A brief account of the Role of parasites in human neoplastic disorders

1. By DrANJAN
SARMA
PGT, Microbiology
MD, GMCH
Role of Parasites
in Neoplastic
Disorders

2.  Cancer : The Problem
 What causes cancer ?
 IARC and the agents
 Parasites & Associated
Malignancy
 Blood Flukes : Schistosomiasis
 Liver Flukes
 Malaria & Burkitt’s
Lymphoma
 S. Stercolis & Cancer
 Dual Impact of T.Cruzi
 A few other associations

3. • Cancer isoneof theleading causesof morbidity
and mortality worldwide, with approximately 14
million new casesin 2012.
• Thenumber of new casesisexpected to riseby
about 70% over thenext 2 decades.
• It isnow thesecond leading causeof death
globally, and wasresponsiblefor 8.8 million deaths
in 2015. Globally, nearly 1 in 6 deathsisdueto
cancer.
Cancer : The Problem

4. What causes cancer?
• Cancer arisesfrom thetransformation of normal cellsinto tumour
cellsin amultistageprocessthat generally progressesfrom apre-
cancerouslesion to amalignant tumour. Thesechangesaretheresult
of theinteraction between aperson'sgenetic factorsand 3 categories
of external agents, including:
 physical carcinogens, such asultraviolet and ionizing radiation;
 chemical carcinogens, such asasbestos, componentsof tobacco
smoke, aflatoxin (afood contaminant), and arsenic (adrinking water
contaminant); and
 biological carcinogens, such asinfectionsfrom certain viruses,
bacteria, or parasites.
• International Agency for Research on Cancer (IARC) from 2008 to
2009 havereassessed and classified human carcinogensinto
"discrete" groupsincluding infectiouspathogens.

6. The Agents…
• Infectionswith eleven speciesof pathogensassociated with cancers
areclassified asGroup 1 carcinogens, definitely “carcinogenic to
humans”, by theIARC. Theseagentsinclude:
 Helicobacter pylori,
 hepatitisB virus(HBV),
 hepatitisC virus(HCV),
 Opisthorchis viverrini,
 Clonorchis sinensis,
 Schistosoma haematobium,
 human papillomavirus(HPV),
 Epstein- Barr virus(EBV),
 human T-cell lymphotropic virustype1 (HTLV- 1),
 human herpesvirustype8 (HHV-8) and
 human immunodeficiency virustype1 (HIV-1)

7. Parasites and Carcinomas
• How can wedetermineacarcinogenic parasitic infection?
a. association between parasiteinfection and risk of cancer;
b. development of tumor/cancer assecondary event to theinfection;
c. aparasitewasfound in thecancer tissueby microscopic
examination.
Theexclusion can bedoneby
a. infection asasecondary event of neoplasia,
b. an infection mimicking cancer,
c. asingle-point prevalencebetween cancer and aparticular parasite
(unclear about relationship).

8. Parasitic pathogens and infection-associated malignancy
Parasitic
pathogens
Disease Endemic
areas
Associated cancer Proposed mechanism
of carcinogenesis
BLOOD FLUKES (Schistosoma)
S. haematobium Schistosomias sub-Saharan
Africa
Urinary Bladder Cancer,
Adenocarcinoma,
Squamous Cell Carcinoma
Inflammation, oxidative
stress caused by
parasite-derived
molecules
S. japonicum Schistosomias sub-Saharan
Africa
Colorectal Cancer,
Rectal Cancer,
Squamous Cell Carcinoma
MembranousNephropathy
Metastatic Lung Cancer
Inflammation, oxidative
stress caused by
parasite-derived
molecules
S. mansoni Schistosomias sub-Saharan
Africa
Adenocarcinoma,
Colorectal Cancer,
Hepatocellular Carcinoma
Inflammation, oxidative
stress caused by
parasite-derived
molecules
LIVER FLUKES
Opisthorchis
viverrini
Opisthorchiasis Southeast
Asia
Cholangiocarcinoma Inflammation, oxidative
stress caused by parasite-
derived molecules, cell
proliferation, H. pylori
mediated induction

9. Parasitic
pathogens
Disease Endemic areas Associated cancer Proposed mechanism of
carcinogenesis
LIVER FLUKES
Clonorchis
sinensis
Clonorchiasis China, Korea, N.
Vietnam
Cholangiocarcinoma Inflammation, oxidative stress
caused by parasite-derived
molecules, cell proliferation
Opisthorchi
s felineus
Opisthorchiasis Europe and
Russia
Cholangiocarcinoma Inflammation, oxidative stress
caused by parasite-derived
molecules, cell proliferation
OTHERS
P. falciparum
P. vivax
P. ovale
P. malariae
P. knowlesi
Malaria sub-Saharan
Africa, Southeast
Asia
Burkitt Lymphoma
(indirect
carcinogenicity)
Expansion of the EBV-infected B
cell population,
Suppression of EBV-specific T-cell
immunity,
Reactivation of EBV,
AID-dependent genomic
translocation
Strongyloides
stercoralis
Strongyloidi
asis
sub-Saharan
Africa,
South and Central
America
Southeast Asia
HTLV-1 induced
lymphomas/leukemia
s
(indirect
carcinogenicity)
Colon
adenocarcinoma
Stimulate HTLV-1 replication,
Oligoclonal expansion of HTLV-1-
infected lymphocytes
Trypanosoma
cruzi
Chagas'
disease
South and Central
America
Gastrointestinal
cancer, Uterine
leiomyoma
Unknown

10. Blood Flukes (SCHISTOSOMIASIS)
• A neglected tropical disease(NTD) caused by infection with blood fluke
trematodesof thegenusSchistosoma.
• Out of 207 million casesof schistosomiasiscurrently estimated
worldwide, 90% occur in sub-SaharanAfrica.
• Thefivespeciesof Schistosomathat infect humansare:
 S. haematobium,
 S. mansoni,
 S. japonicum,
 S. intercalatum,
 S. mekongi.
• Most human infectionsaredueto S. haematobium, S. mansoni, and S.
japonicum.
• Theprevalenceof schistosomiasisisassociated with exposure-related
factors, in particular with afavourableenvironment for theimperative
intermediatehost snails, sub-optimal sanitation infrastructure, and host
genetic factors.

