phylum: apicomplexa class: sporozoea sub-class: coccidia order: eucoccidia sub-order:...

Phylum: Apicomplexa Phylum: Apicomplexa Class: SporozoeaClass: Sporozoea Sub-class: CoccidiaSub-class: Coccidia Order: EucoccidiaOrder: Eucoccidia Sub-order: HaemosporinaSub-order: Haemosporina Family: Plasmodidae Family: Plasmodidae Genus: PlasmodiumGenus: Plasmodium Sub-genus: Plasmodium Laverania Vinckeia …....Sub-genus: Plasmodium Laverania Vinckeia …....

Species: vivax falciparum bergheiSpecies: vivax falciparum berghei malariaemalariae ovaleovale

Causal Agents:Causal Agents:

There are approximately 156 named species There are approximately 156 named species of of Plasmodium Plasmodium which infect various species which infect various species of vertebrates. of vertebrates. 

Four are known to infect humans:. Four are known to infect humans:. 

P. falciparumP. falciparum, , P. vivaxP. vivax , , P. ovaleP. ovale  and   and P. P. malariaemalariae. .

P. vivaxmost widespread

P. falciparumprimarily tropics and subtropics

P. malariaesimilar range as P. falciparum

P. ovale

Distribution of Malarial Parasites

Malaria typesMalaria types

Malignant Tertian MalariaMalignant Tertian Malaria

Benign Tertian MalariaBenign Tertian Malaria

Benign Benign QuartanQuartanMalariaMalaria

Benign Tertian MalariaBenign Tertian Malaria

Anopheles mosqouiteAnopheles mosqouite

Vectores in IranVectores in Iran

An. StephenciAn. Stephenci ایران در ناقل پایدارترین و ایران مهمترین در ناقل پایدارترین و مهمترین

An. SuperpictusAn. Superpictus ایران در آنوفل ایران فراگیرترین در آنوفل فراگیرترین

An. DethaliAn. Dethali ایران شرق جنوب در آنوفل ایران کوچکترین شرق جنوب در آنوفل کوچکترین

An. CulicifaciesAn. Culicifacies

An. FluviatilisAn. Fluviatilis فالسیپاروم ناقل فالسیپاروم خطرناکترین ناقل خطرناکترین

An. MaculipennisAn. Maculipennis غرب شمال و درشمال اصلی غرب ناقل شمال و درشمال اصلی ناقل کشورکشور

An. Sacharovi An. Sacharovi کشور غرب شمال و درشمال اصلی کشور ناقل غرب شمال و درشمال اصلی ناقل

Life Cycle:

Life CycleLife Cycle

Geographic DistributionGeographic Distribution

ایران در ماالریا ایران شیوع در ماالریا شیوع

سال سال در کشور 1537815378: : 13811381در در مثبت کشور مورد در مثبت مورد

جمعیت 24/0 24/0 نفر هزار جمعیت در نفر هزار API:API:در

پالسمودیوم 68/1468/14 به مربوط آن پالسمودیوم درصد به مربوط آن درصدفالسیپارومفالسیپاروم

سال سال در منطقه 1161411614: : 13801380در در بیماری منطقه مورد در بیماری مورداست شده گزارش کشور شرقی است جنوب شده گزارش کشور شرقی ..جنوب

•natural (sporozoites/Anopheles)•blood transfusions

• shorter incubation period• fatality risk (P. falciparum)• no relapses possible (vivax/ovale)

•syringe sharing•congenital

• relatively rare although placenta is heavily infected

Malaria Transmission

Pf Pv/Po PmPrepatent Period 6-9 d 8-12d 15-18dIncubation Period 6-25 d 8-27d 16d-8w

Clinical featuresClinical features

History of exposure:(mostly: past travel or residence in disease-endemic areas). (mostly: past travel or residence in disease-endemic areas).  Since untreated malaria can progress to severe forms that may be Since untreated malaria can progress to severe forms that may be

rapidly (<24 hours) fatal, malaria should always be considered in rapidly (<24 hours) fatal, malaria should always be considered in patients who have apatients who have a

ParoxsymParoxsym::

