MICR 483 Midterm

Symbiont

relationship in which both organisms benefit

i.e., lactobacillus in female genital tract

pathobiont

any potentially pathological organism, under normal circumstances is a symbiont

i.e., E. coli

commensal organism

organism that benefits from the host, but is neutral to the host

pathogen

an organism that can and does cause disease

I.e., HIV

Locations of normal flora

oral cavity

nasal cavity

gut

vagina

skin

Benefits of normal flora

prevent colonization of pathogens

participate in physiologic functions

• bacteria in gut aid digestion, prevent C. Diff colonization

reduce cancer, CF, IBD, Crohn’s

metabolizes therapeutics

Gram positive bacteria

stain purple

thick peptidoglycan layer

gram negative bacteria

stain pink

thin peptidoglycan layer

Skin microbiome

staphylococcus aureus (coagulase negative)

Upper respiratory tract microbiome

mouth/oral/nasal cavities

pharynx/larynx

staphylococcus, streptococcus, Haemophilus

Lower respiratory tract microbiome

sterile

GI tract microbiome

enterococci, pseudomonas, Enterobacteriaceae (E. coli, Klebsiella spp., enterobacter), streptococci, lactobacillus, candida

Urogenital tract microbiome

urinary and reproductive

lactobacillus, streptococcus

female: pre-puberty and post menopause, microbiome is same as skin. During reproductive years, lactobacillus is prominent, to produce lactic acid to maintain pH and prevent colonization of bacteria from GI tract

SCFAs and MAMPs

short chain fatty acids, microbe-associated molecular patterns

influence health (lymphocyte maturation, epithelial gut health, neuroendocrine signalling)

Dysbiosis

perturbations of normal flora

caused by diet or antimicrobials

Diet causing dysbiosis

change composition of normal flora

high fat diet, impairs epithelium in gut

consequences of gut barrier impairment

metabolic endotoxemia (LPS/endotoxins in blood)

translocation of bacteria into blood/tissues

Barriers of the innate immune system

urination

mucociliary elevator

cough reflex

lysozyme (antimicrobial enzyme)

lactoferrin (competes with bacteria for iron)

IgA (antibody in Bodily secretions)

normal flora

Steps of inflammation

injury to tissue (cut, scrape) results in release of damage signals

these signals recruit immune cells (neutrophils, phagocytes)

results in innate immune response, and secretion of vasoactive/chemotactic factors

• increases permeability, increases fluid movement into site, vasodilation

neutrophils adhere to endothelial cells, and migrate to infected tissues via extravastion

neutrophils and phagyocytes engulf pathogens

steps of phagocytosis

1 - pseudopods surround mcirobes

2- microbes engulfed into cell

3- vacuoles and lysosome fuse

4- toxic components and lysosomal enzymes destroy microbe

5 - microbial debris released by exocytosis

Adaptive immune response components

Humoral

Cell mediated

Humoral immune repsonse

antibody mediated

• neutralization

• opsonization

• complement activation

cell mediated immune response

mediated through T cells

cytotoxic T cells

• destroy compromised cells by recognizing antigen on cell surface

helper T cells

• release cytokines to support cytotoxic T cells

Treg cells

• suppress activation of T cells

• prevent atherosclerosis

activation of CD8 T cells

1 - CD4 T cells recognize antigen presented by MHC-II

2- CD4 T cells become activated, release cytokines

3- cytokines stimulate CD8 T cells

4- CD8 T cells interact with MHC-I complex, and produce granzymes and perforins

5 - granzymes/perforins lyse infected cell

Treg cells and atherosclerosis

inhibit dendritic cells, and pro-atherogenic T cells

promote differentiation of macrophages into anti-inflammatory macrophages

suppress activation of endothelial cells

inhibit foam cells (macrophages), which ingest LDLs and localize around plaque

lower LDL and VLDLs

Primary lymphoid organs

thymus, bone marrow

where lymphocytes are formed and mature

secondary lymphoid organs

lymph nodes

spleen

where lymphocytes become activated

Biofilm components

can support aerobic and anaerobic populations

require constant nutrient supply

facilitate quorum sensing

biofilm life cycle

adhesion

• one bacteria adheres to surface

accumulation of other bacteria

maturation

dispersion

• when biofilm requires more nutrients, bacteria break off and relocate

Bacterial evasion of immune response

biofilm production

capsules

• prevents phagocytosis

• thick polysaccharide layer

toxin production

• endotoxin (LPS)

