full genome sequencing of samba virus: a giant virus of ... · mimivirus as a pneumonia agent 1-...
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Full genome sequencing of Samba virus:
a giant virus of the Mimiviridae family
isolated from Negro River, in the
Brazilian Amazon rainforest
Jônatas Abrahão
Federal University of Minas Gerais
São Paulo, April - 2014
The Virosphere’s
Gulliver
La Scola, 2003
• Changed our view about
viruses
• Broken paradigms (complexity
of viruses, what viruses code in
their genomes)
The Virosphere’s Gulliver
Mimivirus vs Picornavirus
• Early 1990’s: isolated from a cooling tower in a Bradford’s
Hospital, England
• Identified as a bacteria (Bradford
coccus) – Associated to Amoebas
Mimiviruses: the discovery
Clinical Infectious Disease, 2008
• Was not able to growth in axenic
medium
• Resistant to antibiotics
• 16S RNA - negative
Mimiviruses: the discovery
• IT’S A GIANT VIRUS!!!!
• 800nm
• 1.2 Mb (DNA)
• About 1.100 ORFs
• Genes never seen before in the
virosphere
Pandoravirus and Pithovirus
Pandoravirus dulcis
~1 um
Genome: 2.5 megabase
(DNA)
Pithovirus sibericum
~1.5um
Genome: 0.6 megabase
(DNA)
Philippe N, et al. Science. 2013 Jul 19;341(6143):281-6.
Legendre M, et al. Proc Natl Acad Sci U S A. 2014 Mar 18;111(11):4274-9.
Mimiviruses in Brazil
Vaccinia virus, the largest animal virus ever isolated in Brazil
2008 – Genetic analysis of viral RNA Helicase
~ Mimiviruses
Mimiviruses in Brazil
Prospecting
studies
Resistance
to biocides
Mimivirus vs
Hosts
Relashionships
Genomic
Approaches
Biological
characterization
Mimivirus
Assis, F.
Silva, L.
Serological
surveysDornas, F.
Structural
studies
Silva, L; Dornas, F.
Campos, R; Dornas, F;
Boratto, P
Campos, R.
Campos, R; Andrade,
K; Boratto, P.
Boratto, P.
Rodrigues, R.
Prospecting studies: where are the
Giants?
Rio Negro: 50
samples
Lagoa da Pampulha
(BH): 148 samples
Lagoa Santa
(MG): 200
samples
Parque
Nacional da
Serra do
Cipó (MG:
400 samples
Low efficience of isolation
Prospecting studies: where are the
Giants?
Pre-enrichment with rice medium,
improves in 500% the efficience of
viral isolation
Viral Isolation: 12 positive samples
PCR detection: 25 positive samples
Prospecting studies: where are the
Giants?
They are everywhere!
Prospecting studies: where are the
Giants?
Structural studies
Niemeyer virus Cipó virus
Kroon virusSamba virus
Several
morphological
features
distinct among
Brazilian Giant
viruses: fibrils,
capsid,
membrane
Size:
357 nm – 598 nm
F
F
F
F
C
C
C
C
M
M
M
M
Kroon virus: Structural studies
Kroon virus
has a extra
shell layer –
increased
resistance to
UV light and
temperature
Acanthamoeba castelannii, A. griffin, A. polyphagaand A. lenticulata
Amoebas as Giant Viruses Reservoirs
• Do not grow in Naegleria sp. or Hartmanella sp.
Amoeba Encystment
Trophozoite CystMetabolically activeAble to phagocyteAble to replicate
Very low metabolism activityUnable to phagocyte
Thick cell wallVery resistant (temp, UV, biocides)
Harsh conditions(scarcity of food,
high or lowtemperatures)
Proteases activation
How Mimivirus deals with Amoeba
Encystment
Boratto, submitted to EnvirMicrob, 2014
• Once triggered encystment, mimivirus infection is remarkable hampered
• Once infected, amoebas do not respond to encystment stimulus
Arm wrestling point between virus and amoebas
Mimivirus blocks a amoeba serina
proteinase and prevents encystment
Boratto, submitted to Envir Microb, 2014
Sponge and Corals as Putative
Mimivirus Hosts
Claverie et al., 2009
Horizontal Gene Transfer between APMV and Sponges/Corals
Oysters as Hotspot for Mimivirus
Isolation
Reis, submitted to Envir Microb Reports, 2014
Mimivirus as a Pneumonia Agent
1- First isolated during a pneumonia outbreak (La Scola, 2003)
2- Some studies demonstrated antibodies and viral DNA in pneumonia patients (La Scola, 2005)
3- Viruses have been isolated from respiratory tract of pneumonia patients
4- Mimivirus is able to replicate in human lung monocytes (Ghigo, 2008)5- Mimivirus cause pneumonia in mice intracardiacally infected (Ghigo, 2007)
Mimivirus detection in other Vertebrates
Dornas et al., 2014
Mimivirus interacts with IFN system
Mimivirus is able to replicate in human PBMCs
Mimivirus is able to down regulate ISG(s)
1
2
Silva et al., 2013
Mimivirus is resistant to IFN-alpha, but not to IFN-beta
3
WHY A AMOEBA VIRUS WOULD HAVE GENETIC INFORMATION
ABLE TO EVADE A VERTEBRATE PATHWAY?
Mimivirus interacts with IFN system
Silva et al., 2013
0000000
Samba virus, a Novel Giant Virus
From A Giant Rain Forest, the
Brazilian Amazon
Rafael K Campos, Paulo V Boratto, Felipe L Assis, Eric Aguiar, LorenaC Silva, Jonas D Albarnaz, Fabio P Dornas, Giliane S Trindade, Paulo P
Ferreira, Joao T Marques, Catherine Robert, Didier Rauolt, Erna GKroon, Bernard La Scola and Jônatas S Abrahão
Campos et al., 2014
Campos et al., 2014
UncoatingPhagocytosis
Viral seed
Viral factory
12
3
4
5
Morphogenesis
6
Lysis
Rio Negro, the First Brazilian
Virophage: a Virus “infecting”
another Virus
Campos et al., 2014
Rio Negro, the first BR-Virophage
Campos et al., 2014
Rio Negro, the first BR-Virophage
Campos et al., 2014
Rio Negro, the first BR-Virophage
Campos et al., 2014
How large and complex: Samba
virus Genomic Analysis
Campos et al., 2014
• Larger viral genome
ever sequenced in
Brazil
• Larger Group A
Mimivirus Genome
• ~ 50.000 bp larger
than APMV (the
prototype)
Samba Genomic Analysis
• 938 ORFs (several
ORFs larger than
APMV)
Campos et al., 2014
Campos et al., 2014
Campos et al., 2014
Venn Diagram: 9 “ORFans”!
Campos et al., 2014
Campos et al., 2014
Conclusions
• Mimiviruses are everywhere, regardless the analyzed Brazilian
biome;
• Despite similar, viral samples showed special features, with
remarkable distinctions in particle size, structure and genomic
composition;
• Samba is the first mimivirus isolated in Brazil, and present the
largest Mimivirus Group A genome sequenced until now;
• Rio Negro virus was the first mimivirus virophage isolated from
Americas;
P
Felipe Assis
Graziele Oliveira
Ana Paula Franco-Luiz
Lorena Silva
Ketyllen Andrade
Alexandre Pererira
Carolina Lima
Felipe Peixoto
Fábio Dornas
Paulo Boratto
Rafael Campos
Bernard La Scola
Didier Raoult
Catherine Robert
Erna Kroon
Cláudio Bonjardim
Giliane Trindade
Paulo Ferreira
Prof. Joao T. Marques
Eric Aguiar
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