Flow cytometry to assess the counts and physiological state of cronobacter sakazakii cells after heat exposure

Producción CientíficaCronobacter sakazakii is an opportunistic pathogen that is associated with outbreaks of neonatal necrotizing enterocolitis, septicaemia, and meningitis. Reconstituted powdered infant formulae is the most common vehicle of infection. The aim of the present study is to gain insight into the physiological states of C. sakazakii cells using flow cytometry to detect the compromised cells, which are viable but non-culturable using plate-based methods, and to evaluate the impact of milk heat treatments on those populations. Dead-cell suspensions as well as heat-treated and non-heat-treated cell suspensions were used. After 60 or 65 °C treatments, the number of compromised cells increased as a result of cells with compromised membranes shifting from the heat-treated suspension. These temperatures were not effective at killing all bacteria but were effective at compromising their membranes. Thus, mild heat treatments are not enough to guarantee the safety of powered infant formulae. Flow cytometry was capable of detecting C. sakazakii’s compromised cells that cannot be detected with classical plate count methods; thus, it could be used as a screening test to decrease the risk derived from the presence of pathogenic viable but non-culturable cells in this food that is intended for newborns’ nutrition.Junta de Castilla y León (projects SAN196/VA07/07, SAN673/VA05/08, and SAN126/09

Gas6, una proteína dependiente de la vitamina k implicada en la respuesta a la infección por sars-cov-2

Trabajo presentado en el LXIII Congreso Nacional SEHH/ XXXVII Congreso Nacional SETH, celebrado en Pamplona (Navarra) del 14 al 16 de octubre de 2021

Growth Arrest-Specific Factor 6 (GAS6) Is Increased in COVID-19 Patients and Predicts Clinical Outcome

Producción CientíficaBackground: Growth arrest-specific factor 6 (GAS6) and the Tyro3, AXL, and MERTK (TAM) receptors counterbalance pro-inflammatory responses. AXL is a candidate receptor for SARS-CoV-2, particularly in the respiratory system, and the GAS6/AXL axis is targeted in current clinical trials against COVID-19. However, GAS6 and TAMs have not been evaluated in COVID-19 patients at emergency admission. Methods: Plasma GAS6, AXL, and MERTK were analyzed in 132 patients consecutively admitted to the emergency ward during the first peak of COVID-19. Results: GAS6 levels were higher in the SARS-CoV-2-positive patients, increasing progressively with the severity of the disease. Patients with initial GAS6 at the highest quartile had the worst outcome, with a 3-month survival of 65%, compared to a 90% survival for the rest. Soluble AXL exhibited higher plasma concentration in deceased patients, without significant differences in MERTK among SARS-CoV-2-positive groups. GAS6 mRNA was mainly expressed in alveolar cells and AXL in airway macrophages. Remarkably, THP-1 human macrophage differentiation neatly induces AXL, and its inhibition (bemcentinib) reduced cytokine production in human macrophages after LPS challenge. Conclusions: Plasma GAS6 and AXL levels reflect COVID-19 severity and could be early markers of disease prognosis, supporting a relevant role of the GAS6/AXL system in the immune response in COVID-19.Ministerio de Ciencia, Innovación y Universidades (project RTI2018-095672-B-I00)Instituto de Salud Carlos III - Fondo de Investigación Sanitaria (grants PI15/00531 and PI19/01410)Fundació La Marató TV3 (grants 20153030 and 20153031)Consejo Superior de Investigaciones Científicas (project CSIC-COV19-016/202020E155)Junta de Castilla y León (project 07.04.467B04.74011.0

Gender differences in the plasma concentration of the GAS6-TAM system in COVID-19 patients

Resumen del trabajo presentado en el 4th European Congress on Thrombosis and Haemostasis, celebrado en Gante (Bélgica), los días 14 y 15 de octubre de 2021Background: SARS-CoV-2 induces an immune response with potentially harmful effects for the patient due to an uncontrolled release of inflammatory factors, specially at the capillary wall. The vitamin K-dependent plasma protein GAS6 and the TAM (TYRO3, AXL, and MERTK) receptors play a relevant role among restorative mechanisms that counterbalance pro-inflammatory responses at the endothelial interface. Aims: To study the influence of gender on the effects of SARS-CoV-2 infection in the GAS6/TAM system, as reflected by plasma concentration at patient admittance at the emergency ward. Methods: The plasma content of GAS6, AXL, and MERTK was analyzed in a first group of 132 patients, 68 females and 64 males consecutively admitted to the emergency ward during the first peak of COVID-19. A confirmatory group was studied from the second wave of contagions. An analysis of gender differences in relation to the GAS6/TAM concentrations in plasma was performed on this population. Results: In accordance with recently published GAS6 levels, significantly higher in the SARS-CoV-2 positive than in negative patients, increased progressively with the severity of the disease in SARS-CoV-2 positive individual irrespective of the gender of the patient. In contrast, while soluble AXL exhibited higher plasma concentration in deceased patients and no significant differences were observed in MERTK concentration, differential gender analysis suggest differences in soluble TAM receptors. While a COVID-19 related increase in sAXL was observed in men, this was not the case in women. Oppositely, MERTK differences due to COVID-19 infection were only significant in women. Summary/Conclusion: GAS6-TAM system of ligands and receptors is implicated in the immune response to SARS-CoV-2 in patients from both genders. Plasma GAS6 levels paralleled COVID-19 severity being an early marker of disease prognosis in both sexes. In contrast, soluble TAM receptors presented a gender-specific behavior. Sex-related differences in sAXL and sMERTK expression in COVID-19 patients could affect therapy efficacy deserving further investigation

