medical parasitoloty: Kyle's stuff for test 2

Kinetoplastids

a DNA-containing domain located within the single mitochondrion and associated w/ the flagellar base

possessed by flagellated protozoa

Kinetoplastids are transmitted by

blood-sucking antropods

kinetoplastids live in the

circulatory system and tissue fluids; some invade cells

two medically important kinetoplastidas

Leishmania spp

Trypanosomes

• T. brucei

  • T. cruzi

life cycle stages of kinetoplastida

trypomastigote

epimastigote

promastigote

amastigote

Leishmania is transmitted by ___ ___ + facts about them

sand flies (Phlebotomidae)

are minute diptera

more closely related to mosquitoes

only mosquito nets w/ fin meshwork hold them off

often breed in rodent burrows

can also transmit Bartonella bacilliformis and Pappataci virus

3 complexes of Leishmaniasis

Visceral: Leishmania donovani, infantum, chagasi

Cutaneous: Leishmania tropica, major, aethipica

Mucocutaneous: Leishmania brazieliensis, mexicana, amazonensis

Old World Leishmaniasis

found in Africa, Asia, Middle East, the Mediterranean, and India

close contact of humans and domestic animals = optimal conditions for sand flies and leishmania transmission

urban environment: human to human

rural areas: Leishmaniasis = zoonosis

infection in dogs frequent in the Mediterranean

New World Leishmaniasis

From Parasites in Central and South America

infected while working or hunting in forest

wild animals including rodents, monkeys, and sloths = reservoir for parasite

sylvatic

transmission pattern within population of wild animals that result in occasional infection of humans

Leishmania parasites are ___ and ___

promastigotes and amastigotes

parasite lives in ____ ____ of sandflies as ___ ___.

in mammalian host parasites multiply as ___ ___

digestive tract; extracellular promastigote;

intracellular amastigotes

Promastigote vs amastigote pics

Leishmania infects and thrives in

macrophages

within mammalian host leishmania infects macrophages and replicates as intracellular

amastigote

macrophages are ‘professonial’

phagocytic cells

parasite invades host cell passively by triggering phagocytosis

phagocytosed pathogens are targeted to the ___ and destroyed

lysosome

phagosome usually fuses w/ primary lysosomes to form secondary lysosome

in secondary lysosomes, pathogens are killed and digested by the action of lytic enzymes in an acidic environment produced by membrane proton pumps

generation of oxygen radicals also induce killing

Leishmania enter macrophage by

phagocytosis

several intracellular pathogens have found waves to escape killing such as

Trypansoma cruzi -- induces phagocytosis but then escapes into the cytoplasm

Toxoplasma -- active invasion, parasitophorous vacuole is never part of the endocytic pathway

Mycobacterium tuberculosis -- induce phagocytosis and block lysosomal maturation

How do leishmania survive?

don’t seem to care

Amastigotes thrive in what looks like fully matured lysosome w/ acidic pH and abundant lysosomal hydrolases

Amastigotes rapidly divide and will infect new macrophages after rupture of host cell

dense surface coat covering Leishmania seems to protect parasite from action of lytic enzymes

however, w/ help of T cells, macrophages can be stimulated to kill the parasite

promastigotes considered part of the ___ phase of Leishmania because

extracellular

have not yet developed intracellular amastigotes

after entering the human host during blood meal of female sandfly, Leishmania promastigotes must first evade

complement-mediated lysis in the bloodstream until engulfed by macrophage

binding of complement can

destroy pathogens but also tags them for phagocytosis

complement is a component of the

immune system meant to lyse pathogens

how do Leishmania stimulate the complement process

by binding elements of complement system to surface

after parasite inoculation into skin, ~90% of parasites are killed by the

comoplement system

parasites use __ __ to be ingested by Mo via complement receptor (CR3)

C3 opsonization

Parasite internalization is a ___process; later on, MΦ are capable of restimulating CD4+ T cells via MHC class __.

silent; II

____, in contrast, are activated upon infection, present Leishmania antigen in the MHC class I/II pathways and promote T cell priming.

