University of Maysan

College of Medicine
Department of Microbiology and Parasitology
2023 -2024
Dr. Rasha Khalil Abd Aljalil Alsaad
Ph.D. Parasitology Lecture: 10

Miscellaneous Sporozoa and Microspora

TOXOPLASMA GONDII
History
Toxoplasma gondii is an obligate intracellular coccidian parasite, first described
in 1908 by Nicolle and Manceaux in spleen and liver smears of a small North
African rodent called gondi, Ctenodactylus gundi.
Its importance as a human pathogen was recognized only much later, when Janku
in 1923 observed the cyst in the retina of a child with hydrocephalus and
microphthalmia, Wolf and Cowen in 1937 identified the first
congenital brain infection , and Pinkerton and Weinman recorded postnatal
infection in 1940.
With the discovery in 1948, of the Sabin-Feldman dye test, the first serological
assay for toxoplasma antibody, the scope and extent of the infection became open
for study.
The name toxoplasma is derived from the Greek word Toxon meaning arc or
bow, referring to the curved shape of the trophozoite. Toxoplasmosis is now
recognised as the most common protozoan parasite globally, with the widest
range of hosts spread over 200 species of birds, reptiles and mammals, including
humans.
The life cycle of the parasite became clear only in 1970 when the domestic cat
was identified as its definitive host, only in which it undergoes the sexual
sporogony.
All other species are merely intermediate hosts, in which only asexual
schizogony takes place. Though human infection is very common, perhaps
involving a third of the human race, clinical disease is relatively rare, being
mostly opportunistic in nature.

Morphology
T. gondii occurs in three forms—trophozoite, tissue cyst and oocyst
Toxoplasma gondi
There are only two morphologic forms seen in humans, tachyzoites and
bradyzoites.
The infective form for humans is the oocyst. Thus, all three of these
morphologic forms are discussed in this section.
Tachyzoites
The actively multiplying, crescent- shaped tachyzoites range in size from 3 to
7 μm by 2 to 4 μm . One end of the organism often appears more rounded
than the other end. Each tachyzoite is equipped with a single centrally located
nucleus, surrounded by a cell membrane. A variety of other organelles are
present, including a mitochondri and Golgi apparatus; however, these structures
are not readily visible.
Single nucleus usually situated towards the blunt end. It appears purplish/red.
Blue cytoplasm.

FIGURE : Toxoplasma gondii. (A) Smear from peritoneal fluid of infected

mouse,
showing crescentic tachyzoites—extracellular trophozoites and intracellular form
within macrophage. (B) Thick-walled tissue cyst containing rounded
formsbradyzoites.(C) Oocyst containing two sporocysts with sporozoites inside

