*path bac exam 3

constitutive vs non-constitutive

**constitutive:** genes that are always expressed

* typically perform housekeeping functions
* tibosomal RNAs and proteins involved in protein synthesis

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**non-constitutive:** genes that are only expressed when their products are required

* subject to regulation
* virulence genes

why are some genes regulated

avoid host defenses - antigenic + phase variation - turn off genes to evade immune system

energy - don’t make components that aren’t needed

central dogma

DNA - transcription - RNA - translation - protein

DNA - gene expression - protein

regulation of gene expression

transcriptional level: DNA → RNA + post-transcriptional levle

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translational level: RNA → protein + post-translational level

transcriptional regulation - 5

transcriptional regulators

two component systems

sigma factors

supercoiling

nucleoid associated proteins

gene structure 6

**Promoter** – Where RNA polymerase binds to DA

**Transcription start site** – Where transcription begins

**Shine and Dalgarno site** – AKA RBS – Where the ribosome binds the mRNA

**Translation start site** – Where translation begins – ATG start codon

**Translation stop site** – Where translation stops – 3 possible codons

**Transcription stop site** – Where transcription terminates

gene structure - ecoli and RNA pol binding

TTGACA = -35 box

TATAAT = -10 box

the optimal inter-base distance cannot be more than 1+/- WHY? RNA pol binds 3 dimentionally and if there is more the 1 bp change then TATAAT will be on the other side of the DNA helix and RNA cannot bind

operon

group of 2+ genes together on the same transcription unit

transcription activator v repression

activator: required to turn on the expression of one or more genes

repressor: required to turn off the expression of one or more genes

transcriptional activators

typically bind to the DNA in the vicinity (mostly upstream) of the promoter to help recruit RNA polymerase

can overlap with the promoter

transcriptional repressors

most common mechanism of action by repressions is to bind to promoter DNA and prevent RNA polymerase binding

can also compete with activators for the same binding site

bacterial virulence gene regulation

regulon definition

transcription regulator proteins often activate transcription from more than one promoter

regulon: all of the genes under transcription control of the same regulator

two component system

what does it typically do

signals 4

two component signal transduction systems

stimulus-response coupling mechanism

two components:

* sensor protein: bacteria use to sense their environment
* response protein: bacteria use to carry out a response

response is typically alterations in gene expression - response protein is typically a transcription regulator

signals: pH, osmolarity, AMPs, cation concentration

two component signal transduction steps

signal received by histidine kinase (sensor)

autophosphorylation of His

phosphate is transferred to Aspartic acid on response regulator

conformational change → cellular response

reaction: altered gene expression

histidine kinase

composed of: HK domain, variety of other domains

sense a variety of stimuli from a variety of sources

* extracellular: lots of complex loops outside the membrane
* intramembrane: lots of transmembrane domains
* intracellular: transmembrane domains and short external loops

activation of HK

activation of HK: dimerization, autophosphorylation

autophosphorylation ALWAYS occurs on a conserved histidine AA residue

response regulator

typically have 2 domains: N-term receiver domain, c-term output domain

N-term receiver domain: ALWAYS contains as Aspartic Acid residue, receives a phosphate group

C-term output domain: usually has a DNA binding domain, most common is HTH (helix-turn-helix)

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receiver domain is phosphorylated, helix-turn-helix region is exposed, dimerizes, then can bind DNA

NarX-NarL system

simple 2-component system

nitrate signal binds NarX, autophosphorylates NarL, dimerizes, transcriptional activation

Rcs phosphorelay system

complex multi-component system

not all “two component systems” have just TWO components

Rcs system has 5 components

BvgS/R system

some histidine kinases have multiple domains (HK and RR domains)

phosphate group is always transferred ….

H → D and D → H NEVER H → H or D →D

check slide 30 lec 13

what is a sigma factor

sigma factor is a protein that interacts with RNA polymerase and DIRECTS it to promoters

not considered a part of RNA polymerase

not directly required for transcription

once open complex forms sigma factor is released

sigma factor + core RNA polymerase = holoenzyme

bacterial use different sigma factors to direct RNA polymerase to different genes

turns on regulatory networks at different times

* sporulation
* stress response
* virulence

sigma factors

direct RNA polymerase to different subsets of genes depending on environmental conditions

oS = stationary phase genes, starvation

oE = envelope stress genes, something wrong with membrane

oF = flagella genes

oH = heat shock genes

having different sigma factors = use the same RNA pol and direct it to genes that need to be expressed

