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Phylum Apicomplexa
Apical complex, with organelles
involved in Invasion including
Rhoptries,
Micronemes,
Dense granules.
Coccidian Parasites
Plasmodium
Cryptosporidium
Isospora
Subclass:
Coccidiasina
Dr Debasis Biswas
Cyclospora
Sarcocystis
Diarrheal disease
Eimeria
Toxoplasma
Typically asymptomatic;
GI symptoms rare
Overview
Cryptosporidium
Overview
Life Cycle
Single host
Sporulated Oocyst
5/23/2014
Life Cycle
Epidemiological Implications
Sporulated
Oocyst
Readily infectious
Person-to-person transmission
Contrast:
Cyclospora ... unsporulated oocyst (No P2PT)
Epidemiological Implications
Host
response
Proneness to
desiccation
Zoonotic
transmission
Pathogenesis
E nt er o c yt e I nf ec t i o n (S m al l i nt est i ne & p r oxi m al co l o n)
Extent of infection associated with immunosuppression
Pref. loss of mature ep cells
Inflamm. Cytokines, e.g. TNF; IL-1;IFN
at villus tips
Chemokines, e.g. IL-8
Increased Epithelial cell turnover
Up-regulation of COX2 by ep. cells
Influx of inflammatory cells
Prodn. of neuropeptides, e.g. Substance P
Na+ malabsorption
Electrogenic Cl- secretion
Increased Intestinal permeability
Villous atrophy
Crypt hyperplasia
Apoptosis, followed by
Necrosis, of ep. cells
Malabsorption:
Vit B12, Bile acids, Fatty acids
Clinical manifestations
Immunocompetent hosts
Diarrhea: Median durn. 5-10 d
Acute watery diarrhea
Assoc: Cramps; Nausea; Fever
Vomitting: less common
Course: Recurrence in 40%
pts. after initial resolution
Persistence >14 days in 45% of
children
Chronic: Malabsorption;
Malnutrition; Weight loss
Severe
Cryptosporidiosis
Malnutrition
5/23/2014
Extra-intestinal manifestations
Diagnosis
Acalculous cholecystitis
Sclerosing cholangitis
Pancreatitis
Right upper quadrant abd pain
(Intermittent & Colicky)
Enzyme alterations:
(Alk Phos/ AST/ ALT/ Amylase/ Lipase)
USG:
Dilated biliary tracts & Cholecystitis
Respiratory Tract
Asymptomatic
Bilateral Pulm infiltrates
Dyspnea
Diagnosis
Direct IF:
Oocyst-specific monoclonal antibody
Gold standard
Microscopy
Wet mount examn : Oocysts
4 6 diam: Yeast forms
Differential staining: ZN stain
Auramine-O
Stool concn technique: Increases sensitivity
Sedimentn: Formalin ether/ Formalin ethyl acetate
Floatn: Sheathers Sucrose/ Saline
Antigen detectn:
ELISA
Immunochromatography
PCR:
Increased sensitivity
ZN Stain
Auramine-Rhodamine Immunofluorescence
Intestinal Biopsy
Cyclospora cayetanensis
5/23/2014
Wet mount
ZN Stain
Diagnosis
Microscopy
Wet mount examn : Oocysts
2 X size of Cryptosporidium oocyst (8 10 )
Differential staining: ZN stain
Auramine-O
Blue autofluorescence under UV epifluorescence
microscopy
Alt. stains : Safranine/ Lactophenol Cotton Blue
Diagnosis
Histopathology / EM of Jejunal aspirates/ Biopsy:
Villous atrophy
Ac/ Chr inflammn in lamina propria
Cyclospora: supranuclear locn within cytoplasm of
enterocytes, cf . Cryptosporidium: surface of
enterocytes
PCR:
Flow cytometry:
Isospora belli
5/23/2014
Diagnosis
Microscopy
Wet mount examn : Oocysts
Large & Elliptical oocysts (22 33 X 12- 15 )
Differential staining: ZN stain
Auramine-O
Blue autofluorescence under UV epifluorescence
microscopy
Alt. stains : Safranine/ Lactophenol Cotton Blue
ZN Stain
Autofluorescence
Treatment
Diagnosis
Histopathology / EM of Jejunal aspirates/ Biopsy:
Villous atrophy
Infiltration of inflammatory cells, particularly
eosinophils, in lamina propria
Crypt hyperplasia
Isospora: parasitophorous vacuoles of enterocytes
Immunocompetent
hosts
Fluid replacement.
Anti-motility agents.
C. parvum
Efficacy of anti-parasitic
agents not proven.
Nitazoxanide
PCR:
HIV/AIDS pts.
3- drug Anti-retroviral
regimen with protease
inhibitors.
PIs can have anticryptosporidial activity.
