Apresentao do PowerPoint

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Leishmaniasis
Silvia Reni B. Uliana
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Leishmania
Family Trypanosomatidae
Genus Leishmania
Sub-genera Leishmania
Viannia
More than 30 species already described
cutaneous leishmaniasis visceral leishmaniasis
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cutaneous leishmaniasis visceral leishmaniasis
Geographical Distribution
14 million cases each year, distributed in 88
countries (350 million people at risk).
http//www.who.int/emc-documents/surveillance/docs
/whocdscsrisr2001.html
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Visceral leishmaniasis / AIDS
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Life cycle
(MAN)
Assexual
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Promastigotes
Extracellular forms living in the intestine of
the mosquito
In the mosquitos gut they go through a
differentiation process called metacyclogenesis.
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Amastigote
Intracellular forms
Ideal host cell macrophage
Amastigotes are found inside macrophages at
various sites skin, lymph nodes, spleen, liver,
bone marrow.
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Mecanismo de infecção
O macrófago parasitado eventualmente se rompe,
liberando amastigotas que passam a infectar
outros macrófagos, circulantes ou teciduais.
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Clinical syndromes
Cutaneous leishmaniasis L. braziliensis, L.
amazonensis, L. guyanensis L. major Diffuse
leishmaniasis L. amazonensis Mucocutaneous
leishmaniasis L. braziliensis Visceral
leishmaniasis L. chagasi, L. donovani
New World species Old World species
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Cutaneous leishmaniasis
L. amazonensis L. mexicana L. guyanensis L.
braziliensis
Reservoir wild rodents, marsupials (opossum),
sloth, anteater
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Diffuse leishmaniasis
L. amazonensis
Reservoir wild rodents
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Mucocutaneous leishmaniasis
L. braziliensis
Reservoir ? Wild rodents
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Visceral leishmaniasis
L. chagasi L. donovani L. infantum
Reservoir fox, dog, humans
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Some tricks performed by Leishmania
S.C. Ilgoutz, M.J. McConville / International
Journal for Parasitology 31 (2001) 899908 906
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Some tricks performed by Leishmania -LPG
form McConville MJ in Molecular Biology of
Parasitic Protozoa, ed. DF Smith, M Parsons
multiplication inside the gut adherence -
procyclic being available to take the next
ride - metacyclic
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Immune system
Complement
C3bi
Lysis complex
OGarra and Arai (2000) Trends Cell Biol.
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the innate defense mechanisms
LPG and complement
lysis complex
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getting home - the macrophage!
Interaction with C3bi receptor via complement and
gp63
inhibition of the respiratory burst
burst
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shall we make it cosy? the parasitophorous
vacuole
Fusion with lisossomes gp63 is a protease (other
cysteino-proteases) LPG and other GL induce
the production of TGF-? and IL-10 AND inhibit
the production of NO
the macrophage army is ready
DISEASE
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Diagnosis

• Biopsy low sensitivity
• which species ?
• Culture slow
• isoenzymes, monoclonal antibodies
• PCR
• ELISA cross-reaction, sensitivity

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Treatment

• pentavalent antimonials
• anphotericin B

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VECTOR partially effective in urban areas
CONTROL
RESERVOIR - wild
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Clinical syndromes
Cutaneous leishmaniasis L. braziliensis, L.
amazonensis, L. guyanensis L. major Diffuse
leishmaniasis L. amazonensis Mucocutaneous
leishmaniasis L. braziliensis Visceral
leishmaniasis L. chagasi, L. donovani
New World species Old World species
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Briones et al 1992, MBP
Orlando et al 2002 MIOC
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Immune-deficiency associated
Primary infection (with or without symptoms)
Where ? How ?
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VIRULENCE
Ilgoutz and McConville (2001) Int J Parasitol
How do they do all this ?
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Characterisation of the meta 1 gene in L. major

• Single copy gene
• RNA expression upregulated in metacyclic
  promastigotes

10 20 30
40 L.maj MEMKNLLGKH KIVSVNGKPA PAGVTVEFKA
SENSGSVYMH L.don ....H..... .V.L...R..
.......... G......H.. L.ama ....S.I..R
RVL....R.. .......... .....TIQ..
50 60 70 80 L.maj
AKVANIMNGP LKLANRKLSG ALVSTMMLGS DDLMNIENAL L.don
.R.......Q .R.E...... ..........
.......... L.ama .N...F...Q ...G......
T......... .......... 90
100 110 L.maj SQGFMEGMTY TVKDGGKLTL
QSKTHIIMLV PA L.don .......... ..........
.......K.. .. L.ama I....D.... ..H.......
K.N..T.K.. ..

• Gene conserved in both Old and New World
  Leishmania species

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Deduced meta 1 aminoacid sequence

• No significant aminoacid or nucleotide identity
  with any known protein.
• No glycosilation sites.
• Hydrophilicity plot with no distinguishing
  features.
• Predicted molecular mass 11.95 kDa.

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What is the function of the meta 1 protein in
Leishmania ?
A- Lines with double replacement of the meta 1
gene were not viable
ESSENTIAL GENE
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B- Overexpressing the meta 1 protein

• High level expression of the L. major meta 1
  protein was obtained in L. amazonensis
  promastigotes, by stable maintenance of an
  episome.
• The levels of expression were high in both log
  phase and stationary phase promastigotes.

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C- Testing the recombinant lines for a phenotype

• No change in growth curves
• No differences in in vitro infection of
  macrophages

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Meta 1 overexpressor in vivo
BALB/c 107 promastigotes
A
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Lesion Index
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10
5
4
6
8
10
12
14
16
Weeks
B
BALC/c 106 amastigotes
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Lesion Index
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15
10
5
4
6
8
10
12
14
16
Weeks
C
C57Bl/6 106 promastigotes
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Lesion index
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WT
HE
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3
2
1
6
12
24
Weeks
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L. major Chromosome 16