S. aureus Flashcards

Question

What does E-Cadherin do and what does its cleavage cause? When is it cleaved?

Answer 1

E-Cadherin is involved in adherens junctions between epithelial cells S. aureus destroys adherens junctions between epithelial cells to disrupt epithelial barrier; Can get into tissues Alpha hemolysin interacts with ADAM10 (protease) which enhances its activity and it cleaves E-Cadherin

Answer 2

Has the ability to activate T cells non-specifically; Massive T-cell activation can cause scarlet fever and toxic shock syndrome

Answer 3

Superantigen Affects immune system by stimulating massive non-specific T cell response S. aureus causes food poisoning via preformed toxin

Answer 4

Superantigen Induces proinflammatory cytokines; Causes fever, shock and death

Answer 5

Proteases Destroy epithelial layer, leading to sloughing of skin (scalded skin syndrome)

Answer 6

Destroy epithelial layer, leading to sloughing of skin (scalded skin syndrome)

Answer 7

Small peptides Associated with a community acquired MRSA strain

Answer 8

MRSA strain that has not developed antibiotic resistance due to the use of antibiotics

Answer 9

Virulence factors that cause blood clots S. aureus is a skin organisms along with many other species which are more abundant - After swabbing the skin and culturing, you need to determine whether S. aureus is present - If coagulase is produced, that is a sign of S. aureus - Take supernatant from culture and see if it clots plasma, showing that coagulase is present

Answer 10

Coa and vWbp Form complex that interacts with prothrombin to form staphylothrombin Staphylothrombin converts fibrinogen (ECM protein) into fibrin Fibrin prevents neutrophil access and protects bacteria in abscess - Forms fibrin coat outside abscess

Answer 11

Necessary to produce the right set of determinants, at the right amount in the right place Also needs the ability to rapidly change according to its rapidly changing environment

Answer 12

Responds to environment via a coordinated response that allows it to be fit as possible within that environment Human are good at coping with organisms like S. aureus; If its going to survive and proliferate then it’s going to have to be in a very interactive process with its environment

Answer 13

Produces several specific virulence determinants at specific times during its lifecycle A pathogens lifecycle changes drastically; Very different from when it first infects and when its established an abscess Many determinants allows it to adapt to a specific niche

Answer 14

During the infection process, the niche the pathogen occupies changes Change is brought about by toxins from the pathogen, but also changes induced by the host in response to pathogen

Answer 15

Has components which can control more than 1 virulence determinant at the same time There is a growth phase dependent virulence determinant production

Answer 16

Exponential: - Many surface proteins - Few exoproteins Stationary: - Few surface proteins - Many exoproteins

Answer 17

Spa, Fnbp etc. Often involved in immune evasion and adhesion

Answer 18

Hla, TSST etc. Toxin enzymes that break down host tissues

Answer 19

Surface proteins are adhesins involved in immune evasion; Happens at the beginning of infection, as they need to avoid the immune system and stick to host proteins There’s an abundance of nutrients in the host, so after using up all the nutrients in an abscess, the pathogen switches to stationary phase in order to produce toxins to break down tissues for nutrients The toxins allow pathogen dispersal and tissue damage

Answer 20

194 Large range of signalling processes were all these components funnel in and interact with each other - All are inhibiting and promoting each other

Answer 21

Complex divergon; 2 divergently transcribed operons with linked functions in the switch process Central regulator involved in switch between surface and exoproteins - +ve regulator of toxin production, upregulating many different toxins and exoproteins - -ve regulator of surface proteins

Answer 22

+ve regulator of agr expression Binds agr promoter region This was discovered using transposon mutagenesis, looking for S. aureus mutants with reduced toxin production/haemolysis

Answer 23

AgrA AgrC AgrD AgrB Hld

Answer 24

Pre-peptide (45 aa) of extracellular density dependent signalling molecule (8 aa) This is autoinducing peptide (AIP)

Answer 25

Secreted signalling molecule to which the organism responds by switching from making surface proteins to exoproteins Bacteria responding to the component which they make; Quorum sensing Several different AIPs according to the strains of S. aureus which make them - All have a common cystine residue that is post-translationally cyclised by a thioester linkage

Answer 26

Encodes AgrD processing enzyme AIP sequence is clipped from AgrD pre-peptide by AgrB Cyclisation then occurs and MroQ clips it at N-Terminal end to form AIP (8 aa)

Answer 27

P2 (leftward) P3 (rightward)

Answer 28

Constitutively expressed at a low level during exponential phase (early on in abscess formation, plenty of nutrients) Slow build up of AIP As bacterial cell density increases, doe does AIP conc. Reaches threshold level of AIP and expression from P2 goes up (mirrors growth phase) This is called density dependent signalling

Answer 29

Correlating increase in AIP conc., so the bacteria must signal to switch to producing exoproteins/toxins Signal is transduced by agrC and agrA

