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Granulocytic myeloid-derived suppressor cells (G-MDSCs) promote tumor growth and immunosuppression in multiple myeloma (MM). However, their phenotype is not well established for accurate monitoring or clinical translation. We aimed to provide the phenotypic profile of G-MDSCs based on their prognostic significance in MM, immunosuppressive potential, and molecular program. The preestablished phenotype of G-MDSCs was evaluated in bone marrow samples from controls and MM patients using multidimensional flow cytometry; surprisingly, we found that CD11b+CD14−CD15+CD33+HLADR− cells overlapped with common eosinophils and neutrophils, which were not expanded in MM patients. Therefore, we relied on automated clustering to unbiasedly identify all granulocytic subsets in the tumor microenvironment: basophils, eosinophils, and immature, intermediate, and mature neutrophils. In a series of 267 newly diagnosed MM patients (GEM2012MENOS65 trial), only the frequency of mature neutrophils at diagnosis was significantly associated with patient outcome, and a high mature neutrophil/T-cell ratio resulted in inferior progression-free survival (P < .001). Upon fluorescence-activated cell sorting of each neutrophil subset, T-cell proliferation decreased in the presence of mature neutrophils (0.5-fold; P = .016), and the cytotoxic potential of T cells engaged by a BCMA×CD3-bispecific antibody increased notably with the depletion of mature neutrophils (fourfold; P = .0007). Most interestingly, RNA sequencing of the 3 subsets revealed that G-MDSC–related genes were specifically upregulated in mature neutrophils from MM patients vs controls because of differential chromatin accessibility. Taken together, our results establish a correlation between the clinical significance, immunosuppressive potential, and transcriptional network of well-defined neutrophil subsets, providing for the first time a set of optimal markers (CD11b/CD13/CD16) for accurate monitoring of G-MDSCs in MM.

Fasting, caloric restriction and foods or compounds mimicking the biological effects of caloric restriction, known as caloric restriction mimetics, have been associated with a lower risk of age-related diseases, including cardiovascular diseases, cancer and cognitive decline, and a longer lifespan. Reduced calorie intake has been shown to stimulate cancer immunosurveillance, reducing the migration of immunosuppressive regulatory T cells towards the tumor bulk. Autophagy stimulation via reduction of lysine acetylation, increased sensitivity to chemo- and immunotherapy, along with a reduction of insulin-like growth factor 1 and reactive oxygen species have been described as some of the major effects triggered by caloric restriction. Fasting and caloric restriction have also been shown to beneficially influence gut microbiota composition, modify host metabolism, reduce total cholesterol and triglyceride levels, lower diastolic blood pressure and elevate morning cortisol level, with beneficial modulatory effects on cardiopulmonary fitness, body fat and weight, fatigue and weakness, and general quality of life. Moreover, caloric restriction may reduce the carcinogenic and metastatic potential of cancer stem cells, which are generally considered responsible of tumor formation and relapse. Here, we reviewed in vitro and in vivo studies describing the effects of fasting, caloric restriction and some caloric restriction mimetics on immunosurveillance, gut microbiota, metabolism, and cancer stem cell growth, highlighting the molecular and cellular mechanisms underlying these effects. Additionally, studies on caloric restriction interventions in cancer patients or cancer risk subjects are discussed. Considering the promising effects associated with caloric restriction and caloric restriction mimetics, we think that controlled-randomized large clinical trials are warranted to evaluate the inclusion of these non-pharmacological approaches in clinical practice.

This study aims to identify pathway involvement in the development of cisplatin (cis-diamminedichloroplatinum (II); CDDP) resistance in A549 lung cancer (LC) cells by utilizing advanced bioinformatics software. We developed CDDP-resistant A549 (A549/DDP) cells through prolonged incubation with the drug and performed RNA-seq on RNA extracts to determine differential mRNA and miRNA expression between A549/DDP and A549 cells. We analyzed the gene dysregulation with Ingenuity Pathway Analysis (IPA; QIAGEN) software. In contrast to prior research, which relied on the clustering of dysregulated genes to pathways as an indication of pathway activity, we utilized the IPA software for the dynamic evaluation of pathway activity depending on the gene dysregulation levels. We predicted 15 pathways significantly contributing to the chemoresistance, with several of them to have not been previously reported or analyzed in detail. Among them, the PKR signaling, cholesterol biosynthesis, and TEC signaling pathways are included, as well as genes, such as PIK3R3, miR-34c-5p, and MDM2, among others. We also provide a preliminary analysis of SNPs and indels, present exclusively in A549/DDP cells. This study's results provide novel potential mechanisms and molecular targets that can be explored in future studies and assist in improving the understanding of the chemoresistance phenotype.

