PID final lectures Flashcards
a chemical substance that is produced by microorganisms and has the capacity in dilute solutions to selectively inhibit the growth of or to kill other microorganisms
antibiotic
a substance of natural, semisynthetic, or synthetic origin that kills or inhibits the growth of microorganisms/bacteria but causes little or no damage to the host
antimicrobial
T/F
all antibiotics are antimicrobials
TRUE
Are all antimicrobials also antibiotics
no
when diseased animals are treated to cure infection
therapeutic antimicrobial use
when healthy herds or animals are treated to prevent infection
prophylactic antimicrobial use
when diseased herds are treated to cure
infection in some individuals and prevent infection in others
metaphylactic antimicrobial use
when healthy animals are treated with low concentrations in feed to improve growth rate and efficiency of feed utilization and improve reproductive performance
growth promotion antimicrobial use
chemically designed by man
synthetic
chemically altered natural compounds
semi-synthetic
produced by fungi and bacteria
natural (antibiotics)
Name 2 synthetic classes of antibacterial agents
- quinolones
2. sulfonamides
bacteriocidal drugs
kills the bacteria
bacteriostatic drugs
inhibits growth
Bacteriostatic or bacteriocidal?
Aminoglycosides
cidal
Bacteriostatic or bacteriocidal?
Cephalosporins
cidal
Bacteriostatic or bacteriocidal?
Glycopeptides
cidal
Bacteriostatic or bacteriocidal?
Lincosamides
static
Bacteriostatic or bacteriocidal?
Macrolides
static
Bacteriostatic or bacteriocidal?
Penicillins
Cidal
Bacteriostatic or bacteriocidal?
Phenicols
static
Bacteriostatic or bacteriocidal?
Quinolones
Cidal
Bacteriostatic or bacteriocidal?
Rifamycins
cidal
Bacteriostatic or bacteriocidal?
Sulfonamides
static
Bacteriostatic or bacteriocidal?
Tetracyclines
static
drugs that have an activity restricted to a few bacterial groups
narrow spectrum
drugs that have an activity against a wide range of different bacterial organisms
broad-spectrum
the drugs spectrum is assessed before marketing by what official document?
Summary of Product Characteristics (SPC)
Name the 3 classes of B-lactams
- Penicillins
- Cephalosporins
- Carbapenems
1st generation cephalosporins spectrum
MAINLY GRAM +
broadest spectrum cephalosporins
3rd generation – includes pseudomonas
Gram + penicillin with a narrow spectrum
Penicillin G / V
gram - penicillin
aminopenicillins
two Carbapenems that are highly resistant to B-lactamases and are injectable
Imipenem and meropenem
Drugs with the best activity against Non-penicillinase producing gram+ cocci
Penicillins and Aminopenicillins
Non-Penicillinase producing gram+ cocci
enterococcus and streptococcus
Drugs with the NO activity against Non-penicillinase producing gram+ cocci
metronidazole
Drugs with the best activity against penicillinase producing staphylococcus
amoxicillin/clavulanate cephalosporins sulfonamides aminoglycosides fluoroquinolones
Drugs with the NO activity against penicillinase producing staphylococcus
penicillins
aminopenicillins
metronidazole
glucose fermentative gram negative rods
escherichia enterobacter klebsiella proteus pasteurella
drugs with BEST activity against glucose fermentative gram negative rods
amoxicillin/clavulante cephalosporins sulfonamides fluoroquinolones aminoglycosides
drugs with NO activity against glucose fermentative gram negative rods
Metronidazole
penicillins
lincosamides
macrolides
Anaerobe examples
actinomyces clostridium bacteroides fusobacterium prevotella
drugs with BEST activity against anaerobes
penicillins
aminopenicillins
lincosamides
metronidazole
drugs with NO activity against anaerobes
aminoglycosides
5 antimicrobial modes of action
- inhibition of cell wall synthesis (penicillins)
- inhibition of protein synthesis (tetracyclines)
- inhibition of DNA synthesis (fluoroquinolones)
- inhibitors of cell membrane integrity (colistin)
- inhibitors of other cell metabolic activities (sulphonamides)
4 drivers of antimicrobial resistance
- antibiotics
- metals
- biocides
- genes
refers to the co-existence of multiple genes or mutations encoding resistance to different drugs within the same strain or genetic element
Co-Resistance
the selection of multiple resistance genes when one of these genes is selected
co-selection
drugs that effect RNA elongation
actinomycin
drugs that effect DNA gyrase
quinolones
novobiocin
3 drug classes not authorized for veterinary use
- carbapenems
- glycopeptides
- oxazolidinones
What class do chloramphenicol and florfenicol belong to?
