oppportunistic parasitic infection

1. TOXOPLASMA
GONDI

2. • Toxoplasma gondii is an obligate intracellular parasite affecting a wide
range of mammals and birds including humans.
• Causes Opportunistic infections in immunocompromised persons and
congenital infection in fetus.
• Habitat
• Obligate intracellular parasite found inside the reticuloendothelial cellsand
other nucleated cells of the host( muscles and intestinal epithelium)

3. Morphology
• It exists in three morphological forms—two asexual
forms (tachyzoite and tissue cyst) and a sexual form
(oocyst).
• Tachyzoite
• actively multiplying form, seen in acute infection.
• Crescent shaped, having a pointed anterior end and a
blunt posterior end
• Has a ovoid nucleus situated between center and
posterior end
• Found commonly in skeletal muscle, cardiac muscle, and
brain.
• Can invade all types of mammalian cells except RBCs.
• Inside the host cell, they multiply with in vacuole by a
process called as endodyogeny by which daughter
trophozoites are formed within the parent cell. They
often form rosettes surrounding the host nucleus

4. • Tissue cyst
• It is the resting stage of the parasite,
usually seen in chronic infections.
• Found in any organ however commonly
found in brain, skeletal and heart
muscles
• The cyst contains many crescent
shaped PAS positive slowly multiplying
trophozoites called as bradyzoites,
surrounded by a cyst wall.
• Bradyzoites are smaller than
tachyzoites and they multiply by
endodyogony
• Tissue cysts vary in size : spherical (40-
50mm in diameter)

5. • Oocyst
• Oocyst is the sexual form of the
parasite found in cats and other
felines.
• It measures 10–12 μm in size,
surrounded by a refractile and
resistant double layered colorless
cyst wall
• Unsporulated oocyst excreted in cat’s
feces is noninfectious. In the
environment, they transform into
sporulating oocyst that contains two
sporocysts (8 μm × 6 μm) each
containing four elongated sporozoites
(6–8 μm × 1–2 μm).

6. Life cycle
• Host: The life cycle involves two hosts:
• 1. Definitive hosts are cat and other felines; where the sexual cycle
takes place
• 2. Intermediate hosts are man and other mammals (goat, sheep, pig,
cattle and certain birds); where the asexual cycle takes place.
• Transmission and infective form: T. gondii in all the three
morphological forms can transmit the infection. Transmission to man
occurs by:
• Ingestion of sporulated oocysts from contaminated soil, food, or water (most
common route)
• Ingestion of tissue cyst containing bradyzoites from undercooked meat
• By blood transfusion, needle stick injuries, organ transplantation,
transplacental transmission or laboratory accidents. Tachyzoites are the
infective form.

8. • Asexual Cycle or Exoenteric Cycle (The Human Cycle)
• Transform into tachyzoites : In the intestine, sporozoites are released
from sporulated oocyst and bradyzoites are released from the tissue
cyst. They invade the intestinal epithelium and transform into
tachyzoites.
• Transform into tissue cyst: Tachyzoites multiply actively by
endodyogeny and spread locally to the mesenteric lymph node.
Subsequently, they also spread to distant extraintestinal organs like
brain, skeletal and cardiac muscles, eye, liver, etc. where they
transform into bradyzoites which multiply slowly to form tissue cysts.

9. • Sexual Cycle or Enteric Cycle (The Feline Cycle)
• Cat and other felines acquire infection by ingestion of tissue cysts in the
meat of rodents and other animals.
• Bradyzoites are released from the tissue cysts, which invade the intestinal
epithelium, undergo several cycles of asexual generations (schizogony)
before the sexual cycle begins
• Sexual cycle (gametogony) begins when the parasite differentiates to form
male and female gametocytes which then transform into male and female
gametes respectively
• Fertilization of male and female gamete results in formation of zygote
which later gets surrounded by a thin, resistant rigid wall to form oocyst
• Oocysts are released in cat’s feces which are unsporulated and
noninfective. The maturation takes place 2–3 days later, in the humid
environment. The mature sporulated oocyst containing two sporocysts is
infectious to man for about 1 year

10. Pathogenicity and Clinical features
• Toxoplasmosis is one of the most common parasitic zoonotic infections. Its
prevalence in humans varies from 5–75% and depends on various risk
factors like:
• The geographical area (cold area, hot arid climatic conditions, high altitudes are
associated with a low prevalence)
• Age: It commonly affects older age and fetus
• Exposure to cat and cat’s feces
• Food habits: Ingestion of uncooked cat and other animal meat (seen in countries like
France)—at higher risk
• Immune status: Patients associated with HIV, malignancies and other
immunocompromised conditions are at high risk
• Patients undergoing blood transfusion, and organ transplantations are at higher risk.

11. • Toxoplasmosis in Immunocompetent Patients
• Here both the humoral and the cellular immune responses control
the infection. Hence, acute toxoplasmosis is usually asymptomatic
and self-limited
• Lymphadenopathy: The most common manifestation of acute toxoplasmosis
is cervical lymphadenopathy. Other lymph nodes may also be affected like sub
occipital, supraclavicular and inguinal nodes
• Other symptoms include headache, malaise, fatigue and fever
• Rare complications are maculopapular rash, pneumonia, myocarditis and
encephalopathy
• Acute infection usually resolves within several weeks, although the
lymphadenopathy may persist for some months.

