Protozoa
Guanling Wu, Prof. in Dept. Pathogen Biology,
Nanjing Medical University, Najing, Jiangsu,
China
jiminjun@[Link]
General Properties of protozoa
Single-cell animal – monocellular or unicellular
organisms with full vital functions
Structurally, protozoa resemble single metazoan cells, while functionally each
is equivalent to a whole metazoan animal (clearly animal-like in nature).
The protozoan cell has a full complement of cellular organelles: nucleus,
mitochondria, endoplasmic reticulum and Golgi apparatus togather with
specialized organelles in some groups, such as rhoptries, micronemes and the
apical complex.
Protozoa range in size from 2 μm to more than 200 μm.
Species – total named species:65,000;
parasitic: around 10,000; medical protozoa: 40
Detailed classification of the protozoa can be
found in textbooks. It is sufficient here to note that
protozoa with the importance of medicine can be broadly
divided into three large groups, generally accoded the
rank of phyla:
Sarcomastigophora: forms whose main method of locomotion by
means of temporary extensions of the cell, known as pseudopodia
flagellates and amebas( for example Entamoeba histolytica), or by
the beating of long whip-like structure called flagella ( for example
Leishmania ,Trichomonus vaginalis, Giardia, Trypanosoma etc.)
Apicomplexa: united by having in some stages of the life cycle a
complex set of organelles, the apical complex, at the anterior pole
of the cell ( for example Plasmodium , Toxoplasma)
Ciliophora: possession of large numbers of short hair-like
extensions of the cell, cilia ( for example Balantidium coli )
Others : Cryptosporidium, Microsporidia
Life cycle patterns
One-host form (direct)
1. One stage form – Trophozoite
2. Two stage form – Trophozoite & Cyst
Two-host form (indirect)
1. Mammals --- mammals
2. Mammals --- insect vectors
Mode of Reproduction
Asexual Reproduction
Binary fission – result in 2 daughter cells
Schizogony – multiple fission result in multiple
daughter cells
Budding
Exogenous budding - by external budding result in
multi- cells
Endodyogony - by internal budding result in 2 cells
Sexual Reproduction
Conjugation – exchange of nuclear material of 2
Gametogony – sexually differentiated cells unite
-- zygote
Pathogenesis
Host Resistance Opportunistic & Accidental
– Innate immunity (protozoa) infections
– Acquired immunity
Parasite Invasion
– Toxin
– Mechanically damage
– Immune impair
• Immune inhibition
• Hypersentivity （超敏反应）
Causative organism of Amoebiasis and
amoebic dysentery
Entamoeba histolytica
Acanthamoeba
Naegleria
Epidemiology
• 4th leading cause of death from parasitic diseases worldwide
Organism # of deaths/yr # infected
Entamoeba ~75,000 ~300 million
Ascaris ~200,000 ~480 million
Schistosoma ~750,000 ~200 million
Plasmodium 2-3 million ~500 million
(Malaria)
• Amoebiasis is not restricted to the tropics and subtropics, it
also occurs in temperate and even in arctic and antarctic
zones
Contaminated water is a source of infection.
Infection is common in developing countries where sanitation is poor.
Amoeba in alimentary tract
Entamoeba
– E. histolytica (pathogenic)
– E. dispar (non-pathogenic)
– E. coli (big sister)
– E. hartmani (little brother)
– E. gingivalis (oral)
Endolimax nana (occasionally pathogenic)
Iodamoeba butschlii
Morphology
Entamoeba histolytica
Cysts Trophozoites
Thick wall Plasmalemma (thin)
1-4 ring-like nuclei 1 ring-like nucleus
Chromatoid body (blunt) Lacking
Round, 10-16 μm Irregular, 10-60 μm
Concentrated Labile
形态
溶组织内阿米巴
包囊 滋养体
(Cysts) (Trophozoites)
壁厚 壁质膜性 (薄)
1-4 个圆形核 1 个圆形核
棒状拟染色体 无拟染色体
圆形, 10-20 μm 不规则形, 10-40 μm
在外界存活时间长 在外界存活时间短
E. histolytica trophozoite Morphology
Ingested
RBC
Nucleus with central
karyosome and
finely divided
chromatin granules
Pseudopod
Morphology
Trophozoites
Single nucleus with a central,
dot-like karyosome
Mature E. histolytica Cyst Morphology
1-4 ring-like nuclei
with finely divided
peripheral chromatin
Cyst wall and
round shape
Morphology
Micrograph of a trophozoite ingesting a red blood cell derived from its host.
