WND e le altre Patologie Emergenti in Italia e nella
Pianura Padana
Maria Grazia Ciufolini,
Department of Infectious,
Parasitic Immuno-mediated Diseases
Istituto Superiore di Sanità, Rome
Rovigo 03/04/2009
WN Disease e Patologie emergenti
nella Pianura Padana
Arbovirus:
ARthropod BOrne Viruses
Arboviruses are viruses which are maintained in
nature principally, or to an important extent, through
biological
transmission
between
susceptible
vertebrate hosts by hematophagous arthropods:
they multiply and produce viremia in vertebrates,
multiply in the tissues of arthropods, and are passed
to new vertebrates by the bites of arthropods after a
period of extrinsic incubation.
W.H.O. 1967
Arbovirus:
ARthropod BOrne Viruses
Virus trasmessi da artropodi
Zanzare
Zecche
Altri
Sono alcune centinaia (>500
nel catalogo del 1985)
Causano gravi problemi
sanitari a livello globale
Causano milioni di casi ogni
anno
Migliaia di morti ogni anno
Forte impatto sull’economia
Seguono spesso
cambiamenti climatici
Uragani
Riscaldamento globale
“Emergenza” degli Arbovirus
L'emergenza può verificarsi in seguito a:
variazioni ambientali (cambiamenti climatici /ecologici
che favoriscono lo sviluppo dei vettori e degli animali
serbatoi aumentando la possibilità di un loro contatto
con l'uomo)
insorgenza di varianti virali favorite dalla natura stessa
del loro genoma ad RNA segmentato (mutazioni del
genoma virale, riassortimento, selezione naturale,
evoluzione).
variazioni dovute a comportamento umano (scambi
commerciali, comportamenti sociali).
Ri-emergenza di epidemie causate
da arbovirus, 1990-2006
CE
SIN
TBE
WN
CCHF
SIN
WN
WN
DEN
WN
WN EEE
WN
SLE LAC SLE
WN
EEE
DEN
VEE WN DEN
WN
DEN WN
ORO VEE DEN
DEN VEE
WN MAY
DEN
ORO
DEN
DEN YFDEN ORO
YF YF
DEN
WN
BF - Barmah Forest
CE - California Encephalitis
CHIK - Chikungunya
CCHF - Congo-Crimean Hemorrhagic Fever
DEN - Dengue
EEE - Eastern Equine Encephalitis
JE - Japanese Encephalitis
KFD - Kyasanur Forest Disease
LAC - LaCrosse Encephalitis
TBE
WN
WN
CCHF
WN
WN
DEN
DEN
WN RVF KFD
CCHF
CCHF
DEN
YF
RVF
CHIK
RVF
DEN
YF
WN
JE
DEN
RVF
DEN ONN
CHIK
DEN
WSL
WN
MAY - Mayaro
MVE - Murray Valley Encephalitis
ONN – O’nyong-nyong
ORO - Oropouche
RVF - Rift Valley Fever
RR - Ross River
SLE - St. Louis Encephalitis
SIN - Sinbis
TBE- Tick-Borne Encephalitis
DEN
JE
DEN
DEN JE DEN
CHIK CHIK
JE DEN
DEN
DEN
RR
BF
MVE
JE
DEN
JE
DEN
VEE - Venezuelan Equine Encephalitis
WEE - Western Equine Encephalitis
WN - West Nile
WSL - Wesselsbron
YF - Yellow Fever
Italy
TBE
Arboviruses in Italy
Chikungunya
West Nile
TBE
West Nile
Bahig
Matruh
Tahyna
TBE
Tribec
Toscana
Toscana
Arbia
Radi
Toscana
Arbia
Bhanja
Radi
TBE
Toscana
Tribec
West Nile
Toscana
Toscana
SFN
Toscana
Toscana
Bhanja
West Nile (?)
SFS
Thogoto
Toscana
Arbovirus causa di malattia
nell’uomo in Italia
Trasmessi da zanzare (Culex et Aedes.):
West Nile: Flaviviridae, Flavivirus
Chikungunya : Togaviridae, Alphavirus
Trasmessi da zecche (I.ricinus):
TBE: Flaviviridae, Flavivirus
Trasmessi da flebotomi ( P.perniciosus e P.perfiliewi):
Virus Toscana: Bunyaviridae, Phlebovirus
Mosquito-borne virus
isolated in Italy
Family/Genus
Sources of isolation
Virus
Year
Place
Species
Bunyaviridae
Bunyavirus
Tahyna
1967
Friuli
Aedes spp.
