Streptococcus Pneumoniae

Problems with Koch's postulates

-need to find the right, susceptible host - not a certainty that we'll cause pneumonia by putting pneumococcus in an individual (people don't know they're infected with them)

Koch's Postulates

1. Agent must be regularly associated with the disease and its characteristics 2. Agent must be isolated form the diseased host and grown in culture 3. Disease must be reproduced when a pure culture of agent is introduced into a healthy, susceptible host 4. The same agent must be re-isolated from the experimentally infected host

Capsule Synthesis:

A primed sugar (made by NDP) is added to a lipid (fatty acid) by the initiating glycosyltransfease. more sugars added by other glycosyltransferases, lipid linked oligosaccharide is flipped to other side of membrane by transporter. polysaccharide polymerase then adds a growing chain to the oligosaccharide. Finished polysaccharide is usually anchored to the peptidoglycan.

where is s. pneumoniae mainly found?

Africa and southeast Asia

complement cascade

C1 (initial compliment) recognizes antibody and a series of cleaving proteins build a pore which kills cell. the by-products of cleaved proteins act as signaling molecules, macrophage recognizes it to come back to clean parts of the killed cell.

two component system

ComD (sensor kinase) and ComE (response regulator)

ComD and ComE

ComD senses environment, partially outside the cell and binds to CSP. ComE functions as the activator and stimulates high levels expression of early genes.

DNA uptake machinery

ComGC recognizes dsDNA, dsDNA binds to cell, digest dsDNA to ssDNA, single strand uptake by competence system, homologous recombination (get variants of genes you already have)

job of Wzy polymerase?

adds oligosaccharides to one another

penicillin

antibiotic that prevents chain crosslinking

vancomycin

binds to substrate to prevent it from being recognized by enzyme (blocks crosslinking). resistance occurs by replacing the terminal D-ala with either lactate or D-serine

how does capsule influence pathogenesis?

capsule covers peptidoglycan. is inhibits the binding of antibodies and complement to the bacterial cell and recognition of bound antibodies and complement by phagocytic immune cells. bacteria who progress to lower repitatory tend to have less capsule.

quorum sensing

cells indirectly sense their own population density and respond by changes in gene expression. cells directly sense an extracellular signaling molecule called "autoinducer". this typically binds to a histidine protein kinase with an affinity that requires a particular environmental concentration. this is useful for bacteria recognizing DNA. high population density = gene on.

protein component of peptidoglycan

chains cross-linked by amino acids.

CSP

competence stimulating peptide, once out of cell is builds up in growth medium until it reaches a critical concentration and is bound by the sensor kinase ComD. it is an autoinducer that turns on the ComDE 2 component system.

RNA Polymerase

consists of a complex of 4 proteins. 5th subunit (sigma factor) determines which promoters RNA polymerase binds to and controls specificity.

how is pneumococcus spread?

coughing and sneezing

when is peptidoglycan made?

during cell division. point of division is where all peptidoglycan is made

ComX

encodes a sigma factor. sigma factor recognizes -35 and -10. ComX doesn't want sigma factor on all the time so it turns on particular set of genes when it wants certain things. ComX turns on late gene operon, this encodes DNA uptake and recombination machinery. ComX also secreted lysins.

lysozyme

enzyme that cleaves beta-linkage between sugars of the chain. Antibacterial enzyme, NOT antibiotic

job of NDP-sugar biosynthesis?

enzymes involved in adding nucleodiphosphides to sugar. make different sugars available for addition.

what is the benefit of competence (ability of cell to take up extracellular naked DNA from environment)?

for pneumococcus, the advantage is obtaining new capsule genes to evade an antibody response, and share or pass antibiotic resistance.

rough pneumococcus colonies

found to be not virulent, and do not possess a polysaccharide capsule. However, they were transformed to smooth and virulent in the presence of heat-killed smooth bacteria. doesn't cause problems

physical characteristics of streptococcus pneumoniae

gram positive, spherical, polysaccharide (sugar) capsule, naturally transformable.

smooth pneumococcus colonies

highly virulent and have capsule. caused problems with mice.

problems with vaccines for pneumococcus

immune response to pneumococcus is frequently t-cell independent so there is little to no immune memory. repeat infections, repeat vaccinations needed. since capsule is major antigen seen by immune system and there are atom of capsule types, immunity to one pneumococcal strain does not result in immunity to others.