  • not secreted

• exotoxin

  • gram positives

  • include enterotoxin (gut), neurotoxins, and cytotoxins

metabolic dormancy

• inhibit growth

• inhibit immune system and antibiotic targeting

Impacts of antibiotics

Positive: reduction of bacterial infections, prophylaxis, reduced morbidity/mortalityn

negative: antibiotics can exacerbate symptoms, and lead to C. Diff

Major antibiotic groups

cell wall synthesis inhibitors (B-lactams, glycopeptides)

protein synthesis inhibitors (aminoglycosides, tetracyclines, macrolides)

nucleic acid inhibitors (fluorquinolones)

metabolic inhibitors (folate inhibitor)

B-lactam antibiotic types

penicillins

• narrow spectrum

cephalosporins

• 1st gen: narrow spectrum, gram positive

• 2nd gen: increased activity against gram negative

• 3rd gen: broad spectrum, good against negative, less good against positive

carbapenems: used in severe/multidrug resistant infections

monobactams

B-lactam mechanism

binds transpeptidase enzyme complex (PBP)

prevents formation of NAM-NAG crosslinks

weakens cell wall, leads to death

glycopeptide antibiotics

inhibit cell wall synthesis by preventing cross linking of NAM D-Ala-D-Ala subunit

act on gram positive

include vancomycin

drug of choice for MRSA

Protein synthesis inhibitors

Bacteriostatic!!

Tetracycline: bind 30S ribosomal subunit, block attachment of new tRNA

Macrolides: Bind 50S subunit, inhibit peptidyl transferase

aminoglycosides: bind 30S, prevents proofreading

nucleic acid inhibitors

inhibit DNA replication

bactericidal

fluoroquinolone

• inhibit topoisomerase (separation of daughter strands) and DNA gyrase (supercoils)

• 1st gen: gram negatives (not used in Canada)

• 2nd gen: Pseudomonas aeruginosa (gram positive), first line for community-acquired pneumonia

• 3rd gen: not in canada, treat streptococcus pneumoniae

• 4th gen: used to treat TB

Metabolic inhibitors

bacteriostatic

folate inhibitor

• inhibit tetrahydrofolic acid, so bacteria cannot synthesize DNA and protein

• trimethoprim: inhibits dirhydrofolic acid, preventing synthesis of THF

• sulphonamide: inhibits synthesis of dihydropteroic acid, prevent synthesis of THF

Developing antibiotic resistance

Acquired: bacteria become resistant when exposed to antibiotic. develops over several generations due to random mutations

inherited: bacteria in future generations after resistance has been acquired, can inherit from their parent

Bacterial cost of antibiotic resistant

slower replication, use more energy, are less fit

types of resistance phenotypes

always on

inducible

Mechanisms of antibiotic resistance

enzymatic inactivation (i.e., B-lactamases)

efflux pumps (pump abtibiotics out)

altered binding sites (so antibiotic cannot bind)

• MRSA = alter PBP to PBP-2a so cannot be recognized by B-lactams

reduced permeability of plasma membrane

• Carbapenem-resistant organisms

• porin loss, narrow channels, decreased expression of porins

causes of multi drug resistant bacteria

high antibiotic use (selective pressure)

poor knowledge (overuse, use for viral infections, incomplete prescriptions)

elderly (poor immune response, more prescriptions, cohort)