Flaviviridae
Flavivirus
West Nile
1969
1998
2008
Lazio
Tuscany
Emilia
Romagna
Ticks
Horse
Emilia
Romagna
Human
Aedes albopictus
Togaviridae
Alphavirus
Chikungunya
2007
Pica pica
Columbia livia
Lancet, 1 dicembre 2007
Castiglione di Cervia
Castiglione di Ravenna
West Nile virus
Virus West Nile:
Isolato per prima volta nel 1937 in Uganda dal
sangue di una donna con sindrome febbrile
Famiglia: Flaviviridae
Genere: Flavivirus
Complesso antigenico: Encefalite giapponese
Il complesso antigenico include i virus: Alfuy(ALF),
Encefalite Giapponese(JE), Cacipacore(CPC), Koutango
(KOU), Kunjin, Encefalite di Murray Valley(MVE),
Encefalite di St. Louis(SLE), Yaounde (YAO), Usutu
(USU), e West Nile(WN).
Tutti i virus sono trasmessi da zanzare, molti di
loro causano malattia febbrile nell’uomo, a volte
fatale.
Virus West Nile:epidemie
Dagli anni ‘60 casi sporadici e gravi epidemie sia
nell’uomo sia nel cavallo, in Africa, Europa e
Medio Oriente.
Dalla metà degli anni ’90 incremento del numero
di epidemie e aumento della gravità della
sintomatologia
Nel 1999 focolai a New York, e dal 2003:
ufficialmente endemica negli USA, ha raggiunto
il Canada e il Sud America.
Virus West Nile distribuzione:epidemie
Israele 1951/1954,1957
Francia 1962
Sud Africa 1974
Nord Africa 1994,1996
Romania 1996,1997
Repubblica Ceca 1997
Algeria 1994
Marocco 1996
Tunisia 1997
Ucraina 1980
Italia 1998
Russia 1999
USA dal 1999…
Israele 1998, 1999
Francia 2000, ‘03, ‘06
Italia 2008
Diffusione del virus West Nile negli
USA
1999
2001
2003
2008
Epidemics caused by West Nile virus, 1937–2006. The red stars indicate epidemics that
have occurred since 1994 that have been associated with severe and fatal neurologic
disease in humans, birds, and/or equines.
Gubler DJ. Clin Infect Dis. 2007 Oct 15;45(8):1039-46.
Filogenesi dei ceppi di virus West
Nile in USA
Tutti i ceppi
americani sono
identici >99.8%
(nucleotidi)
Egypt 1951
France 1965
Albero Filogenetico
del virus West Nile
2001
(Envelope gene)
Differenza <3
amino acidi tra due
isolati qualsiasi
1
Kunjin
India
2
South Africa
Israel 1952
Romania 1996 M
Kenya 1998
Senegal 1993
Morocco 1996
Italy 1998
Volgograd 1999
New York 1999
Israel 1998-A
NY2000 3282
NY2000 3356
NY 1999 equine
NY 1999 hum
Conn 1999
MD 2000
NJ 2000
Israel 1999 H
C.Afr.Rep 1989
Senegal 1979
Algeria 1968
C.Afr.Rep 1967
Iv.Coast 1981
Kunjin 1960
Kunjin 1973
Kunjin 1984b
Kunjin 1991
Kunjin 1984a
Kunjin 1966
Kunjin 1994
India 1955a
India 1980
India 1958
India 1955b
Kenya
Uganda
Senegal 1990
Uganda 1937
C.Afr.Rep 1972a
C.Afr.Rep 1983
Uganda 1959
C.Afr.Rep 1972b
Madagascar 1988
Madagascar 1986
Madagascar 1978
JE SA 14
US/
LIN-1
Israel
LIN-2
Virus West Nile:
la distribuzione in Europa
Isolamento da
zanzare o vertebrati
Casi umani o
equini
Anticorpi nei
vertebrati
Virus West Nile:
ciclo di trasmissione
Vettore
Zanzare del complesso
Culex pipiens
Infezioni occasionali
Altri mammiferi
Trasmissione rara
Senza vettore
In utero
Latte materno
Esposizione
occupazionale
Trasfusione
Trapianto
Vertebrati serbatoio
Trasmissione
diretta
Virus West Nile: entomologia
Isolato da più di 60 specie di zanzare
Specie Culex:
Altre zanzare di altri generi:
Cx. univittatus, Cx. perixiguus, Cx. pipiens,
Cx. modestus, Cx. quinquefasciatus, Cx.