Phenicols
What class do sulfadiazine and sulfadiazine/trimethoprim belong to?
Sulfonamides
3 drugs in the tetracycline class
- tetracycline
- oxytetracycline
- doxycycline
Class that these drugs and combos belong to:
penicillin G/V
Ampicillin/amoxicyllin
amoxicillin/clavulanic acid
Penicillins
What class do these belong to:
oxolinic acid
enrofloxacin/marbofloxacin
pradofloxacin/norfloxacin
Quinolones
What class do these belong to:
colistin
polymixin
polypeptides
what class do these belong to:
streptomycin
gentamicin/neomycin
amikacin
aminoglycosides
what class do these belong to: lyncomycin/clindamycin
lincoamindes
What 3 drugs belong to the Macrolides class?
- erythromycin
- tiamulin
- tilmicosin
What class do these belong to? cefalexin/cephadroxil cefpodoxime/ceftiofur cefovecin cefquinome
cephalosporins
Class that linezolide belongs to
oxazolidinones
Gylcopeptides
vancomycin/teicoplanin
What class do these belong to: imipenem/meropenem ertapenem/doripenem
carbapenems
Is vancomycin authorized for vet use?
NO
Is linezolide authorized for vet use?
NO
Is erythromycin authorized for vet use?
YES
what is the first and most critical step for accurate diagnostic testing
proper specimen collection
what are some critical things to do when collecting a specimen for testing
- avoid contamination
- collect as specifically as possible
- collect before antibiotics are used and before the acute phase
what is an acute phase
very pronounced symptoms, typically a strong immune reaction and active replication of pathogen
common places for BACTERIAL infections in a dog
skin ear urine wounds blood
SAME AS CAT
common places for BACTERIAL infections in a cat
wounds ear skin blood urine
SAME AS DOG
common places for BACTERIAL infections in a cow
milk
common places for BACTERIAL infections in a HORSE
nasal
wound
common places for BACTERIAL infections in food animals
post mortem tissue
it can be hard to distinguish the difference between bacterial infections and ________ or _______
normal flora or environmental contamination
specimen collection sites for PARASITE infections
feces sputum urine blood vomit muscle skin
What does parasite specimen collection depend on?
a suspected parasite collection depends on the symptoms and the life cycle stage of the suspected parasite
common collection sites for VIRAL infections
nasal eye swab feces blood post mortem tissue
phenotypical testing methods for parasites
- direct blood smear and visualize
2. concentration techniques such as centrifuge/filtration or flotation of feces
success of parasite diagnosis by phenotypic techniques depends on these 4 factors
- stage/life cycle or type and sex of parasite
- animal age and species
- technique procedure
- number of eggs shed
CAN HAVE MANY FALSE NEGATIVES AND NEED RETESTED
disadvantages of cytology
mild/chronic are not readily detected
not all samples are appropriate
can magnification cytology see viruses?
