. Overall, the Biotyper system generated significantly more accurate identifications than the Vitek MS IVD system (P ‫؍‬ 0.016; 177 versus 167 out of 181 specimens). The Biotyper system identified the minority species among polymicrobial blood cultures. We also compared the performance of an in-house extraction method with that of the Sepsityper on both MALDI-TOF MS systems. The in-house method generated more correct identifications at the genus level than the Sepsityper (96.7% versus 93.5%) on the Biotyper system, whereas the two methods exhibited the same performance level (88.0% versus 88.0%) on the Vitek MS IVD system. Our study confirmed the practical advantages of MALDI-TOF MS, and our in-house extraction method reduced the reagent cost to $1 per specimen, with a shorter turnaround time of 3 h, which is highly cost-effective for a diagnostic microbiology service. Bloodstream infection is a condition associated with high morbidity and mortality (1, 2). The rapid identification of bloodstream pathogens is critical to clinical management and the choice of appropriate antibiotic treatments (3, 4). Currently, microbiologic diagnosis of bacteremia relies on subculture of positive blood culture broths on solid medium for an 18-to 24-h incubation period followed by biochemical tests or an automated preformed enzyme assay for identification of the bacteria (5). In general, laboratory diagnosis of common pathogens requires 18 to 48 h, while diagnosis of fastidious organisms requires longer incubation and identification procedures. Although the use of modern techniques such as fluorescence in situ hybridization and PCR can shorten the identification time (6), these operations require specialized equipment and technical expertise and the targeted pathogens are limited in a single run (7). Therefore, the introduction of a simple, rapid, broad-spectrum, and cost-effective system for the identification of blood culture pathogens is imperative.Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) was developed in the 1980s, and this technique had been widely used for biomolecule analysis in the chemical industry for over 10 years. In recent years, MALDI-TOF MS was introduced for bacterial identification, which revolutionized the diagnostic microbiology service (8). Two manufacturers, bioMérieux (Marcy l'Etoile, France) and Bruker Daltonics (Bremen, Germany), are marketing efficient MALDI-TOF systems, the Vitek-MS and the Microflex LT, respectively, that allow the identification of bacteria and yeasts in a few minutes instead of the hours required by traditional methods. The practical use of MALDI-TOF MS for microorganism identification directly from positive blood culture specimens has been substantiated by a number of studies (9-13). Generally, studies have shown high identification rates for various types of organisms with the use of MALDI-TOF MS . However, various organism isolation methods from blood were used in those studies and therefore method standardization is necessary. The...

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Background The role of subclinical severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections in perpetuating the COVID-19 pandemic is unknown because population seroprevalence data are absent. We aimed to establish the sensitivity and specificity of our enzyme immunoassay and microneutralisation assay, and the seroprevalence of SARS-CoV-2 in Hong Kong before and after the pandemic, as well as in Hong Kong residents evacuated from Hubei province, China. Methods We did a multicohort study in a hospital and university in Hong Kong. We evaluated the sensitivity of our enzyme immunoassay and microneutralisation assay with RT-PCR data from patients positive for SARS-CoV-2 and the specificity of our enzyme immunoassay and microneutralisation assay with archived serum samples collected before 2019. We compared the seropositivity of the general population of Hong Kong before and after the pandemic had begun, and determined the seropositivity of Hong Kong residents evacuated from Hubei province, China, in March, 2020. Findings Between Feb 26 and March 18, 2020, we assessed RT-PCR samples from 45 patients who had recovered from COVID-19 to establish the sensitivity of our enzyme immunoassay and microneutralisation assay. To establish the specificity of these assays, we retrieved archived serum. The sensitivity was 91·1% (41 of 45 [95% CI 78·8–97·5]) for the microneutralisation assay, 57·8% (26 of 45 [42·2–72·3]) for anti-nucleoprotein