The trophozoite and tissue cyst represent stages in asexual multiplication

(schizogony), while the oocyst is formed by sexual reproduction (gametogony or
sporogony). All three forms occur in the domestic cat and other felines which are
the definitive hosts and which support both schizogony and gametogony.
Only the asexual forms, trophozoites and tissue cysts are present in other
animals, including m humans, and birds, which are the intermediate hosts.
Both oocysts and tissue cysts are infective by ingestion.
Tissue Cyst
The tissue cyst is formed during the chronic phase of the infection and can be
found in the muscles and various other tissues and organs, including the brain.
The parasite multiplying slowly within the host cell, produces a cyst wall within
the host cell membrane.
The cyst wall is eosinophilic and stains with silver, in contrast to the
‘pseudocyst’. With periodic acid Schiff stain, the cyst wall stains weakly and the
parasites inside deeply. The slowly multiplying parasites within the cyst are
called bradyzoites.
The cyst is round or oval, 10 to 200 μm in size and contains numerous
bradyzoites. Cysts remain viable in tissue for several years. In immunologically
normal hosts, the cysts remain silent, but in the immunodeficient subjects they
may get reactivated, leading to clinical disease.
It is relatively resistant and when meat containing the cysts is eaten raw or
undercooked, infection occurs. The cyst wall
is disrupted by peptic or tryptic digestion and the released parasites initiate
infection by invading intestinal epithelial cells. They reach various tissues and
organs through blood and lymphatic dissemination. Cysts are susceptible to
desiccation, freezethawing and heat above 60°C.
Bradyzoites
Although there is evidence to support an antigenic difference, the typical
bradyzoite basically has the same physical appearance as the tachyzoite, only
smaller These slow-growing viable forms gather in clusters inside a host cell,
develop a surrounding membrane, and form a
cyst in a variety of host tissues and muscles outside the intestinal tract. Such
cysts may contain as few as 50 and up to as many as several
thousand bradyzoites. A typical cyst measures from 12 to 100 μm in diameter
Oocyst
The typical infective form of Toxoplasma gondii, The round to slightly oval form
measures 10 to 15 μm long by 8 to 12 μm wide.
The transparent oocyst contains two sporocysts, each with four sporozoites. The
organism is bordered by a clear, colorless, two-layered cell wall.
Oocysts develop only in definitive hosts—in the intestines of cats and other
felines.
When cats get infected by ingestion of either tissue cysts or oocysts, the parasites
develop in the intestinal epithelial cells, where both schizogony and gametogony
take place. Male and female gametocytes develop and after fertilisation, the
zygote gets surrounded by a thin, but extremely resistant wall. This is the oocyst,
which is spherical or ovoid. about 10 to 12 μm in size and contains a sporoblast.
Cats shed millions of oocysts per day in faeces for about two weeks during the
primary infection.
The freshly passed oocyst is not infectious. It becomes infectious only after
development in soil or water for a few days. During this state of sporulation, the
sporoblast divides into two sporocysts and four sporozoites develop inside each
sporocyst. The mature oocyst containing eight sporozoites is the infective form.
It is very resistant to environmental conditions and can remain infective in soil
forabout a year. When the infective oocyst is ingested, it releases sporozoites in
the intestine, which initiate infection.
Life Cycle
The life cycle of the parasite consists of 3 stages as follows—(a) Tachyzoites, the
rapidly multiplying trophozoites which invade and multiply within cells,
(b) bradyzoites, the slowly multiplying forms inside tisssue cysts, seen during
latent and chronic infection, and (c) sporozoites inside oocysts, which are shed in
cat feces and remain in the environment.
In cats, which are the definitive hosts. both schizogony and gametogony take
place in the epithelial cells of the small intestine. This is known as the enteric
cycle.
The oocysts produced by gametogony are shed in faeces. They develop into
infective forms in soil or water. They may be ingested by felines to repeat the
cycle.
When ingested by other man ,animals or birds. which are intermediate hosts.
The oocysts release sporozoites which infect the intestinal epithelial cells. Here,
they multiply by endodyogeny to form tachyzoites. The host cell ruptures
releasing numerous trophozoites which spread through blood and lymph
infecting any type of nucleated cell in various tissues and organs. This is known
as the exoenteric cycle.
Primary infection with the parasite may be asymptomatic, acute or chronic. In
chronic infections tissue cysts are produced within muscles and other tissues.
When other intermediate hosts ingest these tissue cysts, the asexual cycle is
repeated. When cats ingest the tissue cysts they become infected and in them
both asexual and sexual cycles are repeated.
Human infection is a dead end for the parasite. Human infection is obtained in
the following ways:
a. Eating uncooked or undercooked infected meat containing tissue cysts.
b. Ingestion of mature oocysts through food, water or fingers contaminated with
cat feces directly or indirectly.
c. Intrauterine infection from infected mothers to babies.
d. Rarely by blood transfusion or transplantation from infected donors.
FIGURE: Life cycle of Toxoplasma gondii.