Dendritic cells

a __ response is required for parasite control and healing

TH1

Stimulation with different cytokines leads to the development of two types of T-cells specialized for different immune responses

Th1 and Th2 strongly ____ each other

downregulate

non healing Leishmania infections are characterized by strong ___ respnse

TH2

healing infections are characterized by

Th1

parasite seems to manipulate this

balance of Th1 and Th2

3 syndromes associated w/ Leishmania infection in humans

Visceral, Cutaneous, and Mucocutaneous

Visceral Leishmaniasis or Kala Azar

darkening of skin on extremities and abdomen; common symptom of indian form

what is Kala azar caused by

L. donovani complex

systemic infection of reticulo-endothelial cells (mostly macrophages) in multiple internal organs and the blood

weeks to months incubation period

lead symptom of Kala Azar

Abdominal swelling due to hepato- and splenomegaly

high fever (oscillates w/ peak every second day)

progressive weight loss (cachexia)

mortality of untreated disease 75-95%

post kala-azar dermal leishmaniasis

sequel visceral leishmaniasis = may manifest years after successful treatment and resolution

dermal lesions may contain parasites in great numbers

Cutaneous Leishmaniasis (Old world oriental sore)

usually self-limiting

Old world oriental sore (Cutaneous) caused by

L. tropica complex (similar disease in new world caused by L. mexicana)

chronic but self-limiting dry ulceration at site of bite

ulceration starts months after infection

parasites NOT found outside of lesions

granuloma formed = healing leaving depressed scar

nearly absolute resistance to reinfection

inoculation to vaccination long been practiced

Espundia or Mucocutaneous Leishmaniasis

cause by L. braziliensis

~20% of infected patients develop ulcers of oral and nasal mucosa

progression of ulceration = slow but steady; ultimately destroying ALL soft parts of nose, lips, and soft palate

death can occur via secondary bacterial infection

Diagnosis

demonstration of parasites in blood or lymph (kala azar) or scrapings of ulcer (cutaneous versions)

Serological tests for visceral

treatment for visceral Leishmaniasis

India: amphotericin B in its various liposomal prep

East Africa: antimonials (sodium stibogluconate and paromomycin)

Miltefosine = first oral treatment for disease; better tolerated

• reduced hospitalization costs and outpatient distrubtion = option

  • disadvantages = evidence of reduced efficacy after decade of use and it is teratogenic

Cutaneous treatment

amphotericin B (liposome formulation = better tolerated but more expensive)

available drugs = significant side affects and resistance

__ co-infection further complicates treatment

HIV

paramomycin

new hope for better treatment

an old off-patent antibiotic which shows promising activity in studies by One World Health

Intravenous injection = significant cure in Kala-azar patients in India

developed drug for kala-azar together w/ Indian drug maker

infected macrophages taken up w/ blood meal and amastigotes released by digestion transform into

procyclic promastigotes; attach to midgut epithelium

Attached promastigotes divide rapidly

are procyclics infective to mammals?

no

metacyclic (infective) promastigotes

cease replication, detach, and pass forward into the pharynx and regurgitate into bite site

parasite interactions w/ sand fly midgut

promastigotes attach and detach to midgut endothelium

Attachment helps remain in insect gut when blood meal passed

later, need to detach to move to pharynx and proboscis for infection

how do they detach?

Lipophoshoglycans (LPGs)

LPGs role in Leishmania pathogenesis

LPG = dominant molecule on surface of Leishmania

not a protein but glycolipid

anchored in membrane by lipid to which long chain of hydrophilis sugar-phosphate repeats are attached

structure of LPG changes over life cycle to adapt

LPG = pathogenesis factor in mammalian host and important for life cycle within sandfly

LPG structural modifications during sand fly cycle

modified during developmental process of metacyclogenesis (stop replication and begin to pre-adjust to mammalian host)

LPG metacyclics has 2-3 times number of repeat units

side chains w/ terminal galactose = downregulated; remaining galactose residues = capped w/ another terminal sugar (arabinopyranose)

only LPG from ___ attaches to midgut

procyclics

phosphoglycans isolated from procyclics and metacyclics were labeled w/ a dye

opened sandfly midguts were incubated w/ PG from procyclics (A/B) and metacyclics (C/D) and detected w/ antibody

only procyclic phosphoglycan binds to midgut

PG binds to the ___ of sandfly midgut epithelium

microvilli

LPG binds to a species specific ___ in sandfly midgut

galectin

gene for abundantly expressed galactose binding protein or lectin (galectin) was indentified in sandfly sequencing project

specific antibody against the galectin reacts w/ midgut of sandfly species from which it was isolated

high resolution microscopy shows that protein (red) is found on luminal side of midgut

Modification of LPG modulate

interaction of parasite and midgut galectin

Which form of leishmaniasis is characterized by skin sores and lesions?

