[Retrospective analysis of the performance of the SARS-CoV-2 rapid antigen detection test compared to the reference RT-PCR test]. / Analyse rétrospective des performances du test de détection rapide antigénique du SARS-CoV-2 comparé au test de référence RT-PCR.

BACKGROUND: Discovered in 2019 in the region of Wuhan, China, the Severe Acute Respiratory Syndrome coronavirus 2 (SARS-CoV-2) rapidly established itself as a major pathogenic agent of morbidity and mortality. France has implemented a strategy to fight this virus which relies essentially on widespread RT-PCR virological testing in order to isolate positive patients. Antigenic tests have recently been made available to help the diagnostics. We have conducted a retrospective study to determine the sensitivity of these antigenic tests, comparing them to the reference RT-PCR method. METHOD: Between December 7, 2020 and January 31, 2021, each patient we received in our laboratories for an RT-PCR test was enrolled. Out of 271,649 patients, 4,881 had been submitted to an antigenic test (TDR) in the preceding 24 hours. Comparing the data resulting from both tests, we established the sensitivity and the specificity of the antigenic tests. For our analysis we included the parameter of symptom and/or the value of Cycles threshold (Ct) in our parameters. RESULTS: The sensitivity of the TDRs compared to all the positive RT-PCR tests is 56%. We further demonstrate the correlation between the symptom duration and the reduction of the nasopharyngeal viral load. Based on this data, we have established that the sensitivity of the TDRs decreases very rapidly after symptom onset, contrary to the estimated viral load in the RT-PCR. Indeed, less the 24 hours after clinical symptom onset, the sensitivity of the TDRs decreases from 74% to 60%. By including the Ct value in our parameters, we have established that, despite a high viral load and clinical symptoms since 7 days or less, the sensitivity of the TDRs is 66%. Although, a high number of asymptomatic patients among carriers of SARS-CoV-2, we have estimated a specificity of 93% for our test. CONCLUSIONS: Performance in terms of sensitivity and specificity of the TDR, as assessed in practice, are inferior to those given by the manufacturer, which raises several questions. What is the impact of falsely negative results for patients carrying a high viral load? Are the implemented measures sufficient to prevent the epidemic?

Microbiological Outcomes Associated With Low Leukocyte Counts in Cerebrospinal Fluid.

The significance of low leukocyte counts in cerebrospinal fluid (CSF) remains unclear. We performed a 2-year retrospective study to examine microbiological outcomes associated with CSF leukocytes at 6-10/mm3. Of the 178 samples examined, we detected positive results for 11 samples, including 5 cases of tick-borne encephalitis virus infection.

Immunoserological Diagnosis of Human Borrelioses: Current Knowledge and Perspectives.

Spirochetes of the genus Borrelia are divided into relapsing fever borreliae and Lyme disease borreliae. Immunoserological assays have been poorly developed for relapsing fever borreliae, where direct detection methods are more adapted to the pathophysiology of these infections presenting with massive bacteraemia. However, emergence of the novel agent of relapsing fever B. miyamotoi has renewed interest in serology in this context. In Lyme disease, because direct detection methods show low sensitivity, serology plays a central role in the diagnostic strategy. This diagnostic strategy is based on a two-tier methodology involving a first test (ELISA) with high sensitivity and acceptable specificity and a second, more specific test (western blot) for diagnostic confirmation. The most frequent limitations and pitfalls of serology are cross reactions, false IgM positivity, a seronegative window period at the early time of the infection, and serologic scars with a suspicion of reinfection. International guidelines have thus been proposed to avoid these difficulties with interpretation. Finally, unconventional diagnostic tests have been developed recently in the context of a highly publicized disease, with widely varying results, some of which have no available evidence-based data. New two-tier testing strategies using two ELISA tests (C6 and WCS for example) to replace immunoblot are currently proposed by some authors and guidelines, and promising new tests such as CXCL-13 in CSF are promising tools for the improvement of the diagnosis of Lyme borreliosis.

Evaluation of three commercially available hepatitis C virus antibody detection assays under the conditions of a clinical virology laboratory.

BACKGROUND: Most studies evaluating antibody detection assays are conducted on samples from healthy blood donors but not on samples of hospitalized patients which can show non-specific reactions. OBJECTIVES: To compare the performance of three commercial automated assays for the detection of hepatitis C virus (HCV) antibodies, Monolisa anti-HCV Plus version 2, Axsym anti-HCV 3.0 and Vitros anti-HCV, on a population of hospitalized patients. STUDY DESIGN: The specificity of the assays was prospectively evaluated in 2020 routine serum samples. In order to assign the serostatus of each sample, those giving positive or discordant results were further tested by three immunoblots and by RT-PCR (Roche). Moreover, the sensitivity was evaluated on eight commercial HCV seroconversion panels. RESULTS: The Monolisa, Axsym and Vitros assays showed specificities of 99.64%, 99.12% and 99.33%, respectively. Concerning the sensitivity, among 49 samples, the number of positive results was 21, 24 and 24 for the Monolisa, Axsym and Vitros kits, respectively. The differences were not statistically significant at an alpha risk of 5%. CONCLUSIONS: All assays appeared to be reliable for routine screening, but there were a surprising number of indeterminate samples that could not be resolved by confirmatory tests.