11. Distribution of Schistosomiasis

12. Schistosomes That Parasitize Humans
Schistosomes Definitive host Intermediate host (various
genera of snail)
Distribution
African schistosomes
S. haemato bium Man, monkey,
chimpanzee
Bulinus Africa and Middle East
S. manso ni Man, monkey,
chimpanzee and
dog
Biomphalaria Africa and South America
and Caribbean
S. intercalatum Man Bulinus West and Central Africa
Asian schistosomes
S. japonicum Man, dog, cat and
rodent
Oncomelania China and Philippines
S. malayensis Man and
rodent
Robertsiella Malaysia
S. mekongi Man and dog Neotricula Laos and Thailand

13. SCHISTOSOMES
Left: Biomphalaria sp., the intermediate host for S. mansoni.
Right: Bulinus sp., the intermediate host for S. haematobium and S. intercalatum.
Center: Adults of S. mansoni. The thin female resides in the gynecophoral canal of the
thicker male.
Theabsenceof snailsof Bulinus sp and resistanceof common snailsto miracidiaof human
schistosomesareimplicated asthemain reason of relativeabsenceof human schistosomiasisin
India.

14. Life-Cycle of SCHISTOSOMIASIS
IARC
S. haematobium,- Group 1 carcinogen
S. japonicum - Group 2B carcinogen
1. Eggs are eliminated with feces
or urine .
2. Under optimal conditions the
eggs hatch and release
miracidia ,
3. which swim and penetrate
specific snail intermediate
hosts .
4. The stages in the snail include
2 generations of sporocysts
5. and the production of
cercariae .
6. Upon release from the snail,
the infective cercariae swim,
penetrate the skin of the
human host ,
7. and shed their forked tail,
becoming schistosomulae .
8. Circulation
9. The schistosomulae migrate
through several tissues and
stages to their residence in the
veins,
10. Adult worms in humans reside
in the mesenteric venules in
various locations, which at
times seem to be specific for
each species

15. ADULT S. haematobium S. mansoni S. japonicum
Body surface of male Finely tuberculate Grossly tuberculate Non-tuberculate,
Smooth
Testes 4-6 in a cluster 6-9 in a cluster 7 in a linear series
Position of ovary Posterior to the middle of the
body
Anterior to the middle of the
body
Posterior to the middle
of the body
Number of eggs in uterus 20-30 1-4 50-300
EGG
Size and Shape 110-18 um long 40-70 um wide
terminal spine
114-175 um long 45-68 um
wide lateral spine
70-100 um long 50-65
um wide central spine
CERCARIA
Cephalic glands Two pairs , oxophilic Two pairs , basophilic Fours pairs , oxophilic
Schistosoma
haematobium
(A)terminal spined egg;
(B) fork tailed cercaria
A
B
(C) Lateral
spined egg of
Schistosoma
mansoni
C
(D)Egg of
Schistosom
a japonicum
D

16. SCHISTOSOMIASIS RESIDENCE
S. japo nicum Inferior mesenteric vein & superior mesenteric vein
S. manso ni Inferior mesenteric vein.
S. haemato bium Terminal venulesin thewall of thebladder, thegenitourinary
system and thepelvic plexuswithin thedistribution of theinferior
venacava. However, it can also exist in perirectal venulesexcreting
theeggsin thestool.
SCHISTOSOMIASIS BLADDER CANCER
S. haemato bium asso Squamouscell carcinoma, with an earlier ageof onset and
generally sparing thetrigoneof bladder.
Non- S.haematobium asso Transitional cell typeoccurring in theolder agegroup.
SCHISTOSOMIASIS IMMUNOPATHOLOGY
S. Japo nicum
S. manso ni
S. haemato bium
Granulomaformation around theeggsdeposited in thetissuesand
isamanifestation of delayed hypersensitivity reaction.
Leadsto pylephlebitis, peripylephlebitis, portal hypertension,
splenomegaly, oesophageal varices, haematemesisand death.

