Safety and efficacy of convalescent plasma for severe COVID-19: a randomized, single blinded, parallel, controlled clinical study.

BACKGROUND: Convalescent plasma (CP) has been widely used to treat COVID-19 and is under study. However, the variability in the current clinical trials has averted its wide use in the current pandemic. We aimed to evaluate the safety and efficacy of CP in severe coronavirus disease 2019 (COVID-19) in the early stages of the disease. METHODS: A randomized controlled clinical study was conducted on 101 patients admitted to the hospital with confirmed severe COVID-19. Most participants had less than 14 days from symptoms onset and less than seven days from hospitalization. Fifty patients were assigned to receive CP plus standard therapy (ST), and 51 were assigned to receive ST alone. Participants in the CP arm received two doses of 250 mL each, transfused 24 h apart. All transfused plasma was obtained from "super donors" that fulfilled the following criteria: titers of anti-SARS-CoV-2 S1 IgG ≥ 1:3200 and IgA ≥ 1:800 antibodies. The effect of transfused anti-IFN antibodies and the SARS-CoV-2 variants at the entry of the study on the overall CP efficacy was evaluated. The primary outcomes were the reduction in viral load and the increase in IgG and IgA antibodies at 28 days of follow-up. The per-protocol analysis included 91 patients. RESULTS: An early but transient increase in IgG anti-S1-SARS-CoV-2 antibody levels at day 4 post-transfusion was observed (Estimated difference [ED], - 1.36; 95% CI, - 2.33 to - 0.39; P = 0.04). However, CP was not associated with viral load reduction in any of the points evaluated. Analysis of secondary outcomes revealed that those patients in the CP arm disclosed a shorter time to discharge (ED adjusted for mortality, 3.1 days; 95% CI, 0.20 to 5.94; P = 0.0361) or a reduction of 2 points on the WHO scale when compared with the ST group (HR adjusted for mortality, 1.6; 95% CI, 1.03 to 2.5; P = 0.0376). There were no benefits from CP on the rates of intensive care unit admission (HR, 0.82; 95% CI, 0.35 to 1.9; P = 0.6399), mechanical ventilation (HR, 0.66; 95% CI, 0.25 to 1.7; P = 0.4039), or mortality (HR, 3.2; 95% CI, 0.64 to 16; P = 0.1584). Anti-IFN antibodies and SARS-CoV-2 variants did not influence these results. CONCLUSION: CP was not associated with viral load reduction, despite the early increase in IgG anti-SARS-CoV-2 antibodies. However, CP is safe and could be a therapeutic option to reduce the hospital length of stay. Trial registration NCT04332835.

The Impact of Ethnicity and Genetic Ancestry on Disease Prevalence and Risk in Colombia.

Currently, the vast majority of genomic research cohorts are made up of participants with European ancestry. Genomic medicine will only reach its full potential when genomic studies become more broadly representative of global populations. We are working to support the establishment of genomic medicine in developing countries in Latin America via studies of ethnically and ancestrally diverse Colombian populations. The goal of this study was to analyze the effect of ethnicity and genetic ancestry on observed disease prevalence and predicted disease risk in Colombia. Population distributions of Colombia's three major ethnic groups - Mestizo, Afro-Colombian, and Indigenous - were compared to disease prevalence and socioeconomic indicators. Indigenous and Mestizo ethnicity show the highest correlations with disease prevalence, whereas the effect of Afro-Colombian ethnicity is substantially lower. Mestizo ethnicity is mostly negatively correlated with six high-impact health conditions and positively correlated with seven of eight common cancers; Indigenous ethnicity shows the opposite effect. Malaria prevalence in particular is strongly correlated with ethnicity. Disease prevalence co-varies across geographic regions, consistent with the regional distribution of ethnic groups. Ethnicity is also correlated with regional variation in human development, partially explaining the observed differences in disease prevalence. Patterns of genetic ancestry and admixture for a cohort of 624 individuals from Medellín were compared to disease risk inferred via polygenic risk scores (PRS). African genetic ancestry is most strongly correlated with predicted disease risk, whereas European and Native American ancestry show weaker effects. African ancestry is mostly positively correlated with disease risk, and European ancestry is mostly negatively correlated. The relationships between ethnicity and disease prevalence do not show an overall correspondence with the relationships between ancestry and disease risk. We discuss possible reasons for the divergent health effects of ethnicity and ancestry as well as the implication of our results for the development of precision medicine in Colombia.

COVID-19 convalescent plasma composition and immunological effects in severe patients.

