cvcv

1
cvcv
cvcv
cvcv
cvcv
cvcv
Clinical
Summary
Introduction
Epidemiology
Disease biology
Amoebiasis
E-Learning Module
Click here to begin
Matt Pugh
2
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary
Introduction Welcome to the amoebiasis
e-learning module. Amoebiasis is a parasitic
protozoan disease that affects the gut mucosa and
liver, resulting in dysentery, colitis and liver
abscess. The causative agent, Entamoeba
histolytica, is a potent pathogen that is spread
via ingestion of contaminated food and water.
Globally, amoebiasis is highly prevalent, and is
the second leading cause of death to parasitic
disease. This resource will outline the disease
biology, epidemiology and clinical principles of
amoebiasis.
3
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary

• Learning Outcomes
• The learning outcomes for this moudule are
• Understand the biology, life cycle and mode of
  transmission of the amoebiasis causative
  organism, Entamoeba histolytica.
• Understand the pathological processes that lead
  to disease in amoebiasis
• Know the distribution of amoebiasis worldwide,
  and which geographical, cultural and economic
  factors can pre-dispose to the disease
• Know how the disease presents according to the
  infected organ system.
• Be able to outline the treatment and management
  of symtomatic and asymtomatic patients.
• Outline the key features of the developing
  gal-lectin vaccine.

4
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary
How to use this module The module is split into
five main sections introduction, disease
biology, epidemiology, clinical and summary. The
information required to complete the learning
outcomes are contained within the disease
biology, epidemiology and clinical sections. At
the end of each of these sections there will be a
self assessment section. You will require a pen
and paper to write down your answers. How to
navigate
Navigate through the module using the blue arrows
to go back and forth. You may skip straight to,
or back to a section by clicking the tabs at the
top of screen. Additionally, you may skip through
the sub-sections by clicking on the tabs at the
side of the screen.
5
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary
Causative Organism

• The causitive orgainism is parasitic protazoan,
  called Entamoeba histolytica.
• What was once thought to be a single entity, is
  now recognised as two morphologically identical
  but genetically distinct forms E. histolytica
  (pathogen) and E. dispar (commensal).
• This has affected our understanding of amoeba
  distribution. Many suspected cases of E.
  histolytica carrier, may simply have been E.
  dispar colonisation
• The WHO recommendes that E. histolytica
  colonisation should be treated, however,
  treatment is unnecessary for E. dispar
  colonisation

Causative Organism
Life Cycle and transmission 1
Life Cycle and transmission 2
Pathogenesis 1
Pathogenesis 2
Self Assessment
E. Histolytica
6
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary
Life cycle and transmission 1

• Entammoeba histolytica has a biphasic life
  cycle, existing in two forms as an infectious
  cyst and an amoeboid trophozoite

Causative Organism
Life Cycle and transmission 1
Life Cycle and transmission 2
Pathogenesis 1
Pathogenesis 2
Self Assessment
7
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary
Life cycle and transmission 2

• Cysts (10-15µm) are ingested via contaminated
  food or water. A refractile wall containing
  chitin, allows the cyst to survive stomach acid.
• In the terminal ileum or colon, the parasite
  excysts and begins the trophozoite stage.
• Trophozoites (10-50µm) are highly motile and
  pleomorphic. They are unable to survive outside
  the human gut.
• Energy is derived from the ingestion of bacteria
  and food particles. No mitochondria are present
  in trophozoites. Respiration enzymes are
  prokaryotic in origin and are anaerobic,
  converting
• glucose pyruvate
  ethanol
• Trophozoites reproduce by binary fission and
  encyst in the colonic wall. Cysts are passed in
  the stool where they become infectious.
• The signal for encystation is thought to be via
  epithelial galactose/N-acetylgalactosamine
  specific lectin (gal-lectin) binding protein.

