key: cord-1012929-twpe452c authors: Bourgoin, Pénélope; Soliveres, Thomas; Barbaresi, Alexandra; Loundou, Anderson; Belkacem, Inès Ait; Arnoux, Isabelle; Bernot, Denis; Loosveld, Marie; Morange, Pierre‐Emmanuel; Michelet, Pierre; Malergue, Fabrice; Markarian, Thibaut title: CD169 and CD64 could help differentiate bacterial from CoVID‐19 or other viral infections in the Emergency Department date: 2021-02-08 journal: Cytometry A DOI: 10.1002/cyto.a.24314 sha: 30b87e29b1e5a5f923b5fca4b2af2bea03bafc29 doc_id: 1012929 cord_uid: twpe452c The identification of a bacterial, viral, or even noninfectious cause is essential in the management of febrile syndrome in the emergency department (ED), especially in epidemic contexts such as flu or CoVID‐19. The aim was to assess discriminative performances of two biomarkers, CD64 on neutrophils (nCD64) and CD169 on monocytes (mCD169), using a new flow cytometry procedure, in patients presenting with fever to the ED during epidemics. Eighty five adult patients presenting with potential infection were included during the 2019 flu season in the ED of La Timone Hospital. They were divided into four diagnostic outcomes according to their clinical records: no‐infection, bacterial infection, viral infection and co‐infection. Seventy six patients with confirmed SARS‐CoV‐2 infection were also compared to 48 healthy volunteers. For the first cohort, 38 (45%) patients were diagnosed with bacterial infections, 11 (13%) with viral infections and 29 (34%) with co‐infections. mCD169 was elevated in patients with viral infections, with a majority of Flu A virus or Respiratory Syncytial Virus, while nCD64 was elevated in subjects with bacterial infections, with a majority of Streptococcus pneumoniae and Escherichia coli. nCD64 and mCD169 showed 90% and 80% sensitivity, and 78% and 91% specificity, respectively, for identifying patients with bacterial or viral infections. When studied in a second cohort, mCD169 was elevated in 95% of patients with SARS‐CoV‐2 infections and remained at normal level in 100% of healthy volunteers. nCD64 and mCD169 have potential for accurately distinguishing bacterial and acute viral infections. Combined in an easy and rapid flow cytometry procedure, they constitute a potential improvement for infection management in the ED, and could even help for triage of patients during emerging epidemics. In the management of febrile syndrome, diagnostic guidance toward an infectious etiology is essential [1] . The characterization of a viral, bacterial or other infectious or noninfectious cause allows early appropriate patient management. Nevertheless, the data in the literature report the complexity of triage and diagnostic guidance [2] . The integration of the clinical examination is essential, but unfortunately is often not enough [3] ; diagnostic elements are therefore needed as quickly as possible in the emergency department (ED), especially in epidemic contexts such as flu or CoVID-19 [4] [5] [6] . Biological markers such as procalcitonin (PCT) and C-reactive protein (CRP) are commonly used in clinical practice. These markers have bacterial specificity but share a wide range of values with viral infections and do not make it possible to exclude or to confirm definitively the diagnosis [7] [8] [9] . Their triage capacity in ED is thus regularly challenged [10] . Measuring multiple specific biomarkers simultaneously, with a simple technique and rapid time-to-results, would be better compatible with the needs of triage in emergency medicine [11] . We developed a rapid flow cytometry assay, able to measure leucocytes biomarkers expressions within 10 min [12] , and demonstrated promising results for the triage of patients with fever at the Emergency Department [13, 14] , with CD64 on neutrophils (nCD64), increased in case of bacterial infections [15] , and CD169 on monocytes (mCD169), increased in case of viral infections [16] . In these previous studies, one limitation was the low number of infected patients, especially those with viral diseases. In this new study, we have thus included more patients, and focused on those with infectious symptoms during the flu season, and later added a cohort of CoVID-19 patients. The main goal was to confirm the relevance of CD64 and CD169 for discriminating between bacterial and viral infections in such epidemic contexts. pandemic. The inclusion criteria were the presence of fever greater than 38 C or hypothermia less than 36.5 C, and any potential respiratory (cough, sputum, and dyspnea), urinary (potential urinary infection), abdominal (pain syndrome, diarrhea), cutaneous (erysipelas), or neurological (meningitis) infectious clinical signs. The exclusion criteria were incomplete clinical files, traumatized patients or patients presenting with a known inflammatory or autoimmune disease, neoplasia, chronic