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The laboratory confirmation of Zika virus (ZIKV) infection, and the differential diagnosis from other flavivirus infections such as dengue virus (DENV), often requires the use of several diagnostic test types. Cross-reactions and secondary infections complicate the serological diagnosis and specific viral RNA detection assays are often needed for confirming the diagnosis. The aim of this study was to validate serological and molecular methods for diagnosing ZIKV infection. This included the evaluation of a ZIKV RT-qPCR assay for diagnostics that was previously set up for research use and to compare the ZIKV, DENV and TBEV EIA methods. External and in-house controls and pre-characterized sample panels were tested, and also automated and manual nucleic acid extraction methods were compared. A total of ten Finnish traveler patients were diagnosed with acute ZIKV infection during 2015-2017 including one suspected dual DENV and ZIKV infection. These samples along with panels of DENV and tick-borne encephalitis virus (TBEV) infections were used to test the cross-reactive properties of ZIKV, DENV and TBEV IgM assays. Additionally, the diagnosed acute ZIKV patient samples were tested using commercially available diagnostic DENV NS1 antigen assay and a ZIKV NS1 antigen assay intended for research use. The ZIKV RT-qPCR assay was demonstrated to be both specific and sensitive (one genome per reaction) and suitable for routine diagnostic use utilizing automated nucleic acid extraction. Of the tested IgM tests the NS1 antigen-based ZIKV IgM (Euroimmun) assay performed with least cross-reactivity with a specificity of 97.4%. The DENV IgM assay (Focus Diagnostics) had specificity of only 86.1%. The results are in line with previous studies and additionally highlight that also acute TBEV patients may give a false positive test result in DENV and ZIKV IgM assays., (Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.)

The aim of the study was to develop a real-time RT-PCR for the detection of enteroviruses (EVs) and rhinoviruses (RVs) and to assess the performance of the xTAG RVP Fast assay in comparison to a direct fluorescent assay (DFA), a real-time RT-PCR assay for the detection of respiratory syncytial virus (RSV) and human metapneumovirus (hMPV), and the EV/RV RT-PCR assay developed in this study. The performance of the RVP Fast assay was assessed in the analysis of 373 nasopharyngeal samples. For the viruses of the DFA panel, detection rates of 27.6% and 23.8% were obtained by RVP and DFA, respectively, in analysis of a set of 297 samples collected in 2009-2010. These results show statistically significant superiority of the RVP Fast assay (P=0.049). For RSV, hMPV, EV, and RV, detection rates of 48.0% and 45.2% were achieved by RVP and RT-PCR, respectively. For individual targets, increased detection of EV/RV (P=0.043) and decreased detection of influenza A virus (P=0.004) by RVP in comparison to real-time RT-PCR was observed. The results of the present study imply the need to adjust the InfA component of the RVP Fast assay to also cover the InfA(H1N1) 2009 virus., (Copyright © 2012 Elsevier B.V. All rights reserved.)

A quantitative HHV-6 PCR (qPCR) assay was developed and compared to an "in-house" qualitative PCR and to the commercial quantitative Argene CMV, HHV6, 7, 8 R-gene™ test. Clinical specimens consisting of 127 whole blood and 57 cerebrospinal fluid (CSF) specimens were tested using the two qPCRs and the qualitative PCR in parallel. When the qualitative PCR was used as a "gold standard," the sensitivities of the qPCRs for the blood samples were 86% for the "in-house" qPCR and 76% for the Argene's test and the specificities were 96% and 92%, respectively. With CSF specimens the sensitivities were 92% and 80% and the specificities 98% and 82%, respectively. Furthermore, the two qPCRs were compared in the monitoring of liver transplant patients and retrospectively correlated to HHV-6 antigenaemia. In total, 223 blood specimens were tested. HHV-6 antigenaemia had been found in 21/36 (58%) patients and HHV-6 DNAaemia was demonstrated in 18/36 (50%). Viral loads by the "in-house" test varied from 280 to 19700 copies/ml (median 1200) and by Argene's test from 120 to 24070 copies/ml (median 458). The correlation of viral loads between the two qPCRs was good (R=0.94, p<0.01). The new in-house test was found to be reliable for the detection and quantitation of HHV-6 DNA in clinical specimens., (Copyright © 2012 Elsevier B.V. All rights reserved.)

A novel methodology is introduced for rapid serological diagnosis. This methodology combines the antibody bridging assay principle with the measurement of antibody avidity. The combination allows the determination of the infection phase with a single dilution of a single sample of serum. This is a significant improvement on current serological techniques which often require either paired-sample testing (IgG/IgM serology) or testing of the sample in several dilutions (IgG avidity testing). Assay methods were developed on two immunoassay platforms; the heterogeneous time-resolved fluoroimmunoassay and the separation-free two-photon excitation fluorometry. The new methods were compared to conventional class-specific IgG/IgM and IgG avidity techniques. The major findings were that the avidity results of the new methodology were independent of the sample dilution (specific antibody concentration in serum) and consistent between immunoassay platforms. This new methodology is simple, rapid, and quick to perform. It provides the possibility of running serodiagnostic tests at point-of-care with bench-top random-access analyzers., (Copyright 2010 Elsevier B.V. All rights reserved.)

The seroprevalence of human herpesviruses is high and reactivations occur frequently. A microarray was designed and tested for the detection of IgG and IgM antibodies for Puumala hantavirus (PUUV) and IgG antibodies against four herpesviruses. Initially, a microarray platform was set up using an unrelated in-house antigen, PUUV recombinant nucleocapsid protein, to optimize the protocol for the detection of antibodies. Detection of the four herpesviruses was set up in a microarray using the recombinant proteins of herpes simplex virus (HSV) glycoprotein G1 and G2, varicella-zoster virus (VZV) glycoprotein E, and cytomegalovirus (CMV) pp150 phosphoprotein. The results of the PUUV panel were in good agreement with the PUUV IgG immunofluorescent assay and IgM enzyme immunoassay (EIA). Seropositive and negative clinical reference panels were tested for herpesviruses by the serological microarray, and the results were compared to those of individual EIAs used for standard diagnostic purposes. The serologic microarray for HSV, VZV and CMV antibody detection gave good specificities for IgG. However, sensitivities of the assay varied depending on the herpesvirus detected. The serological microarray showed potential for screening purposes. The microarray based analyses were easy to perform, and HSV-1, HSV-2, VZV, and CMV antibodies could be detected on the same microarray.