Convalescent plasma (CP) has emerged as a treatment for COVID-19. However, the composition and mechanism of action are not fully known. Therefore, we undertook a two-phase controlled study in which, first the immunological and metabolomic status of recovered and severe patients were evaluated. Secondly, the 28-day effect of CP on the immune response in severe patients was assessed. Nineteen recovered COVID-19 patients, 18 hospitalized patients with severe disease, and 16 pre-pandemic controls were included. Patients with severe disease were treated with CP transfusion and standard therapy (i.e., plasma recipients, n = 9) or standard therapy alone (n = 9). Clinical and biological assessments were done on day 0 and during follow-up on days 4, 7, 14, and 28. Clinical parameters, viral load, total immunoglobulin (Ig) G and IgA anti-S1-SARS-CoV-2 antibodies, neutralizing antibodies (NAbs), autoantibodies, cytokines, T and B cells, and metabolomic and lipidomic profiles were examined. Total IgG and IgA anti-S1-SARS-CoV-2 antibodies were key factors for CP selection and correlated with NAbs. In severe COVID-19 patients, mostly interleukin (IL)-6 (P = <0.0001), IL-10 (P = <0.0001), IP-10 (P = <0.0001), fatty acyls and glycerophospholipids were higher than in recovered patients. Latent autoimmunity and anti-IFN-α antibodies were observed in both recovered and severe patients. COVID-19 CP induced an early but transient cytokine profile modification and increases IgG anti-S1-SARS-CoV-2 antibodies. At day 28 post-transfusion, a decrease in activated, effector and effector memory CD4+ (P < 0.05) and activated and effector CD8+ (P < 0.01) T cells and naïve B cells (P = 0.001), and an increase in non-classical memory B cells (P=<0.0001) and central memory CD4+ T cells (P = 0.0252) were observed. Moreover, IL-6/IFN-γ (P = 0.0089) and IL-6/IL-10 (P = 0.0180) ratios decreased in plasma recipients compared to those who received standard therapy alone. These results may have therapeutic implications and justify further post-COVID-19 studies.

Convalescent plasma in Covid-19: Possible mechanisms of action.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is responsible of the coronavirus disease 2019 (COVID-19) pandemic. Therapeutic options including antimalarials, antivirals, and vaccines are under study. Meanwhile the current pandemic has called attention over old therapeutic tools to treat infectious diseases. Convalescent plasma (CP) constitutes the first option in the current situation, since it has been successfully used in other coronaviruses outbreaks. Herein, we discuss the possible mechanisms of action of CP and their repercussion in COVID-19 pathogenesis, including direct neutralization of the virus, control of an overactive immune system (i.e., cytokine storm, Th1/Th17 ratio, complement activation) and immunomodulation of a hypercoagulable state. All these benefits of CP are expected to be better achieved if used in non-critically hospitalized patients, in the hope of reducing morbidity and mortality.

Population Pharmacogenomics for Precision Public Health in Colombia.

While genomic approaches to precision medicine hold great promise, they remain prohibitively expensive for developing countries. The precision public health paradigm, whereby healthcare decisions are made at the level of populations as opposed to individuals, provides one way for the genomics revolution to directly impact health outcomes in the developing world. Genomic approaches to precision public health require a deep understanding of local population genomics, which is still missing for many developing countries. We are investigating the population genomics of genetic variants that mediate drug response in an effort to inform healthcare decisions in Colombia. Our work focuses on two neighboring populations with distinct ancestry profiles: Antioquia and Chocó. Antioquia has primarily European genetic ancestry followed by Native American and African components, whereas Chocó shows mainly African ancestry with lower levels of Native American and European admixture. We performed a survey of the global distribution of pharmacogenomic variants followed by a more focused study of pharmacogenomic allele frequency differences between the two Colombian populations. Worldwide, we found pharmacogenomic variants to have both unusually high minor allele frequencies and high levels of population differentiation. A number of these pharmacogenomic variants also show anomalous effect allele frequencies within and between the two Colombian populations, and these differences were found to be associated with their distinct genetic ancestry profiles. For example, the C allele of the single nucleotide polymorphism (SNP) rs4149056 [Solute Carrier Organic Anion Transporter Family Member 1B1 (SLCO1B1)∗5], which is associated with an increased risk of toxicity to a commonly prescribed statin, is found at relatively high frequency in Antioquia and is associated with European ancestry. In addition to pharmacogenomic alleles related to increased toxicity risk, we also have evidence that alleles related to dosage and metabolism have large frequency differences between the two populations, which are associated with their specific ancestries. Using these findings, we have developed and validated an inexpensive allele-specific PCR assay to test for the presence of such population-enriched pharmacogenomic SNPs in Colombia. These results serve as an example of how population-centered approaches to pharmacogenomics can help to realize the promise of precision medicine in resource-limited settings.

