evasion of toxins

• adherence ->adhesine

• invasion -> invasine

• evasion -> etabline

• damage -> Toxine, Wirtsreaktion

• antatomical barriers

• complement system

• phagocytosis

• iron limitation

• hidden AGs

• AG variation

• immunsuppression

• immediate defense against pathogens in a non-specific manner

• no long-lasting or protective immunity to the host

major functions:

• acting as a physical and chemical barrier to infectious AGs - most particular against bacti

• activation of the compliment system to identify/destroy bacteria and to promote clearance of dead cells or AB complexes

• recruiting immune cells to sites of infection/production of specialized chemical mediators (cytokines)

• identification and removal of foreign substances by specialized white blood cells.

• Activation of the adaptive immune sytsem by AG presentation

• epithelal surfaces: physical barrier, impermeable to most infectious (passive) agents

• removing pathogens by movement of the epithelial surface due to peristalsis or cilia movement (gastrointestinal and respiratory tract) (active)

• flushing of the eyes and mouth. tears and salvia contain antimicrobial proteins eg. lysozyme, lactoferrin

• oral and gut microflora can prevent the colonization of pathogenic bacteria. (secreting toxic substances or competing for nutrients or attachment to cell surfaces)

pic 2 -3s

• opsonization: bacti are coverd with complemt proteins. stronger / forced recognition - enhancement of phagocytosis by coating with C3b

• cytolysis: loss of cellular contents through transmembrane channel formed by membrane attack complex C5-C9

• inflammation: increase of blood vessel permebaility and chemotactic attraction of phagocytes (Mast cell with histamin into bloodstream)

C5a pepitdase:

• - streptococcus pygenes, div. strptococcus sp.

• prolytic cleavage of complemt protein C5a

• C5a is the most potent peptide mediator of inflammation. it serves as a chemo-attractant to recruit phagocytes to the site of infection.

• C5a peptidase: anti-phagocytic virulence factor

Factor H binding Protein:

- Yersinia enterolytica, neisseria meningitidis

• factor H is a regulator of complement system activation

• fHBP enables bacteria to evade killing by the complement system

• fHBP is already in use as a component of a recombinant vaccine against N.meningitidis

• recognition and ingestion of microbial pathogens into plasma membrane-derived vesicles (phagosome)

• receptor recognize precise molecular patterns associated with microorganisms (non-opsonic or opsonic receptors)

• non-opsonic receptors identify pathogen-associated molecular patterns (recognize directly)

• opsonic receptors bind to host produced molecules (opsonins)

• opsonins bind to microorganisms and mark them for ingestion (e.g.ABs, complement proteins)

• the plasma membrane covers microorganisms, closes at the distal end and forms a vacuole, the early phagosome.

• the early phagosome fuses with endocytic vesicles, leading to the late phagosome => formed from ER & Golgi

• the late phagosome turn into a microbial vacuole, the phagolysosome, by fusiing with lysosomes (phagolysosome maturation)

• results of this process:

  • remodeling of the membrane

  • progessive acidification of the phagosome ->pH drops down

  • creation of an oxidative and degradative milieu

Antimicrobial effectors inside the phagolysosome:

• low pH

• myeloperoxidas can transform H2O2 into hypoclorous acid HOCI

• NO radicals are produced by the inducible NO synthase

• NO reacts with O2 to form perocynitrite

• lactoferrin captures Fe2+

• defensis (Antimicrobial peptides)

• Cathepsis (lysosomal proteases)

• Lysozyme (degrades peptidoglycan)

• Type 2A phospholipase A2 degrades anionic phospholipids

by passing phagocytosis

• encapsulated bacteria:

  • capule prevents contact to phagocytes

  • e.g. s. pneumoniae; b.anthracis

• grows as a yeast (unicellar) and replicates by budding

• proce a characteristic polysaccharide capsule

• mutants impaired in capsule biosynthesis are attenuated or completely avirulent

cryptococcosis is an opportunistic infection for immunocompromised patients (AIDS)

In humans, C. neoformans causes 3 types of infection:

• wound or cutaneous cryptococcosis

• pulmonary cryptococcosis

• cryptococcal meningitis

• can induce clotting

• secreted coagulates activate prothrombin(!) to generate fibrine

• coagulate activitiy is essential for:

  • the formation of s. aureus-fibrin-platelet microaggregates

  • the homing of s. aureus to the vascular wall under flow

• Clumping factors A und B (ClfA and ClfB), facilitate S.aureus binding to fibrinogen and lead to agglutination of clumping of cells

cellwall proteins -> sticks to fibronogen

-> to big for phagocytosis

toxin production bacteria:

• killing of phagocytes by bacterial toxins e.g. leukocidin

• streptococcus spp; Staphylococcus aureus

secreted pore forming toxins:

• leukocidin ED, y-haemolysin AB, HlgCB, LukAB, Phenol-soluble

ProteinA - s. aureus

Protein G - streptoc. spp

binding Fc region in IgG

LH recognizes pathogen surface

HC recoginizes receptors

• Protein A can promote bacterial aggregation and the formation of biofilms

• inhibitiom of phagocytosis - Protein A binds also to VWF

• Protein A binds to human von Willebrand factos, an important glycoprotein that mediated platetet adhesion at the site of endothelial damage. if would: binding of blood platelets

• Protein A-VH3 IgM complexes on B lymphocytes stimulate proliferation and apoptosis, reducing the repertoire of potential AB-secreting B cells in the spleen and bone marrow

• Protein A can stimulate inflammation in the lung by binding to receptos for tumor necrosis factor 1 that is widely distributed on the airway epithelium

• phagocyte recruitment is limited by binding of CHIPS (chemotaxis inhibitory protein of staphylococci) to chemokine receptors on leukocyte

• Golden carotenoid pigment provides an antioxidant shield

• catalase detoxifies hydrogen peroxide

• s. pyogenes - leading human pathogen

• the M protein is a key virulence factor in GAS

• The M protein is a fibrillar coiled-coil dimer that extends from the bacterial cell wall

• GAS classified on the basis of the antigenically variable surface M protein (encoded by the emm gene) >200 genotypes

• M-protein helps the bacterium overcome innate immunity

• is multifunctional and interacts with numerous host proteins like: albumin, C4BP, CD46,….