1-Chills ( Cold stage)1-Chills ( Cold stage) 2-Fever ( Hot stage)2-Fever ( Hot stage)3- Sweating stage3- Sweating stage

Other clinical features:

splenomegaly, anemia, thrombocytopenia, hypoglycemia, pulmonary splenomegaly, anemia, thrombocytopenia, hypoglycemia, pulmonary or renal dysfunction, and neurologic changes.or renal dysfunction, and neurologic changes.

cold stage• feeling of intense cold• vigorous shivering, rigor• lasts 15-60 min

hot stage• intense heat• dry burning skin• throbbing headache• lasts 2-6 hours

sweating stage• profuse sweating• declining temperature• exhausted, weak sleep• lasts 2-4 hours

• paroxysms associated with synchrony of merozoite release

• temperature is normal and patient feels well between paroxysms

• falciparum may not exhi-bit classic paroxysms• continuous fever• 24 hr periodicity

Malaria Paroxysm

tertian malariaquartan malaria

RelapseRelapse Hypnozoite (Resting stage)Hypnozoite (Resting stage) RecrudescenceRecrudescence Induced malariaInduced malaria HemozoinHemozoin

ها واژه ها توضیح واژه توضیح

Stippling dotsStippling dots

1) Schuffner’s dots1) Schuffner’s dots 2) Maurer’s dots2) Maurer’s dots 3) Ziemann’s dots3) Ziemann’s dots 4) Jame’s dots4) Jame’s dots

Sticky phenomen( knobs)Sticky phenomen( knobs)

Malignant Tertian MalariaMalignant Tertian Malaria Infections caused by Infections caused by P. falciparumP. falciparum can progress to severe; can progress to severe; 1-Cerebral malaria 1-Cerebral malaria 2-Black water fever ( severe anemia)2-Black water fever ( severe anemia) 3-Acute renal failure3-Acute renal failure 4- Algid malaria 4- Algid malaria 5- Respiratory distress syndrome (pneumonic malaria)5- Respiratory distress syndrome (pneumonic malaria) 6- Gastro-intestinal mlaria6- Gastro-intestinal mlaria Complications of Complications of P. vivaxP. vivax malaria include splenomegaly malaria include splenomegaly

(with, rarely, splenic rupture),(with, rarely, splenic rupture), and those of and those of P. malariaeP. malariae include nephrotic syndrome. include nephrotic syndrome.

Karunaweera et al (1992) PNAS 89:3200

• TNF = tumor necrosis factor- ()• proinflammatory cytokine (produced

in response to malarial antigens?)

rigor

sweating

Immunity• slow to develop• short lived

• ‘premunition’• non-sterilizing

• lower parasitemia• less symptoms

Anti-Parasite Immunity• immune response prevents

merozoite invasion, eliminates infected erythrocytes, etc.

Anti-Disease Immunity• eg., neutralization of exo-

antigens or toxic effects

Immunity in MalariaImmunity in Malaria

A) Natural immunityA) Natural immunity 1-Innate immunity1-Innate immunity 2- Genetic immunity2- Genetic immunity

B) Acquired immunityB) Acquired immunity 1-Exo-Erythrocytic stage1-Exo-Erythrocytic stage Premonition: prevents of super-infection no re-infection (stage Premonition: prevents of super-infection no re-infection (stage

specific).specific).

2- Erythrocytic stage2- Erythrocytic stageconcomitant immunity (stage specific & strain specific ):concomitant immunity (stage specific & strain specific ):- Ab mediated immunityAb mediated immunity- Ab dependent cell cytotoxicity (ADCC)Ab dependent cell cytotoxicity (ADCC)- Ab dependent phagocytosisAb dependent phagocytosis- Cellular immunityCellular immunity

Stable or Endemic Malaria

• ~constant incidence over several years• includes seasonal transmission

• immunity and disease tolerance correlates with level of endemicity (especially adults)

Unstable or Epidemic Malaria

• periodic sharp increase in malaria• little immunity• high morbidity and mortality