Clinical Features
Most human infections are asymptomatic. Clinical toxoplasmosis may be
congenital or acquired.
Asymptomatic
Many patients infected with T. gondii remain asymptomatic, especially children
who have passed the neonatal stage of their lives. Although well adapted to its
surroundings,
T. gondii appears to only cause disease in humans when one or more of the
following conditions have been met: (1) a virulent strain of the organism has
entered the body; (2) the host is in a particularly susceptible state (e.g., those
suffering from AIDS); and (3) the specific site of the parasite in the human body
is such that tissue destruction is likely to occur.
Congenital Toxoplasmosis
Congenital toxoplasmosis results when infection is transmitted transplacentally
from mother to foetus. This occurs when the mother gets primary toxoplasma
infection, whether clinical or asymptomatic during the pregnancy. The risk of
foetal infection rises with the progress of gestation, from 25 per cent when the
mother acquires primary infection in the first trimester, to 65 per cent in the third
trimester.
Conversely the severity of foetal damage is highest when infection is transmitted
in early pregnancy. Mothers with chronic or latent Toxoplasma infection
acquired earlier do not ordinarily infect their babies, but in some women with
latent or chronic infection, the tissue cyst may be reactivated during pregnancy
and liberate trophozoites which may reach the fetus in utero.
Most infected newborns are asymptomatic at birth and may remain so
throughout. Some develop clinical manifestations of toxoplasmosis weeks,
months or even years after birth.
The manifestations may be chorioretinitis, strabismus, blindness, deafness,
epilepsy or mental retardation.
A few are born with manifestations of acute toxoplasmosis, which may include
fever, jaundice, diarrhoea, petechial rashes, hydrocephalus, microcephaly,
cerebral calcifications, microphthalmia, cataract, glaucoma, chorioretinitis, optic
atrophy, lymphadenitis, pneumonitis, myocarditis and hepatosplenomegaly.
Acquired Toxoplasmosis
Infection acquired postnatally is mostly asymptomatic. Clinical toxoplasmosis
may present in different forms. The most common manifestation of acute
acquired toxoplasmosis is lymphadenopathy, the cervical lymph nodes being
most frequently affected. Fever, headache, myalgia and splenomegaly are often
present.
The illness may resemble mild ‘flu’ or infectious mononucleosis and is self-
limited, though the lymphadenopathy may persist.
In some there may be a typhus-like exanthem,with pneumonitis, myocarditis and
meningoencephalitis, which may be fatal.
Another type of toxoplasmosis is ocular. Approximately 35 per cent of cases of
chorioretinitis in the USA and Europe have been reported to be due to
toxoplasmosis. While most of these follow congenital infection, it may
sometimes be due to postnatal infection.
Some cases may be so severe as to require enucleation. Toxoplasmosis primarily
involving the central nervous system is usually fatal and often found in AIDS.
Toxoplasmosis is particularly severe in the immunodeficient, particularly in
AIDS patients, whether it be due to reactivation of latent infection or to new
acquisition of infection. In them brain involvement is common.
Host defence against toxoplasma infection involves both humoral and cellular
responses. Specific IgG antibody can lyse extracellular trophozoites. But
activated T cells and natural killer cells appear to be more important in
containing the infection and preventing clinical disease.
Diagnosis
Laboratory diagnosis may be made by microscopic demonstration of the parasite,
by its isolation or by serological tests.
Giemsa stained impression smears of lymph nodes, bone marrow, spleen or brain
may occasionally show the trophozoites, which can be readily identified by their
morphology. Tissue sections may show the cyst forms.
Isolation may be intraperitoneally is made by injecting body fluids or ground
tissues into cell cultures or immunosuppressed mice. Peritoneal fluid and spleen
smears may show the trophozoites after 7 to 10 days.
Serial blind passages may often be necessary for isolation. Sera of inoculated
animals may also be tested for antibodies.
The most common method of laboratory diagnosis is by serology. Several
serological tests are available. These include the Sabin-Feldman dye test, indirect
immunofluorescence, indirect haemagglutination, complement fixation and
ELISA.
The Sabin-Feldman dye test is based on the specific inhibition by antibody of the
staining of the trophozoite by alkaline methylene blue.
Toxoplasma trophozoites
propagated in mice peritoneal cavity are used. An accessory factor present in
fresh

The CF test
becomes positive only 3 to 8 weeks after infection rises in titre over the next 2
to 8 months and declines to low or undetectable levels within a year. The dye
test was the first serological test for toxoplasmosis and remained the gold
standard
for many decades. But as the test required live toxoplasma, it could be done only
in select laboratories. It is seldom done now because of its complexity and as
better
tests like ELISA are avialable. The standard test used now is ELISA, separately
for IgM and IgG antibodies.
ELISA provides
better information than a sinlge test.
Epidemiology
The infection is worldwide, being found wherever there are cats. Numerous
species of mammals, reptiles and birds are naturally infected. The full natural
cycle is maintained predominantly by cats and mice. The mice eat materials
contaminated with oocysts shed by cats. Mice get infected and develop cysts in
their tissues. When such mice are eaten by cats they get infected. Infected cats
shed oocysts in faeces.
Besides this cycle, several others have been documented.
Human toxoplasmosis is a zoonosis. It is acquired through food or water
contaminated with mature oocysts or by ingestion of raw or undercooked meat
containing tissue cysts. Pork and beef frequently have tissue cysts. Flies and
cockroaches may act as mechanical vectors by contaminating food with oocysts
fromsoil.
Infection may be water borne when the source of water is contaminated with
cat faeces. Rarely infection may be transmitted through blood or leucocyte
transfusion or organ transplantation.
Toxoplasmosis may also be acquired by laboratory infection.
The incubation period is usually 1 to 3 weeks.
The outcome of infection depends on the immune status of the infected person.
Active progression of infection is more likely in immunocompromised
individuals.
Toxoplasmosis has acquired great importance as one of the major fatal
complications
in AIDS.
The incidence of congenital toxoplasmosis is estimated as approximately 1 in
1000 live births.Because of the public health importance of congenital
toxoplasmosis,
serological surveys for toxoplasma antibodies are conducted in many advanced
countries in women of childbearing age, antenatal women and newborns.
Prevention
Eradication of toxoplasmosis appears unlikely because it is so widely
disseminated in nature. But some simple measures may reduce the risk of
infection. These include proper cooking of meat and washing of hands before
eating to safeguard against soil contamination of fingers.
Treatment
Combined treatment with pyrimethamine and sulphonamides or cotrimoxazole
may lead to clinical cure, though the parasites may not be eliminated.
Spiromycin and clindamycin have also been used.
Treatment is effective only against trophozoites and not against cysts.