Answer 30

2 component sensor-regulator transduction mechanism (very common regulatory mechanism) AgrC senses AIP and it eventually reach threshold Causes conformational change in agrC, becoming phosphorylated and active It transfers the signal, phosphorylating its cognate regulator, agrA (regulatory protein/transcriptional activator); Pi goes from histidine to aspartate residue AgrA is activated and causes a change in transcription

Answer 31

Upregulates expression from P2 and P3 - Controlling its own expression; Autoregulatory circuit P2 expression is increased as they go into post-exponential phase P3 expression requires activated agrA; Once it is produced in post-exponential growth phase, this expression is massively increased

Answer 32

RNAIII transcript expression (rightward) Single polypeptide; δ-Hemolysin (phenol soluble modulator i.e. Toxin) - If the ribosome binding site for this toxin is removed, the switch still occurs - What is actually causing the changes in expression? RNAIII is a regulatory effector RNA molecule and causes the switch

Answer 33

Upregulates production of many different toxins Downregulates production of many different surface proteins

Answer 34

Transcription – Mechanism unknown - Can see differences in transcription of many genes - Most likely works by binding to one or more DNA binding proteins - RNA molecules are ‘sticky’ so hard to show specificity Translation e.g. spa, hla - RNAIII controls translation of many virulence determinants - Single molecule that upregulates and downregulates different components at same time - Antisense RNA; Binds mRNAs which encode virulence determinants – Controls translation from specific transcripts

Answer 35

Negative regulator of spa transcript (encodes surface proteins A) when going into stationary phase RNAIII binds antisense to spa and covers the ribosome binding site so it can’t be translated This binding also alters the mRNAs structure, forming a stem loop, which tags the mRNA for destruction by RNAse

Answer 36

5’ region of hla transcript complementary bases form stem loop, tagging it for degradation and there is no translation - Stem loop also covers ribosome binding site, preventing translation The organisms make the transcript, even though it can’t be translated at times, as it allows them to respond and make the switch very quickly

Answer 37

RNAIII binds antisense to hla transcript, stopping the stem loop from forming, and leaving the ribosome binding site free to allow for translation RNAIII positively regulates hla

Answer 38

Binds agr promoter region and upregulates its transcription Represses proteases - Depending on where it binds, it either upregulates or represses Acts via and independently of agr

Answer 39

Degrading host tissues as well as S. aureus proteins (virulence determinants) upon response to environmental changes Proteases control virulence determinant stability by cleaving S. aureus surface proteins to allow them to detach and move elsewhere - Allows swift adaptation by turnover of adhesins

Answer 40

Transcription, translation and stability This allows it to respond to the environment and adapt to a wide range of niches and be a versatile pathogen

Answer 41

S.aureus not only interacts with humans, but also the microbiome found in the nose where it inhabits, even other S. aureus strains Mediated by 3 main groups of AIPs - AIP-1 induces production of toxin group 1 but inhibits toxin production in group 2 and 3 (AIP-2 turns on toxin group 2 and so forth) - This signalling mechanism allows organisms to compete with each other - The inhibition of toxin production in other groups gives a competitive advantage in mixed infections

Answer 42

Organisms meet these antibiotics in their natural environments – Preformed resistance mechanisms Penicillin resistance is a plasmid mediated resistance using β-lactamase enzyme which breaks down penicillin ring

Answer 43

Methicillin MRSA; S. aureus lives in an environment with the organisms that make this type of antibiotic Not many people die of MRSA in UK anymore due to large amounts of screening

Answer 44

Thought that bacteria cannot become resistant to it Toxic meaning precise dosage are needed to kill pathogen without seriously damaging host Vancomycin binds to a structure in the bacterial cell wall that was though to be incapable of alteration

Answer 45

VISA/GISA (vancomycin/glycopeptide insensitive strain) Full vancomycin resistance appeared in 2002; Has not become a pandemic

Answer 46

Originating in a hospital

Answer 47

Resistant to all β-lactams (penicillins, cephalosporins, carbapenems, penems etc.)

Answer 48

Resistant to erythromycin, tetracycline, streptomycin, disinfectants etc.

Answer 49

PBPs catalyse transpeptidation reactions to cross link peptidoglycan; Needed for growth and division This occurs on the outside of the cell in the environment; PBPs are good drug target

Answer 50

Gene encoding PBP2A This has low affinity for β-lactam so it less likely to be inhibited Only present in MR isolates; Essential Picked up environmentally; Horizontal gene transfer from S. sciuri MecA is on the mec element, a 30-50kb region flanked by insertion sequences - Primarily transferred by transduction

Answer 51

PBPs are inhibited and don't work Cells only express MecA in presence of β-lactams Gene is switched off when they are not present; Regulation of mecA expression

Answer 52

MecI and MecR - MecR is a membrane protein which senses β-lactam in the outside environment In absence, mecA expression is repressed by mecI dimer on the promoter In presence of β-lactam, it binds mecR, activating it mecR is a metallo-protease, so when it is activated it hydrolyses mecI This allows for mecA transcription