12. Toxoplasmosis in Immunocompromised Patients
• The clinical manifestations are more severe
• The tachyzoites are disseminated to a variety of organs, particularly
lymphatic tissue, skeletal muscle, myocardium, retina, placenta and
the central nervous system (CNS)
• The parasite infects host cells, replicates, leading to cell death and
focal necrosis surrounded by an acute inflammatory response

13. • Toxoplasmosis in patients with HIV:
• Infection occurs either due to reactivation of latent infection or as a newly
acquired infection from an exogenous source such as blood or transplanted
organs
• It mainly targets CNS leading to Toxoplasma encephalitis (TE)
• Toxoplasma encephalitis: −− Most common areas involved are the
brainstem, basal ganglia, pituitary gland and corticomedullary junction
• TE develops when the CD4+ T cell count falls below 100/μL
• Pathogenesis is due to the direct invasion by the parasite leading to necrotizing
encephalitis and also due to secondary pressure effects on the surrounding area of
the CNS
• Patients may present with altered mental status, seizures, sensory abnormalities,
cerebellar signs and focal neurologic findings including motor deficits, cranial nerve
palsies and visual-field loss
• Apart from CNS infections, other manifestations include pulmonary
infections and chorioretinitis.

14. Congenital Toxoplasmosis
• Mother acquiring Toxoplasma infection in pregnancy is usually
asymptomatic. However she can transmit the infection to the fetus.
• Gestational age is the main factor influencing the fetal outcome. As the
gestation proceeds, the chance of transmission increases but the
severity of the infection declines
• If the mother becomes infected during the first trimester, the incidence of
transplacental infection is lowest (15%), then the disease is most severe
• If maternal infection occurs during the third trimester, the incidence of
transplacental infection is maximum (65%), the infant is usually asymptomatic at
birth
• Retinochoroiditis, cerebral calcifications, and convulsions are the classic
triads
Ocular involvement: Most frequently it causes focal necrotizing
chorioretinitis leading to profound visual impairment. Other ocular
manifestations include blurred vision, pain, scotoma, and photophobia

15. Laboratory Diagnosis
• Specimens:
• peripheral blood, body fluids, lymph node aspirate, bone marrow aspirate,
cerebrospinal fluid (CSF) and bronchoalveolar lavage for HIV infected patients,
biopsy material from spleen, liver and brain
• Direct Microscopic Examination
• stained with Giemsa, PAS, silver stains, immunoperoxidase stain
• Comma shaped Tachyzoites in acute infection
• tissue cyst in chronic infection

16. • Direct fluorescent antibody test (DFA):
• Tachyzoites can be detected by using fluorescein conjugated antibody against
T. gondii surface antigens
• Comma shaped tachyzoites are detected in the smear made from blood, body
fluid and tissue and it indicates acute infection.
• Tissue cyst containing strongly PAS positive bradyzoites can be detected in
various tissues like brain or muscle. This denotes the presence of infection but
cannot differentiate acute and chronic infection.
• Antibody detection
• Several methods are employed for detecting specific anti T. gondii antibodies
like Sabin-Feldman dye test, enzyme-linked immunosorbent assay (ELISA),
indirect fluorescent antibody test (IFA), indirect hemagglutination test (IHA)
and latex agglutination test.

17. • Sabin-Feldman dye test
• Gold standard antibody detection method, done in the reference laboratories.
• It is a complement mediated neutralization test that requires live tachyzoites
• live tachyzoites are incubated with complement and test serum. Alkaline
methylene blue dye is added and mixture is reincubated
• Toxoplasma Abs in the test serum bind to the antigens present on live
tachyzoites which are killed as a result of complement mediated lysis.
• Killed tachyzoites are thin, distorted and colorless (not stained)
• The dilution of the test serum at which 50% of the killed tachyzoites is
reported as antibody titer of the test serum
• The dye test is highly sensitive and specific with no false positives reported so
far
• Drawbacks of this test:
• Difficulty in maintaining the live tachyzoites
• It detects immunoglobulin G (IgG) antibodies, hence cannot differentiate recent or past
infection

18. • Detection of Toxoplasma Antigens
• ELISA is available to detect specific Toxoplasma antigens in blood or body
fluids or amniotic fluid. Detection of antigen indicates acute infection.
• Molecular diagnosis
• Polymerase chain reaction (PCR) can be employed to detect Toxoplasma
specific DNA from various clinical samples like blood or body fluids or
amniotic fluid. PCR is highly sensitive, specific; can be used to diagnose TE or
congenital infections in resource—poor settings.
• Animal inoculation
• T. gondii can be isolated from mice by intraperitoneal inoculation of the
clinical samples into the healthy (T. gondii free) laboratory maintained mice.
Mice die in 7–10 days and peritoneal fluid and spleen aspirate smears show
tachyzoites. If death doesn’t occur, then the mice are observed for 6 weeks
and tail blood is screened for Toxoplasma antibodies.

19. • Tissue culture
• T. gondii can be isolated by inoculating into murine alveolar and peripheral
macrophage cell line.
• Imaging methods
• Computed tomography (CT) scan or magnetic resonance imaging (MRI) of
brain can be done to demonstrate multiple ring enhancing lesions in basal
ganglia or corticomedullary junction to diagnose TE in HIV patients.

20. Treatment
• Pyrimethamine, sulphadiazine, clindamycin, atovaquone and
azithromyzin – effective against tachyzoites, but not bradycysts