Parasitology
PhD, Irene RICCI
School of Biosciences and Biotechnology
University of Camerino
Biosciences & Biotechnologies
(III year)
2013-2014
Protozoa
• Unicellular
• Widely dispersed
• Around species 65,000 (of which around
10,000 are parasites)
Reproduction
• Asexual reproduction
– Binary fission: results in 2 daughter cells.
– Schizogony (multiple fission): results in multiple cells.
– Endodyogony (by internal budding): results in 2 cells.
• Sexual reproduction
– Conjugation: exchange of nuclear materials
– Gametogony: sexually differentiated cells unit (zygote)
…about Protozoa mobility
• A flagellate is an organism with one or more whiplike organelles called flagella.
• Amoeba does not have a definite shape and moves
by using pseudopodia or "false feet". Pseudopodia
are formed by the amoeba by throwing out the
cytoplasm, followed by endoplasm flowing inward.
• The ciliates are characterized by the presence of
hair-like organelles called cilia, which are identical in
structure to eukaryotic flagella, but typically shorter
and present in much larger numbers with a
different undulating pattern than flagella.
Species of Medical Concerns
PROTOZOA
4 phyla based upon motility
Sarcodina
(amoeba)
Mastigophora
(flagellates)
Entamoeba
Tripanosoma
Toxoplasma
Leishmania
Plasmodium
Tricomonas
Giardia
Ciliophora
(ciliates)
Sprorozoa
-no organelles of
motility-
Entamoeba histolytica
(Sarcodina – amebae)
Fecal-oral route (cysts are resistant to
external surroundings).
E. histolytica
E. histolytica
• Cysts and trophozoites are passed in feces.
– Cysts are typically found in formed stool, whereas trophozoites in
diarrheal stool.
• Infection by E. histolytica occurs by ingestion of mature cysts in fecally
contaminated food, water, or hands.
• Excystation occurs in the small intestine and trophozoites are released,
which migrate to the large intestine.
• The trophozoites multiply by binary fission and produce cysts, and both
stages are passed in the feces.
• Because of the protection conferred by their walls, the cysts can survive
days to weeks in the external environment and are responsible for
transmission.
• Trophozoites passed in the stool are rapidly destroyed once outside the
body, and if ingested would not survive exposure to the gastric
environment.
E. histolytica
• In many cases, the trophozoites remain confined to the intestinal lumen
(non invasive infection) of individuals who are asymptomatic carriers,
passing cysts in their stool.
• In some patients the trophozoites invade the intestinal mucosa (intestinal
disease), or, through the bloodstream, extraintestinal sites such as the
liver, brain, and lungs (extraintestinal disease), with resultant pathologic
manifestations.
• It has been established that the invasive and non invasive forms represent
two separate species, respectively E. histolytica and E. dispar.
• These two species are morphologically indistinguishable unless E.
histolytica is observed with ingested red blood cells (erythrophagocytosis).
• Transmission can also occur through exposure to fecal matter during
sexual contact (in which case not only cysts, but also trophozoites could
prove infective).
E. histolytica
Erythro-phagocytosis
• E. histolytica (pathogenic species) is morphological
identical to E. dispar (non-pathogenic species)
• Clinics:
– Non symptomatic carriers
– Intestinal amebiasis: dysentery, colitis
– Extraintestinal amebiasis: liver abscess
Epidemiological aspects
•
•
•
•
Cosmopolitan
China: 3%~10%; Rural area>urban
Source of infection: Carriers
Transmit route: fecal-oral (water
contamination)
• Insects (fly, cockroaches) can play an active
role as mechanical vectors
How to prevent and control
• Treatment of patients and carriers:
– Intestinal amoebiasis by metronidazole
– Extra amoebiasis by diloxanide
• Water & soil control
• Insect vector control
• Personal hygienic health education
Trypanosome
(Mastigophora –Flagellates)
• Transmission route: Vectorial Transmission
(Biological vector)
Tsetse fly (genus Glossina)
Triatomine or “kissing” bug
(Triatomine vector species belong to
the genera Triatoma, Rhodinius and
Panstrongylus)
Trypanosome
(Mastigophora –Flagellates)
Trypanosome
(Mastigophora –Flagellates)
• Trypanosoma brucei (African trypanosome)
 T. brucei gambiense
(geographic distribution: is found in foci in large areas of West and
Central Africa)
 T. brucei rhodesiense
(geographic distribution: is limited with the species found in East
and Southeast Africa)
• Trypanosoma cruzi (American trypanosome)
African Trypanosome
African Trypanosome
• During a blood meal on the mammalian host, an infected
tsetse fly injects trypomastigotes into skin tissue.