Answer 53

Both contain active site serine motif; Binds the β-lactam in the same way

Answer 54

Large molecule (≈1000 Da) that inhibits peptidoglycan synthesis Binds the substrate rather than the enzyme; Binds terminal D-Ala - D-Ala residues on peptidoglycan peptide stem, preventing PBP from binding

Answer 55

VISA strains can develop in multiple ways - Change in permeability; Vancomycin is large and has to get through the cell wall to get to the precursor - Thicker cell wall; More binding capacity for vancomycin and harder for it to get in

Answer 56

Acquired from Enterococcus, with the first case being someone with a co-infection of these 2 bacteria Likely has associated fitness costs

Answer 57

Plasmid mediated system Contains vanA which encodes D-Ala – D-Lac ligase Instead of having a terminal D-Ala – D-Ala for vancomycin to target, the pentapeptide stem is altered to D-Ala – D-Lac, preventing vancomycin from binding

Answer 58

- Expensive to discover new antibiotics; $1 billion and 10-12 years for 1 drug to get to sale - This is for the testing of all the compounds that funnel down to the final 1 product - Toxicity and pharmacokinetics are a big issue; Drug needs to get to the place where they work, at a sufficient concentration - Clinical trials are lengthy, and drugs can be discovered to not work - Less incentive as new effective antibiotics want to be rarely used to limit resistance development; Less profit compared with other drugs (e.g. Ozempic) For these reasons most large pharma companies have moved away from antibiotic development

Answer 59

Cost effective Effective against resistance mechanisms Broad spectrum; If someone presents symptoms, time is of the essence so diagnostics to find out the specific bacteria take too long; Broad spectrum also allows it to be used in more cases Use in L and MICs

Answer 60

Empirical - Screening of environments to find organisms which will inhibit growth of chosen marker/pathogen Rational - Use existing targets e.g. Methicillin which was a semi-synthetic β-lactam based on penicillin - Use novel targets; Analysis of genomes to discover new targets and drugs

Answer 61

Analogues of existing drugs that are stable to degradation e.g. amikacin (kanamycin analogue), which is stable against the degradative mechanism which provides resistance to kanamycin Use analogues which aren’t effluxed e.g. glycylcyclines (tetracycline analogues); Useful for when targets are in the cytoplasm as efflux pumps provide resistance Use analogues that bind to modified target e.g. carbapenem derivates that bind and inhibit mecA Inhibit inactivating enzymes e.g. clavulanic acid (β-lactamase inhibitors) Inhibit resistance mechanism to make the bacteria sensitive again

Answer 62

Daptomycin (a.k.a Cubicin) - Cylic lipopeptide - Membrane active; Insert into membrane and allows release K+ - Prevents production of ATP and organism dies - Believed to bind to intermediates involved in peptidoglycan synthesis; Multiple modes of action Linezolid (a.k.a Zyvox) - Oxazolidinone - Binds ribosome 50s - Prevents initiation complex formation, preventing translation Resistance has appeared to both of them

Answer 63

Transpeptidase activity (joins pentapeptide stems), carried out by PBPs, and transglycosylase activity (joins disaccharide units) to join PG molecules PBP2A has an active site and an allosteric site - Active site contains serines and has low affinity for the antibiotics - Allosteric site can bind substrate, which exposes substrate binding site for β-lactam, allowing inhibition

Answer 64

Ceftobiprole - Binds active site; This is only open when the conformation has changed, so not that useful Ceftaroline - Binds active site and allosteric site; β-lactam can bind, preventing PBP2A from being activated Resistance appeared for both; Mechanisms of resistance often don’t involve changes to the enzymes, but rather changes to the cellular physiology

Answer 65

Complex natural antibiotic compound Not likely due to complicated chemistry, but derivatives might

Answer 66

Screened soil using methods like iChip to find organisms that produce antibiotics iChip used to grow bacteria that can't normally grow in media; Allows it to grow in its natural environment with the necessary nutrients They then put S. aureus in each well with different bacteria, and identify wells where S. aureus growth is inhibited Find teixobactin, produced by Elephtheria terrae, a gram-negative proteobacterium - This bacteria could then be trained so that it could grow in vitro

Answer 67

Produced by 2 enzymes - Each contains several different domains which provide different enzyme activities to make it Multiple enzyme activities so hard to produce in the lab

Answer 68

Large molecule (1242 MW) Depsipeptide

Answer 69

Don't kill cells as effectively as they do compared to exponential phase Teixobactin effectiveness not as reduced Causes cell lysis where others don't

Answer 70

Binds and inhibits both lipid II (carrier of muropeptide for peptidoglycan biosynthesis) and lipid III (teichoic acid biosynthesis) Cell wall synthesis is then inhibited

Answer 71

Target isn't a protein so mutations in a gene and therefore protein structure will not affect it

Answer 72

Bacteria have it but we don’t Open to the environment in gram-positive; Teixobactin only effective in gram-positive – Harder to make antibiotic for gram-negative due to the outer membrane Difficult to change as it’s a basic carrier molecule and is not a protein

S. aureus Flashcards

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