Evaluation of cytokines as robust diagnostic biomarkers for COVID-19 detection

Producción CientíficaAntigen tests or polymerase chain reaction (PCR) amplification are currently COVID-19 diagnostic tools. However, developing complementary diagnosis tools is mandatory. Thus, we performed a plasma cytokine array in COVID-19 patients to identify novel diagnostic biomarkers. A discovery–validation study in two independent prospective cohorts was performed. The discovery cohort included 136 COVID-19 and non-COVID-19 patients recruited consecutively from 24 March to 11 April 2020. Forty-five cytokines’ quantification by the MAGPIX system (Luminex Corp., Austin, TX, USA) was performed in plasma samples. The validation cohort included 117 patients recruited consecutively from 15 to 25 April 2020 for validating results by ELISA. COVID-19 patients showed different levels of multiple cytokines compared to non-COVID-19 patients. A single chemokine, IP-10, accurately identified COVID-19 patients who required hospital admission (AUC: 0.962; 95%CI (0.933–0.992); p < 0.001)). The results were validated in an independent cohort by multivariable analysis (OR: 25.573; 95%CI (8.127–80.469); p < 0.001) and AUROC (AUC: 0.900; 95%CI (0.846–0.954); p < 0.001). Moreover, showing IP-10 plasma levels over 173.35 pg/mL identified COVID-19 with higher sensitivity (86.20%) than the first SARS-CoV-2 PCR. Our discover–validation study identified IP-10 as a robust biomarker in clinical practice for COVID-19 diagnosis at hospital. Therefore, IP-10 could be used as a complementary tool in clinical practice, especially in emergency departments.Instituto de Salud Carlos III (grant COV20/00491)Consejo Superior de Investigaciones científicas (grant CSIC-COV19-016/202020E155)Junta de Castilla y León (project COVID 07.04.467B04.74011.0)IBGM excellence programme (grant CLU-2029-02

COVID-19 vaccine failure

COVID-19 affects the population unequally with a higher impact on aged and immunosuppressed people. Hence, we assessed the effect of SARS-CoV-2 vaccination in immune compromised patients (older adults and oncohematologic patients), compared with healthy counterparts. While the acquired humoral and cellular memory did not predict subsequent infection 18 months after full immunization, spectral and computational cytometry revealed several subsets within the CD8+ T-cells, B-cells, NK cells, monocytes and CD45RA+ CCR7- Tγδ cells differentially expressed in further infected and non-infected individuals not just following immunization, but also prior to that. Of note, up to 7 subsets were found within the CD45RA+ CCR7- Tγδ population with some of them being expanded and other decreased in subsequently infected individuals. Moreover, some of these subsets also predicted COVID-induced hospitalization in oncohematologic patients. Therefore, we hereby have identified several cellular subsets that, even before vaccination, strongly related to COVID-19 vulnerability as opposed to the acquisition of cellular and/or humoral memory following vaccination with SARS-CoV2 mRNA vaccines.This study has been funded through Programa Estratégico Instituto de Biología y Genética Molecular (IBGM Junta de Castilla y León. Ref. CCVC8485), Junta de Castilla y León (Proyectos COVID 07.04.467B04.74011.0) and the European Commission – NextGenerationEU (Regulation EU 2020/2094), through CSIC's Global Health Platform (PTI Salud Global; SGL21-03-026 and SGL2021-03-038)N

SARS-CoV-2 susceptibility and COVID-19 disease severity are associated with genetic variants affecting gene expression in a variety of tissues

Variability in SARS-CoV-2 susceptibility and COVID-19 disease severity between individuals is partly due to genetic factors. Here, we identify 4 genomic loci with suggestive associations for SARS-CoV-2 susceptibility and 19 for COVID-19 disease severity. Four of these 23 loci likely have an ethnicity-specific component. Genome-wide association study (GWAS) signals in 11 loci colocalize with expression quantitative trait loci (eQTLs) associated with the expression of 20 genes in 62 tissues/cell types (range: 1:43 tissues/gene), including lung, brain, heart, muscle, and skin as well as the digestive system and immune system. We perform genetic fine mapping to compute 99% credible SNP sets, which identify 10 GWAS loci that have eight or fewer SNPs in the credible set, including three loci with one single likely causal SNP. Our study suggests that the diverse symptoms and disease severity of COVID-19 observed between individuals is associated with variants across the genome, affecting gene expression levels in a wide variety of tissue types

A first update on mapping the human genetic architecture of COVID-19

peer reviewe

A first update on mapping the human genetic architecture of COVID-19

The COVID-19 pandemic continues to pose a major public health threat, especially in countries with low vaccination rates. To better understand the biological underpinnings of SARS-CoV-2 infection and COVID-19 severity, we formed the COVID-19 Host Genetics Initiative1 . Here we present a genome-wide association study meta-analysis of up to 125,584 cases and over 2.5 million control individuals across 60 studies from 25 countries, adding 11 genome-wide significant loci compared with those previously identified2 . Genes at new loci, including SFTPD, MUC5B and ACE2, reveal compelling insights regarding disease susceptibility and severit

COVID-19 Host Genetics Initiative. A first update on mapping the human genetic architecture of COVID-19

The COVID-19 pandemic continues to pose a major public health threat, especially in countries with low vaccination rates. To better understand the biological underpinnings of SARS-CoV-2 infection and COVID-19 severity, we formed the COVID-19 Host Genetics Initiative1. Here we present a genome-wide association study meta-analysis of up to 125,584 cases and over 2.5 million control individuals across 60 studies from 25 countries, adding 11 genome-wide significant loci compared with those previously identified2. Genes at new loci, including SFTPD, MUC5B and ACE2, reveal compelling insights regarding disease susceptibility and severity.</p