17. Schisto
some
Clinical Manifestations
Acute Chronic
S.
haemato
bium
The invasion of cercariae in the skin
causes, within 24 hrs, dermatitis at
penetration site followed by allergic pruritic
papular lesion.
Migration of schistosomula in lungs causes
cough with mild fever.
Urinary Tract schistosomiais : Occurs months to years after
infection by cercariae d/t to proliferation of the parasites &
inflammation of tissues of the host caused by deposition of eggs
in various organs. Soluble antigens released from the eggs
provoke delayed type of hypersensitivity reaction around them.
This leads to the formation of egg granuloma & urinary mucosa
shows shows glandular metaplasia (cystitis glandularis)
Symptoms : Terminal haematuria, Dysuria, Frequent urination
Compl : Hydroureter, Hydronephrosis, Secondary Microbial
Infections , Uraemia
Advanced infection brings about metaplastic changes in urinary
mucosa may lead to carcinoma of bladder (SCC).
S.
manso ni
Cercarial Dermatitis or Swimmer’s Itch :
when exposed to fresh or salt water , d/t
cercarial invasion maculopapular rash
occurs within 24 hrs .
Katayama Fever: Serum-sickness like
illness, occurs after 4-6 weeks of infection
Symptoms: Fever, Myalgia, Athralgia, Rt
upper Quadrant pain.
GI schistosomiais : Caused by retention of eggs & formation of
granuloma in the intestinal wall.
Symptoms : Abdominal pain, mucosal diarrhoea / dysentry with
passage of eggs, exudate , mucus.
Compl : Periportal fibrosis ( Symmers’ pipestem fibrosis) , Portal
hypertension, GI haemorrhage.
Hepatosplenic schistosomiasis : Condition manifests as
Periportal fibrosis ( Symmers’ pipestem fibrosis) , Portal
hypertension, Ascites, Splenomegaly
Compl : Schistosomal cor pulmonale, Transverse myelitis
S.
japo nicu
m
Katayama Fever: Ocuurs more commonly
and severely than S.mansoni . Serum-
sickness like illness, occurs after 4-6
weeks of infection
Symptoms: Fever, Chills, Weakness,
Myalgia, Headache, Abdm pain,
Diarrhoea, Eosinophilia
GI schistosomiais : Same as S.mansoni infection but symptoms
develop more repidly.
CNS schistosomiais : Because of smaller egg size, it causes
60% of all Schistosoma brain infections, with CNS involvement
seen in2-4 %.
Symptoms : Headache, seizures, myeloradiculopathy, bladder
dysfunction, paresthesia.

18. Parasite Cancer Mechanisms of Carcinigenesis
S.haemato bium
Bladder
Cancer
I. Fibrosis induced by schistosome eggs may induce proliferation,
hyperplasia and metaplasia, all of which are possible precancerous
changes.
II. Chronic urinary bacterial infection and production of nitrosamines
from their precursors in urine, that are well known bladder
carcinogens.
III. Urinary stasis allowing concentration of endogenous carcinogens
leading to their absorption from urine and exposure of the bladder
epithelium.
IV. Raised urinary beta-glucuronidase levels originating from miracidia
and adult schistosomes liberating carcinogenic amines in urine.
S. japo nicum Colorectal &
Hepatocellular
carcinoma
(HCC)
I. Soluble egg antigen (SEA) from S. japonicum, which has a strong
immunogenic activity, may contribute to carcinogenesis through
stimulation of chronic inflammation.
II. Somatic mutations in the p53 gene.
III. Higher frequency of arginine missense mutations.
IV. S. japonicum-derived products may be involved in induction of host
genomic instability.
S. mansoni Adenocarcino
ma &
Hepatocellular
carcinoma
(HCC)
I. Cell mediated response is depressed in active intestinal
schistosomiasis and this immunosuppression increases with
advancement of the disease and development of
hepatosplenomegaly.
II. Anti-idiotype antibodies produced in patients with chronic
schistosomiasis can downregulate specific immune responses.
III. Altered expression of the tumor protein 53 (TP53).

19. Laboratory Diagnosis
Parasite Microscopy Histopathology & Serology
S.
haemato
bium
Urine Microscopy:
Detection of non-operculated terminal
spined eggs in the urine or rarely in feces.
Collection time : 10 am to 2 pm as max.
no. of eggs excreted.
Concentrated by centrifugation or by
membrane filtration and observed under
microscope for the presence of non-
operculated terminal spined eggs.
24 hr collection done to quantitate the
severity of infection.
>50 eggs /10 ml urine: Heavy Infection
<50 eggs /10 ml urine: LightInfection
Egg viability test (Egg Hatching ) : to
assess the effectiveness of Tx. Performed
by mixing the urine with distilled water &
observing for hatching miracidia.
Active Infection : viable eggs will be hatched
out of miracidia
Treated or past Infection : Non-viable eggs.
Demonstration of S. haematobium eggs in
bladder mucosal biopsy confirms the diagnosis.
Ab detection : 2 assays are available to detect serum
antibodies against S. haematobium adult worm microsomal
antigen (HAMA).
HAMA-FAST-ELISA [Falcon assay screening test ELISA] is a
new test with high sensitivity (95%)and specificity (99%).
HAMA-EITB (enzymelinked immuno transfer blot)
Others : Cercarial Huller reaction, indirect fluorescent antibody
test (IFA) and indirect hemagglutination (IHA) test.
Antigen detection : For monitoring Tx response.
Circulating cathodic antigen (CCA) and circulating anodic
antigen (CAA) can be detected in serum and urine by ELISA or
dip stick assays. CCA levels are much higher in urine than
CAA ELISA based assay using specific monoclonal antibodies
against soluble egg antigen (M AbSEA) shows sensitivity of
90% (serum) and 94 % (urine).
S.
mansoni
S.japoni
cum
Stool Microscopy: In acute cases, eggs
with lateral spine can be demonstrated in
stool or rarely in urine
Hatching test: This involves hatching of
motile miracidia when the eggs are diluted
in water and perpendicular beam of light is
passed through the water at the top
Histopathological : Demonstration of lateral spined eggs in
biopsy material from rectal mucosa confirms the diagnosis of
schistosomiasis.
Egg shell of S. mansoni is acid fast and can be stained by
modified Z-N stain.
Ab detection : Similar to S. haematobium, ELISA and EITB
formats are available to detect serum antibodies.
Ag detection : Similar to S. haematobium, various antigens
can be detected like CCA in urine and CAA in serum and
soluble egg antigen (SEA) in serum.