E. histolytica Stages - CYSTS
Usual Infective Stage for humans
Resistant walls maintain viability
– If moist can last several weeks
– Killed by desiccation or boiling
Usual Diagnostic Stage in formed stools
– Can be concentrated and stained easily
– Not seen in liquid (diarrheic) stools or tissues
E. histolytica Stages - TROPHOZOITES
Cause amebiasis (damage tissue)
Spread throughout the body
Rarely transmit the infection to others
are Labile in liquid stools or tissue
must be rapidly found or preserved (quick
fixation & cold storage)
Entamoeba coli
Gut commensal
Trophozoite & cyst
Slow “lazy” movement
Oral-faecal transmission
E. Coli trophozoites Morphology
E. Coli cysts Morphology
E. histolytica v E. coli
Trophozoite
– 10-40um – 15-30um
– delicate nucleus – coarse nucleus
structure structure
Cyst
– 9.5-15.5um – 10-30um
– 4 nuclei – 8 nuclei
– Broad, blunt – thin, sharp
chromatid bodies chromatid bodies
Life cycle
Life cycle
Mode of transmission
Humans acquire E. histolytica by:
– Ingesting cysts (4 nuclei mature) in fecally
contaminated food or drink
– Rarely by directly inoculating trophozoites
into colon or other sites (Anal-oral
transmission due to sexual practice is also a
consideration)
Fecal-Oral transmission (hand to mouth)
Summary of the Life cycle
The basic generation-cycle: cyst –
lumen trophozoites – cyst
Trophozoites may invade intestine
and spread
Cyst formation – essential factors:
environment + time
Infective cysts and trophozoites pass
in feces
Pathogenesis
General Types of Virulence Factors:
• Adherence factors
• 260kDa Gal/GalNAc lectin
• Invasion factors
• Amoeba pores
• Cysteine proteinases
• Endotoxins
Pathogenesis
Trophozoites ...
– Attach to mucosal epithelial cells (MEC)
– Lyse MEC
– Ulcerate and invade mucosa
– Cause dysentery (diarrhea + blood)
– Metastasize via blood &/or lymph to other organs
– Form abscesses in extraintestinal sites ...
Clinical Classification of Amebiasis
Asymptomatic (intestinal) "Cyst Passers or
carriers”
Symptomatic Infection:
– Intestinal Amebiasis: (colon and rectum 盲肠、升结肠、直肠
、乙状结肠和阑尾 )
• Acute Dysentery
• Chronic Non-Dysenteric
– Extra-Intestinal Amebiasis:
• Amebic Liver Abscess (ALA)
• Amebic Pulmonary Abscess
• Other sites (brain, skin, GU…)
Clinical classification
Asymptomatic infection (carrier) >90%
(E. dispar?)
Symptomatic cases <10%
– 8-10% dysentery, colitis, etc
– 2% invasive amoebiasis
– 0.1% deaths
Clinical manifestation
Acute Dysenteric Amebiasis
Amebic Dysentery
Symptoms:
Bloody mucoid diarrhea
RBCs and few WBCs in stools
– Abdominal pain
– weight loss
– bloating, tenesmus( 里急后重 ) and
cramps( 痉挛 )
Clinical manifestation
Amebic Dysentery
Signs:
Fever (33%)
Tender (enlarged) liver
Stools positive for trophozoites, WBC
NOT cysts in loose stools
Clinical manifestation
Pinpoint lesion on
mucous membrane
Flask-shaped crateriform
(volcano)ulcers
Extra-intestinal amoebiasis
Hepatic
(1) acute non-suppurative
(2) liver abscess: right upper quadrant pain, referred
to the right shoulder. tender.
Pulmonary
Brain
Skin, perianal infection
Other extra-intestinal amoebiasis
Clinical manifestation
An Amoebic Liver Abscess Being Aspirated.
Note the reddish brown color
of the pus (‘anchovy-sauce’).
This color is due to the
breakdown of liver cells.
Gross pathology of amebic abscess of liver. Tube
of "chocolate" pus from abscess.
Diagnosis
Pathogenic diagnosis
– Stool examination:
• Direct Fecal Smear (trophozoites)
• Fecal concentration and iodine dye techniques -
(cysts) ZnSO4 or formalin-ether
– Cultivation
– DNA detection
Serologic Tests (for chronic disease): ELISA,
IHA (indirect hemagglutination assay)
Imaging: X-ray; CT
Stool examination
trophozoite cyst
specimen feces feces
method direct smear with normal direct smear with iodine
saline stain
diseases chronic intestinal
amoebic dysentery
amoebiasis or carriers
[Link] must clean
[Link] soon after they [Link] specimen warm.
remarks
have been passed.