The development of advanced gene and cell therapies for the treatment of genetic diseases requires reliable animal and cellular models to test their efficacy. Moreover, the availability of the target human primary cells of these therapies is reduced in many diseases. The development of endonucleases that can cut into specific sites of the cell genome, as well as the repair of the generated break by non-homologous end-joining, results in a variety of outcomes, insertions, deletions, and inversions that can induce the disruption of any specific gene. Among the many methods that have been developed for gene editing, CRISPR-Cas9 technology has become one of the most widely used endonuclease tools due to its easy design and its low cost. It has also been reported that the use of two guides, instead of just the one required, reduces the outcomes of non-homologous end joining mainly to the precise genomic sequences between the cutting sites of the guides used. We have explored this strategy to generate useful cellular and animal models. Different distances between the two guides have been tested (from 8 to 500 bp apart), and using the optimal range of 30–60 bp we have obtained a human primary cellular model of a genetic disease, pyruvate kinase deficiency, where the availability of the target cells is limited. We have also generated an in vivo model of glycolate oxidase (GO) deficiency, which is an enzyme involved in the glyoxylate metabolism following the same strategy. We demonstrate that the use of two-guide CRISPR-Cas9-induced non-homologous end joining is a feasible and useful tool for disease modeling, and it is most relevant to those diseases in which it is difficult to get the cells that will be genetically manipulated., Graphical Abstract, A CRISPR-Cas9 system generates precise deletions (up to 90% efficiency) when two guides are used simultaneously by means of non-homologous end joining. The PAM-in/PAM-in Cas9 orientation is the best combination to generate precise deletions. This gene-editing procedure facilitates the generation of homogeneously genetically defined cell and animal models.

Primary effusion lymphoma (PEL) is an aggressive malignancy with poor prognosis even under chemotherapy. Kaposi sarcoma–associated herpesvirus (KSHV), one of the human oncogenic viruses, is the principal causative agent. Currently, there is no specific treatment for PEL; therefore, developing new therapies is of great importance. Sphingolipid metabolism plays an important role in determining the fate of tumor cells. Our previous studies have demonstrated that there is a correlation between sphingolipid metabolism and KSHV+ tumor cell survival. To further develop sphingolipid metabolism-targeted therapy, after screening a series of newly synthesized ceramide analogs, here, we have identified compounds with effective anti-PEL activity. These compounds induce significant PEL apoptosis, cell-cycle arrest, and intracellular ceramide production through regulation of ceramide synthesizing or ceramide metabolizing enzymes and dramatically suppress tumor progression without visible toxicity in vivo. These new compounds also increase viral lytic gene expression in PEL cells. Our comparative transcriptomic analysis revealed their mechanisms of action for inducing PEL cell death and identified a subset of novel cellular genes, including AURKA and CDCA3, controlled by sphingolipid metabolism, and required for PEL survival with functional validation. These data provide the framework for the development of promising sphingolipid-based therapies against this virus-associated malignancy.

Para la propagación de Solanum tuberosum, los tubérculos son la forma más empleada en la mayoría de las zonas agroecológicas. El empleo de esquejes de tallo juvenil es una práctica exclusiva de viveros y empresas agrícolas que hacen uso de hormonas enraizantes, inaccesibles para los pequeños agricultores por su alto costo. Se ha reportado que la auxina sintética 2,4-diclorofenoxiacético (2,4-D) a muy baja concentración induce al enraizamiento de esquejes y puede ser una alternativa viable para la agricultura a pequeña escala. Ante la necesidad de un mayor conocimiento, se propuso como objetivo de investigación evaluar el efecto enraizante del 2,4-D en esquejes de tallo juvenil de S. tuberosum var. yungay. La fase experimental se desarrolló en el invernadero del Instituto de la Papa y Cultivos Andinos de la Universidad Nacional de Trujillo (Perú), con plántulas procedentes del mismo instituto que se convirtieron en plantas madre donadoras de esquejes de tallo juvenil, los cuales fueron sembrados en cama de enraizamiento con diferentes concentraciones de 2,4-D. El análisis estadístico de los resultados demostró que la concentración de 0,3 % generó un promedio mayor en número de raíces (6,52), longitud de raíces (5,31 cm) y altura de plántula (2,63 cm). Se concluyó que el 2,4-D en concentración del 0,3 % produce un mejor enraizamiento de esquejes de tallo juvenil de S. tuberosum var. yungay.

Chloroquine (CQ) and hydroxychloroquine (HCQ) are derivatives of the heterocyclic aromatic compound quinoline. These economical compounds have been used as antimalarial agents for many years. Currently, they are used as monotherapy or in conjunction with other therapies for the treatment of autoimmune diseases such as systemic lupus erythematosus (SLE), rheumatoid arthritis (RA), Sjögren's syndrome (SS) and antiphospholipid antibody syndrome (APS). Based on its effects on the modulation of the autophagy process, various clinical studies suggest that CQ and HCQ could be used in combination with other chemotherapeutics for the treatment of various types of cancer. Furthermore, the antiviral effects showed against Zika, Chikungunya, and HIV are due to the annulation of endosomal/lysosomal acidification. Recently, CQ and HCQ were approved for the U.S. Food and Drug Administration (FDA) for the treatment of infected patients with the coronavirus SARSCoV- 2, causing the disease originated in December 2019, namely COVID-2019. Several mechanisms have been proposed to explain the pharmacological effects of these drugs: 1) disruption of lysosomal and endosomal pH, 2) inhibition of protein secretion/expression, 3) inhibition of antigen presentation, 4) decrease of proinflammatory cytokines, 5) inhibition of autophagy, 6) induction of apoptosis and 7) inhibition of ion channels activation. Thus, evidence has shown that these structures are leading molecules that can be modified or combined with other therapeutic agents. In this review, we will discuss the most recent findings in the mechanisms of action of CQ and HCQ in the immune system, and the use of these antimalarial drugs on diseases.