20. LIVER FLUKES
• Flatworms inhabiting the human liver :
 Clonorchis sinensis
 Opisthorchis viverrini,
 Opisthorchis felineus
• TheIARC classifiesC. sinensisasaGroup 1 agent (carcinogenic to humans).
• Caused by fish-borneliver flukesof thetrematodefamily Opisthorchiidae.
• >45 million peopleworldwideareinfected by thesepathogens.
• O. felineusisendemic in partsof Europeand Russia; C. sinensisin China, the
Republic of Korea, and northern Vietnam; whileO. viverrini-infectionsoccur in
Southeast Asia.
• Theconsumption of rawfish infested with infectiousmetacercariaeand intensifying
transmission of theparasitesto humansfrom domestic O. viverrini infected animals
contributeslargely to increased incidences.

21. Life Cycle : Clonorchis sinensis
1. Embryonated eggs are discharged
in the biliary ducts and in the stool.
2. Eggs are ingested by a suitable snail
intermediate host.
2a. Each egg releases a miracidia,
2b. which go through several
developmental stages (sporocysts),
2c. Rediae
2d. and cercariae
3. The cercariae are released from the
snail and after a short period of free-
swimming time in water, they come in
contact and penetrate the flesh of
freshwater fish, where they encyst as
metacercariae
4. Infection of humans occurs by
ingestion of undercooked, salted,
pickled, or smoked freshwater fish .
5. and ascend the biliary tract through
the ampulla of Vater
6. Maturation takes approximately 1
month. The adult flukes (measuring 10
to 25 mm by 3 to 5 mm) reside in small
and medium sized biliary ducts. In
addition to humans, carnivorous
animals can serve as reservoir hosts.

22. Clonorchis sinensis (schematic diagram) (A) adult worm; (B) egg; (C) cercaria
larva
A B C
Clonorchis sinensis real image
(A)adult worm (carmine stained);
(B) egg (saline mount)
A
B

23. Life Cycle : Opisthorchis 1.The adult flukes deposit fully developed
eggs that are passed in the feces.
2. After ingestion by a suitable snail (first
intermediate host).
2a. the eggs release miracidia
2b. which undergo in the snail several
developmental stages (sporocysts)
2c. Rediae
2d. Cercariae
3. Cercariae are released from the snail.
4. and penetrate freshwater fish (second
intermediate host), encysting as
metacercariae in the muscles or
under the scales.
5. The mammalian definitive host (cats,
dogs, and various fish-eating
mammals including humans) become
infected by ingesting undercooked
fish containing metacercariae. After
ingestion, the metacercariae excyst in
the duodenum.
6. and ascend through the ampulla of
Vater into the biliary ducts, where
they attach and develop into adults,
which lay eggs after 3 to 4 weeks .
The adult flukes (O. viverrini: 5 mm to 10 mm by 1 mm to 2 mm; O. felineus: 7 mm to 12 mm by 2 mm to 3
mm) reside in the biliary and pancreatic ducts of the mammalian host, where they attach to the mucosa.

24. Opisthorchis viverrini schematic diagram (A) adult worm; (B) egg; (C) cercaria
A
B
C
Opisthorchis viverrini
(A) adult worm (carmine stained);
(B) egg (saline mount)
A
B

25. Pathogenicity & Clinical Manisfestations
Clonorchis sinensis O. viverrini /O. felineus
The migrating larvae and adult worms ,both live and dead ,
produce pathologic lesions. Eggs cause little or no
inflammation in the host tissues.
Pathological changes:
In Liver : Hepatomegaly, mild jaundice, eosinophilia. Irregularly
dilated bile ducts with layer of fibrous tissue and parasite.
In Bile duct : Marked epithelial hyperplasia of bile duct.
Live worms cause adenomatous hyperplasia, periductal
inflammation, periductal fibrosis, increased production of
mucin.
Circulating antibodies produced against parasite metabolites
do not confer any protection against the fluke in the host.
IP : 6-8 weeks . Heavy infection (500-1000 flukes) produces
overt clinical infection.
Acute Clonorchiasis :
serum sickness-like illness with high fever, eosinophilia,
cirrhosis, odema and catarhal cholecystitis.
Chronic Clonorchiasis :
Recurrent pyogenic cholangitis : most frequent complication
of long standing infection. Caused by partial obstruction of the
bile duct , as flukes grow, mature and by secondary bacterial
infections. It is an acute condition that is seen within 1-3 hrs of
heavy meal , perhaps caused by excessive secretion of
pancreatic juice in to partially obstructed bile duct.
Cholangiocarcinoma: a cancerous condition due to chronic
irritation of the bile duct for long duration. Elderly people (60-80
yrs) and pre-existing primary sclerosing cholangitis are risk
factors.
Pathogenicity is related to the multiplication
of adult flukes in the hepatobiliary system that
leads to chronic mechanical obstruction
and inflammation of the bile duct.
Cholangiocarcinoma: occurs due to long
standing Opisthorchis infection. In area
endemic for the disease.
Main causes :
Obstruction and irritation of the bile duct by
the adult flukes.
 Chronic inflammatory changes in the bile
duct leading to ductal epithelial cell dysplasia.
Liver Cancer: Few reports of hepatocellular
carcinoma are also reported mainly from the
Northeastern Thailand.
Other Hepatobiliary Manifestations:
Hepatomegaly
Cirrhosis
Cholecystitis
Cholangitis
Obstructive jaundice
Secondary bacterial infection.