NO
magnification – cytology
shows the morphology of cells, bacteria, and parasites but NOT viruses
staining can enhance the visualization and it gives an immediate analysis and idea of severity
stain that uses one dye, and shows the morphology/cell arrangement but not the individual cell components
simple stain
stain that uses multiple dyes and cell structures and types can be seen
differential stain
common tissue stain that can show viral, bacterial, parasite and fungi
Hematoxylin and eosin stain
which is acidic and which is basic in an H&E stain
H - basic: stains acids purple
E - acidic: stains bases red
can identify sample abnormalities, parasite and bacterial infections
is a multispecimens dye
Romanowsky Stain “Diff-Quick”
can distinguish between Gram negative and Gram positive bacteria
shows cell arrangement and morphology
gram stain
G+ purple – thick PG
G - Pink – thin PG
anaerobic
cant grow in O2
capnophiles
needs CO2 to grow
bacteria that require specific nutrients and culture conditions
fastidious
blood agar
differential media that shows hemolysis
alpha- damaged cell
beta - complete lyse
gamma - no damage
MacConkey agar
differential media
gram negative selective
lactose fermentation = turns pink
mannitol salt agar
differential media
gram positive selective
mannitol fermentation - turns yellow
CLED agar
differential media for urinary bacteriology
cysteine-lactose-electrolyte-deficient
supports growth of common urinary pathogens
lactose fermentation
economical test that generates a code for organism ID
ID test strips/plates
urine paddle test incubation
37 degrees for 18-24 hours
urine paddle test
used in clinics or in labs
one side is gram negative selection and the other side is differential media
it gives a semi-quantitative colony count in UTI and presumptive uropathogen identification
breaks down hydrogen peroxide
catalase
T/F
presence of an antibody means there is an active infection
false
absence of antibody does not = absence of pathogen
can detect pathogen specific antibodies or antigens
immunochemical tests
exploit the principles of pathogen-specific immune response to detect and ID pathogens
immunochemical tests
T/F
Cultures can detect parasites and viruses
FALSE THEY CANT
AMR consequences on animal and public health
increased patient mortality and morbidity
risks of zoonotic transmission
economic consequences of AMR
loss of customers as a vet
longer hospital stays
lab tests
reduced food gain in animals
every year AR causes approximately _____ deaths in the US per year
23,000
resistant bacteria are responsible for approximately ______ infections every year
2 million
Name 2 important superbugs that are present in the hospital and community including animals
Methicillin-resistant S. aureus MRSP
ESBL producing E.coli
MSRA/MSRP mechanisms of bacteria resistance
target modification/protection on the ribosomal sites
ESBP mechanism of bacterial resistance
enzymatic drug inactivation
antibiotic therapy is only recommended for _______ infections
invasive
salmonella resistant phenotype
resistance to cephalosporins and fluoroquinolones
campylobacter resistant phenotypes
resistance to macrolides or fluoroquinolones
resistance phenotype in MRSA MRSP and ESBL
beta-lactam resistance
all are resistant to cephalosporins and are often multi-drug resistant
bacteria that mainly affects dogs
MRSP
what does MRSA stand for
methicillin
resistant
staphylococcus
aureus
WHAT gene was acquired to make MRSA resistant and how does it work
it acquired the resistance gene mecA which encodes a penicillin-binding protein (PBP2A) that has a low affinity for beta lactams
MRSA strand cc8 is most common in what species
horses
MRSA strand cc22 is most common in what species
dogs/cats
MRSA strand cc9 is most common in what species
pigs
MRSA strand cc398 is most common in what species
horses and pigs
this is the “live-stock” associated strand MRSA
How do MRSA carriage rates in the normal populate compare to in vets worldwide?
vets have a significantly higher percentage of carriage for MRSA
What does MRSP stand for
methicillin
resistant
staphylococcus
pseudintermedius
T/F
MRSP and MRSA have acquired the same resistant gene, mecA
TRUE
approximately what percentage of cases of MRSP are from skin and wound infections acquired in the clinic?
AND
what are these pathogens called?
70%
nosocomial pathogens
T/F
antimicrobial choice is hard because MRSP strains may be resistant to all antibiotics licensed for veterinary use
TRUE
T/F
vets make act as a vector for animal infection of MRSP
TRUE
MRSP in humans due to transmission from household pets is _______ but carriage is higher in dog owners and vets
RARE
what does ESBL stand for
extended
spectrum
beta
lactamase
ESBL is an enzyme that _______ most b-lactams produced by ______ bacteria, except for ________
hydrolyses/inactivates
gram negative bacteria
carbapenems
what are the three main classes of ESBL
- CTX-M
- SHV
- TEM
most common type of ESBL in animals/livestock
CTX-M-1
what is a false ESBL, and which one is widespread in small animals/limited to poultry in Europe?
resistant to beta lactamase inhibitors
CMY-2
what is a true ESBL?
they are susceptible to beta lactamase inhibitors such as clavulanic acid
which CTX-M stain is variable across the world but is most prevalent in human e.coli infections
CTX-M-15
how do bacteria acquire resistance?
