Anti-BK virus neutralizing antibody titers before transplantation predict BK virus replication in kidney transplant recipients after transplantation

Anti-BK virus neutralizing antibody titers before transplantation predict BK virus replication in kidney transplant recipients after transplantation

S6 Abstracts / Journal of Clinical Virology 82S (2016) S1–S142 (MR) was introduced in order to analyse the qPCR data without input from the operator...

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Abstracts / Journal of Clinical Virology 82S (2016) S1–S142

(MR) was introduced in order to analyse the qPCR data without input from the operator, accounting at the same time for suboptimal reactions. In the present work, we modified MR to filter out results inconsistent with positive reactions using an assumptionfree approach. We applied this novel algorithm of qPCR analysis to several primer sets targeting a plethora of viruses in order to assess its effectiveness with respect to the CT. Methods: Clinical samples (n = 328) were obtained from residual faecal specimens processed by the Clinical Microbiology and Public Health Laboratory at Addenbrooke’s Hospital (Cambridge, UK). The samples were extracted by QIAsymphony SP and amplified on Custom TaqMan Array 384-well Card by TaqMan Fast Virus 1-Step Master Mix 2× on Viia7 thermalcyclers. The results were issued as either positive or negative by three operators and a consensus classification was generated. A training dataset of 1920 reactions was obtained from a pool of 54 primer sets performed over 50 plates. The resulting MR data were analysed by EM algorithm to obtain a cut-off for the positive/negative results. This filtered MR was then applied to 23 primer sets targeting different viruses for a total of 6038 reactions. MR values below the empirical cut-off were considered negative and the consensus classification was used to assess the accuracy of detection in comparison to CT. Results: Five of the 23 primer set analysed (21.74%) showed a better accuracy and negative predictive value using the MR rather than the CT method, both being in average 0.987 ± 0.013 and 0.996 ± 0.006 for the CT and MR, respectively. The clinical sensitivity for four of these primer sets was in average 0.500 ± 0.136 and 0.764 ± 0.274 for the CT and MR, respectively; for the single primer set where this parameter could not be computed, CT and MR showed ten and none false negative reactions, respectively. In all other instances, CT and MR performed equally. Discussion: The data gathered suggested that MR is a competitive analytical algorithm for qPCR analysis, providing higher accuracy than CT in one fifth of the targets tested while being comparable to CT in all other cases. MR also had the advantage over CT of (a) being assumption-free and (b) taking into account primer specific inhibitions. The use of MR can be beneficial for several qPCR applications by increasing the effectiveness and reproducibility of the assay. MR can also assist the operators during the visual inspection of the individual reactions by highlighting problems in the amplification.

Abstract no: 202 Presentation at ESCV 2016: Oral 11 Anti-BK virus neutralizing antibody titers before transplantation predict BK virus replication in kidney transplant recipients after transplantation M. Solis 1,2,∗ , A. Velay 1,2 , R. Porcher 3 , P. Domingo-Calap 2 , E. Soulier 1,2 , M. Joly 2,4 , M. Meddeb 1 , W. Kack-Kack 1,2 , B. Moulin 2,4 , S. Bahram 2 , F. Stoll-Keller 1,2 , H. Barth 1,2 , S. Caillard 2,4 , S. Fafi-Kremer 1,2 1 Laboratoire de Virologie, Hôpitaux Universitaires de Strasbourg, Strasbourg, France 2 Inserm UMR S1109, LabEx Transplantex, Fédération de Médecine Translationnelle de Strasbourg (FMTS), Université de Strasbourg, Strasbourg, France 3 Centre d’Epidémiologie Clinique (CRESS), UMR1153, Université Paris Descartes, Paris, France 4 Département de Néphrologie – Transplantation, Hôpitaux Universitaires de Strasbourg, Strasbourg, France

