Development and Validation of a Highly Sensitive Multiplex Immunoassay for SARS-CoV-2 Humoral Response Monitorization: A Study of the Antibody Response in COVID-19 Patients with Different Clinical Profiles during the First and Second Waves in Cadiz, Spain.

There is still a long way ahead regarding the COVID-19 pandemic, since emerging waves remain a daunting challenge to the healthcare system. For this reason, the development of new preventive tools and therapeutic strategies to deal with the disease have been necessary, among which serological assays have played a key role in the control of COVID-19 outbreaks and vaccine development. Here, we have developed and evaluated an immunoassay capable of simultaneously detecting multiple IgG antibodies against different SARS-CoV-2 antigens through the use of Bio-PlexTM technology. Additionally, we have analyzed the antibody response in COVID-19 patients with different clinical profiles in Cadiz, Spain. The multiplex immunoassay presented is a high-throughput and robust immune response monitoring tool capable of concurrently detecting anti-S1, anti-NC and anti-RBD IgG antibodies in serum with a very high sensitivity (94.34-97.96%) and specificity (91.84-100%). Therefore, the immunoassay proposed herein may be a useful monitoring tool for individual humoral immunity against SARS-CoV-2, as well as for epidemiological surveillance. In addition, we show the values of antibodies against multiple SARS-CoV-2 antigens and their correlation with the different clinical profiles of unvaccinated COVID-19 patients in Cadiz, Spain, during the first and second waves of the pandemic.

Serum microRNAs targeting ACE2 and RAB14 genes distinguish asymptomatic from critical COVID-19 patients.

Despite the extraordinary advances achieved to beat COVID-19 disease, many questions remain unsolved, including the mechanisms of action of SARS-CoV-2 and which factors determine why individuals respond so differently to the viral infection. Herein, we performed an in silico analysis to identify host microRNA targeting ACE2, TMPRSS2, and/or RAB14, all genes known to participate in viral entry and replication. Next, the levels of six microRNA candidates previously linked to viral and respiratory-related pathologies were measured in the serum of COVID-19-negative controls (n = 16), IgG-positive COVID-19 asymptomatic individuals (n = 16), and critical COVID-19 patients (n = 17). Four of the peripheral microRNAs analyzed (hsa-miR-32-5p, hsa-miR-98-3p, hsa-miR-423-3p, and hsa-miR-1246) were upregulated in COVID-19 critical patients compared with COVID-19-negative controls. Moreover, hsa-miR-32-5p and hsa-miR-1246 levels were also altered in critical versus asymptomatic individuals. Furthermore, these microRNA target genes were related to viral infection, inflammatory response, and coagulation-related processes. In conclusion, SARS-CoV-2 promotes the alteration of microRNAs targeting the expression of key proteins for viral entry and replication, and these changes are associated with disease severity. The microRNAs identified could be taken as potential biomarkers of COVID-19 progression as well as candidates for future therapeutic approaches against this disease.

HIV antibodies level as a marker of HIV persistence: the role of hepatitis C virus coinfection.

Human immunodeficiency virus (HIV) antibodies have been proposed as a measure of the size of the HIV reservoir. The aim of our study is to quantify the anti-HIV antibodies level in a cohort of people living with HIV (PLWH), stratified based on the presence of continuous undetectable HIV viral load and the co-existence of hepatitis C virus infection. A sample of 229 HIV-monoinfected (n = 114) or HIV/HCV-coinfected [either with resolved HCV infection (n = 75) or active HCV coinfection (n = 40)] patients, followed up a median of 34 (IQR 20-44) months, was studied. Anti-HIV index was obtained as the 1:800 dilution of HIV antibodies. CD4+ T cell count, time with undetectable HIV viral load, annual increase of CD4+ T cell count, anti-HCV therapy, and diagnosis of cirrhosis were analyzed. Patients with a continued suppressed HIV viral load had significant lower anti-HIV index compared with those with virologic failure during the follow-up. Significant higher CD4+ T cell increase was observed in those with a lower anti-HIV index. HIV-monoinfected patients showed an anti-HIV index significantly lower than patients with HCV coinfection. Resolved HCV infection after interferon-based therapy, but not with direct acting antivirals, was associated with a lower anti-HIV index. HIV/HCV-coinfected patients showed higher HIV antibodies level when compared with HIV-monoinfected individuals. A decrease in anti-HIV index in HIV/HCV-coinfected patients was detected when a sustained virological HCV response was obtained after interferon-based therapy, in possible relation with the direct or indirect effect of interferon on PLWH CD4 T cells.

Co-Occurrence of mcr-1 and qnrS1 on an IncHI2 Plasmid in Clinical Isolates of Salmonella Typhimurium in Spain.

Salmonella enterica is a well-adapted zoonotic bacterium associated to cases of gastroenteritis and bacteremia with increased morbidity and mortality. In this study, three isolates of Salmonella Typhimurium obtained from human clinical samples, showing colistin resistance and low-level resistance to quinolones, have been genetically characterized. We detected the co-occurrence of mcr-1 and qnrS1 on a single IncHI2 plasmid in isolates of Salmonella Typhimurium obtained from Spanish children without a travel history. The multiresistant region contained numerous resistance genes. Isolates were clonally related, which suggests the presence of these clones in the community and the potential to cause outbreaks affecting the most susceptible population. It is necessary to monitor the presence of these plasmid-mediated resistance genes in human European strains of Salmonella spp. because of the risk of producing outbreaks of community-acquired infections.