- mutation
2. horizontal gene transfer
3 distinct methods of horizontal gene transfer
- transformation
- transduction
- conjugation
what is transformation>
uptake of free DNA
what is transduction
transfer mediated by phage delivery
what is conjugation
cell to cell contact plasmid transfer
T/F
source attribution is easy for ESBL
FALSE
difficult due to involvement of different genes and plasmids
what 3 things can antigens from pathogens be?
- the whole pathogen itself (only as small part on the surface is the antigen)
- a molecule produced by the pathogen
- pathogen molecules presented on the surface of host cells
do you find antigens in the erythrocytes or plasma?
plasma/serum
two common antibodies detected
IgM and IgG
common specimens for IgM
blood
common specimens for IgG
blood and tissue fluids
common specimens for antigens
primarily the area of infection or where the pathogen replicates or antigen is present
ex: for a GI tract infection you can use feces as a sample
what is KEY for accurate immunochemical test results
TIMING
3 indicators of an active or recent infection
- pathogen detection by phenotypic methods or detection of an antigen
- present or recent clinical symptoms of infection
- titre of antibodies (decreases with time)
What is an ELISA test
it does specific Ag/Ab detection and can detect a response to a parasite, virus, fungi, or bacteria
has HIGH specificity and sensitivity and gives a quantitative idea of Ag/Ab present
What is an IDEXX Snap Test
it has lateral flow and can detect Ab/Ag in a specimen
can be used for Bacteria, Parasites, and Virus
ex: whole blood/plasma or fecal tests
Agglutination tests
Ag/Ab clump together due to specificity to detect an immune response
indirect – uses a latex bead with the antigen on it so antibodies will clump to it and make it more visible for presence detection
immunochemical test advantages
- ID when it cannot be cultured
- most have high sensitivity and specificity
- mid to high volume testing is possible
immunochemical test disadvantages
- antibody may not show an active infection
- if tested too early the Ab may not even show up
- some pathogens may have the same antigen and so more than one could be detected by one antibody
ID of markers in the genome or proteome of a pathogen
molecular diagnostics
MALDI-TOF tests are mainly for what
bacterial ID (sometimes fungi)
mass spectrometry test that gives a finger print of the components of a pathogen for identification, but is limited to the database
MALDI-TOF test
T/F
MALDI-TOF test requires a pure culture
TRUE
Test used for high scale ID of many pathogens
multiple PCR or microarrays
detects nucleic acids and amplifies them using tagged oligonucleotide probes in a digested sample to identify present pathogens
Multiple PCR or microarrays
advantages of using molecular diagnostics
it is rapid, highly sensitive, accurate and can do high volume testing
disadvantages of using molecular diagnostics
it is expensive, can have possible false negatives, and only gives yes/no answers (except for real time PCR)
what things make a good diagnostic lab
guidance accredited skilled workers to interpret the data quality assurance measures state of the art techniques
OH NOoOOooO
we dont have a diagnosis…. what happened?
- incorrect sample collection or handling
- incorrect test for the pathogen or infection stage
- BAD timing for parasite life cycle or AB/AG test
** monitor and repeat diagnostic tests
a suspension of antigens administered to induce immunity
vaccine
currently the majority of vaccines are derived from _____
microbial pathogens
what is an adjuvant
it enhances the immune response to antigens
delays the release of Ag from site of infection, induces the secretion of chemokines by leukocytes
2 examples are: aluminum hydroxide and saponin
what makes an ideal vaccine?
cheap long lived immunity no adverse side effects immunological memory range of epitopes stable shelf life proper immune response induced consistent in formation
3 types of infectious vaccines
- live attenuated
- recombinant organism vaccine
- marker vaccine
2 types of non-infectious vaccines
- subunit
2. naked DNA vax
what is a live attenuated vaccine
attenuated yet in tact and viable organism
T/F
live attenuated vaccines have a low level of infection
TRUE
vaccine type that does not induce significant tissue pathology or clinical disease
live attenuated
T/F
live attenuated vaccines require and adjuvant
FALSE
pros to live attenuated vaccines
rapid onset immunity
sustained immunity after a single dose
T/F
live attenuated vaccines have a great shelf life stability
false they are less stable bc live
cons of live attenuated vaccines
they have the potential for reversion to virulence and can cause virulence in immunocompromised people or cause immune suppression
recombinant organism vaccines
carrier organisms do not cause disease in vaccinated animals
adjuvant is NOT required
will NOT revert to virulence
permits discrimination between a vaccinal and an exposure immune response
marker vaccine
example of a marker vaccine?
