Chagas Disease - 3 (6/7) Flashcards Preview

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Flashcards in Chagas Disease - 3 (6/7) Deck (53):
1

Chagas

transmission

  • causative agent is transmitted by Triatomine bugsg
  • 138 different species can transmit the disease
  • most improtant are
    • Triatoma infestans
    • Rhodnius proxilus
    • Panstrongylus megistus
  • common names include assassin bugs or kissing bugs (bite people's faces)
    • retractable proboscis

 

2

Triatomine bugs are

  • relatively large (5-30mm)
  • heamatophagous (blood feeding), at night
  • disease transmitted in insected feces ("stercorarian")
  • insect life cycle: egg → 5 nymphal instars followed by sexually mature, winged adults
  • all stages can spread chagas disease

 

3

Chagas transmission in feces

  • inefficient
  • requires a lot of contact between human and vector
  • high levels of host-vector contact has thatched roofs/adobe walls = ideal habitat for bugs
  • triatomid bugs are suually found in tree canopy → used as thatch material

 

4

Zoonotic disease

animal (any mammal) to human

  • several transmissoin cycles
    • sylvatic cycle = wild animals to humans - eg monkeys, rodents, ground squirrels, opossums, armadillos
    • peri-domicillary cycle = "in/around" house involuntarily - eg rodents, foxes, bats
    • domicillary cycle = pets - eg dogs, cats guinea pigs

 

5

Other routes of transmission

  • ingestion of contaminated water/food
    • sugar cane and fruit juice
  • blood transfusion
    • drugs
  • organ transplantation
  • congenital (mother to child)
  • modern medical practices coupled with population movement has resulted in problems in southerns tates of USA and Spain/portugal
  • 300,000+ infected people live in USA
  • estimated 100,000 Spain/Portugal

6

Chagas

causative agent

disease caused by the protozoan parasite Trypanosoma cruzi

7

Chagas 

major form of the parasite - epimastigote

  • common morphology in insect midgut
  • divides by binary fission
  • non-infectious

 

8

Chagas

major form of the parasite - trypomastigote

  • highly infectious
  • insect → humans = metacyclic trypomastigote
  • humans → insects = bloodstream form (BSF) trypomastigote
  • metacyclics and BSF trypomastigotes can invade mammalian cells
  • can't divide
  • in feces of insect vector

9

Chagas

major form of the parasite - amastigote

  • intracellular form
  • divides by binary fission
  • many (500+) parasites in 1 host cell (within mammalian cell)
  • infectious – can invade mammalian cells

 

10

T. cruzi lifecycle

  1. triatomine bug takes blood meal, metacyclic trypomastigote in through bite or mucus membranes
  2. invades mammalian cells near bite (local tropism)
  3. divide to amastigote (and zoid), which divide by binary fission and communicate to form (in sync)
  4. bloodstream form trypomastigote burst into the bloodstream with tropism for tissues (cardiac, muscles, enteric nerves)
  5. BSF taken up by insect, into midgut to form epimastigote
  6. divide by binary fission
  7. in hindgut form nondoviding metacyclic trypomastigote

11

Life cycle 1. 

Q image thumb

  • as triatomine bug takes a blood meal it defecates
  • metacyclic trypomastigotes in the feces enter through host through bite site or mucosal membrane (eg conjunctiva)
  • person rubs skin at site of bite/defecation
  • also in through mouth, eye’s mucous membrane
  • metacyclic trypomastigotes invade various mammalian cells near bite site (local tropism)
  • initial tropism for local macrophages, fibroblasts, and other mesenchymal tissues
  • the differentiation is unequal: nucleus doesn’t divide with the rest of the organelles so get zoid without a nucleus and an amastigote

12

Life cycle 2. 

Q image thumb

  • inside the mammalian cell, the parasite transforms into the amastigote form
  • genetic exchange reported in amastigote stage (not essential for cell cycle progression)
  • amastigote divides in cell cytoplasm, colonizes the cell
  • amastigotes divide by binary fission
  • amastigotes able to communicate to differentiate at the same time into…

13

Life cycle 3. 