• The parasites enter the lymphatic system and pass into the
bloodstream.
• Inside the host, they are carried to other sites throughout the
body, reach other fluids (e.g. spinal fluid) and continue the
replication by binary fission.
• The tsetse fly becomes infected with bloodstream
trypomastigotes when taking a blood meal on an infected
mammalian host.
African Trypanosome
• In the fly’s midgut the trypomastigotes multiply by binary fission,
leave the midgut, and transform into epimastigotes.
• The epimastigotes reach the fly’s salivary glands and continue
multiplication by binary fission.
– The cycle in the fly takes approximately 3 weeks.
 The entire life cycle of African Trypanosomes is represented by
extracellular stages.
 Humans are the main reservoir for Trypanosoma b. gambiense,
but this species can also be found in animals.
 Wild game animals are the main reservoir of T. b. rhodesiense.
Clinical Features
Infection occurs in 3 stages:
• A trypanosomal chancre can develop
on the site of inoculation.
• This is followed by a hemolymphatic
stage with symptoms that include
fever, lymphadenopathy, and
pruritus.
• In the meningoencephalitic stage (the
sleeping sickness), invasion of the
central nervous system can cause
headaches, somnolence, abnormal
behavior, and lead to loss of
consciousness and coma.
 The course of infection is much more
acute with T. b. rhodesiense than T. b.
gambiense.
American Trypanosome
American Trypanosome
• An infected triatomine takes a blood meal and releases
trypomastigotes in its feces near the site of the bite wound.
– Trypomastigotes enter the host through the wound or through
intact mucosal membranes, such as the conjunctiva.
• Inside the host, the trypomastigotes invade cells near the
site of inoculation, where they differentiate into
intracellular amastigotes.
• Then amastigotes multiply by binary fission and
differentiate into trypomastigotes that can infect other cells
and transform into amastigotes in new infection sites.
• Amastigotes differentiate into trypomastigotes and enter
the bloodstream.
American Trypanosome
• Trypomastigotes infect cells from a variety of tissues.
– Clinical manifestations can result from this infective cycle.
• The bloodstream trypomastigotes do not replicate (different
from the African trypanosomes), replication resumes only
when the parasites enter another cell or are ingested by
another vector.
• The “kissing” bug becomes infected by feeding on human or
animal blood that contains circulating parasites.
• The ingested trypomastigotes transform into epimastigotes
in the vector’s midgut.
• The parasites multiply in the midgut and differentiate into
infective trypomastigotes in the hindgut.
American Trypanosome
• Trypanosoma cruzi can also be transmitted through blood
transfusions, organ transplantation, transplacentally and in
laboratory accidents.
• Geographic Distribution: The Americas from the southern
United States to southern Argentina. Mostly in poor, rural
areas of Mexico, Central America, and South America.
Chronic Chagas disease is a major health problem in many
Latin American countries.
Clinical Features
• The acute phase is usually asymptomatic, but can present with
manifestations that include fever, anorexia, lymphadenopathy, mild
hepatosplenomegaly, and myocarditis.
• Romaña's sign (unilateral palpebral and periocular swelling) may
appear as a result of conjunctival contamination with the vector's
feces.
• A nodular lesion or furuncle, usually called chagoma, can appear at
the site of inoculation.
• Most acute cases resolve over a period of a few weeks or months
into an asymptomatic chronic form of the disease.
Clinical Features
• The symptomatic chronic form may not occur for years or even
decades after initial infection.
• Its manifestations include cardiomyopathy (the most serious
manifestation); pathologies of the digestive tract such as
megaesophagus and megacolon; and weight loss.
