French Polynesia in 2013 [5] was followed by the rapid spread of
the virus to South, Central and North America where transmission
was associated with millions of cases of infection [6]. The majority
of cases of ZIKV infection are asymptomatic, but infection can be
associated with a range of manifestations including rash, fever,
headache, and muscle and joint pain [7]. While ZIKV infection is
normally self-limiting and is often resolved without medical intervention, in some cases more serious consequences can arise, including Guillain–Barre syndrome in adults and major birth defects in
the fetuses of woman infected while pregnant [8]. The significant
effects of ZIKV on fetuses of women who become infected while
pregnant is perhaps the single most important public health aspect
of ZIKV infection and will shape future vaccine development for
this virus.
Vaccine development to protect against infection with members of the genus Flavivirus has been uneven. While there are
excellent vaccines against yellow fever virus [9] and Japanese
encephalitis virus [10], vaccine development against dengue virus
has been much slower and recently the first commercial dengue
vaccine (Dengvaxia) has been associated with adverse effects in
certain vaccinees [11]. A number of approaches toward vaccine
development are available. The most successful approaches to date
have been the development of live attenuated vaccines, as used for
yellow fever virus and Japanese encephalitis virus vaccines and
whole inactivated viruses as used previously in some Japanese
encephalitis vaccines [10]. However, both of these approaches
have some drawbacks, including potential reversion to wild type
for live attenuated viruses and underinactivation for whole inactivated virus vaccines. In particular, neither type of vaccine approach
is desired for use with ZIKV given the association with neonatal
birth defects. Virus-like particles (VLP) are an attractive approach
to vaccine development for Flaviviruses [12]. VLP consist of the
structural proteins of the virus in a native form but without the
associated genetic element. The lack of a genetic element makes
VLP a safer choice than either live attenuated or inactivated vaccines. This protocol can be used to prepare laboratory scale preparations of purified ZIKV VLP for vaccine development through
expression of the VLP in a mammalian cell culture system and
subsequent purification through buoyant density centrifugation.
2 Materials
2.1 Zika VLP Plasmid
Construction
and Preparation
1. The sequence of capsid (C), premembrane/membrane (prM/
M), and envelope (E) genes of a Cambodian isolate of Zika virus
(isolate FSS13025) Genbank number AFD30972.1 was taken
from the NCBI database and commercially synthesized (see
Note 1).
184
Atichat Kuadkitkan et al.
the virus to South, Central and North America where transmission
was associated with millions of cases of infection [6]. The majority
of cases of ZIKV infection are asymptomatic, but infection can be
associated with a range of manifestations including rash, fever,
headache, and muscle and joint pain [7]. While ZIKV infection is
normally self-limiting and is often resolved without medical intervention, in some cases more serious consequences can arise, including Guillain–Barre syndrome in adults and major birth defects in
the fetuses of woman infected while pregnant [8]. The significant
effects of ZIKV on fetuses of women who become infected while
pregnant is perhaps the single most important public health aspect
of ZIKV infection and will shape future vaccine development for
this virus.
Vaccine development to protect against infection with members of the genus Flavivirus has been uneven. While there are
excellent vaccines against yellow fever virus [9] and Japanese
encephalitis virus [10], vaccine development against dengue virus
has been much slower and recently the first commercial dengue
vaccine (Dengvaxia) has been associated with adverse effects in
certain vaccinees [11]. A number of approaches toward vaccine
development are available. The most successful approaches to date
have been the development of live attenuated vaccines, as used for
yellow fever virus and Japanese encephalitis virus vaccines and
whole inactivated viruses as used previously in some Japanese
encephalitis vaccines [10]. However, both of these approaches
have some drawbacks, including potential reversion to wild type
for live attenuated viruses and underinactivation for whole inactivated virus vaccines. In particular, neither type of vaccine approach
is desired for use with ZIKV given the association with neonatal
birth defects. Virus-like particles (VLP) are an attractive approach
to vaccine development for Flaviviruses [12]. VLP consist of the
structural proteins of the virus in a native form but without the
associated genetic element. The lack of a genetic element makes
VLP a safer choice than either live attenuated or inactivated vaccines. This protocol can be used to prepare laboratory scale preparations of purified ZIKV VLP for vaccine development through
expression of the VLP in a mammalian cell culture system and
subsequent purification through buoyant density centrifugation.
2 Materials
2.1 Zika VLP Plasmid
Construction
and Preparation
1. The sequence of capsid (C), premembrane/membrane (prM/
M), and envelope (E) genes of a Cambodian isolate of Zika virus
(isolate FSS13025) Genbank number AFD30972.1 was taken
from the NCBI database and commercially synthesized (see
Note 1).
184
Atichat Kuadkitkan et al.