A) Visceral leishmaniasis B) Cutaneous leishmaniasis C) Mucocutaneous leishmaniasis D) Dermal leishmaniasis

B, C, D

What is the causative agent of visceral leishmaniasis?

A) Leishmania tropica B) Leishmania braziliensis C) Leishmania donovani D) Leishmania major

C

Which of the following is a common symptom of visceral leishmaniasis?

A) Fever B) Skin lesions C) Oral ulcers D) All of the above

A

Leishmaniasis is primarily found in which of the following regions?

A) North America B) Sub-Saharan Africa C) South Asia D) All of the above

D

True or False: Leishmaniasis can be transmitted through direct contact with infected individuals.

False

True or False: The treatment for leishmaniasis includes antimonial drugs.

False

True or False: Cutaneous leishmaniasis can sometimes resolve on its own without treatment.

True

True or False: The sandfly is most active during the daytime.

False

Describe the lifecycle of Leishmania and the role of its vector.

sandflies

What are the main methods used for diagnosing leishmaniasis?

demonstration of parasites in blood or lymph (kala azar) or in scrapings of ulcer (cutaneous)

surgical (visceral)

Discuss the public health strategies that can help prevent leishmaniasis transmission?

fine knotted nets

ammphotericin B (india)

antimonials (east afirca)

Miltefosine (oral)

Paramomycin

Which immune cell type is primarily responsible for controlling Leishmania infection?

a) B cells b) CD4+ T cells c) Neutrophils d) Eosinophils

B

Which of the following is a key mechanism by which Leishmania evades the host immune response?

a) Inducing apoptosis in macrophages b) Producing antibodies c) Inhibiting Th2 responses d) Upregulating MHC class II molecules

A, C, D

Leishmania exists in which morphological form(s)?

a) Epimastigote b) Amastigote c) Metacyclic Promastigote d) Trypomastigote

B, C

Protozoa

one-celled animal-like protists (Eukaryota)

have a true nucleus w/ chromosomes enclosed by a membranous envelope

ubiquitous throughout aqueous environments and the soil

some are parasitic and responsible or serious disease.

some protozoa form ___ for

cysts; protection against harsh conditions, to survive extreme temperatures or harmful chemicals or lack of food, water, or oxygen

cyst is usually important for transmission

Protozoan Flagellates

Giardia

Trichomonas

Trypansomes

Amebas

Entamoeba

NAegleria

Acanthamoeba

Balamuthia

Protozoan Apicomplexa

Toxoplasma

Plasmodium

Babesia

Theileria

Cryptosporidium

Cyclospora

Entamoeba, Giardia, Trichomonas: common characteristics

early-diverging eukaryotes

extracellular parasites (do not invade host cells)

adhere to plasma membrane of host cells

contact-dependent cytotoxicity

mechanical damage to host tissues

asexual reproduction and division within host by binary fission

facultative anaerobes: carbohydrate metabolism is anaerobic

LACK MITOCHONDRIA or have underdeveloped one

all are killed via treatment w/ METRONIDAZOLE (selective for anaerobic cells)

diseases w/ flagellates

Giardia Lamblia: Giardiasis (diarrhea/dysentery) (most frequent cause of non-bacterial diarrhea)

Entamoeba histolytica: Amebiasis (diarrhea/Liver Abscess)

Trichomonas vaginalis: Trichomoniasis (Vaginitis)

who first observed Giardia

van Leeuwenhoek when examining his own feces

who named Giardia

Vilem Lambi : (1859) observed G. intestinalis in stools of children w/ diarrhea

not confirmed as a pathogen until the late 1970s

Giardiasis: infection w/ Giardia duodenalis

G. intestinales & G. lamblia also used

can parasitize the intestinal tract of wide range of vertebrates including humans