26. Mechanisms of Carcinogenisis
Parasites Mechanisms
C.
Sinensis
O.
viverrini
O.
Felineus
Chronic irritation and chronic inflammation caused by the fluke results in hyperplasia and
adenomatous changes of bile duct epithelium.
These hyperplastic cells are vulnerable to carcinogen because the agent can easily
induce DNA damage during active cell proliferation
 Endogenous nitrosation caused by liver fluke infestation has been studied in both
humans and animals.
It is likely that N-nitroso compounds are formed in the area of chronic inflammation
around the bile ducts as the result of local generation of nitric oxide by inflammatory cells.
Therefore, bile duct epithelial cells are exposed continuously to high concentrations of
nitroso compounds leading to neoplastic transformation
Activation of drug metabolizing enzymes: In male hamsters infected by O. viverrini ,
activities of hepatic cytochrome p-450 isoenzyme have been shown to be higher than those
of controls especially in hepatocytes in the area of inflammation.
N-nitrosodimethylamine, one of the products of endogenous nitrosation formed in the
tissue, is significantly metabolized by cytochrome P-450.
The product of this metabolism is a DNA methylating agent that can result in DNA
damage, particularly in proliferating bile duct epithelial cells.
There was a significant reduction in the levels of these enzymes after eradication of
flukes by praziquintal treatment.

27. The chronic inflammation during Clonorchis, Opisthorchis and Schistosoma infections leads to the activation of signaling
pathways including p53, NF-κB, Jak/Stat and Rb that could generate somatic mutations and/or activate oncogenes.
Fluke-derived products and metabolites secreted to the host microenvironment may induce metabolic processes
including oxidative stress that facilitate damage to the chromosomal DNA of proximal epithelial cells, specially
cholangiocytes and urothelial cells for the liver and blood flukes, respectively.
Physical damage of host tissues during the development of parasites together with the active wound healing process lead
to increased cell transformation and proliferation, which also are associated with the DNA damage.
Combined parasite-host interaction events (chronic inflammation, parasite-derived products, and physical damage) and
their combined effects on the chromosomes and fates of cells lead to the modification of the cell growth, proliferation and
survival that in turn initiate and promote malignancy.

28. Laboratory Diagnosis
Parasite Microscopy Serodiagnosis & Molecular Methods
Clonorchis
sinensis
Stool Microscopy:
Demonstration of the characteristic flask
shaped eggs in the stool establishes the
diagnosis.
Microscopy of the duodenal aspirate is
more sensitive than stool microscopy.
Formalin ether concentration should be
done when egg burden is low.
 However, the eggs of C. sinensis are
morphologically similar to that of
Opisthorchis, Heterophyes, and
Metagonimus.
Serology:
Various serological tests like CFT, indirect
hemagglutination test and ELISA are used to
detect the antibodies in the serum.
 A double sandwich ELISA for circulating antigen
in the serum is also available. Detection of
antigen is more useful as it indicates current
/recent infection.
Molecular : A multiplex PCR has been developed
to detect Clonorchis and Opisthorchis
simultaneously. It is rapid with high sensitivity and
specificity.
Skin Test : Immediate hypersensitivity test using
soluble extract form adult C.sinesis.
Used for epidemiological tool in China and Korea.
Opisthorchis
viverrini
Stool Microscopy:
Multiple stool examination can be carried
out to detect the characteristic fl ask
shaped eggs. Sedimentation techniques
are followed for stool concentration. Th e
eggs are similar to that of C. sinensis,
Heterophyes and Metagonimus; hence
reliable diagnosis depends on recovery of
the adult worm, patient history and
geographical area.
DNA hybridization and detection of antigen have
been used to detect the parasite in the stool.

29. Malaria & Burkitt Lymphoma
• Burkitt lymphomaisamonoclonal B cellscancer and thefastest
growing tumor in humansin malariaendemic areasof sub-Saharan
Africa.
• Burkitt lymphomaisclassified into theclinical typesof endemic,
sporadic and immunodeficiency-associated Burkitt lymphoma
• Thechromosometranslocation between thec-Myc oncogeneand
immunoglobulin (Ig) geneloci that leadsto deregulation of c-Myc
expression together with p53 genemutationsareknown to bemost
relevant in thepathogenesisof Burkitt lymphoma.
• Associated Risk factors:
 EBV infection
 HIV/AIDS
 P. falciparum

30. Parasite Mechanism
Malaria
(P.
falciparum )
Expansion of
EBV-Infected B
Cells
P. falciparum-infected erythrocytes directly adhere to and activate B cells through the
CIDR1α domain of P. falciparum erythrocyte membrane protein 1 (PfEMP1).
Interaction of PfEMP1-CIDR1α induces proliferation of B cells, expression of distinct
activation molecules, and differentiation into plasma cells, thereby increasing the
secretion of IgM immunoglobulins and cytokines .
Increasing proliferation of polyclonal B cell populations might enhance the risk of
expansion and transition of EBV-infected B cells, which could lead to the emergence of
a malignant B-cell clone.
Suppression of
EBV-Specific T
Cell Immunity
Failure of EBV-specific T cells to control EBV-infected cells in malaria patients leads
to the expansion and abnormal proliferation of EBV-infected B cells.
Dendritic cells could contribute to inhibition of T cell immunity during malaria, as P.
falciparum-infected erythrocytes are able to adhere to dendritic cells and modulate their
functions through a TLR9-dependent pathway.
These interactions inhibit maturation of dendritic cells (DCs) and their capacity to
activate immune responses and alter the IL-12 and IL-10 secretion patterns.
Reactivation of
EBV Viremia
Induced by
Malaria
The expansion of EBV-infected B cells is associated with higher levels of B cell-
carried EBV-DNA and plasma cell-free EBV-DNA.
Binding between latently EBV-infected B cells and the domain CIDR1α of the PfEMP1
protein directly switches the virus into lytic replication and CIDR1α stimulates EBV
production in peripheral blood mononuclear cells.
cell-free EBV-DNA levels in plasma of children and pregnant women with malaria
were increased compared to those without malaria, showing that EBV can be
reactivated during malaria infection.
AID-Dependent
Genomic
Translocation
Induced by
Plasmodium
falciparum
Malaria is not a direct trigger of cancer, but P. falciparum infection rather modifies the
lymphoma phenotype to favor more mature B cell lymphomas by stimulating prolonged
AID (activation induced cytidine deaminase) expression in germinal center B cells.