sigma factors allow the cell to reuse core RNA polymerase = converses energy

\# sigma factors varies greatly depending on the bactera

genome condensation

to fit the nucleoid (genome) inside the cell bacteria use a combination of

* nucleoid associated proteins
* supercoiling

DNA supercoiling

DNA helix underwound

introduces negative (right handed) supercoils

condenses DNA

degree of negative superhelicity changes according to environmental/physiological conditions and stimuli

how do bacterial control supercoiling levels

DNA gyrase and Topoisomerase 1

Gyrase introduces negative supercoils

Topoisomerase 1 introduces positive supercoils (relaxes DNA)

underwinding/negative supercoiling

overwinding/ positive supercoiling

?? slide 41 lec 13

DNA supercoiling during infection

as salmonella bacteria enters macrophage, salmonella DNA becomes more relaxes (less negatively supercoiled)

DNA relaxation? = signal to activate genes require for survival inside macrophages

promoters become activated by DNA relaxation

slide 42 lec 13

nucleoid associated proteins

functions 3

participate in DNA transactions 4

four major NAPs

regulation of gene expression? 2

class of small DNA binding proteins found associated with the genome (nucleoid)

bind DNA

bend DNA

participate in DNA transactions

* transcription
* recombination
* replication
* integration and excision

functions:

* condense DNA
* DNA structural organization (supercoiling)
* transcriptional regulators

four major nucleoid associated proteins

* H-NS
* HU
* IHF
* Fis

play a role in regulation of gene expression

* structural: i.e. positive interaction bends DNA so activator can bind RNA pol
* as transcription activators/ repressors: typically indirect (don’t contact RNAP), compete for binding with activator/repressors

virulence regulation 3

cofactors and ligands → DtxR

regulatory RNAs

quorum sensing

transcription regulators

positive regulation: activator > repressor

negative regulation: repressor > activator

activation of transcription factors

function

transcription factors may be activated or deactivated

TCS: phosphorylation of response regulator activates protein

interaction with other transcription factors (homo or hetero dimerization)

ligands can bind to TF activating/deactivating the protein

ligand: a substance (usually a small molecule) that forms a complex with a protein to serve a biological purpose

* activate
* deactivate
* modify function

regulation of dtxR and tox

regulation of dipthereia toxin (DT) gene expression is controlled by a transcription regulator DtxR

DtxR is activated when Fe2+ levels are high (binds)

DtxR-Fe2+ binds to the tox gene promoter and represses transcription

DtxR regulates tox gene, binds Iron, becomes active,

lots of Fe2+ = tox not produced

low Fe2+ (like inside host)= tox produced, Fe2+ dissociates from DtxR, DtxR no longer binds to tox promoter → transcription of tox → make DT

regulatory RNAs

aka small RNAs (sRNAs)

heterogeneous class of RNA

diversity of regulatory mechanisms

regulate directly or indirectly multiple genes

can be part of the transcript they regulate (cis-acting)

regulate other transcripts (trans-acting)

regulatory activity typically depends on structure and base pairing

regulatory RNAs types 3

riboswitches and antitermination

cis-acting RNA thermosensors

trans-acting RNAs

riboswitches and antitermination SAM

SAM (S-adenosylmethionine) riboswitch - example of a small molecule regulating its own production

long 5” UTR --------- CDS: gene for protein involved in methionine biosynthesis (pathway to generate SAM)

lots of SAM? binds to RNA structure and stabilizes it, it leads to a terminator stabilized and terminates sequence, does not transcribe CDS

low SAM? antiterminator creater, CDS is transcribed, CDS contains same gene → Low SAM makes more SAM

riboswitches and antitermination T box

Tbox riboswitch

CDS encode tRNA synthetase to charge uncharged-tRNAs

when charged tRNA is present in the cell, does not bind UTR and terminator terminates sequence

when uncharged tRNA present in the cell, base pairs with in UTR and disrupts the terminator, antitermination, CDS made

a charged tRNA cannot disable the terminator, it can only work with uncharged tRNA (the AA gets in the way)

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RNA thermosensors

translation regulation

cis acting

full length RNA transcript is made

RNA temperature sensitive

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at low temp, RNA adopts a stable secondary structure

ribosome binding site (RBS = Shine Dalgarno) is masked or sequesteered

ribosome cannot get access to the mRNA → protein is not translated

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at high temperature RNA secondary structure charges or destabilized