Malaria Epidemiology

EndemicityLevels:• holo-• hyper-• meso-• hypo-

Malaria laboratory diagnosisMalaria laboratory diagnosis

1) Microscopic identification:1) Microscopic identification: -preparing thick and thin biood smear -preparing thick and thin biood smear -comparison of plasmodium species-comparison of plasmodium species

2- Quality Buffy Coat ( QBC)2- Quality Buffy Coat ( QBC)

3) Immunochromatographic methods3) Immunochromatographic methods

4)Antibody Detection4)Antibody Detection

5) Molecular diagnosis techniques5) Molecular diagnosis techniques

Thin Thin Blood Blood SmearSmear

Thick Blood SmearThick Blood Smear

AA:: Immature schizont in a thin Immature schizont in a thin blood smear.blood smear.B:B: Mature schizont Mature schizont

A B

Multiply infected Multiply infected red blood cells red blood cells with appliqué with appliqué

forms in thin blood forms in thin blood smears smears

     A B C

Ruptured schizonts in a thin blood Ruptured schizonts in a thin blood smearsmear..

   C D

Malaria antibody detectionMalaria antibody detection    The IFA procedure The IFA procedure Blood stage Blood stage PlasmodiumPlasmodium species schizonts species schizonts

(meronts) are used as antigen(meronts) are used as antigen

Enzyme immunoassays have also been employed Enzyme immunoassays have also been employed as a tool to screen blood donors,as a tool to screen blood donors,

but are not recommended for clinical diagnosis due but are not recommended for clinical diagnosis due to limited sensitivityto limited sensitivity

serologic testing is not practical for routine serologic testing is not practical for routine diagnosis of acute malaria. diagnosis of acute malaria. 

Antibody detection may be useful Antibody detection may be useful for:for:

1- screening blood donors1- screening blood donors

2- Fever of Unknown Origin2- Fever of Unknown Origin

3- testing a patient who has been recently 3- testing a patient who has been recently treated for malaria but in whom the treated for malaria but in whom the diagnosis is questioned diagnosis is questioned

Species-specific testing is available for the Species-specific testing is available for the four human species: four human species: P. falciparumP. falciparum, , P. vivaxP. vivax, , P. malariaeP. malariae, and , and P. ovaleP. ovale. . 

Cross reactions often occur between Cross reactions often occur between PlasmodiumPlasmodium species and species and BabesiaBabesia species.  species. 

Usefulness of Antibody Usefulness of Antibody Detection in the Detection in the

Diagnosis of Malaria Diagnosis of Malaria ParasitesParasites

Types of Serological Assays Types of Serological Assays MalariaMalaria

Antibody Detection:

Indirect Fluorescent Antibody

Enzyme immunoassays

Antigen Detection:

Immunochromatographic

Antibody DetectionAntibody Detection

=

Antigen-antibodycomplex

Patient’s serumcontains specificand non-specific

antibodies

+

Antigen

Antibody DetectionAntibody Detection

*-labeled antibody to

human antibody

+

Antigen-antibody-

*antibody complex

=

Antigen-antibodycomplex

Indirect Fluorescent Antibody Indirect Fluorescent Antibody (IFA)(IFA)

Microscope slide

Enzyme Immunoassay Enzyme Immunoassay (EIA/ELISA)(EIA/ELISA)

_

+enzymesubstrate

ELISA

Antigen DetectionAntigen Detection

Monoclonal antibody

=

Antigen-antibodycomplex

+

Antigen inpatient’s serum

Antigen DetectionAntigen Detection

=

Antibody-antigen-antibodycomplex

+

Immobilized

monoclonal

antibody

Antigen-antibodycomplex

Antigen DetectionAntigen DetectionMalaria Immunochromatographic Malaria Immunochromatographic

DipstickDipstickOptimal Assay

ControlPlasmodium pan specific

monoclonal antibody

P. falciparum specific

monoclonal antibody

Antigen DetectionAntigen DetectionMalaria Immunochromatographic Malaria Immunochromatographic

DipstickDipstickProblems: Problems: Low sensitivity with parasites density Low sensitivity with parasites density

<100/ml<100/ml

Currently only useful for detection ofCurrently only useful for detection of

P. falciparumP. falciparum infections infections

Diagnostic ToolsDiagnostic Toolsfor Human Infections with for Human Infections with