Q image thumb

  • intracellular amastigotes transform in to BSF trypomastigotes
  • BSF trypomastigotes burst out of the cell entering the bloodstream
  • BSF trypomastigotes can then infect other cells
  • tropism for cardiac and skeletal muscle and enteric nerves (may lead to pathologies)
  • “nest” of parasites
  • becomes chrnic – tropism for certain tissues
  • eg heart, GI tract

 

 

  • BSF trypomastigotes can be taken up again by the bug in blood meal
  • in the bug it goes to the midgut

 

14

Life cycle 4. 

Q image thumb

  • BSF trypomastigotses can be taken up by a triatomine bug taking a blood meal

15

Life cycle 5. 

Q image thumb

  • in the midgut, the BSF trypomastigotes transform into epimastigotes
  • the epimastigotes divide by binary fission
  • colonizes the midgut, into the hindgut

16

Life cycle 6.

Q image thumb

  • in the hindgut, the epimastigotes transform into non-dividing metacyclic trypomastigotes
    • this form can be transmitted back to humans

 

17

Disease progresses through three distinct phases

  1. acute stage
  2. indeterminate (latent) stage
  3. chronic stage

 

18

Acute stage

general

  • 2-4 months
  • generally asymptomatic
  • mortality in non-immunocompromised patients rare (children)
    • but die of meningoencephalitis, acute myocarditis

 

19

Acute stage (2-4 months) 

site of infection → hallmark sign

erythetomatous skin lesion - chagoma

  • if entered through bite
  • immune system responding to acitvely dividing and growing parasites at that site
  • periorbital edema and conjunctivitis
    • local fluid retention
    • swelling around the eye - Romana's sign only if infection occurs via mucous membrane around the eye
      • only if actively dividing at that site
  • inflammation response to localized replicatoin of intracellular parasite
  • infection is active (ie parasites are dividing)

20

Acute stage (2-4 months)

non-specific symptoms

  • other non-specific systemic symptoms (mild) may present 
    • fever
    • malaise
    • swelling of lymph node/liver/spleen/heart
  • 7-14 days after infection 
  • resolves in 3-4 months

 

21

Acute stage (2-4 months)

host immune responses

  • able to control infection but NOT eliminate it
    • parasites decline over time - our immune system responds and able to reduce parasitemia but not actually clear the parasite
  • immunosuppression (AIDs/Chemotherapy) causes disease reactivation
  • AIDS reactivation →
    • high mortality rate
    • meningoencephalitis
    • acute myocarditis

 

22

Acute stage (2-4 months)

division and antibodies

  • unequal cell division 
  • trypomastigote → amastigote have zoid that's broken down by the cell and antigens presented on the cell that are recognized by immune mediators
    • antibody-mediated immune response to these antigens
    • antibodies suppress the parasites as they appear in the bloodstream
    • antibodies are long-lasting (person known to be cured still ahs antibodies 20 years later - so hard to tell if they've cleared the parasite or not because antibodies still in tests)
  • suppress the immune system → parasites break out
  • reactivation has extremely high mortality rate

 

23

Indeterminate stage

general

  • absence of parasites → disappear from blood
    • the parasite is there but at a very ow level - beyond the limits of detection
  • asyptomatic
    • patient acts as reservoir
    • parasite hiding away from the immune system but infection still in background
  • patient remains seropositive
  • sub-clinical level of cardiac involvement as judged by ultrasonic cardiography (echocardiography)
  • 70% of cases - disease doesn't progress

24

Indeterminate stage

question about when it starts/finished

  • in this stage may have issues related to heart function appearing - but those conditions are not life-threatening at this point
  • some people start having heart murmurs
  • potentially the pathologies associated with the next stage may start in this stage depending if parasite in the heart or not
  • for most people this is as far as the disease goes
    • still ahve the parasite but asymptomatic
    • normal life span
  • remaining 30%...