[Link] bloody and [Link] using history.
mucous portion.
Two microscopically indistinguishable Entamoeba spp.
E. histolytica
– invades tissues
– should always be treated
E. dispar
– is non-pathogenic
– should not be treated
Treatment of Amebiasis
For invasive forms: metronidazole
For luminal forms: Iodoquinofonum,
paromomycin, diloxanide
Do not treat asymptomatic
intestinal E. dispar infection
Prevention & Control
Individual measures
• Diagnosis and treatment of E. histolytica patients
– no animal reservoirs (other than humans) are known
• Safe drinking water (boiling or 0.22 µm filtration)
• Cleaning of uncooked fruits and vegetables
• Prevention of contamination of foods
Chemotherapeutic Trial
Prevention & Control
Community measures
– Public services and utilities
• Adequate disposal of human stools
• Safe and adequate water supply
– Primary health care systems
• Health education (washing hands, cleaning and protecting
food, controlling insects)
• Specific surveillance programs and Control programs
integrated into ongoing sanitation & diarrhea control
– Health Regulations
• Control of food vendors and food handlers
• Control of flies and cockroaches
Summary
Entamoeba histolytica: Amebic dysentery; amebic
liver abscess
[Link]: Found worldwide, especially in tropical
areas, Entamoeba histolytica is a U.S. disease
agent. There is no animal reservoir.
[Link] of transmission: Ingestion of cysts. Anal-oral
transmission due to sexual practice is also a
consideration.
C. Pathology: Two-stage life cycle. The trophozoite (ameba
stage) is motile. The cyst stage is nonmotile. Trophozoites
are found in the intestinal and extraintestinal lesions. Cysts
predominate in the stools, with somes trophozoites present.
1. Amebic dysentery: Colonization of cecum & colon by
Entamoeba histolytica is common. Localized necrosis
results in "teardrop" ulcerations. Invasion into the portal
submucosa is progressive after penetration of the
submucosa.
2. Liver abscess: Penetration of the diaphragm can lead to
lung disease. Most liver disease not preceded by dysentery.
D. Clinical Considerations: Abdominal discomfort, flatulence,
tenesmus. Bloody, mucous-containing diarrhea. Liver
abscess characterized by right upper quadrant pain, weight
loss, fever, tender, enlarged liver.
E. Laboratory diagnosis: Trophozoites or cysts in
stool; cysts with 4 nuclei are diagnostic for E.
histolytica. Serology (Ab detection).
F. Treatment: Metronidazole for amebic abscess;
Diloxanide furoate for asymptomatic cyst passers
& noninvasive intestinal forms. Prevention includes
boiling water or filter sterilization. Chlorination is
not effective.
The End
齿龈内阿米巴
Entamoeba gingivalis
jiminjun@[Link]
General Introduction
Discovered in 1849
Little importance was attached to it until just
before World War I, when it was discovered that
it was always present in the plaque of patients
with destructive gum disease
Although there has been interest in this parasite,
from time to time, for 150 years, there has been
no definitive work done on this protozoan until
recently
Morphology
Trophozoits only
10 ～ 20µm
One nucleus with central
karyosome and chromatin
granules
Could ingest bacteria,
leukocyte and occasionally
活滋养 RBC
体
（生理盐水涂
片）
Morphology
活滋养 齿龈组织切 滋养
体 片 体
（生理盐水涂 （示组织内滋养 （铁苏木素染
片） 体） 色）
Clinical Manifestation
Frequently included apparent difficulty in maintaining a
clean mouth, with heavy plaque formation which rapidly
regenerated after brushing, or other plaque control
procedures
An unpleasant taste, an awareness of the gums, which
bled easily; ulcerations; halitosis ， sore, dry or itchy
eyes and a history of generalized malaise, fatigue and
frequent headache
If Entamoeba gingivalis was recovered from apparently
healthy gingival tissue and no treatment was
undertaken, it was found, on subsequent re-
examination, that there had been a periodontal
decline, if the protozoa were still present
Clinical Manifestation
与牙周病的关系
福建医科大学刘光英等 (2001) 研究资料
– 齿龈内阿米巴感染率调查 :
• 正常人 39.0%, 口腔科门诊病人 59.6%, 牙周病人 67.1%
– 牙周病实验动物模型观察 :
• 齿龈内阿米巴诱导组牙周病发生率 94.0%
• 共生菌诱导组牙周病发生率 56.2%
• 生理盐水对照组牙周病发生率 0.6%
Diagnosis
Plaque sample collection and put it
in patients saliva
HE dye may improve the sensitivity
and specificity
Infections with
Free Living Ameba
Naegleria 耐格里属
Acanthamoeba 棘阿米巴
Free Living Ameba Not seen
in humans
Naegleria
8-15 µm (smaller
than A. sp.) with
lobate pseudopod
i
i
Acanthamoeba
15-25 µm with cysts & trophs
filiform pseudopod
are seen in
humans
i
Acanthamoeba spp.