Causative Organism
Life Cycle and transmission 1
Life Cycle and transmission 2
Pathogenesis 1
Pathogenesis 2
Self Assessment
8
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary
Pathogenesis 1

• Amoebic trophozoites invade the colon causing
  colitis. They may also invade the portal
  circulation and travel to the liver, causing
  liver abscess.
• Gastrointestinal Pathology
• The spectrum of colitis in amoebiasis ranges
  from mucosal thickening, to multiple cyst
  formation, to diffuse Inflammation / oedema, to
  necrosis and perforation of colonic wall.
• Binding of E. histolytica to epithelial cells via
  gal-lectin. This molecule shows homologous to
  human CD59, conferring resistance to complement .
  A change in the epithelial permeability is
  induced, probably via the inter-cellular tight
  junctions.
• Cell lysis and apoptosis of mucosa are thought
  to be mediated by amoebapores, peptides capable
  of forming pores in lipid bi-layers.
• Trophozoites invade through to the submucosa
  causing flask shaped cysts .
• Cysteine proteases released by trophozoites
  digest extracellular matrix in liver and colon,
  and induce interleukin-1 mediated inflammation.
  Proteases also cleave IgA and IgG antibodies.
• Neutrophils and macrophages are drawn to invasion
  sites. E. histolytica can lyse neutrophils
  leading to further tissue damage, and
  contributing towards the induction of diarrhoea.
• Inflammation is a significant cause of tissue
  damage, however, innate immunity may be the main
  combatant against the disease.

Causative Organism
Life Cycle and transmission 1
Life Cycle and transmission 2
Pathogenesis 1
Pathogenesis 2
Self Assessment
9
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary
Pathogenesis 2

• Hepatic Pathology
• Trophozoites invading the colonic mucosa may
  enter the hepatic circulation and reach the liver

Causative Organism

• Well circumscribed abscesses are formed in the
  liver containing liquefied cells surrounded by
  inflammatory cells and trophozoites
• Adjacent parenchyma is usually unaffected

Life Cycle and transmission 1
Life Cycle and transmission 2
Amoebic liver abscess
Pathogenesis 1
Pathogenesis 2
Self Assessment
Histological cross section of classical flask
shaped amoebic ulcer in colonic mucosa.
Amoebic colitis with multiple ulcer formation
10
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary
Questions

• 1) Which of the following organisms is the
  pathogenic causitive agent of amoebiasis, and
  which is a commensal?
• Entamoeba histolytica .
• Entamoeba dispar .
• Draw a simple diagram oulining the life cycle of
  entamoeba histolytica.
• Which two organs does E. histolytica primarily
  invade?
• What is the name and mechanism of action of the
  peptide responsible for cell lysis and apoptosis
  in the mucosa?
• What is the name of the enzyme group released by
  trophozoites to digest the extra-cellular matrix

Causative Organism
Life Cycle and transmission 1
Life Cycle and transmission 2
Pathogenesis 1
Pathogenesis 2
Self Assessment
Reveal Answers
11
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary
Answers

• 1) Which of the following organisms is the
  pathogenic causitive agent of amoebiasis, and
  which is a commensal?
• Entamoeba histolytica Pathogen.
• Entamoeba dispar Commensal.
• Draw a simple diagram oulining the life cycle of
  entamoeba histolytica.
• 3) Which two organs does E. histolytica
  primarily invade?
• Colon and liver
• 4) What is the name and mechanism of action of
  the peptide responsible for cell lysis and
  apoptosis in the mucosa?
• Cell lysis and apoptosis of mucosa are
  thought to be mediated by amoebapores, These
  peptides form pores in lipid bi-layers of mucosal
  cells, leading to cell leakage resulting in lysis
  and apoptosis.
• 5) What is the name of the enzyme group
  released by trophozoites to digest the
  extra-cellular matrix ?
• Cysteine proteases

Causative Organism
Life Cycle and transmission 1
Life Cycle and transmission 2
Pathogenesis 1
Pathogenesis 2
Self Assessment
12
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary
Epidemiology

• Amoebiasis is found primarily in developing
  tropical and subtropical countries where
  sanitation is poor, leading to a direct link
  between faeces and ingestion (see Box-1).
  Occasionally cases are reported in non-endemic
  areas e.g. UK and USA. Usually due to travel and
  immigration from endemic areas.
• There are an estimated 40,000-100,000 deaths due
  to amoebiasis worldwide each year.