• Chronic Chagas disease and its complications can be fatal.
Leishmania
(Mastigophora – Flagellates)
• Transmission route: Vectorial Transmission
(Biological vector)
Phlebotomine sandfly
Leishmania (Mastigophora – Flagellates)
Leishmania
(Mastigophora – Flagellates)
• Leishmaniasis is transmitted by the bite of infected female
phlebotomine sandflies.
• The sandflies inject promastigotes (the infective stage) from
their proboscis during blood meals.
• Promastigotes that reach the puncture wound are
phagocytised by macrophages and other types of
mononuclear phagocytic cells.
• Progmastigotes transform in these cells into amastigotes (the
tissue stage of the parasite), which multiply by simple division
and proceed to infect other mononuclear phagocytic cells.
Leishmania
(Mastigophora – Flagellates)
Macrophages infected with amastigotes
Leishmania
(Mastigophora – Flagellates)
• Sandflies become infected
by ingesting infected cells
during blood meals.
• In sandflies, amastigotes
transform into
promastigotes, develop in
the gut and migrate to the
proboscis.
– in the hindgut for leishmanial
organisms in the Viannia
subgenus
– in the midgut for organisms in
the Leishmania subgenus)
Leishmaniasis
• Parasite, host and other factors affect whether the infection
becomes symptomatic and whether cutaneous or visceral
leishmaniasis results.
Leishmaniasis
Geographic Distribution:
• Leishmaniasis is found in parts of about 88 countries (approximately
350 million people live in these areas)!
• Most of the affected countries are in the tropics and subtropics.
• The settings in which leishmaniasis is found range from rain forests
in Central and South America to deserts in West Asia.
• More than 90% of the world's cases of visceral leishmaniasis are in
India, Bangladesh, Nepal, Sudan and Brazil.
• Leishmaniasis is found in Mexico, Central America, and South
America -from northern Argentina to Texas (not in Uruguay, Chile,
or Canada), Southern Europe, Asia (not Southeast Asia), the Middle
East, and Africa (particularly East and North Africa, with some cases
elsewhere).
Leishmania sp.
• Human infection is caused by about 21 of 30 species that infect mammals.
These include:
 L. donovani complex
 L. donovani, L. infantum and L. chagasi
 L. mexicana complex
 L. mexicana, L. amazonensis and L. venezuelensis
 L. tropica; L. major; L. aethiopica;
 the subgenus Viannia
 L. (V.) braziliensis, L. (V.) guyanensis, L. (V.) panamensis and L. (V.) peruviana
• The different species are morphologically indistinguishable, but they can
be differentiated by isoenzyme analysis, molecular methods, or
monoclonal antibodies.
Trichomonas vaginalis
(Mastigophora – Flagellates)
• Transmission route: Sexual contact
T. vaginalis trophozoites
T. vaginalis (Mastigophora – Flagellates)
T. vaginalis (Mastigophora – Flagellates)
• T. vaginalis resides in the female lower genital tract and the
male urethra and prostate, where it replicates by binary
fission.
• The parasite does not appear to have a cyst form, and does
not survive well in the external environment.
• T. vaginalis is transmitted among humans, its only known
host, primarily by sexual intercourse.
• Geographic Distribution: Worldwide. Higher prevalence
among persons with multiple sexual partners or other
venereal diseases.
Giardia duodenalis
(Mastigophora – Flagellates)
• Transmission route: fecal-oral route
Trophozoite
Cyst
G. duodenalis (Mastigophora – Flagellates)
G. duodenalis (Mastigophora – Flagellates)
G. duodenalis (Mastigophora – Flagellates)
• Cysts are resistant forms and are responsible for transmission of
giardiasis.
• Both cysts and trophozoites can be found in the feces (diagnostic
stages).
• The cysts are hardy and can survive several months in cold water.
• Infection occurs by the ingestion of cysts in contaminated water,
food, or by the fecal-oral route (hands).
• In the small intestine, excystation releases trophozoites (each cyst
produces two trophozoites) .
• Trophozoites multiply by longitudinal binary fission, remaining in
the lumen of the proximal small bowel where they can be free or
attached to the mucosa by a ventral sucking disk.
