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

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S28 Abstracts / Journal of Clinical Virology 82S (2016) S1–S142
cal practice by different groups is to be compared and best practice
identified. A programme of standardisation of antibody measure-
ment is needed for a wider range of viruses.
Abstract no: 138
Presentation at ESCV 2016: Poster 14
External in-run controls for gastro-intestinal,
respiratory infections and Zika virus that will
improve assay standardisation
Sarah L. Kempster ∗, Cristina Santirso-Margaretto,
Kathryn Doris, Neil Almond, Rob Anderson
NIBSC, United Kingdom
Nucleic acid amplification is commonly used for rapid pathogen
identification for disease diagnosis and has traditionally identified
a single pathogen in a single sample. The recent development of
syndromic panels that can identify multiple viruses, bacteria and
parasites in a single sample has resulted in increased efficiency
and also time and cost savings to the diagnostic laboratory. Whilst
considerable effort is taken to validate these assays, effective stan-
dardisation is seldom undertaken that would assure the quality of
data as it is generated over time and between different laboratories.
Commercially available and in house qPCR assays vary both
within and between laboratories in their content and sensitivity
due to varying extraction platforms, qPCR design, qPCR reagents
andusers.Many positive controls currently inuse consist of purified
plasmid DNA. This has its limitations as the construct or the prepa-
ration differs between laboratories and furthermore these plasmids
do not mimic a clinical sample that has undergone extraction.
At NIBSC we have developed a multiplex in run control that
contains a total of 20 viruses, bacteria and parasites to facili-
tate standardisation of assays for gastro-intestinal infections and
a second targeting respiratory pathogens containing 15 target
organisms and viruses. The pathogens were selected following
consultation with multiple laboratories. The pathogens are mixed
and freeze dried in a universal buffer that can be reconstituted
in a matrix compatible with the end users requirements and
extracted alongside the clinical samples allowing for standard-
isation of the extraction procedure as well as the qPCR. Greater
nucleic acid stability was obtained by heating bacteria at 99 ◦ C
for 1 h when compared to ethanol treatment. The freeze drying
excipient concentrations (mannitol and trehalose) were also opti-
mised to maintain cake structure, pathogen stability and ensure
commutability.
As part of NIBSC’s response to the Zika virus outbreak in South
America, we have developed a Zika virus reference in plasma that
is being calibrated against the candidate International Standard
and external in-run controls that can be reconstituted in a suitable
matrix and run alongside clinical samples. These materials support
monitoring of intra-laboratory variation on a daily basis and also
allow standardisation of laboratory measurements of virus load.
The availability of these new external in-run controls will con-
tribute to effective standardisation of diagnostic assays and reduce
both intra- and inter laboratory variability of reported results.
Abstract no: 144
Presentation at ESCV 2016: Poster 15
CXCL13 in patients with facial palsy caused by
varicella zoster virus and Borrelia burgdorferi:
A comparative study
Johan Lindström ∗, Daniel Bremell ,
Henrik Zetterberg , Anna Grahn ,
Marie Studahl
1 Department of Infectious Diseases, Institute of
Biomedicine, Gothenburg, Sweden
2 Department of Psychiatry and Neurochemistry,
Sahlgrenska Academy, University of Gothenburg,
Gothenburg, Sweden
3 Clinical Neurochemistry Laboratory, Sahlgrenska
University Hospital, Mölndal, Sweden
4 Department of Molecular Neuroscience, UCL
Institute of Neurology, UK
In an effort to improve diagnostics in central nervous system
(CNS) infections, the chemokine CXCL13 has emerged as a possible
diagnosticmarker of Lyme neuroborreliosis (LNB). Whenmeasured
in the cerebrospinal fluid (CSF), CXCL13 has shown to be signifi-
cantly higher in patients with LNB compared to several other CNS
infections, with the exception of cryptococcosis and neurosyphilis.
Several such studies have used receiver operating characteristic
(ROC) analyses, yielding a variety of suggested cut-off levels for CSF
CXCL13, ranging from 61 pg/mL to 1224 pg/mL. However, patients
included in previous studies presented with a variety of clinical
syndromes, which raises questions on comparability. Additionally,
there is no accepted reference method for CXCL13, so there may
also be method-related explanations for the varying cut-offs. Facial
palsy is a common manifestation of LNB, but can also be caused by
varicella zoster virus (VZV) reactivation, traditionally named Ram-
say Hunt syndrome (RHS). Improved diagnostics, such as VZV PCR
of patient CSF, allows patients with VZV facial palsy a definite diag-
nose regardless of the presence of blisters associated with classical
RHS. A comparison of CXCL13 in such similar patient groups has so
far not been done.
28 patients with VZV facial palsy, diagnosed by detection of
VZV DNA in CSF by PCR, were retrospectively identified. A total
of 21 patients with facial palsy caused by LNB were included from
two patient cohorts previously included in unrelated prospective
studies on LNB. The median number of days between debut of
facial palsy and CSF sampling was 2 (range (−)9 to 10) for VZV
patients and 4 (range 1–35) for LNB patients. A control group
with 52 patients without CNS infection was included. CXCL13 was
measured in stored CSF samples by ELISA (R&D Systems), with a
detection limit of 7.8 pg/mL.
Median CSF concentrations of CXCL13 for facial palsy caused
by LNB were 1808 pg/mL (range 15–36,924), for VZV facial palsy
9 pg/mL (range <7.8 to 437); all control samples but one were
below the detection limit. The differences in CXCL13 concentra-
tions between patients with LNB facial palsy and VZV facial palsy
were highly significant (p < 0.0001). ROC analysis-derived cut-off
level of 34.5 pg/mL yielded a sensitivity of 82.6% and a specificity
of 82.1%.
In this first comparative study on CXCL13 in patients with facial
palsy caused by LNB and VZV, we can confirm significantly higher
concentrations of CXCL13 in CSF of patients with LNB compared to
patients with VZV. However, the previously proposed cut-off lev-
els for CXCL13 would lead to unacceptably low sensitivity in our
material. A cut-off at 61 pg/mL corresponds to a sensitivity of 73.9%,
dropping to as low as 56% with the proposed cut-off at 1224 pg/mL.
Although more than half of the patients with VZV facial palsy had