Epidemiology
Susceptibility
Self Assessment
Box-1. Amoebiasis rates/figures in endemic
regions -Egypt accounts for 38 of patients
presenting with acute diarrhoea in outpatient
clinic. -Mexico1.3 million cases reported in
1996. -Hue, Vietnam 1500 of a 1million
population over 5 years
13
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary
Susceptibility

• Generally considered to affect children and
  adults, of both sexes equally. However, some data
  and anecdotal evidence suggests a male
  predominance.
• Amoebic liver abscesses are most common in males,
  18-55.
• Susceptibility to liver abscess conferred by
  HLA-DR3 and complotype SC01 in the Mexican
  populations
• Other risk factors include oral and anal sex, and
  contact with contaminated enema apparatus.

Epidemiology
Susceptibility
Self assessment
14
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary
Questions

• How many deaths are caused by amoebiasis each
  year?
• 1000 5000 b) 40,000-100,000 c)
  500,000-1,000,000
• Which part of the world is amoebiasis primarily
  found?
• Developed countries b) Tropical and subtropical
  c)Cold climates
• Does amoebiasis affect males or females more?
• Apart from poor sanitation, what other risk
  factor pre-dispose to amoebiasis infection?

Epidemiology
Susceptibility
Self assessment
Reveal Answers
15
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary
Answer

• How many deaths are caused by amoebiasis each
  year?
• 1000 5000 b) 40,000-100,000 c)
  500,000-1,000,000
• Which part of the world is amoebiasis primarily
  found?
• Developed countries b) Tropical and subtropical
  c)Cold climates
• Does amoebiasis affect males or females more?
• Thought to affect both sexes equally,
  however, anecdotal evidence suggests a male
  predominance.
• 4) Apart from poor sanitation, what other risk
  factor pre-dispose to amoebiasis infection?
• Other risk factors include oral and anal
  sex, and contact with contaminated enema
  apparatus.

Epidemiology
Susceptibility
Self assessment
16
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary
Presentation

• Some individuals carry E. histolytica
  asymptomatically. 4 -10 will go on to develop
  the disease within a year.
• Gastroenterological
• Gradual onset (weeks) of bloody diarrhoea,
  occasionally with small volumes of mucoid stool.
  If blood is not visible, stool is usually haem
  positive due to the breach of the mucosa.
• Abdominal pain and tenderness.
• Leucocytes and pus may be present in stool.
  Fever present in lt40 of patients.
• Weight loss and anorexia can be present.
• In more severe cases fulminant amoebic colitis
  develops. Liver involvement is more common in
  these cases, along with paralytic ileus, toxic
  megacolon and mucosal sloughing. Over 75 of
  patients with fulminant colitis develop
  intestinal perforation.
• Local inflammatory masses, amoebomas, may cause
  obstructive symptoms.
• Hepatic
• More common in men
• Liver abscess pan present in conjunction with
  bowel symptoms (10 of cases), or in isolation.
• Sudden onset of upper abdominal pain with fever.
  Pain may radiate to right shoulder or be
  exacerbated by repiratory movements.
• Hepatic tenderness may be present. Jaundice is
  unusual.
• Complicated liver abscess may develop if abscess
  ruptures into the peritoneal, pericardial or
  pleural cavity. Morbidity and mortality is high.
• Rarely, trophozoites may also invade the
  respiratory tract, brain and GU tract

Presentation
Diagnosis
Treatment and Management
Vaccine Development 1
Vaccine Development 2
Vaccine Development 3
Self Assessment
17
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary
Diagnosis

• Clinical history is important. In low resource
  settings this may be the means of diagnosis. A
  good travel history is important as disease may
  develop years after a visit to an endemic area.
• Demonstration of E. histolytica in stool by
  microscopy (old), or ELISA assay for antigen
  detection. Trophozoites only survive for short
  periods of time, therefore, fresh stool samples
  should be used
• Colonoscopy to confirm colitis and tissue biopsy
  for amoeba
• Liver abscess space occupying lesion on CT/USS
  with positive amoebic serology

Presentation
Diagnosis
Treatment and Management
Vaccine Development 1
Vaccine Development 2
Vaccine Development 3
Self Assessment
18
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary
Treatment and Management

• Amoebiasis, in particular with liver
  involvement, can be fatal if not treated.
  Chemotherapy can effectively cure ameobiasis.
• Nitroimidazole (e.g.metronidazole) is used to
  treat the invasive pathogens 800mg t.d.s for 10
  days.
• This is followed by a luminal agent
  (e.g.diloxanide furoate) to eliminate
  colonisation 500mg t.d.s for 10 days. This is
  also suitable for asymptomatic individuals.
• Complicated liver abscesses should be drained
  surgically.
• Prevention
• Boiling water for at least ten minutes kills
  amoebic cysts effectively. Chlorine and iodine
  tablets are not thought to be 100 effective.

