associated with a urinary catheter

how is ventilator-associated pneumonia surveilled?

pneumonia in the patient is related with a ventilator device like an endotracheal tube or trachetomy

what is total surveillance? how is the infection rate determined?

• all of the infections are recorded, analyzed, and risk is assessed

• the infection rate is determine and analyzed and compared to a established baseline

how is SSI rates defined related to total surveillance?

number of infections divided by number of procedures reported as a percent

how is CLABSI and VAP rated defined related to total surveillance?

number of infections divided by 1000 device days

what is targeted surveillance?

• due budget or personnel restraints

• infection rates are monitored and a close watch is kept on specific, high-risk, and high-volume procedures

what are the five ways data gathered and supported in the laboratory?

1. isolates are identifies

2. antimicrobial susceptibility patterns

3. specimen contamination rates

4. number of isolates per site

5. number of isolates per hospital unit

how are trends identified in infections?

finding the normal rate/baseline data of previously found data on infection rates

what is an outbreak related to how trends are identifies?

upward trend of infection above the baseline data which may be associated with specific patients and/or procedures

what are the two reporting requirements for Nevada?

1. requires reporting from a VARIETY of professions like labs, physicians, vets, college admins. and correctional institutes

2. labs must submit SPECIFIC bacterial isolates to the state public health lab (will have specific criteria)

what is sterilization?

destruction of ALL forms of life including sores

what is disinfection?

elimination of a defined scope of microorganisms and a chemical agent is applied to it

what is an antiseptic?

substance that is applied to the skin to eliminate OR reduce the number of bacteria that is present but does NOT kill spores

what are the three factors in disinfection? explain why each factor is important

1. concentration of disinfecting agents

   • make sure its not TOO concentrated or TOO diluted because it is not effective if its too concentrated or too diluted

2. presence of organic material like blood, mucus, pus

   • inactivates the chemical agents and essentially coats the surface that needs to be treated which prevents the interaction between the chemical and the microorganism

3. nature of surface being disinfected

   • will the chemical agent damage the surface or does the time of exposure need to be altered?

what is isopropyl alcohol?

• a flammable, broad spectrum antiseptic but is NOT sporicidal

  • to remove spores, filter through 0.22 micron filters

• inactived by organic material (clean hands first)

• works by denaturing proteins but must be used in 60%-90% concentration but MUST be evaporated from the surface

what is isopropyl alcohol used for?

• used as a handwash

• surgical hand scrub

• patient preoperative skin prep

• housekeeping disinfectant

what is the goal with handwashing?

eliminate transient flora which is contracted from the environment but also protect skin’s resident or normal flora

what is quality control?

measures designed to ensure the medical reliability of lab data

what are the three stages for lab workload? explain them

1. preanalytical = ordering test, collecting specimen, identifying specimen, transporting specimen

2. analytic = sample testing

3. postanalytical = results delivered and action is taken based on the result

what is quality assurance?

taking measures that ensures high-quality patient care

what needs to be checked on daily regarding their temperatures?

• incubators

• heating blocks

• water baths

• refrigerators

• freezers

why are NIST calibrated thermometers used?

has appropriate correction factors and does not use mercury

how is QC susceptibility testing done?

done on american type culture collection control organisms because they give STANDARDIZED results

what are the variables that affect the accuracy of antimicrobial susceptibility QC results?

1. antibiotic potency

2. agar depth

3. evaporation

4. cation content

5. pH

6. instrument failure

7. inoculum concentration

8. difficulty in determining endpoints

what is personnel competency?

• ensures that the individual is competent through verification, documentation, and obtaining continuing education credits each year

• required under CLIA and individual needs to be reverified each year with proof in their file

how are stock cultured used?

utilized to test various media to make sure it is working correctly or competency testing

what are the four sources that stock cultures can come from?

1. commercial sources

2. proficiency testing isolates

3. patient isolates

4. ATCC

how should cultures be grown? what can be done with them?

• should be grown in broth-large batch

• can make single-use aliquots which can be stored in the freezer for a year

  • take a new vial each week to prevent mutations from subculture and subculture twice after thawing to return it to healthy state