31. Plasmodium falciparum infected red blood cells (iRBC) bind to the Epstein-Barr virus (EBV) latently infected B cells through
the CIDR1α domain of P. falciparum erythrocyte membrane protein 1 (PfEMP1) that lead to the expansion of the latently
infected B cell pool and/or lead to the reactivation of EBV.The interaction between iRBCs and EBV-infected B cells in the
germinal center (GC) also results in the increased expression of the Activation- Induced cytidine Deaminase (AID). The
AID in turn contributes to break host DNA at the immunoglobulin (Ig) or/and highly transcribed regions, to activate
oncogenes (c-Myc) and to induce somatic mutations. The AID also induces the chromosomal rearrangement especially
the translocation between Ig regions and c-Myc oncogene.
All these processes lead to the genomic instability that can drive the proliferation and differentiation of B cells in GC and
subsequently lead to the emergence of a malignant B-cell clone. In addition, the binding of iRBC to the dendritic cells (DCs)
could lead to a modification of DC functions that contributes to suppress EBV-specific T-cell immunity (CD8+ and CD4+ T
cells), therefore resulting in the loss of controlling the expansion of EBV-infected B cells including emergent Burkitt
lymphoma clones.

32. Strongyloides stercoralis and Cancer
• Strongyloidesstercoralis, an intestinal nematode, can cause
strongyloidiasisand gastrointestinal ulcer. S. stercoralisinfects
approximately 50–100 million peoplein tropical and subtropical
regions.
• Infection with S. stercoralismay becomplicated by autoinfection,
which resultsin ahyperinfection syndromein part geographically
associated with theoccurrenceof HTLV-1 infections.
• What is HTLV-1 ??
• HTLV-1 causesadult T cell leukaemia/ lymphomaby enhancing
immortalisation and transformation of T cells.
• Classified asaGroup 1 carcinogen by theIARC.
• TheHTLV-1 proteinsTax and HBz areinvolved in many regulatory
processesincluding induction of growth of infected T cellsand
transformation, transcription of cellular genes, and genetic instability.

33. Life Cycle- Strongyloides stercoralis
1.The rhabditiform larvae passed in the
stool
2. can either become infective filariform
larvae or free living adult males and
females
3. that mate and produce eggs
4. from which rhabditiform larvae hatch
5. and eventually become infective
filariform larvae
6. The filariform larvae penetrate the
human host skin to initiate the parasitic
cycle
7. Parasitic cycle: Filariform larvae in
contaminated soil penetrate the human
skin and by various, often random routes,
migrate into the small intestine
8. L3 larvae migrate via the bloodstream
to the lungs, where they are eventually
coughed up and swallowed or L3 larvae
can migrate directly to the intestine via
connective tissues. In the small intestine
they molt twice and become adult female
worms
9. The females live threaded in the
epithelium of the small intestine and by
parthenogenesis produce eggs , pass in
stool or cause
10. Autoinfection : rhabditiform larvae
become infective filariform larvae, which
can penetrate either the intestinal
mucosa (internal autoinfection) or the
skin of the perianal area (external
autoinfection); in either case, the
filariform larvae may disseminate
throughout the body.

34. Strongyloides stercoralis and Cancer
Parasite Mechanism
Strongyloides
stercoralis
 Induce polyclonal expansion of HTLV-1-infected T cells by activation of
the IL-2/IL-2R system suggesting that S. stercoralis is a cofactor for the
development of HTLV-1- induced lymphoid cancers.
 HTLV-1 proviral loads were significantly higher in HTLV-1 carriers with
strongyloidiasis than in HTLV-1 positive individuals without S. stercoralis
infection suggesting that S. stercoralis may stimulate HTLV-1 replication
 A case report described a Korean patient presenting with both S.
stercoralis infection and early gastric adenocarcinoma. Further analysis
revealed that the gastric adenocarcinoma and adenoma tissues were
positive for S. stercoralis suggesting a causative effect of S. Stercoralis.
Also stimulates induction of colon adenocarcinoma
Probably by interacting with the host and/or activating the
Host immune response.

35. Strongyloides stercoralis (A) adult male
(arrow shows spicules); (B) adult female
(containing single row of eggs); (C) filariform
larva; (D) rhabditiform larva
A B
C
D
Lab Diagnosis
Microscopy
The rhabditiform larvae can be demonstrated in stool by direct
microscopy or follo wing concentration techniques. Sometime, the
hookworm eggs may hatch in the stool releasing the rhabditiform
larva which has to be differentiated from that ofS. stercoralis .
Single stool examination is less sensitive (30%) due to irregular
and low output of larvae. Hence repeated stool examination (four
consecutive samples) is required.
Entero-test: Sometime duodenal aspirate can be collected by
entero test and examined for the presence of larva .
Disseminated strongyloidiasis can be readily diagnosed by
examining stool, sputum, other body fluids, and tissues, which
typically contain high numbers of filariform larvae.
Stool Culture
Freshly passed stool samples should be cultured. L3 stage
filariform larvae are formed within 2 days which should be
differentiated from that of hookworm .
Various culture techniques can be used. They are:
Harada Mori filter paper tube method
Petridish (slant culture) technique
Baermann funnel technique
Charcoal culture method
Agar plate technique (more sensitive).
Serology
(ELISA) using crude larval antigens has a greater sensitivity (95%)
and should be used when microscopic examinations are negative.
However, it is less specific because of cross reactivity with other
helminthic infection. More so, antibody detection cannot
differentiate recent and past infection.

