Molecular%20Epidemiology%20and%20Susceptibility%20to%20Malaria%20Infection

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Molecular Epidemiology and Susceptibilityto
Malaria Infection
Douglas Jay Perkins, Ph.D.
University of Pittsburgh Graduate School of
Public Health Department of Infectious Diseases
and Microbiology Centers for Disease Control and
Prevention Division of Parasitic
Diseases-Immunology Branch Molecular Vaccine
Section, Atlanta, GA
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Malaria Transmission Cycle
Pre-erythrocytic
Asymptomatic
Erythrocytic
Clinical symptoms
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Malaria in Humans

• Four species of genus Plasmodium infect humans
  P. falciparum, P. vivax, P. ovale, and P.
  malariae
• Transmitted by female Anopheline mosquito
• 300-500 million clinical cases per year

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Populations at Risk

• Infants, young children, and pregnant women in
  malaria endemic regions
• Greater than 3 million deaths (primarily in
  children less than 5 y/o due to non-immune
  status)
• Non-immune individuals traveling through and/or
  living in malaria endemic regions
• 35 million non-immune individuals travel through
  malaria endemic regions every year

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Clinical Features of P. falciparum

• P. falciparum can cause severe malaria-hyperpara
  sitemia -severe anemia -hypoglycemia-respirat
  ory distress-cerebral malaria
• Molecular determinants that regulate mild versus
  severe disease largely unknown

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Current Situation Major International Health
Problem

• Rapidly expanding number of clinical cases each
  year
• Growing problem of antimalarial drug resistance
  with few novel therapeutics available
• Lack of an effective vaccine

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Potential Solutions

• Gain an understand of the genetic and immunologic
  basis of protective immunity
• Identify novel targets for therapeutic
  intervention
• Determine reliable markers for measuring
  protection and pathogenesis for use in
  pharmacologic and/or vaccine trials

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Genetic Susceptibilityto Malaria

• At least 10,000 years of pressure on the human
  genome from the malaria parasite
• In 1948 J.B.S. Haldane suggested that the high
  frequency of thalassemia in Mediterranean
  populations might confer a heterozygote advantage
  against malaria
• Thalassemias are defects in synthesis of either
  a- or b-globin chains of hemoglobin (hemoglobin
  adult a2b2)
• Mechanism of protection may be related to
  increased binding of antibodies and/or increased
  retention of fetal hemoglobin

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Sickle Cell Gene and Resistanceto Malaria

• Over 400 abnormal hemoglobins but only three
  reach polymorphic frequencies (S, C, E)
• Homozygous state (SS) sickle cell disease
• Heterozygous state (SC) protection from malaria
• Mechanism unknown but red blood cells from (SC)
  individuals have reduced parasite growth and
  impaired invasion under low O2 tension
• In addition to red cell abnormalities, there are
  many other genetic changes..

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Host Response Genes and Susceptibility to Malaria

• In 1993 Murphy compared sequences of human and
  rodent genes and found greater variability among
  host defense genes
• Polymorphisms in cytokines genes (e.g. TNF-a) and
  effector molecules (e.g. nitric oxide, NO) are
  now being investigated
• Study of genetic variation may utilize several
  types of DNA markers to analyze candidate
  susceptibility genes
• Single base pair variations SNPs
• Microsatellite or variable number tandem repeats
  (VNTRs)

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Overview
Part 1. NOS2 (G 954C) in Gabonese Children
with Severe Malarial Anemia Part 2. NOS2 (G
954C) in Tanzanian Children with Cerebral
Malaria Part 3. NOS2 (G 954C) in
Kenyan Children with Severe Malarial Anemia
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Nitric Oxide Biosynthesis
NOS
L-Arginine
L-Citrulline

NO
L-NMMA Aminoguanidine
NO2-
NO3-
NOS Enzyme Assay
Cellular Lysate
14CL-Arg remains
Co-factors
14CL-Arg
14CL-Cit
14CL-Cit flows through
Cation Exchange Column
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Nitric Oxide Synthase
eNOS nNOS NOS3 NOS1
iNOS NOS2
Constitutive Expression
Inducible Expression
- Ca2- and Calmodulin- Dependent
- Ca2- and Calmodulin- Independent
NO Synthesis for Normal Physiologic Function
NO Synthesis in the Setting of Inflammation
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Nitric Oxide Previous Observations in Malaria

• Nitric oxide production is anti-plasmodial in
  vitro and in vivo-(Oswald et al.,Comp Biochem
  Physiol Pharmacol Toxicol Endocrino, 1994
  10811-18)
• Elevated NO metabolites are associated with
  accelerated clinical cure and increased
  parasitologic clearance in Gabonese adults and
  children -(Kremsner et al., Trans R Soc Trop Med
  Hyg, 1996 90 44-47)
• NO appears protective against malaria

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Model of NO Production in Malaria
Monocyte
PRBC
PRBC
Monocyte/Macrophage
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Hypothesis
Increased capacity of the host to generate nitric
oxide is protective against severe malaria