IBR in cows – deletes a gene GpE in cattle so anti-GpE cows have been vaccinated
killed, whole organism vaccines that are antigenically intact
non-infectious vaccines
T/F
Non-infectious vaccines are able to replicate
false
T/F
Non-infectious vaccines can induce pathology or clinical disease
FALSE THEY CANT
chemical killing formalin, alcohols, and alkylating agents are examples of what kind of vaccines
non-infectious
pros of inactivated vaccines
safe
no interference with other vaccines
stable in storage
T/F
inactivated vaccines have a fast onset immunity
false - slow onset immunity
CONS of inactivated vaccines
may need boosters
adjuvant required and may cause adverse effects
lower degree of protection as compared to live vax
vaccine containing immunological structural proteins or metabolites of an organism
subunit vaccines
–can be purified proteins, synthetic peptides, or recombinant proteins
a gene of interest is cloned from a pathogen to a plasmid and is delivered directly to an animal
naked DNA vaccines
the pathogen gene is expressed on the APCs for antigen presentation to create and immune response
NAKED DNA Vax
vaccination = ?
immunization
performed antibodies are administered to a particular antigen
passive immunization
provides immediate immunological protection that is temporary
passive immunization
tetanus antitoxins and antivenoms are an example of what type of immunization?
PASSIVE
immunization type that sensitizes the recipient for a hypersensitive reaction
passive immunization
inhibits the endogenous antibody response of the recipient
passive immunization
3 methods of delivering vaccines
- injection
- internasal
- needle free
antigens are administered to create immunological memory
active immunization
an immune response is generated/induced in the recipient (humoral or cell mediated)
active immunization
adverse side effects to vaccines
type 1 hypersensitivity
— facial or periorbital edema / pruritis
Feline injection site sarcoma
example of an adverse side effect
poor prognosis
why do we need antimicrobial susceptibility testing?
guidance to antimicrobial therapy in labs
surveillance of antimicrobial resistance
antimicrobial drug discovery
2 methods for AST
- dilution method
2. agar diffusion method
the lowest concentration that inhibits complete growth of the test strain
MIC = minimum inhibitory concentration
lowest concentration that kills the test strain
MBC = minimum bactericidal concentration
______ methods enable quantification of antimicrobial susceptibility by determining MIC and MBC
dilution
T/F
MIC and MBC are accurate
false – gives a range
2 broth dilution methods
macrodilution and microdilution
________ gives automated or semiautomated readings
microdilutions
primarily used for research purposes, for example when many strains needs tested in one drug
agar dilution
pros and cons of dilution methods
pro:
has a high reproducibility and robustness
Cons:
very expensive
pros and cons of diffusion methods
pro: medium reproducibility and robustenss
con: cheap
T/F
Standardization and quality control are required for reproducibility of AST
TRUE
laboratory AST procedure steps
- adjust inoculum of bacteria
- add to plate
- incubate at 35 for 16-20 hours
- read plate
a drug specific value to interpret the results of susceptibility testing and determine if an antibacterial in potentially useful in the treatment of a bacterial infection
breakpoint
a strain is defined as ______ by a level of antimicrobial activity associated with a high likelihood of therapeutic success
susceptible
a strain is defined as ______ by a level of antimicrobial activity associated with a high likelihood of therapeutic failure
resistant
a strain is defined as ______ for a species by the absences of any required resistant genes and/or mutations increasing the MIC of the antimicrobial agent
wild type WT
data needed for setting breakpoints
- MIC distribution to determine the MIC value that separates the wild type from resistant populations
- PD/PK index
- clinical and bacteriological outcome data from clinical trials
what is the PD/PK index?