Aplicación farmacogenómica de los genes CYP2C19, CYP2C9 y VKORC1 implicados en el metabolismo de los fármacos clopidogrel y warfarina / Pharmacogenomic application of the CYP2C19, CYP2C9 and VKORC1 genes implicated in the metabolism of clopidogrel and warfarin

Resumen El estudio de las variaciones de las secuencias de ADN y ARN en relación con la respuesta a diferentes fármacos, se ha convertido en un área de estudio particularmente prometedora para la aplicación en genómica clínica y estudios de genomas personalizados. Medicamentos de uso diario en el tratamiento de enfermedades cardiovasculares han demostrado variaciones en la respuesta en función de las variantes genéticas de los individuos. Dos fármacos han concentrado el interés mundial: la warfarina, un anticoagulante oral, y el clopidogrel, un antiagregante plaquetario, los cuales actúan alterando diferentes vías que conforman la cascada de la coagulación, ya sea limitando directamente la producción de trombina o bloqueando otros activadores de la ruta. Los cambios genéticos que se han asociado a la reducción de la actividad enzimática de estos fármacos ocurren en los genes, CYP2C19 para clopidogrel y CYP2C9 y VKORC1 para warfarina. Las variaciones genéticas identificadas para estos genes se relacionan con perfiles genotípicos que determinan la dosis requerida para el paciente. Es allí donde ciencias como la farmacogenómica tienen como fin brindar una ayuda diagnóstica más objetiva al optimizar tiempo y recursos, así como disminuir el riesgo del paciente a sufrir complicaciones que comprometan su vida.

Abstract The study of the variations in DNA and RNA sequencing as regards the response to different drugs has become a particularly promising area for their application in clinical genomics and personalised genome studies. Drugs of daily use in the treatment of cardiovascular diseases have shown variations in the response depending on the genetic variations of the individuals. Two drugs have gathered worldwide interest: warfarin, an oral anticoagulant, and clopidogrel, an antiplatelet drug, which act by altering different pathways that constitute the clotting cascade either by directly limiting the production of thrombin, or by blocking other activators of the pathway. The genetic changes that have been associated with the reduction in the enzyme activity of these drugs occur in the genes, CYP2C19 for clopidogrel, and the genes, CYP2C9 and VKORC1 for warfarin. The genetic variations identified for these genes are associated with genotype profiles that determine the dose required by the patient. It is from there, sciences like pharmacogenomics have as their aim to provide a more objective diagnostic aid in order to optimise time and resources, as well as to reduce the risk of the patient suffering complications that may compromise their life.

Actualidad de la genómica clínica en el área cardiovascular en Colombia / Current state of cardiovascular genomics in Colombia

La genómica clínica ha avanzado de forma exponencial en el mundo durante la última década, debido a la innovación en el campo de las técnicas de secuenciación, que han hecho que los procedimientos sean más cada vez más rápidos y económicos. En la actualidad se plantea un escenario en el cual el médico tiene a su disposición ayudas diagnósticas de última generación en técnicas moleculares y de secuenciación. A continuación se revisan brevemente las ayudas diagnósticas basadas en el conocimiento genético del paciente y su estado actual en el mercado colombiano. Antes de las tecnologías de secuenciación de nueva generación, la alternativa para obtener la información genética de un paciente eran los métodos de secuenciación tradicionales, basados en la tecnología Sanger. Para obtener la secuencia de un gen específico, primero se escogía el gen blanco de interés. El médico debía tener una idea sobre cual o cuales de estos genes podían estar involucrados en la enfermedad de su paciente, algo que puede ser relativamente complejo en el momento de tomar una decisión. La secuenciación de cualquier gen puede ser engorrosa debido a la gran diferencia de tamaños y número de exones entre genes, hecho relevante para entender la forma por la cual se secuencian genes por métodos tradicionales. Para estudios genéticos de un gen, en primer lugar se amplifican todas las partes codificantes del mismo, por medio de la técnica de PCR (polymerase chain reaction). Por ejemplo, el gen SCN5A, posee 28 exones con un tamaño aproximado de 80 kb. Esto implica muchos ciclos de amplificación por PCR, situación en la cual se debe amplificar cada exón de forma independiente, incrementando así costos y tiempo de personal capacitado en el área de la biología molecular. En los casos en los que la secuenciación de un solo gen tenga validez clínica, tomando SCN5A como ejemplo, la complejidad de procesos de laboratorio se opaca con el gran beneficio que tendrán los pacientes al conocer la posible causa genética de su enfermedad o evento cardiovascular. Pero, como las condiciones cardiovasculares no son enfermedades clásicas mendelianas, en la mayoría de los casos no es posible tomar una decisión fácil sobre cual gen específico estudiar.

From NGS assembly challenges to instability of fungal mitochondrial genomes: A case study in genome complexity.