RBS no longer sequestered

binding of ribosome → translation

trans-acting RNAs

mech of action 2

very heterogenous group

2 main mechanisms of action


1. sRNAs that bind to RNA (usually by base pairing)
2. sRNAs that bind to proteins

sRNAs that bind to RNA

typically a method of post-transcriptional regulation

* no translation: sRNA binds RBS, ribosome cannot bind, = no translation
* mRNA degration: sRNA binds RBS, dsRNA made, body assumes it is a virus and degrades the RNA
* translation occurs: mRNA forms secondary structure where RBS is blocked, sRNA binds and releases the base pairing reaction and free’s the RBS? = translation

sRNAs that bind to proteins - Csr

CsrA is a translation repressor protein

binds to the RBS of glgCAP mRNA

prevents translation

crsB is a sRNA containing multiple CsrA binding sites

csrB can titrate CsrA away from glgCAP allowing translation to occur

quorum sensing

other name

aka population density dependent gene regulation

cell to cell signaling mechanism that enables bacteria to collectively control gene expression

this type of bacterial communication is achieved only at **higher cell densities**

at low densities bacteria release molecules called __**autoinducers**__ into the extracellular medium

when the bacteria reach a high density (**quorum**) the concentration of autoinducers exceeds a particular threshold

autoinducer is transported back into the cell and activates expression of a particular set of genes

genes responsible for virulence, competence, light production

quorum sensing controlled processes 5

bioluminescence

biofilm formation

sporulation

competence

virulence gene expression

quorum sensing molecules

2 major types of molecules

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N-acyl-homoserine lactones (AHLs) Gram-negative

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Autoinducer peptides (AIPs) Gram-positive

N-acyl-homoserine lactones

synthesized by

recognized by

what does it do

what changes

mediates quorum sensing in G-

several types depending on their length of acyl side chain

able to diffuse through membrane

synthesized by autoinducer synthase = LuxI

recognized by autoinducer receptor/DNA binding transcriptional activator protein = LuxR

autoinducer peptides

often involves?

recognized by

regulates processes 3

shape

G+ use small peptides as quorum sensing molecules

signaling often involves TCS

peptide is recognized by membrane bound histidine kinase

regulates processes such as: competence, sporulation, virulent gene expression

may be linear or cyclic peptides

restriction enzymes

recognize and cut DNA at specific recognition sites

type 2 RE cleave DNA at/around the site

cleavage may produce sticky ends or blunt ends in the DNA target

hundreds are commercially available

very useful in cloning

most commercially available restriction endonucleases were identified in bacteria

restrictions enzymes are named based on the organism in which they were discovered

HindIII: isolated from *Haemophilus influenzaeI* strain Rd

old formatting: *Hin*d III

new formatting: HindIII

what prevents restriction enzymes from cutting u the host bacteria DNA

methylation via methylases

restriction-modification systems

restriction endonucleases are paired with a second enzyme - methylase

methylase recognizes the same DNA site as the restriction endonucleases

after DNA replication methylase adds a methyl group to the DNA

together the two enzymes represent a Restriction-Modification system

methylated DNA will NOT be recognized and cut by the corresponding restriction endonuclease

Restriction-Modification systems can help protect the bacterial cell from foreign DNA

how does restriction-modification systems protect bacterial call from foreign DNA?

host DNA is methylated by methylase

foreign DNA is not methylated

restriction enzymes cut up the foreign DNA and kills it

host DNA is protected

CRISPR origins

name

originally identified as “phage-like” repetitive DNA sequences on bacterial genomes

unknown function

widespread, found in bacterial and archaeal genomes

NAME: Clustered Regularly Interspaced Short Pallindromic Repeats = CRISPR

CRISPR structure

consists of


1. direct repeats, varying in size from 21-37 bp
2. interspaced by similarly sized non-reptitive sequences (spacers)

repeats are pallindromic repeats: DNA sequence that reads the same forward and backward, can create hairpins

family of homologous genes were found associated with CRISPR

CRISPR associated genes (Cas)

number of Cas genes can vary

CRISPR Cas array

Cas gene(s) + Repeats + spacers

CRISPR yogurt origins

researchers at DANISCO were investing practical questions in yogurt fermentation industry

phage contamination: most serious problem in fermentation industries

phage-resistant strains would emerge after phage pandemics

after viral challenge, bacteria integrated new spacers derived from phage genomic sequences, CRISPR + Cas provided resistance against phages