 

25

Chronic stage

general

  • ~30% of cases disease reactivates (why/how?)
  • reactivation may occur 40 years later after acute phase
    • reactivation - evades immune mechanisms directly or because the immune system starts falling apart and enters the chronic phase
  • cardiac and gastrointestinal complications
  • fatal
    • die of cardiac or GI tract complicatoins
    • probably host factors contributing to whether this disease breaks out 
      • not parasite per se but the person's body not working as it should

 

 

26

Chronic stage

cardiac

  • cardiac problems can affect all parts of the heart
    • endocardium - innermost layer of tissue that lines chambers of the heart
    • myocardium - muscular tissue of the heart
    • pericardium - fluid filled sac that surrounds the heart
  • autonomic ganglia/cardiac nerves frequently damaged
    • nervous system associatd with the heart (helps it beat and function normally) can also be infected

 

27

Chronic stage

cardiac problems include

  • arrhythmias
  • conduction defects

 

  • symptoms include rapid heart rate, light-headedness, dizziness, fainting

 

  • cardiomegaly
  • thrombo-embolic events

28

Chronic stage 

arrhythmias

 

  • irregular beating
  • electrocardiogram (ECG) abnormalities 2-fold higher in seropositive patients of seropositive patients comapred to seronegative persons

 

29

Chronic stage

conduction defects

 

  • blockage of blood vessels serving the heart
  • reduced bloodflow to cardiomyocyted downstream

 

30

Chronic stage

cardiomegaly

  • progressive enlargement of the heart
  • hypertrophy/dilation - increase in volume of the heart due to elargement of the component cells
    • muscle cells getting larger
    • dilation of the whole heart or parts of the heart
  • light bulb test - bulb behind the heart to see if light ocmes through 
    • normal heart is thick solid mass
    • here dilation leads to thinning of the heart
    • can get thinning of the heart walls at key points
    • apex thinning so it bursts - get aneurisms
      • observed in 20-30% of autopsies

31

Chronic stage

thrombo-embolic events

  • blood clots form 
  • can be disseminated to clog up other organs as well - embolisms on other organs
  • leading cause of heart disese in affected countries eg Brazil (now other causes for heart failure)

 

32

Chronic stage

histopathology of infected heart

what causes these defects?

all comes down to localized "nest" of parasites

  • when the nests burst immune systems respond to get inflammation in certain sites
    • released parasites promote infiltration by host immune cells (lymphocytes and macrophages)
  • burst + inflammation associated with infiltration often damage myocardial fibers
  • extensive fibrosis occurs
    • replacement of damaged cardiac muscle with connective tissue

 

  • immune response/inflammation doesn't stop at the parasites but also causes necrosis surrounding the parasite → aberrant side effects to local tissues, necrosis building up
    • our response to that is to fill the gaps with connective tissue eg collagen
    • causes weak links → arrhythmias
    • degree of fibrosis
    • connection between cardiac symptoms and degree of fibrosis in the person's heart
  • as the chronic stage progresses, some of these may start in indeterminate
  • cardiac conditions get worse as you accumulate more damage from the immune system that lead to death
  • pathologies → what we see at the cellular level

33

Chronic stage

heart histoplathology continued

  • all cardiomyopathies due to progressive destruction of myocardium + conduction system
  • cardiomyopathies accumulate with time (starts in latent period?)
  • patient progressively worsens
  • cardiac symptoms correlate with fibrosis
  • cellular damage leads to arrhythmias, cardiomegay, and thrombo-embolic events
  • death usually due to arrhythmia, congestive heart failure (can no longer pump enough blood) or thrombo-embolic events

34

Chronic stage

GI problems

  • associated with dilation of the digestive tract
  • organs commonly affected - esophagus and colon
    • become so dilated/large you have hypertrophy, cells becoming larger
      • megaesophagus (stained with barium)
      • megacolon
      • independently or in conjunction
      • if in conjunction, megaesophagus proceeds megacolon (esophagus first)
      • in many patients, megaesophagus proceeds cardiac problems (esophagus before heart)

35

Chronic

GI 

dilation →

  • autonomic neuronal dysfunction
  • autopsy shows reduction in the number of ganglia
  • denervation - parasite infection or autoimmune disease?
  • cause/mechanism unknown

 

36

Chronic

GI 

megoesophagus

clinical symptoms include:

  • difficult/painful swallowing
  • food regurgitation
  • speech impairment

 

37

Chronic

GI

megacolon

clinical symptoms include

  • prolonged constipation
  • pain 
  • constant physical distress

 

38

Diagnosis

suspected in patients:

exhibiting symptoms

  • in chronic stage look for lower digestive tract problems

having epidemiological history of possible exposure

  • resident in sub-standard housing
  • receipt of unscreened blood products fro endemic areas
  • geographical connection with symptom

 

x-rays and other imaging techniques to detect enlargement of affected organs (in chronic stage)