IgG, 66·7% (30 of 45 [51·1–80·0]) for anti-spike protein receptor binding domain (RBD) IgG, and 73·3% (33 of 45 [58·1–85·4]) for enzyme immunoassay (either positive for anti-nucleoprotein or anti-RBD IgG). The specificity was 100% (152 of 152 [95% CI 97·6–100·0]) for both the enzyme immunoassay and microneutralisation assay. Among the Hong Kong general population, 53 (2·7%) of 1938 were enzyme immunoassay positive, but of those who were positive, all 53 were microneutralisation negative, and no significant increase was seen in the seroprevalence between April 12, 2018, and Feb 13, 2020. Among asymptomatic Hubei returnees, 17 (4%) of 452 were seropositive with the enzyme immunoassay or the microneutralisation assay, with 15 (88%) of 17 seropositive with the microneutralisation assay, and two familial clusters were identified. Interpretation Our serological data suggest that SARS-CoV-2 is a new emerging virus. The seropositivity rate in Hubei returnees indicates that RT-PCR-confirmed patients only represent a small proportion of the total number of cases. The low seroprevalence suggests that most of the Hong Kong and Hubei population remain susceptible to COVID-19. Future waves of the outbreak are inevitable without a vaccine or antiviral prophylaxis. The role of age-related cross reactive non-neutralising antibodies in the pathogenesis of COVID-19 warrants further investigation. Funding Richard and Carol Yu, May Tam Mak Mei Yin, Shaw Foundation (Hong...

Using paired serum samples obtained from patients with illness associated with increases in anti-human coronavirus OC43 (HCoV-OC43) or anti-HCoV-229E antibodies, we examined the possibility of false-positive results detected in a recombinant severe acute respiratory syndrome (SARS)-associated coronavirus (SARSCoV) nucleocapsid protein immunoglobulin G enzyme-linked immunosorbent assay (ELISA). Three of the 21 and 1 of the 7 convalescent-phase serum samples from persons with increases in antibodies against HCoV-OC43 and HCoV-229E, respectively, tested positive by the recombinant SARS-CoV nucleocapsid protein-based ELISA. None of these samples were found to contain a specific antibody in the recombinant SARS-CoV spike polypeptide-based Western blot assay.Severe acute respiratory syndrome (SARS), caused by SARS-associated coronavirus (SARS-CoV), has affected 30 countries in five continents, with more than 8,000 cases and 750 deaths (7-11). As for the detection of antibodies against SARS-CoV, at the moment, the most widely used methods are antibody detection in acute-and convalescent-phase sera by indirect immunofluorescence assay and enzyme-linked immunosorbent assay (ELISA) using cell culture extracts (4,8,10). However, antibody detection by these methods may be less reproducible, more difficult to standardize, and more laborintensive than ELISA-based antibody detection tests using recombinant antigens. Furthermore, producing the infected cell lines for coating the ELISA plates and the slides for indirect immunofluorescence requires cultivation of the SARS-CoV, for which biosafety level 3 laboratory facilities are required. Such facilities are not available in most clinical microbiology laboratories.ELISA-based antibody detection tests using recombinant antigens are well known to offer higher reproducibility and to be easier to standardize and less labor-intensive than antibody detection by indirect immunofluorescence assay and ELISA using cell culture extract, and they do not require cultivation of the SARS-CoV (1, 2, 14, 18). Recently, we have reported the use of recombinant SARS-CoV nucleocapsid protein ELISAbased antibody tests for serodiagnosis of SARS-CoV pneumonia and the study of the seroprevalence of nonpneumonic SARS-CoV infections (15, 16). In addition, others have also reported the use of recombinant-protein-based immunoassays for serodiagnosis of SARS-CoV pneumonia (3, 17). However, in our studies, we have also shown that false-positive reactions were detected if the recombinant SARS-CoV nucleocapsid protein-based ELISA was used alone for antibody detection (15). In this study, using paired serum samples obtained from patients with increases in anti-human CoV OC43 (HCoV-OC43) or anti-HCoV-229E antibodies, we examined the possibility of false-positive results detected by the recombinant SARS-CoV nucleocapsid protein-based ELISA. The importance of using Western blot assays, with the nucleocapsid protein and spike polypeptide of SARS-CoV, for confirmation was also determined.Paired serum samples collecte...