CRISPR steps

acquisition: spacer acquisition

expression: CRISPR expression crRNA biogenesis

interference: or immunity

adaptation

invasion of the cell by foreign genetic elements (bacteriophages or plasmids)

spacers: corresponding to foreign DNA sequences are acquired and inserted into the CRISPR locus between repeat sequences

if the bacterial cell survives infection with bacteriophage it will recognize that phage in the future and be resistant

CRISPR expression

CRISPR repeat-spacer arrays are transcribed into long primary transcripts

precursor CRISPR RNA = pre-crRNA

subsquently processed into a set of short crRNAs

each crRNA contains a single spacer and a repeat fragment

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3 major types of CRISPR

type 1 and 3 are similar, type 2 is different

(end products are the same, mechanisms slightly different)

type 1 CRISPR + mech

CRISPR array: multiple Cas genes + repeats + spacer

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promoter generates long pre-crRNA transcript, processed into individual crRNA’s (broken into 1 spacer + 1 repeat), palindromes form hairpins,

Cas genes encode a nuclease - an enzyme complex that cuts DNA, combines with crRNA, crRNA programs Cas nuclease to cut spacers, spacers come from viruses, so when virus infects again the spacer+nuclease can bind to viral DNA and cut + destroy virus

type 2 CRISPR

CRISPR array: tracer RNA gene + 1 Cas gene + repeats + spacers

there is ONE Cas gene: nuclease

contains tracer gene

transcribes genes → tracer gene, pre-crRNA, Cas gene

pre-crRNA → crRNA into single spacer/repeat pieces

tracer + crRNA = guide RNA

guide RNA binds to Cas to program it

tracrRNA binds repeat

CRISPR interference

crRNAs bond with Cas proteins to form an effector complex

recognizes the target sequence in the invasive nucleic acid by base pairing

induces sequence-specific cleavage and degradation of foreign DNA

prevents proliferation and propagation of foreign genetic elements

type 1 vs 2 CRISPR

type 1:

* only 1 RNA = crRNA
* Cas nuclease typically many protein subunits

type 2:

* 2 RNAs: tracrRNA + crRNA = guide RNA
* Cas nuclease typically consists of one protein (most common is Cas9)

in developing the CRISPR-Cas9 system

bacterial vs simplified

bacterial type 2 CRISPR:

* 3 components: 2x sRNAs + 1 protein (Cas9)
* bacterial defense system against infection

simplified type 2 CRISPR:

* 1 sRNA (single guide RNA)
* 1 protein Cas9
* humans use bc its simple, easy system and very programmable

uses of CRISPR-Cas9 system

inserts, deletions/ knockouts, change the gene sequence,

gene is disrupted

gene has new sequence

increase gene expression: attached to an activator protein which activates gene expression = visualization

why CRISPR good?

precision and control

CRISPR history, nobel

identified in 1987, 2007 first experimental evidence,

2020 Nobel Prize in Chemistry Emmanuelle Charpentier and Jennifer Doudna

restriction modification systems and CRISPR-Cas used for?

restriction modification systems: bacterial innate immunity

CRISPR-Cas: bacterial adaptive immunity

disinfectant, antiseptic, antibiotic

antimicrobial compounds applied to inanimate objects and surfaces

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antimicrobial compounds applied to the skin/outside the body

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antimicrobial compounds that can be taken inside the body that either inhibits or kills microorganisms

disinfectants types and mode of killing 7

alcohols: denature proteins

alkylating agents: form epoxide bridges that inactive proteins

halides: oxidizing agents

heavy metals: bind SH denature proteins

phenols: denature proteins

QACs: disrupt cell membranes

UV radiation: blocks DNA replication

some features of antimicrobial drugs 5

selective toxicity

antimicrobial action

spectrum of activity

pharmacokinetic properties

adverse effects

selective toxicity

how to select - 2

= efficacy vs toxicity - how good is it at killing vs huch will it harm human host

cause greater harm to microorganisms than to host

how can we select for this?