G. Duodenalis (Mastigophora – Flagellates)
• Encystation occurs as the parasites transit toward the colon.
• The cyst is the stage found most commonly in non-diarrheal feces .
• Because the cysts are infectious when passed in the stool or shortly
afterward, person-to-person transmission is possible.
• Wild animals are infected with Giardia, their importance as a reservoir
is unclear.
• Geographic Distribution: Worldwide, more prevalent in warm
climates, and in children.
Toxoplasma
(Philum: Sporozoa)
• Transmission routes:
 row (undercooked) meat ingestion,
 vegetables ingestion,
 contact with cat stool,
 mother > fetus
Toxoplasma
(Philum: Sporozoa)
• Apicomplexa
• Motility: sliding
Toxoplasma (Philum: Sporozoa)
Tissue cyst bradyzoites
Oocyst
Tachyzoites
Toxoplasma (Philum: Sporozoa)
• The only known definitive hosts for Toxoplasma gondii are members
of family Felidae (domestic cats and their relatives).
• Unsporulated oocysts are shed in the cat’s feces.
• Oocysts take 1-5 days to sporulate in the environment and become
infective.
• Intermediate hosts in nature (including birds and rodents) become
infected after ingesting soil, water or plant material contaminated
with oocysts .
• Oocysts transform into tachyzoites shortly after ingestion.
• These tachyzoites localize in neural and muscle tissue and develop
into tissue cyst bradyzoites.
• Cats become infected after consuming intermediate hosts
harboring tissue cysts.
Toxoplasma (Philum: Sporozoa)
• Humans can become infected by any of several routes:
 eating undercooked meat of animals harboring tissue cysts
 consuming food or water contaminated with cat feces or by
contaminated environmental samples (such as fecalcontaminated soil or changing the litter box of a pet cat)
 blood transfusion or organ transplantation
 transplacentally from mother to fetus.
• In the human host, the parasites form tissue cysts, most commonly
in skeletal muscle, myocardium, brain, and eyes; these cysts may
remain throughout the life of the host.
• Diagnosis is usually achieved by serology, although tissue cysts may
be observed in stained biopsy specimens.
• Diagnosis of congenital infections can be achieved by detecting T.
gondii DNA in amniotic fluid using molecular methods such as PCR.
Toxoplasmosis diagnosis
Plasmodium
(Philum: Sporozoa)
• Transmission route: vectorial transmission
(biological vector)
Mosquito (Genus: Anopheles)
Plasmodium (Philum: Sporozoa)
4 species infecting human (malaria):
– Plasmodium falciparum
– Plasmodium vivax
– Plasmodium malariae
– Plasmodium ovale
Plasmodium (Philum: Sporozoa)
Plasmodium (Philum: Sporozoa)
• The malaria parasite life cycle involves two hosts.
• During a blood meal, a malaria-infected female Anopheles mosquito
inoculates sporozoites into the human host.
• Sporozoites infect liver cells and mature into schizonts (I), which
rupture and release merozoites (I).
– In P. vivax and P. ovale a dormant stage [hypnozoites] can persist in the liver
and cause relapses by invading the bloodstream weeks, or even years later.
• After this initial replication in the liver (exo-erythrocytic
schizogony), the merozoites undergo asexual multiplication in the
erythrocytes (erythrocytic schizogony).
• The ring stage trophozoites mature into schizonts (II), which
rupture releasing merozoites (II).
– Blood stage parasites are responsible for the clinical manifestations of the
disease.
• Some parasites differentiate into sexual erythrocytic stages, the
gametocytes.
Ring stage of trophozoites and gametocytes
of P. faciparum in human blood
Plasmodium (Philum: Sporozoa)
• The gametocytes, male (microgametocytes) and female
(macrogametocytes), are ingested by an Anopheles mosquito during
a blood meal.
• The parasites’ multiplication in the mosquito is known as the
sporogonic cycle.
• While in the mosquito's stomach, the microgametes penetrate the
macrogametes generating zygotes.
• The zygotes in turn become motile and elongated (ookinetes) which
invade the midgut wall of the mosquito where they develop into
oocysts.