36. Paradoxical Dual Impacts of Chagas Disease in
Carcinogenesis
Chagasdisease(CD), aparasitic diseasecaused by theflagellated
protozoan Trypanosomacruzi, occursthroughout South and Central
America, and affectsapproximately 15 million people.
Successful transmission of T. cruzi primarily occursthrough
triatomineinsects(kissing bugs).
Peoplebecomeinfected when fecesof thekissing bug containing
thetrypomastigotestageof T. cruzi aredeposited on thehuman
skin whiletheinsect feedson blood; theT. cruzi containing insect
fecescontaminatemucousmembranes, conjunctivae, or skin
breaks, and initiatehuman infection.

37. Life Cycle - Trypanosoma cruzi1. An infected triatomine insect vector (or
"kissing" bug) 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
2. Inside the host, the trypomastigotes
invade cells near the site of inoculation,
where they differentiate into intracellular
amastigotes
3. The amastigotes multiply by binary fission
4. and differentiate into trypomastigotes,
and then are released into the circulation
as bloodstream trypomastigotes .
5. Trypomastigotes infect cells from a
variety of tissues and transform into
intracellular amastigotes in new infection
sites. 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
6. The ingested trypomastigotes transform
into epimastigotes in the vector’s midgut
7. The parasites multiply and differentiate in
the midgut
8. differentiate into infective metacyclic
trypomastigotes in the hindgut

38. Trypanosoma cruzi and its Dual Impact
Parasite Disease Mechanisms
Trypano
soma
cruzi
Esophageal
leiomyosarcoma
Chagasic
megacolon
Colon cancer
An increase of gastroesophageal reflux into the megaesophagus
Aneuploidies of chromosomes 7, 11, and 17 in 60% and the deletion of
the oncogene p53
Point mutations in exonic regions of p53, FHIT (fragile histidine triad
gene) and CDKN2A (cyclin-dependent kinase Inhibitor 2A) genes or
genomic imbalances
a silent mutation in exon7 of the FHIT gene and copy numbers of the
CDKN2A and CEP9 (C-terminally encoded peptide 9)
Anticancer
Activity
Surface cellular antigens and an inhibiting or lysing factor of T. cruzi
contribute to anticancer activities.
Immunization with T. Cruzi epimastigote lysate strongly inhibited tumor
development in vivo by inducing the activation of both CD4(+) and CD8(+)
T cells as well as by increasing numbers of CD11b/c(+) His48(−) MHC II(+)
cells, which correspond to macrophages and/or dendritic cells.
Antibodies against T. cruzi lysate recognized human tumor cell types
such as colon and human breast cancer cells and thusmediate tumor cell
killing through antibody-dependent cellular cytotoxicity (ADCC).
A parasite chaperone molecule, the T. cruzi calreticulin (TcCRT) directly
interact with human endothelial cells through a receptor-dependent
mechanism and to inhibit their proliferation, migration and capillary
morphogenesis.
TcCRT plays a central role during the host-parasite interplay by
interactingwith complement proteins such as complement factor
(C1),mannose-binding lectin (MBL), and ficolins to inhibit activation of the
complement system that leads to increased infectivity of the parasite and
thus inhibits growth and metastasis of tumors.

39. T. cruzi has antitumor effects by inducing host immunity against tumor. T. cruzi expresses a calreticulin (T. cruzi
calreticulin, TcCRT) that can directly interact with endothelial cells and inhibit their proliferation, migration and
capillary morphogenesis as well as inhibit tumor cell growth. The DNA mismatch repair protein (TcMSH2), a
central component of the mismatch repair (MMR) machinery in T. cruzi, allows T. cruzi to respond effectively to
the oxidative stress during infection. The oxidative stress mediated by alkylating agents and hydrogen peroxide
leads to carcinogenesis by damaging DNA. The TcMSH2 protein of T. cruzi may also contribute to protect host
chromosomes from oxidative stress during infection, and therefore consequently inhibit tumorigenicity.
On the other hand, T. cruzi may also cause cancer by inducing somatic mutation and genomic imbalance
during chronic inflammation. However, the molecular details of this latter phenomenon are yet not understood

40. Trypanosoma cruzi (A) trypomastigote form (thin blood smear stained with Giemsa) (B)amastigote forms in
heart tissue stained by hematoxylin and eosin (C) indirect fluorescent antibody test showing trypomastigote
forms
A B C
Lab Diagnosis of T.Cruzi
Peripheral blood Microscopy:
In acute Chagas’ disease, the trypomastigote are frequently found in peripheral blood
which can be detected by:
Wet mount preparation of anticoagulated blood or buffy coat can be done to see the
rapid movements of trypomastigotes.
Thick and thin smear: (Stained by Giemsa stain) thick smear is more sensitive in
detecting the parasite whereas the thin smear helps in differentiating T. cruzi with
morphologically similar looking T. rangeli.
Blood concentration techniques like microhematocrit method and Strout method of buffy
coat preparation may be employed if the parasite count is low.
Amastigotes can be demonstrated in heart tissue obtained at autopsy stained by
histopathological stain.