1. antibiotic target is ONLY found in the bacteria
2. antibiotic target in the bacteria is sufficiently different from homologue in humans

antimicrobial action

bactericidal: kill bacteria

bacteriostatic: slow down or inhibit growth of bacteria

spectrum of activity

antibiotics vary with respect to the range of bacteria they kill or inhibit

broad-spectrum antibiotic: active against a wide range of bacteria, both Gram-positive __*and*__ Gram-negative

narrow-spectrum antibiotic: limited range, may only be effective against either Gram-positive __*or*__ Gram-negative bacteria

pharmacokinetic properties

pharmacokinetics: examines the fate of a drug from the moment that it is administered up to the point at which it is completely eliminated from the body

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important parameters are:

* absorption - The process of a substance entering the blood circulation
* distribution - The dispersion or dissemination of substances throughout the fluids and tissues of the body
* metabolism - The drug is recognized by the body and broken down
* excretion - The removal of the substances from the body

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bioavailability – How much of the administered dose gets into the system

adverse effects

allergic reactions: some people develop hypersensitives to antimicrobials

suppression of normal microbiota: when normal microbiota are killed other pathogens may be able to grow to high numbers (cdiff)

antibiotic development timeline

antibiotic drug discovery

types of AB drugs

10+ years, $50 million dollars

less antibiotics being created bc money

natural products, semisynthetic compounds, synthetic compounds

natural products

by definition antibiotics are compounds __produced by microbes__ that kill or inhibit growth of other microbes

most antibiotics come from soil microbes (bacteria)

antibiotics are compounds produced by microorganisms typically as **secondary metabolites** or as a response to stress  = antibiotics

Why do microorganisms produce compounds that kill other microorganisms?

must compete with other microbes for habitat

soil microbes?

less than 1% can be grown in lab

scientists grew unculturable bacteria in agar in the soil

semisynthetic compounds

chemical modification of naturally occurring antibiotics leads to semisynthetic antibiotics

penicillin G and V = natural

ampicillin, amoxicillin, methicillin = semisynthetic

synthetic compounds

synthetic antibiotics are compounds that are __**not found in nature**__

they are completely synthesized in a lab

how many chemical structures are possible?

two major pathways in synthetic drug design:


1. combinatorial chemistry
2. rational drug design

combinatorial chemistry

begin with a core compound

synthesize thousands of derivatives by adding groups at specific positions (e.g. R1 and R2)

combine different compounds together in groups (1,000’s of compounds in each group)

pool 1,000’s of compounds together

test pooled compounds for antibacterial activity

when activity is detected determine which compound is responsible by __**deconvolution**__

breaking large group into smaller groups and re-testing

HTS – High Throughput Screening

(creating sub-libraries to search for activity)

rational drug design

rational drug design is a targeted approach to antibiotic development

select a specific bacterial target (typically one that is required for survival/infection)

using a variety of approaches design a molecule that will inhibit that target

depends heavily on computer based modeling of target:inhibitor

HTS also used to optimize compounds

(doesn’t work as well as we hoped)

11 major classes of antibiotics

β-lactams

Glycopeptides

Aminoglycosides

Tetracyclines

Macrolides/lincosamides

Streptogramins

Fluoroquinolones

Rifampin

Trimethoprim/sulfonamides

Metronidazole

Oxazolidinones

antibiotic targets 4

cell wall

protein synthesis

DNA/RNA

tetrahydrofolic acid biosynthesis

cell wall synthesis inhibitors

1 AB: 4 subgroups

β-lactam antibiotics

* penicillins
* cephalosporins
* carbapenems
* monobactams

kill bacteria by inhibiting the last step in peptidoglycan biosynthesis

transpeptidation: crosslinking peptide side chains

carried out by penicillin binding proteins = transpeptidase

PBP’s inhibited by beta-lactam antibiotics

(PBP’s crosslink peptidoglycan)

protein synthesis inhibitors 5

euk ribosome v pro

ribosomes in pro are very different that euk

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eukaryotes –   __80S Ribosome__

  40S -Subunit

  60S –Subunit

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prokaryotes –   __70S Ribosome__

  30S -Subunit

  50S -Subunit

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pro ribosome is sufficiently different than eukaryotic, compounds that inhibit prokaryotic ribosome do not inhibit eukaryotic (or do so poorly)

makes prokaryotic ribosome an attractive target

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types:

* aminoglycosides - block 30
* tetracycline - block 30
* macrolides - block 50
* azithromycin
* lincosimides - block 50

aminoglycosides

structure

functional against

function

3 examples

trisaccharide structure

against many Gram- and some Gram+

bind to specific sites in the 30S subunit

block protein synthesis

* gentamicin – Used for acute, life-threatening Gram- infections
* neomycin – Too toxic for internal use.  Used topically for skin infections
* streptomycin – Active against Mycobacterium tuberculosis

tetracyclines

four fused cyclic ring structure

broad-spectrum

bind to specific sites in the 30S subunit

generally bacteriostatic

low toxicity

Tet binds to Ca in growing bones and teeth and can discolor teeth

should be avoided in children < 8 yrs old

DNA/RNA synthesis inhibitors

metronidazole: inhibits NA synthesis by disrupting the DNA of microbial cells, interacts with alcohol = projectile vomiting