The presence of repetitive or non-unique DNA persisting over sizable regions of a eukaryotic genome can hinder the genome's successful de novo assembly from short reads: ambiguities in assigning genome locations to the non-unique subsequences can result in premature termination of contigs and thus overfragmented assemblies. Fungal mitochondrial (mtDNA) genomes are compact (typically less than 100 kb), yet often contain short non-unique sequences that can be shown to impede their successful de novo assembly in silico. Such repeats can also confuse processes in the cell in vivo. A well-studied example is ectopic (out-of-register, illegitimate) recombination associated with repeat pairs, which can lead to deletion of functionally important genes that are located between the repeats. Repeats that remain conserved over micro- or macroevolutionary timescales despite such risks may indicate functionally or structurally (e.g., for replication) important regions. This principle could form the basis of a mining strategy for accelerating discovery of function in genome sequences. We present here our screening of a sample of 11 fully sequenced fungal mitochondrial genomes by observing where exact k-mer repeats occurred several times; initial analyses motivated us to focus on 17-mers occurring more than three times. Based on the diverse repeats we observe, we propose that such screening may serve as an efficient expedient for gaining a rapid but representative first insight into the repeat landscapes of sparsely characterized mitochondrial chromosomes. Our matching of the flagged repeats to previously reported regions of interest supports the idea that systems of persisting, non-trivial repeats in genomes can often highlight features meriting further attention.

The complex task of choosing a de novo assembly: lessons from fungal genomes.

Selecting the values of parameters used by de novo genomic assembly programs, or choosing an optimal de novo assembly from several runs obtained with different parameters or programs, are tasks that can require complex decision-making. A key parameter that must be supplied to typical next generation sequencing (NGS) assemblers is the k-mer length, i.e., the word size that determines which de Bruijn graph the program should map out and use. The topic of assembly selection criteria was recently revisited in the Assemblathon 2 study (Bradnam et al., 2013). Although no clear message was delivered with regard to optimal k-mer lengths, it was shown with examples that it is sometimes important to decide if one is most interested in optimizing the sequences of protein-coding genes (the gene space) or in optimizing the whole genome sequence including the intergenic DNA, as what is best for one criterion may not be best for the other. In the present study, our aim was to better understand how the assembly of unicellular fungi (which are typically intermediate in size and complexity between prokaryotes and metazoan eukaryotes) can change as one varies the k-mer values over a wide range. We used two different de novo assembly programs (SOAPdenovo2 and ABySS), and simple assembly metrics that also focused on success in assembling the gene space and repetitive elements. A recent increase in Illumina read length to around 150 bp allowed us to attempt de novo assemblies with a larger range of k-mers, up to 127 bp. We applied these methods to Illumina paired-end sequencing read sets of fungal strains of Paracoccidioides brasiliensis and other species. By visualizing the results in simple plots, we were able to track the effect of changing k-mer size and assembly program, and to demonstrate how such plots can readily reveal discontinuities or other unexpected characteristics that assembly programs can present in practice, especially when they are used in a traditional molecular microbiology laboratory with a 'genomics corner'. Here we propose and apply a component of a first pass validation methodology for benchmarking and understanding fungal genome de novo assembly processes.

The eukaryotic genome, its reads, and the unfinished assembly.

In recent years, readily affordable short read sequences provided by next-generation sequencing (NGS) have become longer and more accurate. This has led to a jump in interest in the utility of NGS-only approaches for exploring eukaryotic genomes. The concept of a static, 'finished' genome assembly, which still appears to be a faraway goal for many eukaryotes, is yielding to new paradigms. We here motivate an object-view concept where the raw reads are the main, fixed object, and assemblies with their annotations take a role of dynamically changing and modifiable views of that object.

The multiple autoimmune syndromes. A clue for the autoimmune tautology.

The multiple autoimmune syndromes (MAS) consist on the presence of three or more well-defined autoimmune diseases (ADs) in a single patient. The aim of this study was to analyze the clinical and genetic characteristics of a large series of patients with MAS. A cluster analysis and familial aggregation analysis of ADs was performed in 84 patients. A genome-wide microsatellite screen was performed in MAS families, and associated loci were investigated through the pedigree disequilibrium test. Systemic lupus erythematosus (SLE), autoimmune thyroid disease (AITD), and Sjögren's syndrome together were the most frequent ADs encountered. Three main clusters were established. Aggregation for type 1 diabetes, AITD, SLE, and all ADs as a trait was found. Eight loci associated with MAS were observed harboring autoimmunity genes. The MAS represent the best example of polyautoimmunity as well as the effect of a single genotype on diverse phenotypes. Its study provides important clues to elucidate the common mechanisms of ADs (i.e., autoimmune tautology).