  • to show dilation

 

  • all of these don't get to the root cause of what's underlying
  • have to go low-tech - look for the parasite where possible

 

39

Diagnosis

direct methods

  • microscopy of fresh blood smear - coupled with stain (Giemsa)
  • eg in chronic see BSF in the bloodstream
    • staining technique
    • trypomastigote in blood
  • concentration may be employed 
    • examination of buffy coat
    • like HAT
  • xenodiagnoses
    • get uninfected bug - let parasite develop then sacrifice the insect to see if the parasite is there
    • take a long time though - not used in a clinical context

 

40

Diagnosis

confirmation by laboratory testing

indirect methods

  • PCR 
    • amplify the kinetoplast or nuclear genome
    • lots of good targets to use
    • not used in clinical labs - only research labs
  • serological tests
    • antibody-related techniques
    • indirect fluorescence assay
    • indirect haemagglutination
    • enzyme-linked immunosorbent assay (ELISA)
    • may be cured but still test positive (lots of false positives)

41

Treatments for Chagas disease

  • nifurtimox
  • benznidazole

42

Chagas treatment

nifurtimox

  • too dangerous to use in (Southern) Latin America
    • skin falls off
  • used in central America

 

  • 5-nitrofuran pro-drug
    • furan ring (5) with a nitro group
    • nitro group is responsible for the activity of the drug initially
    • lipophilic - goes through membranes
      • can cross blood/brain barrier
  • introduced in 1964 by Bayer-Lampit

 

  • administered orally
  • 8-10 mg/kg/day - 2 or 3 tablets/day
  • treatment lasts for 90-120 days
    • adds to cost
  • other complications frequently result in giving up the course of treatment

 

 

43

Chagas treatment

nifurtimox

uptake

  • uptake by activation of the nitro-group
  • readily taken up by cells
  • crosses blood/brain barrier (lipophilic)
  • metabolized by liver cytochrome P450/cytochrome P450 reductase
    • metabolized
    • short half-life = take more to keep at therapeutic levels
  • not excreted in urine
  • half-life ~3 hours
  • uptake by passive diffusion

44

Chagas treatment

nifurtimox

uptake - detailed

 

  • activation of nitro-group
  • parasites (brucei and cruzi) have type I nitroreductase
    • absent from humans (only in bacteria and SOME protozoan parasites)
    • contains flavin mononucleotide as a cofactor
    • activity not affected by oxygen
    • can metabolize a whole range of different nitro- and quinone-based compounds
  • can purify these enzymes from E. coli and do in vitro work with enzymes to ID what compounds are metabolized by the enzymes
    • in vitro take purified enzymes in parasite mitochondrion
    • use nifurtimox as substrate for the parasite enzyme see what's produced
    • nitro-group through 2 electron reductions to hydroxylamine derivative
    • electronic switch - the nitro group is like a vacuum sucking electrons from furan ring but when convert to hydroxylamine the hydroxylamine acts as a blower (blows electrons onto the furan ring)
    • pushing electrons on and sucking off = ring falls apart/breaks (weak bond breaks) to generate anunsaturated open-chain nitrile that's sticky and will
      • unsaturated form is toxic to both pathogen AND host, reacts with proteins, nucleic acids, thiols
      • stick to biological molecules and form a conjugate
  • that's the basis for the selectivity of the drug because if you play around with the copy number of the type I nitroreductase in the parasite (reduce the number of copies of the gene) (diploid, copy number of 1 to make haploid) → those cells become slightly more resistant to the drug
  • culture the parasites on nifurtimox → lost one of the alleles with the NTR gene 
    • haploid for NTR gene = reduction of activity leads to resistance
  • converse experiment - overexpres the enzyme within the parasite to make the cells more susceptible to the drug (10-15 fold more susceptible)
    • initial activation event is key

45

Chagas treatment

nifurtimox

side effects

  • can fail
    • different strains display different types of susceptibilities to this drug
  • usually used when asymptomatic in acute phase
  • skin falls off
  • mutagenic
  • carcinogenic
  • can fail to clear parasitemia
  • limited efficacy (acute phase only) 
  • resistant strains

 

 