Sharing of resistance mechanisms

Conjugation

• pilus contact → F+ donor forms plus with F-, transfer of mobile genetic elements

transformation

• naked DNA taken up

• via free DNA fragments or plasmid

• cell must be competent to take up genetic material

Transduction

• bacteriophage

MRSA

dysbiosis leads to skin infections, bacteraemia, endocarditis, pneumonia

resistance gene MecA results in change in PBP to PBP-2a, become resistant to methicillin (B-lactam)

susceptible to vancomycin

MRSA screening

most infections in hospitals and institutions (i.e., prisons)

screen via nasal swabs and perineum

prevention: hand hygiene, surveillance

VRE

vancomycin resistant enterococci

in dysbiosis, leads to UTIs

resistant to many drugs: vancomycin, cephalosporins, ciprofloxacin, folate inhibitors, ampicillin

usually occurs in immunocompromised patients

VanA resistance gene changes D-Ala-D-Ala to D-Ala-D-Lac

VRE screening

rectal swabs

not always done, since VRE not very pathogenic

ESBLs

extended spectrum B lactamases

inactivate B-lactams

sensitive to carbapenems

AmpC genes: chromosomal, only passed vertically

• occur in SPICE organisms

ESBL resistance risk factors

use of antibiotics

travel

hospitalization

carriage of SPICE organisms in GI tract

Carbapenem-resistant organisms (CROs)

any gram negative resistant to Carbapenem

degrade carbapenems, or porin mutations prevent Carbapenem uptake

CRO genes: KPC-2, NDM-1, OXA-48

KPC-2 location

USA, Greece, Israel

also North America, Asia, Europe, africa

NDM-1 location

europe, USA, India, pakistan

OXA-48 location

Turkey, Middle East, North Africa, Europe

classifications of parasites

endoparasites

• protozoa - single celled

• helminths - multicellular

  • cestodes

  • trematodes

  • nematodes

ectoparasites

• arthropods

Endoparasites

live within host, cause infections

typically permanent until medical intervention

undergo specialization to maximize nutrient absorption

protected from outside environment, but must combat immune system

Protozoa

single celled eukaryotic organisms

live in blood, tissues, intestines

include:

• plasmodium

• giardia

• toxoplasma

Helminths

multicellular parasites

infect GI tract

do not proliferate in hosts

Cestodes

flatworms

type of helminth

infect intestines

transmitted via pork, beef, fish

trematodes

flatworms

type of helminth

infect blood vessels, lungs, GI, liver

nematodes

roundworm parasites

type of helminth

transmitted via contaminated water or food

enterobius

Extoparasites

live on host

i.e., fleas, lice

temporary, intermittent, or permanent

do not have to deal with immune system, but must get nutrients via adaptations i.e., puncturing skin

arthropods

have exoskeleton

6 (fleas, lice, flies) or 8 legged (ticks, mites)

some hematophagous (blood eating) or histophagous (tissue eating)

i.e., scabes

Primary host

where parasite reaches adult stage

able to undergo sexual replication

secondary host

asexual reproduction, develops larvae

reservoir host

harbours infectious parasite

Direct life cycle

only require one host, easier to control with antiparasitics

example for trichostronylus (nematode)

1- once in host, parasite lays eggs

2- under favourable conditions, eggs hatch and release larvae

• larvae in rhabditiform stage (developing, not infective) grow in feces or soil (outside body)

3- larvae become infective filariform larvae

4- contact human host, penetrate feet, and carried via blood to heart/lungs, ascend lungs until pharynx, where they are swallowed

5- larvae reach intestine, where they mature and lay eggs

Indirect life cycle

require one or more intermediate hosts. more complex, more virulent

i.e., toxoplasma

1- unsporulated oocysts shed in cat feces

2- oocyst sporulates, becomes infective. intermediate hosts (birds, rodents) Infected upon ingesting contaminated soil/water/plants

3- oocysts transform into tachyzoites after ingestion by intermediate host, localize in neural/muscle tissue, develop into bradyzoites