pneumolysin (PLY)

it is a cholesterol dependent cytolysin that associates with cholesterol and oligomerizes to form a large pore. it is important for invasion into bloodstream and hemolysis. at low, sublytic concentrations it can activate the classical complement pathway in the absence of antibody causing lysis of host cells and increased inflammation.

peptidoglycan

it is a hybrid of protein and polysaccharide (protein and sugar).

bacteriology of s. pneumoniae

it is an aerotolerant anaerobic bacterium, non-motile, hemolytic, grown in 5% CO2 on blood agar,

what is transformation and how does it happen?

it is the permanent genetic change of a cell/organism generally resulting in a phenotypic change. we think of it as the uptake of exogenous DNA which requires the bacteria to be in a competent state for transformation.

what is the DNA binding site called?

operator

non-invasive diseases

otitis media, sinusitis, bronchitis . HIGH INCIDENCE. occurs in non-sterile environments in the upper respiratory tract

how does penicillin inhibit peptidoglycan synthesis?

penicillin binding protein binds to the sugar linked peptides and catalyzes crosslink with interpepride bridge, releasing an alanine. it then dissociates from the cell wall and penicillin comes in to bind in its active site. the beta-lactam ring reacts with the serine group of the PBP active site becoming covalently linked.

association of pneumococcus with influenza?

pneumococcus was responsible for 40% of deaths in the 1918 flu pandemic. increase incidence in flu led to increase in bacterial pneumonia.

invasive diseases

pneumonia, meningitis, febrile bacteremia (sepsis). HIGH MORTALITY. occur when the bacteria is spread to previously sterile body compartments such as the lower respiratory tract, the blood stream, and the CNS.

sugar component of peptidoglycan

polysaccharide polymer of NAG (N-acetylglucosamine) and NAM (N-acetylmuramic acid) joined by beta-linkage to form chains.

lysins

produced and secreted by ComX. they kill neighboring bacterial cells.

ComM

protects the competent cell from lysis

sigma factors

proteins that bind specific promoter sequences and instruct RNA polymerase where to begin transcribing.

repressors

proteins that bind to specific DNA sequence and inhibit gene transcription

activator

proteins that bind to specific DNA sequences and activate expression of downstream genes. they stabilize RNA polymerase interaction with promoter. in absence of environmental stimulus, weak activation.

t-helper cells

recognize antigen presented by antigen presenting cells such as macrophage. when activated, they secrete cytokines that stimulate b-cells and cytotoxic t-cells to cause an immune response. t-helper cells do not directly or indirectly kill pathogens, they amplify a specific response.

function of ComAB

secretes PreCSP/CSP

job of Wzx transporter?

takes sugars from inside cell and flip it to be outside cell

problem with pneumococcus and macrophage

the thick capsule prevents recognition of bound antibody by the macrophage to it limits uptake. the antigen needs to be frequently covered in antibody or complement or be a product of apoptosis but thick capsule prevents that. capsule is taken up as an antigen but won't be recognized by t helper cells because they recognize peptide, not sugars.

Promoters

these are DNA SEQUENCES that indicate where to start transcription.

difference between PCV 7 and PCV 13

they both have polysaccharide conjugated to diptheria toxoid CRM197 (kicks on t h cells). In US, we use PCV 13 because it covers more capsule types than PCV 7.

glycosyltransferases?

they determine the types of sugars to be added and the linkages of those sugars.

B-cells

they make antibody, differentiate into plasma cells and memory B-cells when activated by t helper cells.

gram positive cell architecture

thick external layer of peptidoglycan, inner plasma membrane. Cell wall is usual target for penicillin.

MHC-2 Antigen

this gives presentation of peptide. gives rise to a t-helper cell response

operons

this is how most bacterial genes are organized. when multiple genes are expressed from a common promoter. each gene has its own ribosome binding sequence, translational start, and translational stop. they are linked to one another transcriptionally but they are independent components so translation is not linked.

CiaRH

turns system off. it's a 2 component system that is expressed when competence begins (as soon as gene turns on) and eventually over-rides the competence system and represses early gene expression. environmental input leads to transcription being turned off completely.