46

Chagas treatment 

nifurtimox

WHO/clinical trials

  • added to List of Essential Medicines by WHO as part of NECT against West African trypanosomiasis
  • phase II clinical trials against pediatric neuroblastoma

47

Chagas treatment

nifurtimox

humans lack the type I nitroreductase but...

can process with type II nitroreductase

  • one-electron reduction to form nitro anion radical
  • type II nitroreductases are: ubiquitious
    • FAD- or FMN-containing activity affected by O2
    • include glutathione (or trypanothione) reductase, lipamide dehydrogenase, cytochromoe P450 reductase
  • nitro anion radical in oxygen environment → reacts with O2 toform superoxide anions
    • re-forms the drug 
      • futile cycle not doing anything
    • oxygen is converted to superoxde that's normally removed by superoxide dismutatses (SOD) but if this is initial reaction occurs at rate to produce lots of O2-
      • at high levels, SOD activity swamped
      • cells undergo oxidative stress
        • kills tumours
    • accounts for activity within neuroblastoma cell lines
  • whole basis is how the drug at the nitro group is reduced

48

Chagas treatment

nifurtimox

parasites vs humans

in parasites → causes production of cytotoxic reduction products

 

in humans (and parasites) → causes oxidative stress

 

nifurtimox selectivity against trypanosomes based on mechanism of activation

49

Chagas treatment

benznidazole

  • 2 nitroimidazole prodrug
  • introduced in 1970s by Hoffman-La Roche

 

  • oral administration
  • 5-7 mg/kg/day - 2 tablets/day
  • treatment lasts for 30-60days

 

  • found equally in plasma and cells
  • 70% excreted in urine
  • metabolized in liver (mechanism?)
  • half-life ~12 hours

50

Chagas treatment

benznidazole

side effects

 

  • pharmokinetics similar to nifurtimox, similar side effects
  • mutagenic
  • carcinogenic
  • can fail to clear parasitemai
  • limited efficacy
    • under question - BENEFIT
    • recent work in BENEFIT trial in Venezuela - looking at people with cardiac complications in chronic phase suggest that this can be used against that particular pathology
    • initial data indicate that this is effective in patients with cardiac problems associated with the chronic stage
  • side effects not bad as bad as nifurtimox, drug of choice in Brazil, etc.
  • resistant strains

51

Chagas disease

benznidazole

uptake/mode of action - detailed

lipophilic – taken up by cells in passive diffusion

  • type I nitroreductase plays a role in metabolizing it

similar pathway

  • nitro group drawing electrons from imidizaole ring
  • nitro group reacts with type-I nitroreductase, get hydroxylamine derivative formed that pushes electron onto the ring
  • as with nifurtimox, the switch causes fragmentation of the ring backbone through intermediates that can be detected with LC-MS/MS
  • forming dialdehyde-glyoxyl
  • glyoxal binds to biological molecules, leading to damage

52

Chagas treatment 

benznidazole

uptake/mode of action - general

  • 2-nitroimidazole prodrug
  • activated by nitro-reductoin
  • specific enzyme implicate (type I nitroreductase)
  • converts nitro group to hydroxylamine (via intermediate)
  • hydroxylamine unstable
  • imidazole ring becomes hydroxylated
  • imidazole ring breaks releasing glyoxal
  • hydroxylamine, nitrenium, and glyoxal form adducts with proteins, nucleic acids, thiols

53

Chagas Disease

prevention

vector control strategies

  • mosquito nets
  • insecticide sprays and paints
  • improving housing and sanitary conditions in the rural areas
  • control strategies employed and used effectively to reduce/eliminate the disease

 

Southern cone initiative 

  • 72% reduction
  • Chile, Uruguay, 10 Brazilian states, 12 Argentinian provinces free of disease (2001)
  • cleared the disease in humans, largely
  • parasite and vectors still there
  • if don't maintain these strategies it can readily come back
  • problem in rural population

 

other initiatives started, looking at different regions of pacific coast but less political will to drive the initiatives → not taken off int he same way as the Southern Cone initiative

  • Initiative of Central American Countries to Interrupt Vectoral and   Transfusional Transmission of Chagas Disease (1997)
  • Initiative of the Andean Countries to Control Vectoral and Transfusional   Transmission of Chagas Disease (1997)
  • Initiative of the Amazon Countries for Surveillance and Control of   Chagas Disease (1997)