Presentation
Diagnosis
Treatment and Management
Vaccine Development 1
Vaccine Development 2
Vaccine Development 3
Self Assessment
19
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary
Vaccine Development 1

• Amoebiasis incidence could be vastly reduced with
  simple sanitation and hygiene measures. However,
  given the current political and economic climate,
  this seems unlikely in the near future.
  Furthermore, with developing drug resistance in
  E. histolytica, vaccine development could be
  effective.
• Why vaccinate?
• Could prevent development of amoebic disease and
  associated sequelae.
• Humans only host for E. histolytica, therefore
  eradication vaccine would eliminate E.
  histolytica from the carrier pool.
• Which target?
• A number of potential targets have been
  identified including cysteine proteases, LPGs and
  peroxiredoxins. The two most promising antigens
  identified are Serine-Rich E. histolytica Protein
  (SREHP) and Galactose/N-acetylgalactosamine
  lectin (Gal-lectin). Here the potential
  Gal-lectin vaccine will be described

Presentation
Diagnosis
Treatment and Management
Vaccine Development 1
Vaccine Development 2
Vaccine Development 3
Self Assessment
20
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary
Vaccine Development 2

• Gal lectin and the immune response
• Gal-lectin is a 260kDa complex protein which
  consists of disulphide linked light (35 kDa) and
  heavy subunits (170kDa). The heavy chain is
  cysteine rich and is thought to be a target for
  immune responses, inducing a Th1 cytokine cell
  mediated immune response
• Macrophages induced by cytokines
  interferon(INF)-? have amoebocytic activity, as
  do T-cells exposed to INF-? exposed or TNF.
• Trophozoite killing by macrophages is done via
  nitric oxide (NO). Gal-lectin can directly
  activate macrophages to release NO and induce
  mRNA transcription of Th1 cytokines, thereby
  enhancing the cell mediated immune response.
• Monoclonal antibodies (MAbs), antiserum and IgA
  secreted from the gut mucosa against the
  Gal-Lectin antigen, have the ability to inhibit
  E. histolytica adherence to colonic mucosa in
  vitro.

Presentation
Diagnosis
Treatment and Management
Vaccine Development 1
Vaccine Development 2
Vaccine Development 3
Self Assessment
21
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary
Vaccine Development 3

• Both Gal-lectin classical and DNA based vaccines
  have been tested in murine models.
• Gal-lectin DNA vaccine
• DNA of heavy gal-lec subunit used as vaccine
  (see Fig-5)4
• Induced Th1 mediated anti-body specific response
  greater than control (nothing), however response
  was small. Vaccine moderately inhibited
  trophozoite adherence in vitro via anti-body
  action.

Protection conferred from purified and
recombinant vaccines
Presentation
Gal-lectin classical vaccine Purified (lectin)
and recombinant gal-letin (LecA) have been
trialled, showing good efficacy in preventing E.
histolytica pathogenesis. Immunisation were
intra-nasal and intra-peritoneal in order to
stimulate the gastrointestinal immunity.
Diagnosis
Protection
Treatment and Management
Vaccine Development 1
Vaccine Development 2
Production of DNA gal-lectin DNA vaccine in
murine model.
Vaccine Development 3
170
35
Self Assessment
Gene coding for portion of heavy gal-lec subunit
isolated
Transfected into plasmid
Muscle cells take up and incorporate gal-lec
sequence in DNA
Plasmid injected intra-muscularly
Protein expressed and immune response induced
22
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary
Questions

• What are the symptoms of gastrointestinal
  amoebiasis?
• What are the symptoms of hepatic amoebiasis?
• Why is a good travel history important in
  diagnosis of amoebiasis?
• What investigations can be performed to confirm a
  diagnosis?
• Name two drugs and dosage regimes that can be
  used to treat amoebiasis.
• Is the following statement true or false?
• chlorine and iodine can be used to decontaminate
  water of E.histolytica with 100 effectiveness
• 7) Does Gal-lectin induce a Th1 or Th2 cell
  mediated immune response?