tritaeniorhynchus, and Cx. vishnui
Aedes, Aedeomyia, Anopheles, Coquillettidia,
Mansonia, Mimomyia
Isolato da zecche:
Argas, Ornithodoros
Amblyomma, Dermacentor, Hyalomma,
Rhipicephalus,
West Nile : caratteristiche del virus
Famiglia
Flaviviridae
Trasmissione
Zanzare
Particelle
virali
Sferiche,
40-60 nm
envelope
Proteine
strutturali
Genoma
C, M, E
Ss-RNA, non
segmentato,
polarità positiva
West Nile : il virus
VIRIONE IMMATURO
VIRIONE MATURO
proteina prM
proteina M
Proteina E
NUCLEOCAPSIDE
INVOLUCRO
PERICAPSIDICO
(RNA e proteina C)
Genoma dei Flavivirus
NC
ns4a
ns2a
5'- C prM
furin
E
NS5
NS3
NS1
ns2b
NC
ns4b
pr + M
7 geni per proteine non strutturali
3 geni per
proteine
strutturali
-3'
L’albero evolutivo dei Flavivirus
COMPLESSO SIEROLOGICO
DENGUE
Omologia degli aminoacidi della proteina E (%)
DEN
3 1 2 4
ENCEFALITE
GIAPPONESE
WN MVE SLE
KUN JE
FEBBRE
GIALLA
YF
ENCEFALITE
TRASMESSA DA ZECCHE
POWLGTLI TBE
CEE RSSE
100
96
91-94
93
90
85-89
80
82
77
70
60
77-78
77-78
72-74
69
62
46-53
50
40
40-44
Virus West Nile:
la malattia nell’uomo
Periodo di incubazione: 2-15 gg
Nella maggior parte dei casi:
Sindrome febbrile, simil-influenzale con
inizio improvviso
Febbre moderata-alta
Mal di testa, mal di gola, dolori alla schiena,
mialgia, artralgia, affaticamento
Esantema, linfoadenopatia
Raramente: pancreatite, epatite,
miocardite
Virus West Nile:
la malattia nell’uomo
Malattia neurologica grave
Meningite
Encefalite
Meningoencefalite
Paralisi flaccide acute
West Nile virus in Italia
Between August and October 1998 an
outbreak of equine encephalomyelitis
occurred in Tuscany Region. 14 cases
were registered, with 6 deaths.
A strain of West Nile virus was isolated
from the brain of a horse.
“At risk” humans investigated for
West Nile virus infection
Positive/tested
Locality
N°
HI
ELISA
PRNT
Pisa
9
6(±)/9
0/9
0/6
Lucca
3
0/3
0/3
n.d.
Pistoia
114
6/81
9/114
4/9
Total
126
Viremia e cinetica anticorpale durante
l’infezione da Flavivirus neurotropi
Sintomi
neurologici
IgM
IgG
Viremia
RNA
virale
Infezione
Comparsa
febbre
Malattia
2-4 mesi
>1 anno
Diagnostic problems
All flaviviruses are serologically correlated.