• The oocysts grow, rupture, and release sporozoites, which make
their way to the mosquito's salivary glands.
• Inoculation of the sporozoites into a new human host perpetuates
the malaria life cycle .
Plasmodium development in the mosquito
15 min
1 hour
12 hour
8-20 day
24-36 hour
Malaria
• Each year, there are approximately 350–500 million cases of malaria, killing
between one and three million of people, the majority of whom are young
children in sub-Saharan Africa.
• Currently malaria is considered the second deadly infectious disease after
HIV.
• The unbearable burden of malaria is increasing worldwide and the
projection of global malaria shows a spreading in non endemic regions in
the next decades.
Scanning electron micrographs of
Plasmodium-infected red blood cells
One cell has burst open,
releasing merozoites.
Erythrocyte, parasitized by
Plasmodium falciparum,
showing surface knobs.
Pathogenesis
• Primary attack
– Infected erythrocyte rupture →
– products of schizont, stimulate the release of cytokines (TNF) →
paroxysm (shiver, fever, sweat)
• Relapse
– It is a recurrence that taken place after complete initial clearing of
the erythrocytic infection and implies re-invation of the blood
stream by merozoites from activated hypnozoites in liver (P. vivax
and P. ovale).
• Recrudescence
– It is a recurrence of symptoms in a patient whose blood stream
infection has previously been at such a low level as not to be
clinically demonstrable or cause symptoms.
Complications
• Anemia
– Hemolysis of infected erythrocytes
– Hypersplenism
– Autoimmunization of uninfected erythrocytes
– TNF-α
•
•
•
•
Splenomegaly
Malarious nephrosis
Cerebral malaria
Death
Immunity
• Evasion of immunity
– An ability of malaria parasite to evade host immunity.
• Possible mechanism of evasion:
 Antigenic variation
 Sequestration (avoiding exposure to immune effector
mechanisms)
 Poor immunogenicity of its antigens (analogy exists
between parasitic antigens and host molecules)
Diagnosis
• Parasitological diagnosis:
– Parasite; Species; Density
 Thin blood films (species identification)
 Thick blood films
Diagnosis
• Immuno-diagnosis:
 Specific antibody detection
– past malaria
 Antigen detection
• Specific DNA or RNA detection
Treatment
Classes of antimalarial drugs:
1) Blood schizonticides: Quinine, chloroquine, artemisinin,
mefloquine, sulfadoxin-pyrimethamine.
 Effect on erythrocytic stage (use for acute attack).
2) Tissue schizonticides: Primaquine.
 Effect on the stages in liver (including hypnozoite), use for
prevent relapse (radical cure) of P. vivax or P. ovale malaria.
Choice of drugs
1) Treatment of P. vivax, P. malariae, P. ovale and
chloroquine-sensitive P. falciparum malaria:
chloroquine.
2) Radical cure of P. vivax or P. ovale malaria:
chloroquine + primaquine.
3) Treatment of chloriquine-resistant P.
falciparum malaria: artemisinin or mefloquine
or quinine.
Distribution of malaria in the world
♦ Elevated occurrence of chloroquine- or multi-resistant malaria
♦ Occurrence of chloroquine-resistant malaria
♦ No Plasmodium falciparum or chloroquine-resistance
♦ No malaria
Malaria vectors
Transmission and Prevention
Factors of transmission
 Infected human (gametocyte-bearing).
 Suitable species of Anopheles (60 species are
considered to be vectors of malaria!).
 Resistance of Anopheles and Plasmodium to
insecticides and antimalarial drugs respectively.
 Other transmission mode: by transfusion, syringe,
congenital transmission.
Prevention: breaking the humanmosquito-human cycle
 Control of the source of infection by chemotherapy
 Control of transmission route:
 residual insecticides, avoidance of infected mosquitoes
(bed nets impregnated with permethrin; mosquito
repellents as diethyl-metatoluamide)
 Chemoproplylaxis taking suppressive drugs,
beginning one week before travel to endemic area
and continuing until 6 weeks after return.
 Malaria vaccines (an effcient vaccine is not yet
available).