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

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Abstracts / Journal of Clinical Virology 82S (2016) S1–S142 S67
Abstract no: 151
Presentation at ESCV 2016: Poster 92
Memory T cells specific for HBV enumerated by
a peptide-based cultured enzyme-linked
immunospot assay in healthy HBV-vaccinated
subjects
I. Cassaniti ∗, S.A. Calarota ,
K.M.G. Adzasehoun , A. Chiesa , G. Comolli ,
M. Parea , F. Baldanti
1 Molecular Virology Unit, Microbiology and Virology
Department, Fondazione IRCCS Policlinico San
Matteo, Via Taramelli 5, 27100 Pavia, Italy
2 Experimental Research Laboratories, Biotechnology
Area, Fondazione IRCCS Policlinico San Matteo, Viale
Golgi 19, 2, Italy
3 Department of Clinical, Surgical, Diagnostic and
Pediatric Sciences, University of Pavia, 27100 Pavia,
Italy
Background: Hepatitis B vaccine is the most effective strategy
to control hepatitis B virus (HBV) infection and disease. It is consid-
ered that an anti-HBs (antibodies against HBV surface antigen) titer
>10mIU/ml, measured shortly after a complete vaccination sched-
ule, provides protection against infection. Approximately 4–10% of
healthy individuals fail to respond to three doses of vaccine. Addi-
tionally, an estimated 13–60% of initial responders to HBV vaccine
(>10mIU/ml after three doses)may lose anti-HBs after several years
post-vaccination. The aim of this study was to evaluate the long-
term HBV-specific memory T-cell response in healthy vaccinated
subjects.
Materials and methods: We quantified HBV-specific expand-
able memory T cells by using a peptide-based cultured IFN-gamma
enzyme-linked immunospot following 10 days stimulation (cul-
tured ELISPOT). Response to an overlapping peptide pool (15-mers
overlapping by 11 amino acids) representing the complete L (large)
HBV envelope polypeptide was evaluated in 46 healthy subjects
(mean age of 36.24 years, standard deviation (SD) = 10.48; 12males
and 34 females). Forty-one subjects (89.1%) were vaccinated for
HBV about 15–20 years earlier. Plasma samples were tested for
anti-HBs.
Results: We observed that vaccinated subjects had signifi-
cantly higher HBV-specific T-cellular response than unvaccinated
(p = 0.0002). HBV-specific memory T-cell response quantified by
cultured ELISPOT was mainly mediated by CD4+ T cells. No con-
cordance was found between cultured ELISPOT and anti-HBs data
in vaccinated subjects. Thirty-one (76%) vaccinated subjects were
responders (anti-HBs >10mIU/ml), while 10 (34%) were non-
responders (anti-HBs <10mIU/mL). Nineteen (46%) vaccinated
subjects were considered to be responders in the HBV-specific cul-
tured ELISPOT assay. Twenty-two (54%) vaccinated subjects were
considered non-responders in the HBV-specific cultured ELISPOT;
five of them (23%) were also humoral non-responders. About 12%
of healthy HBV-vaccinated subjects were both humoral and cel-
lular non-responders. Thus, these subjects may be at risk for HBV
infection and disease, especially health care workers.
Conclusion: In conclusion, the evaluation of HBV-specific T-cell
response by cultured ELISPOT may represent a new tool to mon-
itor memory immunity to HBV vaccine in immunocompromised
patients, such as hemodialyzedpatients or patientswho underwent
solid organ transplantation, that are at high risk for infection.
Abstract no: 159
Presentation at ESCV 2016: Poster 93
Impact of the genotyping method on the
distribution of hepatitis C virus subtypes of
genotype 1
C. Van den Borre ∗, W. Schuermans ,
H. Van Vlierberghe , A. Geerts , X. Verhelst ,
E. Padalko
1 Department of Clinical Chemistry, Microbiology
and Immunology, Ghent University and Hospital,
Ghent, Belgium
2 Department of Gastroenterology and Hepatology,
Ghent University and Hospital, Ghent, Belgium
Background:At least 6 major hepatitis C virus (HCV) genotypes
(1–6) andmultiple subtypes (e.g. a, b, c) have been identified, based
on the sequence differences. Distribution of certain genotypes in
the patient population is highly dependent on geographical region
but also differs by gender, ethnicity, age and mode of transmis-
sion. Differences in the patient population regarding subtypes of
HCV genotype 1 have also been reported: e.g. patients older than
65 years are more likely and injecting drug users are less likely to
harbour subtype 1b than 1a. In the present study we investigated
the potential impact of the change in the laboratory method used
for HCV genotyping on the subtype distribution of HCV genotype
1.
Materials and methods: Samples submitted for HCV geno-
typing at the Ghent University Hospital from January 2002 until
December 2014 were included in the current study, based on the
retrieval of results through the laboratory information system. HCV
genotyping results during this period of 12 years were obtained by
the reverse hybridization based Versant HCV Genotype Assay but 2
different versions of the same assay were used: from January 2002
till March 2007 Versant HCV Genotype 1.0 was used and from April
2007 till December 2014 Versant HCV Genotype 2.0. The improve-
ment of the second generation assay, including the core region
probes, lies mainly in the distinction between subtype a and sub-
type b of genotype 1. Therefore, we focused on the possible changes
in distribution for subtypes of genotype 1.
Results: From January 2002 till December 2014, 1631 serum
samples were determined positive for HCV genotype 1. For 853
genotype 1 positive samples analyzed with Versant HCV Genotype
1.0 from January 2002 till March 2007, 23 (2.7%) samples were
attributed to subtype 1a; 716 (83.9%) samples to 1b and 114 (13.4%)
genotype 1 positive samples could not be subtyped further. For 778
genotype 1 positive samples analyzed with Versant HCV Genotype
2.0 from April 2007 till December 2014, 271 (34.8%) samples were
attributed to subtype 1a; 493 (63.4%) samples to 1b and only 14
(1.8%) genotype 1 positive samples could not be subtyped further
with this advanced version of the same assay.
Conclusions: In the current study we demonstrate that the
switch of the version of the reverse hybridization based Versant
HCV Genotype Assay had a major impact on the distribution of sub-
types of HCV genotype 1 bringing subtype 1a from less than 3% of
genotype 1 samples in the first version – based only on the analysis
on 5 -UTR – to more than one third of the genotype 1 samples, as
analyzed by the advanced version of the same assay where core-
region analysis was added. The number of non-subtyped genotype
1 samples has been reduced by version switch frommore than 13%
to less than 2%. Evaluation of the potential impact of other factors
that can influence change in distribution of subtypes of HCV geno-
type 1 as age of the patients and the mode of transmission still has
to be performed. However, the current use of the improved ver-