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quinolone: nalidixic acid

fluoroquinolones: norfloxacin, ciprofloxacin

both inhibits DNA replication by binding to and inhibiting DNA gyrase after gyrase has introduced double stranded nicks in DNA

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rifampin: inhibits RNA polymerase by binding to the beta subunit and preventing transcription

tetrahydrofolic acid inhibitors

tetrahydrofolic acid is an essential co-factor in bacteria

required for synthesis of nucleic acids and formylmethionine (fMet)

bacteria make their own via the FH4 biosynthesis pathway

humans require FH4 but do not make it

makes it an ideal target for antibiotics

* **Sulfonamides**
* **Trimethoprim**

bacteria can make FH4 but not uptake, humans cannot make and must uptake

resistant

susceptible

\*get wording right

somewhat arbitrary designation that implies that an antimicrobial **WILL NOT** inhibit bacterial growth at clinically achievable concentrations

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somewhat arbitrary designation that implies that an antimicrobial **WILL** inhibit bacterial growth at clinically achievable concentrations

MIC

MBC

MBEC

MIC: minimal inhibitory concentration, lowest concentration of antimicrobial that completely inhibits growth of bacteria, common in clinical lab (no growth in tube)

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MBC: minimal bactericidal concentration, concentration of an antimicrobial that completely kills bacteria, used only in special circumstances (no growth on plate)

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MBEC: minimum biofilm eradication concentration, concentration of an antimicrobial agent required to kill a bacterial biofilm (typically have highest concentration)

antibiotics vs AB resistance

happening forever

increase in resistance with increase in AB use (humans)

bacteria usually become resistant in 3-5 years loosely

antibiotic misuse

recognized for several decades that up to 50% of antimicrobial use is inappropriate

given when they are not needed/at the wrong dose

continued when they are no longer necessary

discontinued prematurely = resistant bacteria grows

broad spectrum agents are used to treat very susceptible bacteria

agricultural misuse: additional of sub-therapeutic levels of antibiotics in livestock feed results in 10% increase in body mass, unknown why, banned in europe, 70-80% antibiotics sold in US used for animal production

antimicrobial action

bactericidal: kill bacteria

bacteriostatic: slow down or inhibit growth of bacteria (immune system then kills bacteria)

mechanisms of antibiotic resistance

intrinsic resistance

acquired resistance

intrinsic resistance

intrinsic resistance is the innate ability of a bacterial species to resist activity of an antibiotic through its inherent structural or functional characteristics

organisms that are intrinsically resistant have __**never been susceptible**__ to that particular drug

* lack of __affinity__ of the drug for the bacterial target
* inaccessibility of the drug into the bacterial cell
* extrusion of the drug by chromosomally encoded active exporters
* innate production of enzymes that inactivate the drug

organisms, natural resistance against, mechanism

tolerance/ persister cells

persister cells are a small sub-population of dormant bacteria

metabolically inactive

because they are not growing they are able to tolerate antibiotics

not true resistance

not a spore, everything inside cell is shut down

how they survive? AB target protein synthesis/DNA/RNA but cannot affect persister cells bc they are shut down

normally AB would kill persister cells

unknown length of dormancy

always maintain a subset of persister cells, incase a colony gets wiped out the persister cells can regrow when AB leaves

biofilms with persister cells: AB kills all but persister cells, ECM is left behind, persister cells can regrow and regrow biofilm. In the same ECM as usual

acquired resistance

acquires resistance occurs when a particular microorganism obtains the ability to resist the activity of a particular antimicrobial agent to which it was previously susceptible

this can result from 2 different genetic processes:


1. mutation of genes
2. the acquisition of foreign resistance genes (most common - HGT)

mutations of genes

random changes in the DNA

acquired resistance

mech of action 3

acquisition of foreign resistance genes

HGT: resistance bacteria transfer DNA to sensitive bacteria

3 mechanisms of acquired resistance:


1. inactivation of the antibiotic
2. target modification
3. removal of antibiotic