relationship between drug concentrations, the MIC value, and antimicrobial effects
category that is also considered a buffer zone in AST
intermediate
the strain is inhibited at blood concentrations achieved by standard dosage of the drug
susceptible
clinical efficacy is possible if the strain infects body sites where the drug concentrates or if the dosage can be increased compared to standard dosage
intermediate
the strain is not inhibited at blood concentrations achieved by standard dosage of the drug
resistant
drugs with no breakpoints available
topical drugs
most frequent bacterial contaminants
- coagulase-negative staphylococci (SKIN)
- bacillus spp (soil)
- enterococci (fecal)
problems interpreting AST results
mixed cultures
absence of antimicrobials used in clinical practice
presence of antimicrobials not used in clinical practice
frequent with otitis, UTI, and wound infections
mixed cultures
used in the lack of approved clinical breakpoints to predict susceptibility to other drugs belonging to the same class
surrogate antimicrobials
ampicillin surrogate
amoxicilin
sulfamethoxazole surrogate
sulfadiazine
lincomycin surrogate
clindomycin
tertracyclin surrogate
doxytetracycline
what 2 drugs are used for the detection of MRSA/MRSP and why
- oxacillin
- cefoxitin
because their methicillin resistant gene mecA is poorly expressed in laboratory media
T/F
invitro susceptibility testing does not take into account in vivo factors
TRUE
T/F
if more tetracycline is used, it is more likely that there will be resistant e.coli stains
true
what species uses the most antimicrobials in Denmark and therefore has more bacteria with resistant strains
pigs
T/F
antimicrobial resistance can be eradicated
FALSE
use of antimicrobial agents aimed at maximal therapeutic efficacy while minimizing the risks of developing resistance, in the strain causing infection and the patient’s commensal flora
rational antimicrobial use
provides international guidelines on major infections in companion animals
International Society for Companion Animals Infectious Diseases (ISCAID)
are national or international guidelines better?
national
what should all clinics have in place
infection-specific antibiotic formularies
what are the 3 last resort drugs for human use?
carbapenems
linezolid
vancomycin
what are some conditions in which a laboratory culture should be used?
- no response to therapy
- previous antibiotic treatment
- immunocompromised patients
- life threatening infections
- patients at risk of multiple drug resistance
- long treatment course
T/F
doing in-clinic cultures is a great idea
FALSE – avoid kitchen microbiology
“shoot” regular – dose
uses time dependent drugs such as beta lactams at regular intervals and owners must know the importance of compliance
prescribe high doses for concentration dependent drugs like fluoroquinolones and aminoglycosides
“shoot” high dosage
MPC
mutant prevention concentration
Do humans or animals typically get treated for longer periods by antimicrobials
animals
latest trend in human medicine is that……
unnecessary treatment should be avoided after clinical resolution of symptoms
4 ways to control AMR transmission
- hospital infection control
- farm biosecurity
- slaughterhouse hygiene
- educated consumers and owners
3 ways to control AMR antimicrobial use
- legal interventions like bans and penalties
- preventative vet med
- antimicrobial stewardship
refers to coordinated interventions designed to improve and measure the appropriate use of antimicrobials by promoting the selection of the optimal antimicrobial drug regimen, dose, duration of therapy, and route of administration
antimicrobial stewardship
where are antimicrobial stewardship programs normally underdeveloped ?
vet clinics
what 3 things does the establishment of antimicrobial stewardship programs require?
- commitment of staff
- collaboration with microbiology lab
- infectious disease specialist or educated clinician in antimicrobials and resistance
the discipline concerned with preventing nosocomial or health-care related infections
hospital infection control
regardless of type or size, every vet clinic should have these three things
- formal infection control program
- written manual
- infection control practitioner
what does ICP stand for
infection control practitioner
how long and when should you wash your hands…..