Acanthamoeba
trophozoites with
acanthopodia
Primary Amebic Meningoencephalitis PAME
An acute suppurative infection of the brain
and meninges that is rapidly fatal and
usually not diagnosed antemortem
– Caused by Naegleria fowleri
– Headache, lethargy and olfactory problems
– Sore throat, runny nose, severe headache,
vomiting, stiff neck, confusion leading to ...
– Coma and death
DIAGNOSIS PAME
Pt. History (child)
– Prior Health Excellent
– Recent Swimming (fresh water/pools)
– Cases peak during HOT months
Symptoms/Signs
– Sore throat, runny nose, headache,
vomiting, stiff neck, mental confusion,
olfactory problems, lethargy, coma and
death
Treatment PAME
– None effective - few patients survive
– Amphotericin B +/- ?
Granulomatous Amebic Encephalitis GAE
A more slowly progressive, chronic form of the
disease not associated with swimming (except
in hot tubs)
cause: Acanthamoeba castellanii
subcutaneous nodules, eye or skin infection,
progressive nasal congestion, headache ...
CNS lesions with negative serology for
toxoplasmosis
in debilitated / immuno-compromised Pts with CD4+
T cells <200/mm3
disseminated infection: skin, sinuses, lungs,
CNS/CSF
Pathology GAE
Abscesses / lesions (tissues) have
– granulomatous inflammation
– hemorrhagic necrosis and vasculitis
– trophozoites & cysts with wrinkled-walls!
ameba rarely seen in CSF
Treatment GAE
No satisfactory or effective Rx ?
– amphotericin B
Acanthamoeba Keratitis AK
Corneal infection with Acanthamoeba
spp. trophozoites & cysts
Ulcerations & “Ring Infiltrate” of
cornea
Induced by
– trauma to eye, exposure to
contaminated H2O
– contact lens wear with tap water
rinsing
AK
Diagnosis
– Examine corneal scrapings or smear
– Histopathologic examination of cornea
Treatment
– Triple Antiamoebic Therapy
• neomycin-polymyxin-gramicidin /propamidine/
miconazole
– Penetrating keratoplasty (cadaver cornea)
Summary
E. histolytica
– Life cycle
– carrier
– Morphology differences with E. coli
E. gingivalis
– Morphology
– Current advances in pathogenesis
Free living amoeba
– prevention
How will one get amoebiasis and What are the consequences?
What are the researches need to be done for E. gingivalis?
Homework
Entamoeba histolytica: Amebic dysentery; amebic
liver abscess
[Link]: Found worldwide, especially in tropical
areas, Entamoeba histolytica is a U.S. disease
agent. There is no animal reservoir.
[Link] of transmission: Ingestion of cysts. Anal-oral
transmission due to sexual practice is also a
consideration.
C. Pathology: Two-stage life cycle. The trophozoite (ameba
stage) is motile. The cyst stage is nonmotile. Trophozoites
are found in the intestinal and extraintestinal lesions. Cysts
predominate in the stools, with somes trophozoites present.
1. Amebic dysentery: Colonization of cecum & colon by
Entamoeba histolytica is common. Localized necrosis
results in "teardrop" ulcerations. Invasion into the portal
submucosa is progressive after penetration of the
submucosa.
2. Liver abscess: Penetration of the diaphragm can lead to
lung disease. Most liver disease not preceded by dysentery.
D. Clinical Considerations: Abdominal discomfort, flatulence,
tenesmus. Bloody, mucous-containing diarrhea. Liver
abscess characterized by right upper quadrant pain, weight
loss, fever, tender, enlarged liver.
E. Laboratory diagnosis: Trophozoites or cysts in
stool; cysts with 4 nuclei are diagnostic for E.
histolytica. Serology (Ab detection).
F. Treatment: Metronidazole for amebic abscess;
Diloxanide furoate for asymptomatic cyst passers
& noninvasive intestinal forms. Prevention includes
boiling water or filter sterilization. Chlorination is
not effective.