• inhibits phagocytosis of GAS in the absence of opsonising ABs

• Ferrous Iron (+2) intracellulary mostly bound to ferritin and hemoglobin (oxygen transport)

• Ferric Iron (+3) extracellularly bound to high-affinity binding proteins, e.g. Transferrin and lactoferrin ->electron transfer reaction -enzymes

• transferrin and lactoferrin show very high equilibrium constants (10^36) and both proteins are ususally not completely saturated (only 30-40%)

  • not freely solube iron in the body

  • iron concentration required to sustain bacterial growth +/- 1µM

Fe3+: Transferrin, lactoferrin

Heme: haemoglobin, Haemophore

siderophore: Ferrichrome, Enterobacti

• Free heme is scavened by albumin and/or hemopexin

• free hemoglobin is tightly bound by haptoglobin and subsequently cleared by tissue macrophages

• hemophore =bacterial heme binding proteins

• hemopexin = human heme binding protein (recycing/liver)

• haptoglobin = hemeglobin binding protein

• second layer of nutritional immunity

• siderocoalin (lipocalin-2 or neutrophil gelatinase-associated lipocalin (NGAL))

• secreted by neutrophils in response to infection

• binds enterobacti, the primary siderophore of many enteric bacteria

• sequesters the siderophore-iron complex, preventin bacterial uptake

• Regulation (stabe arrangement of genes; no regulation): temporary, adaptive, reversible, complex, induction/repression, function of genes

• Mutation: rare, randomly, undirected

• variation (phase-AG-): pre-adaptive, specific sequencey, reversible (in parts)

AG Variation - temp. different in body while infection; also nutriens different -> bacti has to adapt

• lteration of (surface) AGs that are presented to, and targeted by, the host’s adaptive immune system

• subpopulations of antigenically distinct organisms arise within a population that are temporarily not recognized by the primary adaptive immune response

• expression of alternativ forms of a particular AG

Phase - Variation

• genes encoding surface AGs are switched “on/off”!!

  (expression of an AG alters between 2 stages of expression)

• expression of one multiple antigenic forms of a specific protein

no stable arrangements bc of changes in the genome

How does phase variation come to play? -> AS proteins which influence recognition

• high spontaneous mutation frequency, typically in viruses (e.g. HIV-1, influeza virus, often calles AG drift

• reassortment of genome parts or segments, typically in viruses, referred to as AG shift

• specific modification of DNA sequences at defined genomic loci by homologous DNA recombination

• lytic enzymes : Proteases, Phospholipases

• pore forming enzymes (a-toxin -> porenbildung und austritt von Zellinhalt führt zu Zelltod)

• thiol activated toxins:

  • gram + bacteria

  • receptor: Cholesterol

  • high homology, cross-reactive ABs

  • active at low pH (in parts)

  • polymerization, building very large pores up to 35nm, release of solutes and macromolecules

  • streptolysin O, Listeriolysin u.a.

    
    

ADP-Ribosylating toxins

glycosidaes

Botulinium

Tetanus

• gram - bacti

• “Repeats in Toxins: Tandemrepeats 9 aa

• ca-dependent pore formin toxins

• encoding genes: A strucutral genes; B/D secretion; C activator

• E. coli: a Hemolysin

• A. pleuropneumoniae: Apx Toxins

Most potent toxins target the NS

• botulinum neurotoxins, causing botulism, caused by clostridium botulinum

• tetanus neurotoxin, causing tetanus, caused by clostridium tetani

BoNT and TeNT have the same molecular architecture (-150-kDa protoxin) with 3 functional domains

• LC, N terminal 50kDa, a zink-dependent protease

• HC 100kDa, contains…

  • an N-terminal Domain, responsible for delivering the LC into the cytosol

  • C-terminal domain responsible for recognizing specific cell surfaces

• target and enter motor nerve terminals at neuromuscular junctions via receptor-mediated endocytosis

• acidification triggers conformational changes of the toxins that result in transfer of the LC into the cytosol

• in the cytosol LC bocks neurotransmitter release from nerve terminals by cleaving SNARE proteins

….but a distinct destination in the Body which determines their symptoms

Snaoe proteins constiture the core complex that mediates membrane fusion events in the eukaryotic cells

BoNT and TeNT specifically cleave SNARE proteins that mediate fusion of synaptic vesicle membranes to the presynaptic membrane in neurons

Botulism is usually caused by food poisoning due to ingestion of BoNTs

BoNTs target and enter peripheral motor nerve terminals

the typical symptom of botulism is flaccid paralysis - the inability to contract skeletal mucles. ultimatley, patients die from respiratory failure caused b diaphragm paralyis

• spores contaminate deep wounds. TeNT is produces in situ and directly enters the circulatory system

• TeNT is transported retrogradely along the motor neuron axon axon to the soma

• TeNT is released from motor neurons and reenters connecting inhibitory neurons, where it blocks neurotransmitter release.

• losing inhibitory input leads to hperactivity of motor neurons, resulting in spastic paralysis