Presentation
Diagnosis
Treatment and Management
Vaccine Development 1
Vaccine Development 2
Vaccine Development 3
Reveal Answer
Self Assessment
23
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary
Answers

• What are the symptoms of gastrointestinal
  amoebiasis?
• Gradual onset (weeks) of bloody diarrhoea,
  abdominal pain and tenderness, fever present in
  lt40 of patients, weight loss and anorexia,
  amoebomas, may cause obstructive symptoms.
• What are the symptoms of hepatic amoebiasis?
• Sudden onset of upper abdominal pain with fever.
  Pain may radiate to right shoulder or be
  exacerbated by repiratory movements.
• Hepatic tenderness may be present. Jaundice is
  unusual
• 3) Why is a good travel history important
  in diagnosis of amoebiasis?
• A good travel history is vital to ascertain
  whether a patient has visited an endemic area.
  The disease may develop over a year after travel.
• What investigations can be performed to confirm a
  diagnosis?
• Demonstration of E. histolytica in stool by
  microscopy (old), or ELISA assay for antigen
  detection. Colonoscopy may be performed to check
  for colitis and biopsy. Check for liver abscess
  with USS or CT.
• Name two drugs and dosage regimes that can be
  used to treat amoebiasis.
• Nitroimidazole (e.g.metronidazole) 800mg t.d.s
  for 10 days. This is followed by a luminal agent
  (e.g.diloxanide furoate) 500mg t.d.s for 10 days.
  
• 6) Is the following statement true or false?
• chlorine and iodine can be used to decontaminate
  water of E.histolytica with 100 effectiveness
• Boiling is the most effective methos for water
  decontamination
• Does Gal-lectin induce a Th1 or Th2 cell mediated
  immune response?
• Th1 cell mediated response

Presentation
Diagnosis
Treatment and Management
Vaccine Development 1
Vaccine Development 2
Vaccine Development 3
Self Assessment
24
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary
Summary

• Amoebiasis is a major global cause of mortality
  and morbidity, due to dysentery. The causative
  organism, E. histolytica.
• E. histolytica has a biphasic life cycle and
  exists as an infective cyst and pathological
  trophozoite.
• The disease is spread via contaminated food and
  water, usually due to poor sanitation.
• The disease is found in tropical and sub-tropical
  parts of the world.
• Every year, 40,000-100,000 people die from
  amoebiasis
• Certain genetic traits pre-dispose to certain
  pathologies.
• Patients usually present with abdominal pain,
  bloody stools and fever. Hepatic symptoms are
  more acute with upper abdominal pain and
  radiation to the right shoulder.
• Treatment is with Nitroimidazole
  (e.g.metronidazole) and a luminal agent. Spread
  can be prevented by boiling water.
• A potential gal-lectin vaccine is currently in
  development. Good results have been yielded with
  native gal-lectin vaccines, and moderate results
  with a DNA based vaccine. Immunity appears to be
  mainly via a Th1 cell medicated response and
  secretory IgA

Summary
References
25
cvcv
cvcv
cvcv
cvcv
cvcv
Introduction
Disease biology
Epidemiology
Clinical
Summary
References and further reading

• Gaucher D., Chadee K. (2003). Prospect for an
  Entamoeba histolytica Gal-lectin-based vaccine.
  Parasite Immunology. 25, 5558 (review)
• Gaucher D., Chadee K. (2002). Construction and
  immunogenicity of a codon-optimized Entamoeba
  histolytica Gal-lectin-based DNA vaccine.
  Vaccine. 20, 3244-3253
• Houpt E., Barroso L., Lockhart L., Wright R.,
  Cramer C., Lyerly D., Petri W.A. (2003)
  Prevention of intestinal amebiasis by vaccination
  with the Entamoeba histolytica Gal/GalNac lectin.
  Vaccine. 22, 611617
• Kelly P, Farthing M (2005) Protozoal
  gastrointestinal infections Medicine 33 4 ,
  81-83.
• Stanley S.L. (2003) Amoebiasis. The lancet.
  361,1025-1034 (review)

Summary
References