‘Beaver fever’

trophozoite and cyst forms

disease prevalent in children at daycare

domestic dogs and certain wild animals serve as hosts

Transmission predominantly by ingestion of food or water contaminated w/ cysts

Life cycle of Giardiasis

only ONE host; direct life cycle

cysts ingested and in small intestine, excystation release trophozoites (each cyst = two trophozoites)

trophozoites multiple by longitudinal binary fission, remaining in lumen of proximal small bowel where they can be free or attached to mucosa by ventral sucking disk

encystation occurs as parasites transit toward colon

cyst is stage most commonly found in non-diarrheal feces

Giardiasis clinical symptoms

incubation period: 1-2 weeks; majority of ppl asymptomatic

diarrhea w/ loose, fool-smelling stools

flatulence

abdominal cramps and bloating

nausea, anorexia

malaise

weight loss

prolonged symptoms

malabsorption

Giardiasis pathogenesis

many show no evidence of disease

mechanism not well defined

colonizes upper small intestine; attachment to intestinal epithelial cells may cause inflammation. Lymphocytic infiltration

Malabsorption: interfere w/ absorption of nutrients like fats and some vitamins

villous blunting

no tissue invasion-high number of trophozoites present in duodenal crypts

Giardiasis diagnosis

identification of cysts or trophozoites in feces

detection of giardia antigens in feces (ELISA test)

molecular methods (PCR)

giardiasis treatment

Metronidazole: not FDA approved but used

tinidazole: not approved in US

Furazolidone: approved by FDA

paromomycin: used for pregnant women

Nitazoxanide: approved for giardiasis and cryptosporidiosis in children

Giardia transmission and epidemiology

resistant cyst responsible for transmission

cysts survive several months in cold water

reservoir-human

infectious dose: 10-25 cysts (feces, defective piping, flies, soiled hands)

—

worldwide distribution

common in areas w/ poor sanitation

seasonal peak during late summer

prevalence: 2-5% in indust. countries; 20-30% in developing

risk groups: travelers, campers, children in daycares and homosexual men

cell biology of Giardia: The Trophozoite

active metabolizing, motile form, lives in upper two-thirds of small intestine

12 - 15 um long, shaped like pear

2 nuclei: equal DNA content

• both transcriptionally active

• both divide at same time

4 pairs of flagella = 8 flagella

ventral adhesive disc

reproduction by binary fission-asexual rep

Encystation

as trophozoite travels to lower levels of SI on way to colon gets exposed to biliary components, which trigger the encystation process

giardia cyst

after cyst formation the parasite undergoes 1 round of nuclear division w/o cytokinesis = 4 nuclei located at anterior end

flagella and adhesive disk = lost; but axonemes (Ax) and median bodies (MB) persist

cysts are oval shaped, and measure 11-14 um in lenth and 6-10 um wide

have well-defined wall (CW) often set apart from cytoplasm of parasite

cysts passed w/ feces and can survive up to 3 months

Giardia Mitosome

organelle of mitochondria origin found in ‘amitochondrial’ eukaryotes

found in Entamoeba histolytica, giardia and some microsporidia

suggests that amitochondriates harbored mitochondria and are not lineages that evolved before acquisition of mitochondria

mitochondrial origin of mitosome supported by

presence of 2 membranes

localization of iron sulfur cluster assembly machinery in these organelles

targeting of proteins into mitosome by an N-terminal leader sequences

FeS cluster assembly activity identified in mitosome-rich cell fraction

Giardia metabolism

G. lambila: aerotolerant anaerobe

remnant mitochondria or mitosome

tricarboxylic acid and cytochrome system is absent

parasite uses glucose and stores glycogen

pyruvate ferredoxin oxidoreductase (uses a ferredoxin and converts pyruvate to acetyl CoA)

can produce H under strict anoxic conditions

plasma membrane of Giardia trophozoite is covered by

Variant Specific Protein (VSP) for antigenic switching

200-300 different VSP genes

coupled to encystation/excystation

• encystation: ‘old’ VSP expression

  • excystation: ‘new’ VSP on surface

what may explain common occurrence of repeated Giardia infections

differentiation-associated switching of antigens

evidence of VSP during encystation/excystation

Trichomonas vaginalis

cause of trichomoniasis

common cause of vaginitis

found in urogenital tract

extracellular: adheres to epithelium

trophozoite

transmission by direct contact: sexually transmitted

Life cycle of Trichomonas vaginalis

microaerophilic and resides on the epithelium of urogenital tract where it replicates by binary fission

does not survive well in external environment

transmitted among humans, only known host

cysts??

cysts form of Trichomonas vaginalis??

recent report of ‘cyst like structures’ (CLS)

formation includes:

• rapid induction under acidic conditios

• developed in stationary phase culture

• survive osmotic lysis of trophozoites

• survive treatment w/ detergent

  • chitin-containing outer covering