BK virus-associated nephropathy (BKVN) is the most frequent BKV-associated disease after renal transplantation, with BKV reactivation occurring in up to 80% of kidney transplant recipients (KTR). Virological diagnosis of BKVN relies on the detection and quantification of viral load in urine and blood by real-time PCR techniques, allowing preemptive immunosuppressive therapy adaptation. However, the delayed nature and incomplete success of this preemptive strategy underscore the need for prognostic markers of BKV reactivation. Neutralizing antibodies (Nabs) against BKV genotypes were analyzed in a prospective KTR cohort to investigate whether Nabs titers may predict BKV replication. Blood and urine samples were prospectively collected from 168 KTR the day of transplantation, weekly the first month post-transplantation then monthly during 96 weeks. Using the BKV pseudovirus system (Pastrana et al., J Virol 2013), anti-BKV Nabs titers were measured on the day of transplantation and at additional time points post-transplantation. BKV DNA load was quantified in urine and blood samples using a commercial qPCR kit (BK virus R-gene® , Biomérieux, France). BKV strains of KTR displaying viruria and/or viremia were genotyped as previously described (Solis et al, JCM 2016). Anti-BKV Nabs were positive in 164 (97.6%) patients before transplantation. Hundredten (67.1%) KTR harbored higher Nabs titers against genotype I, while 16 (9.8%) and 7 (4.3%) KTR showed higher Nabs titers against genotype II and genotype IV, respectively. Twenty eight KTR harbored higher titers for two genotypes (17 for genotype I and II, 5 for genotype I and IV and 6 for genotype II and IV). Three harbored similar Nabs titers against the 3 genotypes. BKV viruria was detected in 52 (31%) patients 1 to 78 weeks (median 5 weeks) after transplantation. BKV viremia was observed in 28 (16.7%) patients 5–75 weeks (median 18 weeks) after transplantation, among them 13 (7.7%) developed BKVN 10–76 weeks (median 17 weeks) after transplantation. In BKV-replicating KTR, BKV genotype I, genotype II and genotype IV were identified in 45 (86.5%), 1 (1.9%) and 6 (11.5%) patients, respectively. The risk of developing viruria was higher for patients with lower Nabs titers before transplantation against their subsequently-replicating genotype (HR (95% CI) = 0.44 (0.25–0.76; p = 0.003). The replicating BKV is acknowledged to be of donor origin. Indeed, donor/recipient mismatches in regard to genotypic neutralization profiles and replicating strains were found to be greater in BKV-replicating KTR (p < 0.05). Anti-BKV Nabs titer before transplantation may represent a valuable prognostic marker of BKV replication after

Abstracts / Journal of Clinical Virology 82S (2016) S1–S142

transplantation. Determination of anti-BKV Nabs titer in donors and recipients before transplantation may allow for better-suited induction and maintenance immunosuppressive therapy as well as adapted viral monitoring. Abstract no: 341 Presentation at ESCV 2016: Oral 12 A longitudinal study on dynamics of plasma neutralising antibodies and its determinants in HIV-2 infected individuals G. Ozkaya Sahin 1,∗ , S. Karlson 2 , F. Mansson 3 , A. Biague 4 , H. Norrgren 5 , M. Jansson 2 1 Clinical Microbiology, Laboratory Medicine Skane, Lund, Department of Laboratory Medicine, Lund University, Lund, Sweden 2 Department of Laboratory Medicine, Lund University, Lund, Sweden 3 Department of Clinical Sciences, Lund University, Malmo, Lund, Sweden 4 National Public Health Laboratory, Bissau, Guinea-Bissau 5 Division of Infection Medicine, Department of Clinical Sciences, Lund University, Lund, Sweden