After infection antibodies are produced that
cross-react in ELISA tests with other flaviruses
Vaccination against YF and JE, or Dengue
infection, produce antibodies cross-reacting,
non-neutralising that give positive ELISA test
results
In these cases specific humoral immunity
response against flaviviruses virus can be
determined only with neutralization test
Human cases investigated for
West Nile virus infection (2008)
Positive/tested
Region
N°
ELISA
HI
PRNT
Veneto
26
17/26
26/26
6/26
5
5/5
5/5
3/5
31
22/31
31/31
9/31
Emilia
Romagna
Total
Phylogenetic analysis of West Nile virus isolated in Italy in 2008 G Savini, F
Monaco, P Calistri, R Lelli. Eurosurveillance, Volume 13, Issue 48, 27
November 2008
Tick-borne viruses
Tick-borne viruses
isolated in Italy
Family/
Genus
Sources of isolation
Virus
Year
Place
Species
Flaviviridae
Flavivirus
TBE
1978 ,1980
1994, 2004
1998, 2000
2005
Toscana
Trentino
Veneto
Friuli
Ixodes ricinus
Bunyaviridae
Ungrouped
Bhanja
1967
1973
Lazio
Toscana
Haemaphysalis
punctata
Orthomyxoviridae
Orthomyxovirus
Thogoto
1969
Sicilia
Rhipicephalus
bursa
Reoviridae
Orbivirus
Tribec
1972
1977
Friuli
Toscana
Ixodes ricinus
Virus TBE
Famiglia: Flaviviridae
Genere: Flavivirus
TBEV : species in the Mammalian
group of tick- borne flavivirus
TBEV species include 3 sub-types:
Far Eastern (RSSE)
Siberian (West Siberian)
Western European ( CEE)
Antigenically related
Louping ill virus (LIV),
Langat virus (LGTV),
Powassan virus (POWV),
Omsk Hemorrhagic fever virus
(OHVF),et al. (TBEV serocomplex):
19
75
19
79
19
80
19
82
19
83
19
86
19
87
19
91
19
92
19
93
19
94
19
95
19
96
19
97
19
98
19
99
20
00
20
01
20
02
20
03
20
04
20
05
20
06
20
07
TBE cases diagnosed in Italy
20
15
10
5
0
Florence
Bolzano
Trento
Friuli
Belluno
Distribution of TBE cases and
virus isolation in Italy
Trentino Alto Adige
(1992-2007)
50 human cases
2 virus isolation*
Friuli Venezia Giulia
(1998-2007)
49 human cases
virus isolation
Toscana
(1975-2004)
14 human cases
2 virus isolations*°
* From Ixodes ricinus ticks
° From Apodemus silvaticus mouse
Veneto
(1992-2007)
137 human cases
4 virus isolations*
1280
NT80
antibody
titres
640
320
160
80
40
HI
antibody
titres
20
10
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
Antibody titres (reciprocal of
serum dilution)
Hemagglutination inhibition (HI) and neutralizing (NT80: reciprocal of
the serum dilution that gave an 80% reduction of the number of plaques in
the neutralization test) antibody titers.
patients
Humoral immunity and correlation between ELISA hemagglutination inhibition, and neutralization
tests after vaccination against tick-borne encephalitis virus in children
G. Venturi , R . Mel, A. Marchi , S. Mancuso,,F. Russino , G. Da Pra,N.Papa,G.Bertiato
C.Fiorentini , M.G Ciufolini Journal of Virological Methods 134 (2006) 136–139
Comparison
Comparison of
of the
the results
results obtained
obtained with
with HI
HI and
and PRNT
PRNT tests
tests in
in anti
anti
TBE
TBE positive
positive ELISA
ELISA serum
serum samples
samples of
of the
the 33 study
study groups
groups and
and aa
population
population of
of vaccinated
vaccinated children
children
HI test:
positive/tested (%)
PRNT:
positive/tested (%)
Acute TBEV
infection.
43/43 (100)
30/42 (71.4 )
Healthy population
residing in a TBE
risk area.
24 /27(88.9)
19/23 (82.6)
Wild ruminants
(red deer, roe deer
and chamois) from
a TBEV risk area.
11/18 (61.1)
11/12 (91.6)
36/36 (100)
36/36 (100)
Vaccinated
children (Venturi
et al., 2006).
Sand fly-borne viruses
Sand fly-borne viruses
isolated in Italy
Family/
Genus
Bunyaviridae
Phlebovirus
Rhabdoviridae
Vesiculovirus
Sources of isolation
Virus
Year
Place
Species
Arbia
19811994
Tuscany
Marche
P. perniciosus
P. perfiliewi
Sandfly fever
Naples
1944
Campania
Human
Sandfly fever
Sicilian
1943
Sicily
Human
Toscana
19712002
Radi
19831994
Central and P. perniciosus
Southern
P. perfiliewi
Italy
Human
Tuscany
P. perniciosus
Marche
P. perfiliewi
TOSv: caratteristiche generali
Famiglia
Genere
Trasmissione
Particelle
virali
Proteine
strutturali
Genoma
Bunyaviridae
Phlebovirus
artropodi vettori
(flebotomi)
Sferiche,
80-120 nm
Glicoproteine: G1, G2
Nucleoproteina: N
Ss-RNA,
trisegmentato,
polarità negativa
Struttura del virione
= RNA polimerasi
L
= G1
M
= G2
S
= Nucleocapside
80 – 120 nm
Natural cycle of Toscana virus
Venereal
transmission
Transovarial
transmission
?