TMP-SMX: BD X 7 - 10 days
+ Suppressive therapy thrice
weekly is recommended to
prevent relapse
Phylum Apicomplexa
Plasmodium
Subclass:
Coccidiasina
Cryptosporidium
Isospora
Cyclospora
Sarcocystis
Toxoplasma gondii
Eimeria
Toxoplasma
Congenital Infection
5/23/2014
4 Parasitic stages
3 Infective Stages
Tachyzoites
Tachyzoites
Excreted in
the stool of
Definitive hosts:
Felines
Four
Sporozoites
Invasive form
Rapidly dividing
Macrophage
Sporulated Oocyst
Unsporulated Oocyst
Two
Sporocysts
Tissue Cyst
Tissue Cyst
Environmental form
Survive > 1 yr in moist env. with Bradyzoites
Water, Soil, Vegetable & Fruit
Contamination
Four
Sporozoites
Latent form
Slowly dividing
Long-term survival
Sporulated Oocyst
Tachyzoites
Oocyst
Humans:
Int. Hosts
Tachyzoite
Tissue Cyst
5/23/2014
Pathogenesis
Pathogenesis
REACTIVN
Mostly asymptomatic
May mimic Infectious Mononucleosis (Epstein Barr Virus)
Fever Cervical Lymphadenopathy
Monocytosis in peripheral smear
Rarely: Chorioretinitis
Severity related to genotype of the strain
Type II:
Western Europe & North America Less severe manifestns
Other genotypes:
Africa & South America . Higher incidence of Chorioretinitis
or other serious manifestns like meningoencephalitis,
pneumonitis, myocarditis, etc.
Microphthalmia Cataract
Strabismus Increased IOP
Optic neuritis
Retinal necrosis
Uveitis
Chorioretinitis
Blindness
Hepatosplenomegaly Pneumonitis
Anemia Thrombocytopenia
Diagnosis
A. Parasite- based
Demonstration of parasite:
Tachyzoites in tissue sections/ smears from body fluids
Tissue Cyst in tissues, + surrounding inflammation/ necrosis
Isolation of parasite:
Mouse inoculation
Cell culture
Demonstration of parasitic DNA:
PCR of blood/ body fluids/ tissues
B. Host response- based
Antibody to parasite: Serologic tests
5/23/2014
Diagnosis
Diagnosis
A. Parasite- based
Demonstration of parasite:
Tachyzoites in tissue sections/ body fluids (CSF/ Amn fl/ BAL)
Tissue Cyst in tissues, + surrounding inflammation/ necrosis
Encephalitis/ Lymphadenitis/ Pneumonitis/ Myocarditis
Wright- Giemsa stain: Cheap; Quick; Less sensitive
Fluorescent Antibody staining
Immunoperoxidase staining
Diagnosis
A. Parasite- based
Isolation of parasite:
Blood/ Body Fluids: Suggests acute infection
Placenta/ Fetal tissues: Suggests congenital infection
Mouse inoculation: More sensitive
Cell culture: Quicker .. 3-6 days
Demonstrn of parasite- laden cells: Plaques
containing tachyzoites
Diagnosis
A. Parasite- based
Demonstration of parasitic DNA:
PCR: targeting multi copy B1 gene or REP-529 gene
Increased sensitivity before or within the 1st week of t/t.
Whole Blood/ Buffy Coat: Disseminated Toxoplasmosis
Amniotic Fluid: Intrauterine Infection
Placenta/ Fetal tissues: Congenital Toxoplasmosis
CSF: Toxoplasma Encephalitis
Appear
IgA
IgM
IgG
Plateau
1 week
1 month
4-6 weeks
2-3 months
Disappear
9 months
1 year
Lifelong
5/23/2014
IgG +
IgM -
Distant infection
No Follow/ up
Monthly Follow/ up
Till 2-3 weeks after delivery
Subsequent
Seroconversion
+ve
T/t switch to
Pyrimethamine + Sulfonamide
Abortion proposed in presence
of echographical signs
-ve
IgG +/IgM +
IgG Avidity
High
Low
Rising
Titers
Dinagnosis: Immunocompetent
Serologic tests:
To determine the immune status
Pregnant lady in early stage of pregnancy
Uveitis/ Retinochoroiditis w/o h/o congenital infection
Organ Donors/ Transplant recipients
D/D of Fever Lymphadenopathy: CMV/ EBV/ HIV
Parasite isolation/ demonstration: Less common
> 33 wks
Postnatal dg:
PCR: Placenta/ Cord blood
Cord blood serology (IgM/ IgA)
Western Blot of mother-infant
paired serum
Infant serum (IgM/ IgA)
Diagnosis: Immunocompromised
Parasite detection: Cornerstone
Cerebral: CSF/ Blood
Disseminated: Particularly Transplant recipients
Blood/ Bone Marrow Aspirate/ BAL/ CSF
PCR
Isolation: Cell Culture/ Mouse inoculation
Histology
Serologic tests:
To exclude Toxoplasmosis
To monitor disease reactivation
Rising/ Very high IgG titers
Reappearance of IgM
Diagnosis: Retinochoroiditis
Typically Based on:
Ophthalmoscopic exam:
Typical white focal lesions with vitreous
inflammatory reaxn
Seropositivity for Toxoplasma
Response to Anti- Toxoplasma t/t
In case of atypical lesions or inadequate response to t/t:
PCR of AH/ VH
Serologic tests: Local ab production
GWC: Goldmann-Witmer coefficient
Anti- Toxo IgG in AH/ Anti- Toxo IgG in serum
Total IgG in AH/ Total IgG in serum