Development and laboratory evaluation of a new recombinant ELISA for the serodiagnosis of Lyme disease.

OBJECTIVE: The use of recombinant proteins for serologic testing represents a modern approach for the improved laboratory diagnosis of Lyme disease (LD). The aim of the present study was to develop and evaluate a new recombinant ELISA (RE) for the detection of specific IgG and IgM antibodies against Borrelia burgdorferi. MATERIALS AND METHODS: The RE (Biotest AG, Dreieich, Germany) uses mixtures of recombinant p100, OspC, p18 of Borrelia afzelii and a fusion protein of recombinant internal fragments of the flagellum protein (p41) of Borrelia garinii strain PBi and Borrelia afzelii strain PKo. Serologic testing was performed on a commercially available ELISA processor without pre-absorption of sera. The sensitivity of the RE was determined by testing 226 sera obtained from patients suffering from Lyme disease (stage I: n = 148, stage II: n = 35, stage III: n = 43). Specificity of the RE was evaluated in 1107 sera from healthy blood donors and 275 sera from patients with other infectious diseases or autoimmune illnesses (leptospirosis: n = 53, syphilis: n = 70, toxoplasmosis: n = 60, herpes simplex virus: n = 30, HIV: n = 30; rheumatoid-factor positive: n = 32). In addition, 394 routine samples were prospectively tested in comparison with a well established whole-cell lysate extract ELISA (ENZYGNOST Borreliosis, DadeBehring, Germany) for relative sensitivity and specificity. RESULTS: Overall specificity, determined in 1107 healthy blood donors and 275 sera from patients with other diseases, was 94% for IgG and 91% for IgM. The overall sensitivities in 226 sera obtained from patients suffering from different stages of LD were 67-95%. Moreover, as revealed by prospective testing of 394 routine samples, the relative sensitivity of the RE in comparison with an established whole-cell lysate extract ELISA in the detection of seropositive samples was 81.1% for IgG and 86.5% for IgM with a relative specificity of 98.5% and 93% respectively. The ELISAs showed an overall agreement of 97% for IgG and 92.4% for IgM test results. CONCLUSION: The RE proved to be a reliable and specific screening test in the routine serodiagnosis of LD. In addition, the RE is easy to perform and requires no pre-absorption.

Early evolution of hepatitis C virus (HCV) quasispecies after liver transplant for HCV-related disease.

BACKGROUND: End-stage liver disease as a result of chronic hepatitis C virus (HCV) infection is the main indication for liver transplant (LT), but allografts are systematically infected with HCV soon after transplant. Viral quasispecies are poorly described during the early posttransplant period. METHODS: For 17 patients who received an LT for HCV disease, plasma viral quasispecies evolution was determined by sequence analysis of hypervariable region 1 of the E2 envelope gene before transplant (BT), after 7 days (D7), and after 1 month (M1). T helper (Th)1/Th2 cytokine levels were determined concomitantly. RESULTS: HCV quasispecies showed a significant decrease in amino acid diversity at D7 and M1, compared with BT (P<.05). A correlation was observed between low plasma tumor necrosis factor-alpha levels at D7 and decreased quasispecies amino acid complexity at the same date. Nucleic acid diversity was lower for genotype 1 than for genotype 3 infection (P<.05). The complexity and diversity of amino acids were lower in patients with hepatocellular carcinoma (HCC) BT than in those without HCC (P<.05). Conserved amino acid residues within quasispecies were shared by the whole cohort before and after LT. CONCLUSION: Viral structural and/or host immunological features could favor the emergence of fitter HCV strains after LT.

Monitoring low cytomegalovirus viremia in transplanted patients by a real-time PCR on plasma.

Until recently, human cytomegalovirus (hCMV) infection and anti-CMV treatment in transplanted patients have been monitored essentially by pp65 antigenemia, which is time-consuming and requires experienced operators. For the last two years, pp65 antigenemia levels have tended to be lower than previously in our laboratory, which could be due to better monitoring of CMV-related risk. Results obtained by real-time PCR with a LightCycler instrument or by pp65 antigen assay were compared on 145 serial samples from bone marrow or kidney transplant recipients under the usual conditions of our laboratory. CMV DNA was extracted from plasma and quantified by using primers and probes directed to HXFL4 gene. The plasma CMV DNA load was measured by using a standard curve constructed with a commercially available quantified CMV DNA suspension. Among the 145 samples, 139 showed a pp65 antigen which was negative or lower than 20 positively stained cells per 200,000 leukocytes. In the patients with positive pp65 antigenemia, the corresponding values of CMV DNA copy number/ml were significantly higher than those observed in patients without antigenemia (P < 0.001). CMV DNA was detected from 4 up to 52 days before pp65 antigen. Elsewhere, between two dates at which pp65 antigen was positive, intermediate PCR results could be positive while the pp65 antigen was negative. This real-time quantitative PCR assay is a rapid technique adapted to monitor plasma CMV DNA in transplant setting, even for low viremia.