?
Monthly distribution, virus isolations in
Fermo “focus” (1989-1993)
No. virus isolation
Capture
month
Sandflies/pools
TOS
(MFIR)*
ARB
(MFIR)
RADI
(MFIR)
Total
(MFIR)
Jun
911/21
6 (0.6)
1 (0.1)
3 (0.3)
10 (1.1)
Jul
3,561/74
24 (0.7)
2 (0.1)
3 (0.1)
29 (0.8)
Aug
4,090/84
18 (0.4)
2 (0.05)
10 (0.2)
30 (0.7)
Sep
1,460/34
8 (0.5)
-
1 (0.1)
9 (0.6)
Total
10,022/213
56 (0.5)
5 (0.05)
17 (0.2)
78 (0.8)
*MFIR=Minimum field infection rate/100 P. perfiliewi tested
Virus isolation by sex (1989-1993)
No. virus isolation
Sex
Sandflies/pools
TOS
(MFIR)*
Females
7,642/155
Males
2,380/58
Total
10,022/213
ARB
(MFIR)
RADI
(MFIR)
Total
(MFIR)
48 (0.6) 4 (0.05) 14 (0.2) 66 (0.9)
8 (0.3)
1 (0.04)
3 (0.1)
12 (0.5)
56 (0.6) 5 (0.05) 17 (0.2) 78 (0.8)
*MFIR=Minimum field infection rate/100 P. perfiliewi tested
N° of patients
Monthly distribution of Toscana
virus infections
140
120
100
80
60
40
20
0
J
F
M
A
M
J
J
A
Month
TOS pos
TOS neg
S
O
N
D
Known distribution of Toscana
virus in Italy
Virus isolation from Phlebotominae Sandflies
Human cases
Clinical Infectious Disease 2001, 32: 1241-3
Eur J Epidemiol. 1997 Oct;13(7):761-4.
Evidence of Toscana virus infections without central
nervous system involvement: a serological study.
Braito A, Corbisiero R, Corradini S, Marchi B, Sancasciani N, Fiorentini
C, Ciufolini MG
Neighbour-joining tree for M genomic segment
nucleotide sequences of TosV strains.
Journal of General Virology (2007), 88, 1288–1294
Imported infections
Dengue and Chikungunya
imported cases
N° of cases
120
100
80
60
40
20
0
1990 1991 1992 1993 1994 1995 1996 1997 1998 1999 2000 2001 2002 2003 2004 2005 2006 2007
DEN
Preg.
Undetermined
CHIK
Negative
Distribuzione di Aedes albopictus,
2007
BZ
CO
SO
PN UD
GO
VA
TS
VI
BI
BS
VE
NO MI
VR
VC
LO
PD
CR MN
TO
PV
AT
PC
AL
FE
PR RE
CN
GE
BO
RA
SV
MO
RN
SPMS
FC
IM
LU PT
FI
PU
PO
AR
AN
PI
MC
SI
LI
PG
AP
GR
TE
TR
PE
VT
RI
CH
AQ
RM
CB
FG
FR IS
LT
CE BN
NA AV
OT
PZ
SS
SA
NU
LC BG
AO
Con l’eccezione della Val
d’Aosta, focolai sporadici
sono presenti in tutte le
Regioni, dal livello del
mare fino a 600 m di
altitudine
BL
OR
MD
TN
TV
BA
MT
CS
CA
KR
CZ
VV
RC
Zone libere
LE
OG
CI
Zone infestate
TA BR
ME
TP
PA
EN CT
AG
CL
SR
RG
And many thanks to
Paola Verani
Marco Balducci
Maria Clotilde Lopes
Loredana Nicoletti
Antonella Marchi
Cristiano Fiorentini
Fabio Magurano
Claudia Fortuna
Eleonora Benedetti
Paola Bucci
Thank you for your attention
