Journal of Clinical Virology - Volume 82 - Supplement 1 - September 2016

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S18 Abstracts / Journal of Clinical Virology 82S (2016) S1–S142
such cases, and HPI appear to be an option for treatment of resistant
HSV strains to conventional antivirals (Himaki et al., 2012).
Abstract no: 117
Presentation at ESCV 2016: Oral 32
HIV neutralising antibody delivered by gene
therapy with a stable retroviral vector encoded
in baculovirus expression systems
L. Faqih ∗, P.E. Klapper , T.J. Blanchard ,
E.A. McKenzie Vallely
1 Faculty of Medical and Human Sciences, Institute of
Inflammation and Repair, University of Manchester,
Manchester, United Kingdom
2 Clinical Research Group, Liverpool School of
Tropical Medicine, Liverpool, UK
3 Manchester Institute of Biotechnology, Manchester,
United Kingdom
Introduction: Virus like particles (VLPs) are replication-
incompetent virus shells that represent an intact, non-replicative
virion lacking a genome. They maintain the original antigenic com-
position of the packaging component incorporated into the virion’s
outer membrane. Recently, interest in using VLPs as gene therapy
agents has increased
In this study, we are aiming to develop retrovirus like particles
to serve as a new gene therapy carrier system. Our VLP is of simian
immunodeficiency virus (SIV) origin and to extend the limited cell
tropism inherent in SIV the VLP will be pseudotyped with vesicular
stomatis virus (VSV) glycoprotein. The particlewill deliver IgG1 b12
antibody genes for insertion into the mammalian genome, to pro-
duce long-lasting, high titres of neutralising anti-HIV monoclonal
antibody
Baculoviruses can be used as vehicles to efficiently deliver
and express genes in mammalian cells. BacMam technology uses
a recombinant baculovirus engineered to contain a mammalian
expression cassette for transgene expression in mammalian cells.
The mammalian gene is expressed without baculovirus replication.
VLPs can be produced using this expression system
Methods: Five different target genes have been cloned into five
altered transfer plasmids, to construct five different recombinant
baculoviruses containing the Tat/Gag/Pol genes of SIV, plus the VSV
glycoprotein gene and T7 polymerase. Either CMV or T7-promoters
are driving expression of all genes. Confirmation of cloning was
done by restriction digests using unique restriction enzymes fol-
lowed by sequencing. Recombinant baculoviruses are generated by
homologous recombination between virus baculovirus DNA and
the transfer plasmids. Mammalian cells will be transduced with
recombinant baculoviruses to express proteins of interest. Western
blot and Elisa confirm detection of protein expression.
Results: Thus far, all genes of interest have been cloned success-
fully into a baculovirus transfer vector (pOET6 BacMAM).
Discussion: We believe that the BacMam construct will deliver
SIV genes intomammalian cells and produce SIV like particles psue-
dotypedwithVSVglycoproteins. Psuedotyping the SIV like particles
with VSV-G can eliminate any limitation caused by the use of SIV
envelope genes, by widening cell tropism. Thus IgG1 b12 antibody
genes will be delivered and inserted into the genome of numerous
cell types, to produce long-lasting, high titres of neutralising anti-
HIV monoclonal antibody. Since baculoviruses cannot replicate in
mammalian cells, this system can be used in vivo as well as in vitro.
Reference
[1] C.M. Thompson, et al., Critical assessment of influenza VLP production in Sf9
and HEK293 expression systems, BMC Biotechnol. 15 (2015) 31.
[2] T.A. Kost, J.P. Condreay, D.L. Jarvis, Baculovirus as versatile vectors for protein
expression in insect and mammalian cells, Nat. Biotechnol. 23 (2005) 567–575.
[3] T. Nakajima, K. Nakamaru, E. Ido, Development of novel simian
immunodeficiency virus vectors carrying a dual gene expression system, Hum.
Gene 1874 (2000) 1863–1874.
[4] S. Shukla, C. Schwartz, K. Kapoor, Use of baculovirus BacMam vectors for
expression of ABC drug transporters in mammalian cells, Drug Metab. 304–312
(2012).
Abstract no: 152
Presentation at ESCV 2016: Oral 33
European non-polio enterovirus surveillance
and laboratory detection – Are we prepared to
detect an enterovirus outbreak?
Heli Harvala ∗, Aftab Jasir , Pasi Penttinen ,
Lucia Pastore Celantano , Donato Greco ,
Eeva Broberg
1 Public Health Agency of Sweden and ECDC, Sweden
2 European Centre for Disease Prevention and
Control (ECDC), Sweden
Background: Enteroviruses (EVs) are known to cause large and
severe outbreaks, as recently demonstrated by EV-D68 in USA
and Europe. Another type, EV-A71, is also known for its ability
to cause geographically widespread and clinically significant hand,
foot, and mouth disease (HFMD) outbreaks. Although EV-A71 out-
breaks have beenmostly described in Asia so far, the virus is already
known to circulate in Europe and has been occasionally linked to
the fatal outcomes. We have evaluated the European prepared-
ness for detection and characterisation of non-polio EVs in order
to improve our response for (re)-emergencing EVs linked to severe
disease.
Methods: An on-line survey on non-polio enterovirus surveil-
lance and enterovirus typing/characaterisation was submitted to
all EU/EEA Member States (MS) national coordinating competent
bodies.
Results: A total of 29 MS from 30responded to the survey.
Twenty-seven countries conduct non-polio enterovirus surveil-
lance based on reporting of enteroviruses detected from clinical
specimens, two of them without further characterisation of EV-
positive samples. Almost all countries include typing of EV-positive
samples obtained from individuals with neurological infections
(n = 24) in their EV surveillance, whereas HFMD and respiratory
infections are included in the EV surveillance less frequently (n = 18
and 16, respectively). Three countries have also initiated specific
surveillance for HFMD, and eleven for EV-D68. EV-D68 surveillance
has been established via sentinel influenza surveillance ( n = 7), by
typing EV-positive respiratory samples (n = 10) and/or via acute
flaccid paralysis (AFP) surveillance (n = 5). Virus isolation is per-
formed in all except one country, whereas molecular methods
are used by all. Non-polio enterovirus typing is performed in 26
MS; ten MS type/characterise only culture-positive EV isolates,
whereas the remaining MS subject also PCR-positive samples to
typing/characterisation. Nineteen MS have introduced sequencing
based EV typing, whereas neutralisation assay is used by 16 MS
and seven of them rely entirely on it. Over 5000 EV-positive spec-
imens were successfully characterised/typed in the EU/EEA region
in 2015. The estimated number of typed EV specimens was <50 in
eleven countries (including all seven countries that type by neutral-
isation assay only), whereas sixMS had successfully typed over 300