MalariaMalaria

Blood film Blood film examinationexamination

Serology - IFASerology - IFAPCRPCR

Malaria IFA TestMalaria IFA Test

Sensitivity = 98%Sensitivity = 98%

Specificity = 99.5%Specificity = 99.5%

Sulzer et al, Am J Trop Med Hyg 1969;18:199-Sulzer et al, Am J Trop Med Hyg 1969;18:199-205205

Sulzer et al, Bull Wld Hlth Org 1971;45:375-379Sulzer et al, Bull Wld Hlth Org 1971;45:375-379

P. malariae

P. malariae

P. malariaeP. malariae

P. falciparum

P. falciparum

P. falciparumP. falciparum

A:A: Positive IFA result with Positive IFA result with P. malariaeP. malariae schizont schizont

antigen.antigen.

        A

Malaria antibody detectionMalaria antibody detection for clinical diagnosis is performed using the indirect for clinical diagnosis is performed using the indirect

fluorescent antibody (IFA) test.  fluorescent antibody (IFA) test. 

The IFA procedure can be used as a diagnostic tool to determine The IFA procedure can be used as a diagnostic tool to determine if a patient has been infected with if a patient has been infected with PlasmodiumPlasmodium.  . 

Blood stage Blood stage PlasmodiumPlasmodium species schizonts (meronts) are used species schizonts (meronts) are used as antigenas antigen

Because of the time required for development of antibody and Because of the time required for development of antibody and also the persistence of antibodies, serologic testing is not also the persistence of antibodies, serologic testing is not practical for routine diagnosis of acute malariapractical for routine diagnosis of acute malaria. . 

Enzyme immunoassays have also been employed as a tool to Enzyme immunoassays have also been employed as a tool to screen blood donors, but are not recommended for clinical screen blood donors, but are not recommended for clinical diagnosis due to limited sensitivitydiagnosis due to limited sensitivity

Antibody detection may be useful for:Antibody detection may be useful for:

1- screening blood donors involved in cases of transfusion-1- screening blood donors involved in cases of transfusion-induced malaria when the donor's parasitemia may be below induced malaria when the donor's parasitemia may be below the detectable level of blood film examination the detectable level of blood film examination

2- testing a patient with a febrile illness who is suspected of 2- testing a patient with a febrile illness who is suspected of having malaria and from whom repeated blood smears are having malaria and from whom repeated blood smears are negative ( Fever of Unknown Origin)negative ( Fever of Unknown Origin)

3- testing a patient who has been recently treated for malaria but 3- testing a patient who has been recently treated for malaria but in whom the diagnosis is questioned in whom the diagnosis is questioned

Species-specific testing is available for the four human Species-specific testing is available for the four human species: species: P. falciparumP. falciparum, , P. vivaxP. vivax, , P. malariaeP. malariae, and , and P. ovaleP. ovale. . 

Cross reactions often occur between Cross reactions often occur between PlasmodiumPlasmodium species and species and BabesiaBabesia species. . species. .

A:A: Positive IFA result with Positive IFA result with P. malariaeP. malariae schizont antigen. schizont antigen.

        A

Rapid Diagnostic Tests(dipstickor test strip)Rapid Diagnostic Tests(dipstickor test strip) (basedon the detection of antigens malaria parasites; Histiding- rich protein II) (basedon the detection of antigens malaria parasites; Histiding- rich protein II)

Comparison of Comparison of PlasmodiumPlasmodium Species Species

TreatmentTreatment• ChloroquineChloroquine

• Sulfadoxine-pyrimethamine (Fansidar) Sulfadoxine-pyrimethamine (Fansidar)

• Mefloquine (Lariam)Mefloquine (Lariam) • Quinine Quinine

• DoxycyclineDoxycycline • Artemisin derivativesArtemisin derivatives

Malaria ControlReduce Human-Mosquito Contact

• impregnated bed nets• repellants, protective clothing• screens, house spraying

Reduce Vector • environmental modificaton• larvacides/insecticides• biological control

Reduce Parasite Reservoir• diagnosis and treatment• chemoprophylaxis