Background: Majority of HIV-2-infected individuals survive as elite controllers. Therefore, HIV-2 infection represents a model for the studies of immune responses that may control an HIV infection, and possibly give leads towards a functional cure. Plasma neutralising antibodies (NAb) are thought to play a central role in HIV-2 evolution and pathogenesis. However, due to relatively silent disease course, it has been almost impossible to diagnose HIV-2 seroconversion time, to follow-up the natural history of infection and to investigate the dynamics of the NAb response. Research group in Sweden and Guinea-Bissau has been organised to investigate the long-term epidemiological trends of HIV-2 infection since 1987. Questions: When does broad and potent NAb response develop in HIV-2 infected individuals? What are the modulators of broad and potent NAb response? Materials and methods: Forty-six plasma with known T cell count were obtained from 15 individuals from a cohort of police officers in Guinea-Bissau, between 1992-2010. Participants were classified into two subgroups based on mean CD4+ T cell count/␮l: Immunocompetent group with cell count ≥500 vs immunosuppressed group with cell count <500 for the neutralization assay ghost 3-ccr5 cell line heat-inactivated plasma and five hiv-2 isolates originating from west Africa were used cut-off point was 30. Results: Immunocompetent individuals were HIV-2 seroconverted at an earlier age and displayed higher plasma CD8+ T cell count/␮l compared to immunodeficient group (median age, 28 years vs 38 years, respectively, p < 0.05; median CD8+ T cell count/␮l, 755 vs 334, respectively, p < 0.01). In all participants, NAb response was found to be potent and broad already during the first year of infection. Moreover, this response persisted throughout the whole follow-up period. Interestingly, at the end of follow-up period, NAb response was significantly broader and more potent in the immunocompetent group compared to immunodeficient group (breadth 4.3 vs 2.9, p < 0.05; potency 200000 vs 25000, respectively, p < 0.05). In both groups, age at seroconversion correlated negatively with CD4+ and CD8+ T cell count (r = −0.64 and −0.41, respectively, p < 0.05). In the immunocompromised group, CD4+ and CD8+ T cell counts tended to decrease with infection duration (Spearman’s r: −0.62 and −0.88, respectively, p < 0.05).


Interestingly, decreasing number of cellular immunity cells correlated negatively with potency of NAb response (r: −0.55 and −0.78, p < 0.05). In the immunocompetent group, both breadth and potency of NAb response tended to increase with infection duration (r: 0.53 and 0.51, respectively, p < 0.05). Furthermore, potency of NAb response correlated positively with CD4+ T cell count (r: 0.72, p < 0.05). Discussion and significance: This study represents the most diverse longitudinal primary infection cohort studied to date for HIV-2 neutralization. Broadly p-NAb response in HIV-1 infection arises only in around 15% of patients after 2-4 years of infection. In addition, p-NAb response tends to fluctuate with low potency. Contrarily, here we show that broad and potent p-NAb response develops in all HIV-2 infected participants during the first year of infection and tends to be persistent. These results may provide insights into the role of potently persistent neutralizing humoral immune response on mild outcome of HIV-2 infection. Abstract no: 108 Presentation at ESCV 2016: Oral 13 Implementation of a rapid HIV-1 RNA test in diagnosing acute HIV infections among visitors of the Amsterdam clinic of sexually transmitted infections M. Dijkstra 1,∗ , S. Bruisten 1 , E. Hoornenborg 1 , A. Hogewoning 1 , H. de Vries 1 , M. Schim van der Loeff 1 , B. Berretty 2 , I. Linde 1 , K. Adams 1 , U. Davidivich 1 , G. de Bree 3 , on behalf of the H-TEAM initiative 1

Public Health Service of Amsterdam, The Netherlands 2 Amsterdam Institute for Global Health and Development, The Netherlands 3 Academic Medical Center, Amsterdam, The Netherlands Background: Immediate diagnosis and treatment of acute HIV infection (AHI) is important both from a patient and a public health perspective. Firstly, it can prevent progression to chronic symptomatic HIV disease and thereby improve an individual’s prognosis. Secondly, it can reduce the risk of onward transmission associated with AHI in individuals unaware of being infected and usually having high viral loads. At the sexually transmitted infections (STI) outpatient clinic in Amsterdam, the diagnosis of AHI relied on the routine use of serological antibody and antigen assays. These assays have a window phase of at least 15 days between infection and seroconversion. A new promising avenue is the incorporation of a rapid HIV-RNA test that shortens this period with around 5 days. As part of the HIV-Transmission Elimination AMsterdam (HTEAM) initiative a rapid AHI diagnosis and referral trajectory was implemented at the STI clinic in Amsterdam in 2015. This involved the addition of a rapid HIV-RNA assay to standard HIV testing among men who have sex with men (MSM). We now present our first experiences with this new rapid AHI test and referral trajectory. Methods: MSM were assessed for eligibility at the STI clinic for an AHI test. They were either referred by a media campaign (, with a self-referral screening tool), or by their general practitioner (GP), or if they came for routine STI screening. Eligibility was based on symptoms of AHI in combination with condom-less anal sex with a man within 2 weeks to 3 months preceding the visit. Participants completed questionnaires and pro-