Staphylococcus aureus carriage in older populations in community residential care homes: Prevalence and molecular characterization of MRSA isolates / Colonización por Staphylococcus aureus en pacientes institucionalizados en residencias geriátricas: prevalencia y caracterización molecular de los aislados resistentes a meticilina

Introduction: The epidemiology of S. aureus depends on conditions in specific populations. Few studies of S. aureus colonization in the older population have been performed in Spain. The aim of this study was to determine the prevalence of methicillin-resistant S. aureus (MRSA) colonization and its molecular epidemiological characteristics in an institutionalized population in community residential care homes in Cadiz, Spain. Methods: A cross-sectional epidemiological study was conducted in three residential care homes for older people. Axilla and nostril samples were tested. Identification of S. aureus and antimicrobial susceptibility testing were by MALDI-TOF and MicroScan panels. MRSA strains were subjected to SCCmec typing, multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE). The presence of Panton-Valentine leukocidin (PVL) genes was determined by PCR in all S. aureus strains. Results: A total of 293 residents were included. Fifty-one residents (17.4%) were colonized with methicillin-sensitive S. aureus (MSSA) and 11 (3.8%) with MRSA. Resistance to at least two aminoglycosides was observed in 25.4% of MSSA and 90.9% and of MRSA isolates, and resistance to levofloxacin in 80.3% of MSSA and 100% of MRSA isolates. SCCmecIV was detected in all isolates and all except one (ST-125) were ST-8. None of the S. aureus isolates were positive for PVL. Conclusions: A low rate of S. aureus carriage was detected and the prevalence of MRSA was very low. ST8-MRSA-IVc was the dominant clone, and only one strain belonged to ST125-MRSA-IVc. We found MRSA transmission within the residential care homes and a very high rate of quinolone resistance in MSSA and MRSA

Introducción: La epidemiología de S. aureus depende de las condiciones particulares de cada población. En España se han realizado pocos estudios sobre la colonización por S. aureus en la población geriátrica. El objetivo de este estudio es determinar la prevalencia de colonización por S. aureus resistente a meticilina (SARM) y sus características epidemiológicas moleculares en población institucionalizada en residencias de ancianos en Cádiz, España. Métodos: Se realizó un estudio epidemiológico transversal en 3 residencias de ancianos. Se estudiaron muestras de las fosas nasales y axilas. La identificación y las pruebas de sensibilidad se realizaron utilizando MALDI-TOF y paneles MicroScan(R). En los aislados de SARM se determinó el tipo de SCCmec y se tiparon mediante Multilocus Sequence Typing (MLST) y Pulsed-field Gel Electrophoresis (PFGE). La presencia de genes de la leucocidina de Panton-Valentine (LPV) se determinó mediante PCR en todas las cepas de S. aureus. Resultados: Se incluyeron un total de 293 residentes. Cincuenta y un residentes (17,4%) estaban colonizados por S. aureus sensible a la meticilina (SASM) y 11 (3,8%) por SARM. Se observó resistencia frente al menos 2 aminoglucósidos en el 25,4 y 90,9% y resistencia a levofloxacino en el 80,3 y 100% de los aislamientos de SASM y SARM, respectivamente. Se detectó SCCmecIV en todos los aislados, y todos, excepto uno (ST-125) correspondían al ST-8. Ninguno de los aislados de S. aureus fue positivo para LPV. Conclusiones: Se detectó una baja tasa de portadores de S. aureus, siendo el porcentaje de SARM muy bajo. ST8-MRSA-IVc fue el clon predominante, y solo una cepa pertenecía a ST125-MRSA-IVc. Se objetivó transmisión de SARM intracentro. Se observó una tasa muy alta de resistencia a quinolonas en SASM y SARM

Staphylococcus aureus carriage in older populations in community residential care homes: Prevalence and molecular characterization of MRSA isolates.

INTRODUCTION: The epidemiology of S. aureus depends on conditions in specific populations. Few studies of S. aureus colonization in the older population have been performed in Spain. The aim of this study was to determine the prevalence of methicillin-resistant S. aureus (MRSA) colonization and its molecular epidemiological characteristics in an institutionalized population in community residential care homes in Cadiz, Spain. METHODS: A cross-sectional epidemiological study was conducted in three residential care homes for older people. Axilla and nostril samples were tested. Identification of S. aureus and antimicrobial susceptibility testing were by MALDI-TOF and MicroScan panels. MRSA strains were subjected to SCCmec typing, multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE). The presence of Panton-Valentine leukocidin (PVL) genes was determined by PCR in all S. aureus strains. RESULTS: A total of 293 residents were included. Fifty-one residents (17.4%) were colonized with methicillin-sensitive S. aureus (MSSA) and 11 (3.8%) with MRSA. Resistance to at least two aminoglycosides was observed in 25.4% of MSSA and 90.9% and of MRSA isolates, and resistance to levofloxacin in 80.3% of MSSA and 100% of MRSA isolates. SCCmecIV was detected in all isolates and all except one (ST-125) were ST-8. None of the S. aureus isolates were positive for PVL. CONCLUSIONS: A low rate of S. aureus carriage was detected and the prevalence of MRSA was very low. ST8-MRSA-IVc was the dominant clone, and only one strain belonged to ST125-MRSA-IVc. We found MRSA transmission within the residential care homes and a very high rate of quinolone resistance in MSSA and MRSA.