4- cats eat infected intermediate hosts, become infected

• animals bred for human consumption can also become infected when ingesting sporulated oocytes in environment

5- humans infected by: undercooked meat, food/water contaminated with cat feces, blood transfusion, and transplacentally

Malarial parasite

plasmodium spp.

spread via anopheles mosquito

protozoan

Anopheles mosquito life cycle

egg

larvae (remain underwater)

pupa (inactive, immature)

Imago (adult stage, fly, reproduce)

mosquito must live 10-21 days (minimum) to transmit parasite

Mosquito vector charactics

higher temps = faster parasite growth

humidity/rainfall

anopheles feeding

• anthrophilic: feed on humans, more likely to transmit malaria human → human

• zoophilic: feed on animals (cattle)

Plasmodium life cycle

Infect human with sporozoites (via mosquito blood meal)

asexual reproduction

sexual reproduction

Plasmodium Asexual reproduction

exo-erythrotic phase

• sporozoites infect liver, mature into schizonts, which mature and release merozoites

• merozoites = motile zygotes, invade host cells and reproduce via multiple fission

• may then differentiate into gametocytes, where the cycle repeats

erythrocytic phase

• merozoites undergo asexual reproduction in RBCs

• release, infect other RBCs

Plasmodium sexual reproduction

anopheles mosquito ingests gametocytes from human host (produced in eco-erythrocytic phase)

in mosquito, sexual reproduction via sporogenic cycle

• gametogeny: male and female gametes fuse, form diploid zygote called ookinete

• sporogony: asexual reproduction, form haploid sporozoites which can infect humans

Plasmodium pathophysiology

sepsis: due to uptake of infected RBCs causing macrophage activation in spleen and production of cytokines

parasite sequestration in brain/lungs: vascular endothelial cells express adhesion molecules

endothelial dysfunction: causes leaky vasculature and issues with cellular perfusion

tissue inflammation: leukocyte infiltration of tissue parenchyma

Malaria symptoms

anemia

fever (due to cytokines)

Malaria treatment

anti parasitic drugs:

primaquine (liver, blood, and gametocyte formation stages)

chloroquine (first line of defence)

atovaquone-proguanil (aka malarono, multi-drug regimen)(for resistant forms)

Anti-malarial resistamce

seen mostly in Southeast Asia (Myanmar, Laos, Thailand)

Malaria and SCD

heterozygotes for SCD have RBCs that rupture prematurely, preventing reproduction of plasmodium

leads to enrichment of SCD genes in areas with high malaria

Reducing global burden of malaria

Prevent mosquito biting

• mosquito nets, insecticides

prevent mosquito breeding

• clean up ponds/streams, prevent mosquito reproduction

Stronyloides stercoralis

aka threadworm

roundworm (nematode)

soil transmitted

• use of human excrement as fertilizer (night soil)

  • common in areas with poor sanitation facilities and poor countries

• also transmitted via textiles

Strongyloids life cycle

free living

• dwell in moist soil with warm climates

• adult males and females produce eggs, which hatch and release rhabitiform larvae

• larvae mature into infective filariform larvae, which penetrate skin

Parasitic

• enter host via skin, migrate via blood or lymph to small intestine

• larvae develop into worms, produce eggs, passed in stool

autoinfection

• host reinfected with parasite already present in body

• eggs hatch in small intestine, release larvae, which disseminate through body

Disseminated/hyperinfection Strongyloides

in malnourished or steroid treated hosts

leads to shock, meningitis, renal failure, DIC

90% mortality

Giardia

protozoa

acquired by feral-oral route (contaminated water)