infectious disease (viral, fungal, or bacterial), or antibiotic, antiviral, or immunosuppressive treatment prior to admission, and patients with extensive burns or recent surgery (less than 1 month). Their routine care was not modified, and confidentiality was preserved at all levels. All enrolled patients provided informed consent and no objection authorization, so that their data could be retrieved from their clinical records by a team of emergency department specialists, and could be used in the study. Based on all clinical and biological data, an adjudication committee classified patients from the first cohort in four groups. In some cases, viral agents were found by antigen-based tests or serological assays. Group IV: Subjects diagnosed as having both viral and bacterial infections. The committee was not aware of the flow cytometry results. The adjudication committee was not required for subjects included during CoVID-19 pandemic, since RT-PCR results were used as gold standard. Leftover ethylenediaminetetraacetic acid (EDTA)-treated blood samples were pseudonymized, and processed by flow cytometry according to a newly described one-step procedure [12] . Briefly, a multicolor panel was constituted and dried as a "glassified" layer at An overview of this study workflow is shown in Figure 1 . The Overall, 94 common pathogen species were detected ( Table 2) . The most frequent pathogens were Streptococcus pneumoniae (38%) and Flu A virus (26%). In comparison to non-infected subjects (nCD64 MFI of 1.1 ± 0.5; CD64 on neutrophils and CD169 on monocytes were the two main biomarkers assessed in the study by flow cytometry for discriminating between bacterial versus viral infections. Expressions of these two biomarkers have been shown to be directly induced, within hours, by interferons produced by the body in response to pathogen detection [14] . Ability of nCD64 to discriminate between bacterial and nonbacterial infections has been largely demonstrated for years [17] , whereas mCD169 increase after infection by viruses has only been described recently. mCD169 seems to be a general biomarker of acute viral infections since it has been found in patients with HIV [16, 18, 19] , EBV [20] , RSV [21] , CMV [22] , dengue [23, 24] , Zika [25] , noroviruses [26] , Lassa, and Marburg [27] . Here, high levels of mCD169 have been observed for the first time for Flu A virus. High levels of sensitivity and specificity were found in the study for both biomarkers. Interestingly, nCD64 showed a better sensitivity of 90% than specificity of 78%, whereas mCD169 showed a better specificity of 91% than sensitivity of 80%. These results further demonstrate their valuable use for infection etiology guidance in ED settings for patient triage. Indeed, a biomarker used for bacterial infection identification in ED needs to be as sensitive as possible to detect the majority of cases, demonstrating at least 90% sensitivity, as any missing case could delay patient from receiving appropriate antibiotic therapy, and thus increase their risk of developing sepsis and progression to death. Conversely, a viral marker in ED has to be very specific to ensure the etiology, with at least 90% specificity, as it allows the practitioner to discharge the patient, as well as avoiding the empirical use of antimicrobial drugs in case they are not required [28] . The global health issue of overuse of antibiotics has been illus- As expected in these virally infected patients, CD64 on neutrophils level remained low. A few patients had an increased level that could indicate a bacterial co-infection or could also be related to the severity of the disease, as it has been described an interferon-gamma release during the inflammation phase [31] . This combination of biomarker is thus promising for the triage of patients at the Emergency Department in the routine and also during current epidemics such as flu or CoVID-19, and could probably be directly applied to any new emergent infectious disease. Other promising biomarkers had been included in the flow cytometry panels, and thus analyzed concomitantly: HLA-DR on monocytes, CD64 on monocytes, and HLA-ABC ratio of monocytes/ neutrophils in the first part of the study [13] . They showed weak performances to identify bacterial or viral infections in these cohorts ( Figure S2 ). The authors state that they have followed the principles outlined in the Declaration of Helsinki for all human experimental investigations. The authors certify that this manuscript reports original clinical research data. Individual data that underlie the results reported in this article are available from the corresponding author following publication, including the study report and study protocol. Additional data are available upon reasonable request. 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