41. Lab Diagnosis of T.Cruzi
Culture :
Blood is inoculated in NNN medium or Yager’s liver infusion tryptose medium, incubated at 25°C and observed
for the epimastigote forms for up to 30 days before they are considered negative.
Culture is more sensitive than smear microscopy.
Ab detection :
Chronic Chagas’ disease is diagnosed by the detection of specifi c IgG antibodies against T.cruzi antigens. IgM
antibodies are diagnostic for congenital infection.
Several methods are employed like CFT (Guerreiro Machado test), ELISA, IFA, IHA (indirect hemagglutination
test) and Western blot.
Radioimmunoprecipitation assay (Chagas’ RIPA) is a highly sensitive and specific (confirmatory) method.
Ag detection : T. cruzi specifi c antigens from serum and urine of the infected patients are detected which are
very useful for diagnosing acute infection and congenital transmission. Recently, a chemiluminescence
immunoassay (CLIA) has been developed for blood bank screening and for the monitoring the response to
treatment.
Molecular Methods : PCR is available that detects T. cruzi specific kinetoplast or nuclear DNA in blood. It is
more sensitive than microscopy and serology for the diagnosis of chronic disease. It can detect as low as one
trypomastigote per 20 mL of blood. It is also useful in monitoring the response to treatment and for the diagnosis
of congenital infection.
Animal Inoculation : Blood or CSF of the patients is inoculated intraperitoneally into mice. Trypomastigotes
can be demonstrated from the blood of mice within 10 days of inoculation.
Xenodiagnosis:
The infected patients are exposed to 20 numbers of laboratory maintained nymphs of reduviid bugs daily for 3
days and the dropping of the insects are examined monthly for 3 months for the presence of the epimastigote
forms. This is more sensitive to detect light chronic infection.

42. A few other associations…
Parasite Disease Mechanisms
Trichomonas
vaginalis
Cervical Invasive Neoplasia
(SCC Gr I & II)
Cytopathogenicity mainly due to contact-
dependent mechanism of T. Vaginalis.
The presence of a cell-free product of t.
Vaginalis, cell-detaching factor, involved in
the cytopathic effects and a very low acidic
pH acidic metabolites produced by T.
vaginalis.
Taenia solium
Glioblastoma Multiformae
Cerebral Glioma
Malignant Haematological
Disorder
a) Chronic inflammation leading to release of
nitric oxide in brain which is a potential
carcinogen.
b) The parasite induced modulation of host
immune response leading to inhibition of
tumour suppressor surveillance
mechanisms.
c) The transfer of genetic material from
parasite to host causing DNA damage thus
predisposing to carcinogenesis.

43. A few other associations…
Parasite Disease Mechanisms
Toxoplasma
gondii
Pituitary Adenoma
Ocular Tumours
Meningioma,
Leukemia
Overstimulation of pituitary gland to fight the parasitic
infection, may lead to adenoma formation
Cryptosporidium
parvum
GIT neoplasia
Colonic Adeno Ca
Pancreatic Ca
Colorectal cancer
a. Modifications of the host actin cytoskeleton of intestinal
epithelial cells.
b. A chronic infection and inflammation by this parasite is
thought to be the cause of the malignant transformation.
c. It can modulate host-cell apoptosis. BCL2 and c-Myc
genes shows an altered expression. Apoptosis
prevention probably benefits the parasite by stabilizing
the host cell long enough to permit the completion of the
life cycle and resistance to apoptosis could be an
essential step in the progression to malignancy
d. Higher rates of oocyst shedding seemed to be associated
with an earlier onset and more rapid evolution of
neoplastic lesions.

44. A few other associations…
Parasite Disease Mechanisms
Theileria parva
Theileria annulata
Lymphoproliferative Disease
(Transformation of the
leukocyte infected cells &
uncontrolled proliferation
allowing clonal expansion
It resides freely in the host leukocyte in direct
contact with host-cell cytoplasmic structures,
and
modify host-cell cytoskeleton & alters host-cell
actin dynamics, increases motility and enables
infected host-cell to behave as leukocyte
metastasis by :
a.anti-apoptosis signaling pathway is stimulated
by the activation of the transcription factor NF-
κB.
b.cytokine GM-CSF secretion is enhanced and
itself re-stimulate infected host cell proliferation
via autocrine loops.
c.GM-CSF contributes to the induction of the
factor c-Myc, leading to lymphocyte proliferation

46. References…
• Textbook of Medical Parasitlogy, S. C Parija
• Essentials of Medical Parasitology, Apurba S Sastry,
Jaypee Publications, 2014
• https://www.cdc.gov CDC, Atlanta
• Khurana S, Dubey M L, Malla N. Association of Parasitic
Infections and Cancers. Indian J Med Microbiol 2005;23:74-
9
• Van Tong H, Brindley PJ, Meyer CG, Velavan TP. Parasite
Infection, Carcinogenesis and Human
Malignancy. EBio Me dicine . 2017;15:12-23
• Gilbert RO, Elia G, Beach DH, Klaessig S, Singh BN.
Cytopathogenic Effect of Tricho m o nas vag inalis on Human
Vaginal Epithelial Cells Cultured In Vitro. Petri WA,
ed. Infe ctio n and Im m unity. 2000;68(7):4200-4206.
• Benamrouz S, Conseil V, Creusy C, Calderon E, Dei-Cas E,

47. Thank You….