at least 20 seconds
before and after every patient
building block of nucleic acidz
nucleotidez
T/F
purines have 2 rings one with 5 and one with 6 atoms
TRUE
the sugar is connected to the nitrogenous base at what position
1 – by a glycosidic bond
A matches to T with how many hydrogen bonds
2
C matches to G with how many hydrogen bonds
3
DNA sequence is read in this order
5’ - 3’
has same sequence of mRNA
coding or sense strand
directs synthesis of mRNA via complementary base pairing
antisense or template strand
DNA to RNA
transcription
RNA to protein
translation
T/F
both DNA and RNA can replicate themselves
TRUE
when there is a parent strand and a new strand in a new molecule of DNA
DNA is semiconservative
T/F
DNA is very conservative. modest is hottest
FALSE – only semiconservative
unwinds the DNA double helix
helicase
T/F
the lagging strand is synthesized discontinuously
TRUE
T/F
bacterial transcription and translation all occur in the same location
TRUE
T/F
animal cell transcription and translation occur in the same location
false
transcription in the nucleus
translation in the cytoplasm
what cells have introns
eukaryotic
T/F
eukaryotic cells do not have polyadenylation
TRUE
but eukaryotic cells do at the 3’ end
has a 5’ methylated cap
eukaryotic cells only
how many nucleotides are in each codon
64
T/F
any base pair of DNA can be mutated
TRUE
mutations from the background/locus/generation
spontaneous
mutagens by mutagens
induced
act directly by modifying a particular base or being incorporated into the nucleic acid
mutagen
mutations without an apparent effect
either no change in AA or a change in aa that doesnt change the proteins function
silent mutation
a mutation that changes the codon and therefore changes the amino acid and protein function
missense mutation
change in a coding codon to a terminal codon resulting in premature termination
nonsense mutation
inserts or deletes a number of bp other than a multiple of 3
frameshift mutation
acquisition of new genetic markers by incorporation of added DNA
transformation – a type of genetic transfer in bacteria
mating between two bacteria involving transfer of genetic material
conjunction – a type of genetic transfer in bacteria
transfer of bacterial gene to another bacterium by a phage
transduction – a type of genetic transfer of bacteria
movement of a transposon to a new site in the genome
transposition – a type of genetic transfer of bacteria
electrophoresis
separates molecules by charge on an electrophoretic field – commonly DNA RNA and proteins (negatively charged and move to the positive end)
what determines electrophoresis particle mobility
SIZE – smaller go further/faster
**also structure plays a role.. is it a monomer/trimer/dimer/supercoiled ect
cut DNA at or near specific recognition nucleotide sequences
restriction enzymes
**sequence specific
What do restriction enzymes come from
bacteria to defend from viral infections
restriction fragment length polymorphism
cutting sequence specific DNA
number of restriction enzymes studied
3000
600 available commercially
is EcoRI sticky or blunt end cutting
STICKY
is smaI sticky or blunt end cutting
BLUNT
common uses of RFLP
forensic analysis
parent identification
mixed infections
denatured, single stranded DNA with a probe complementary single strand bound to it
hybridization
T/F
probes have a high degree of specificity
TRUE
fragment of nucleic acids that is used for detecting complementary sequences in samples
Probe
examples of what a probe could be
labeled, radioisotope, enzyme, or chemiluminescence
enzyme-mediated process to synthesize copies of target nucleic acid
target amplification – type of nucleic acid amplification
two types of primers
random
specific
signal amplification
bDNA assays or hybrid capture assays
polymerase chain reaction
isothermal amplification..
has high sensitivity and also could give a false positive – type of nucleic acid amplification
steps of cycle 1 of PCR
- denature to separate DNA strands
- anneal/cool to allow primer to form the hydrogen bond with target sequence
- extension – DNA polymerase adds nucleotides to the 3’ end of each primer
products of PCR
products = 2^n
n= number of cycles run
what determines the size of the PCR product?
the distance between the primer binding sites
singe-stranded DNA fragments, complementary to sequences flanking the region to be amplified
primers (for PCR)
what determines the specificity during PCR?
the PCR primer
what type of enzymes are needed in PCR
THERMOSTABLE polymerases – to survive the denaturation stage heat stage
what determines how long the lag phase is in real time PCR
the starting amount of the material
less = shorter lag phase/cycles
allows quantification of starting material
real time PCR / quantitative PCR
LAMP pros
no thermal cycler needed
fast (less than an hour)
sensitivity > PCR
gives visible results
LAMP cons
design of primer sets complicated