Giardia risk factors

weak immune system

international travel

giardia forms

trophozoite - active form

cyst - inactive form

• persist up to 11 weeks in environment, better at cooler temps

• resistant to chlorination

• eliminated by UV

Giardia life cycle

Ingestion

excystation

• interaction with gastric contents causes trophozoite to emerge into active state of feeding and motility

asexual reproduction

• binary fission

• travel to small intestine, attach to brush border in duodenum

encystation

• trophozoites pass to colon, become cysts in presence of low [bile salt]

excretion via feces

Giardia encystation

trophozoite: sense external signals (low bile salt and cholesterol)

early phase: transcription of cyst wall proteins, transported into encystation vesicles

late phase: arrival of cyst wall proteins enables construction of cell wall that will be part of the cyst, cells become round

cyst: 2 rounds of DNA replication, leads to formation of cyst with 4 nuclei, and genome polity of 4n

pathophysiology of long term Giardia

cellular level

• villi in intestines shorter, narrower, no longer as absorptive

• inhibition of brush border enzymes (disaccharidases)

tissue level

• apoptosis of intestinal mucosal cells

  • loss of tight junctions, increased water and ion loss

  • inflammation and hyper motility of small intestine

body level

• malnutrition (fat, lactose, vitamin A, vitamin B12), failure to thrive

immune system: chronic fatigue, IBS, allergies

hypokalemic myopathy

Prevention of giardia

avoid contact with feces

cook food, peel fruit when travelling

good hand hygeine

don’t drink contaminated water

toxoplasma gondii

protozoan, causes toxoplasmosis

transmitted via:

• undercooked meat, unpasteurized milk (cyst forms)

• blood transfusion, organ transplant (tachyzoite form)

• soil, fruits, cat litter (oocyte form)

toxoplasma tachyzoite ←→ bradyzoite

tachyzoite → bradyzoite

• initial infection = tachyzoites

• convert into bradyzoites to form cysts in body

• caused by: too high or low pH, heat shock, NO

bradyzoite → tachyzoite

• responsible for chronic disease

• reactivation of toxoplasmosis

• caused by: lack of NO/IFN/TNF/T cells/IL-12

Toxoplasma immune evasion

inhibition of lysis: prevent fusion of phagosome and lysosome

proliferation: replicates to large numbers in vacuole, bursts to release cysts

toxoplasmosis clinical manifestation

immunocompetent: 80-90% asymptomatic, non-specific symptoms (chills, fever, etc). Severe infections = hepatitis, myocarditis, etc.

immunocompromised: CNS disease, myocarditis, etc. Common in AIDS patients

occular infections: due to congenital (75-80% of congenital cases) or acquired infection

can cross placenta, risk of fetal abnormalities (especially early pregnancy)

Toxoplasmosis life cycle

1 - cat sheds cysts in feces (or tachyzoites reside in animal tissues)

2- human ingests cysts or tachyzoites (cysts burst, form tachyzoites)

3- tachyzoites multiply, invade host cells

• may or may not have symptoms

• remain in host tissue indefinitely, may become reactivated

  • brain damage, blindness, or death of fetus

Enterobius vermicularis

aka pinworms

helminths

transmit via:

• direct contact (anus to finger to mouth)

• ingestion of contaminated food

• retrograde (anus to intestine)

• fomites

common in schools, and prisons/long term care facilities

enterobius life cycle

direct, only infect humans

1- egg deposition by adult females on perianal forms.

2- auto infection due to scratching perianal area, and ingestion

3- eggs hatch, release larvae into small intestine, adults establish themselves in cecum and appendix

4 - females migrate to anus, lay eggs in perianal folds

enterobius clinical manifestations

perianal itching

insomnia, restlessness, irritability, impetigo, enuresis, vulvovaginitis

enterobius diagnosis

direct observation: worms observed in perianal area 2-3 hours after slee

scotch tape: tape asshole first thing in the morning, view eggs under microscope

microscopy of fingernails

enterobius infection control

difficult to prevent due to asymptomatic carriers, incomplete cure, and easy transmissibiluty

should treat entire household when one person is infected

meds target only adult worms by inhibition of microtubules and causing neuromuscular blockade