Private well water systems in rural areas that are improperly maintained will result in poor drinking water quality, loss of water supply, and pose human health risk. The purpose of this study was to investigate the occurrence of fecal indicator bacteria (FIB) and opportunistic pathogens in private well water in rural areas surrounding New Orleans, Louisiana. Our results confirmed the ubiquitous nature of Legionella (86.7%) and mycobacteria (68.1%) in private well water in the study area, with gene concentration ranged from 0.60 to 5.53 and 0.67 to 5.95 Log10 of GC/100 mL, respectively. Naegleria fowleri target sequence was detected in 16.8% and Escherichia coli was detected in 43.4% of the water samples. Total coliform, as well as Legionella and mycobacteria genetic markers' concentrations were significantly reduced by 3-minute flushing. Next-generation sequencing (NGS) data indicated that the abundance of bacterial species was significantly increased in water collected in kitchens compared with samples from wells directly. This study provided integrated knowledge on the persistence of pathogenic organisms in private well water. Further study is needed to explore the presence of clinical species of those opportunistic pathogens in private well water systems to elucidate the health risk.

Capsicum chinense Jacq. “ají mochero” es catalogado como patrimonio gastronómico de Perú por su agradable grado de picor, que condimenta numerosos platos reconocidos internacionalmente. El boom gastronómico en Perú ha conllevado a la exportación de diferentes insumos nacionales, incluyendo el ají mochero, por lo que se requieren estudios agronómicos que brinden información sobre el estado productivo de los frutos y las semillas de este cultivo. El estudio buscó determinar la morfometría de los frutos y las semillas de C. chinense “ají mochero”. Para ello, se recolectaron frutos procedentes del distrito de Moche (provincia de Trujillo, departamento de La Libertad) y en el laboratorio se evaluó el peso del fruto y de la pulpa, así como las dimensiones y el número de semillas. Se realizó un análisis estadístico descriptivo con 50 repeticiones por cada componente evaluado. El peso promedio del fruto fue de 2,4 g y el peso promedio de la pulpa, de 2,1 g. La semilla registró una longitud promedio de 4,3 mm y un ancho promedio de 1,4 mm, y se reportaron 16,4 semillas por fruto. Se identificó un alto coeficiente de variación y una alta correlación (p

Adeno-associated virus (AAV) is a highly promising gene transfer vector, yet major cellular requirements for AAV entry are poorly understood. Using a genome-wide CRISPR screen for entry of evolutionarily divergent serotype AAVrh32.33, we identified GPR108, a member of the G protein-coupled receptor superfamily, as an AAV entry factor. Of greater than 20 divergent AAVs across all AAV clades tested in human cell lines, only AAV5 transduction was unaffected in the GPR108 knockout (KO). GPR108 dependency was further shown in murine and primary cells in vitro. These findings are further validated in vivo, as the Gpr108 KO mouse demonstrates 10- to 100-fold reduced expression for AAV8 and rh32.33 but not AAV5. Mechanistically, both GPR108 N- and C-terminal domains are required for transduction, and on the capsid, a VP1 unique domain that is not conserved on AAV5 can be transferred to confer GPR108 independence onto AAV2 chimeras. In vitro binding and fractionation studies indicate reduced nuclear import and cytosolic accumulation in the absence of GPR108. We thus have identified the second of two AAV entry factors that is conserved between mice and humans relevant both in vitro and in vivo, further providing a mechanistic understanding to the tropism of AAV gene therapy vectors.

tRNA-derived fragments (tRFs) have been defined as a novel class of small noncoding RNAs. tRFs have been reported to be deregulated in cancer, but their biologic function remains to be fully unders...

Introduction: Tuberculosis continues to be a public health priority. Indigenous peoples are vulnerable groups with cultural determinants that increase the risk of the disease. Objective: To determine molecular epidemiology and phenotypical features and of Mycobacterium tuberculosis isolates from indigenous people in Colombia during the period from 2009 to 2014. Materials and methods: We conducted an analytical observational study; we analyzed 234 isolates to determine their patterns of sensitivity to antituberculosis drugs and their molecular structures by spoligotyping. Results: The isolates came from 41 indigenous groups, predominantly the Wayúu (13.10%) and Emberá Chamí (11.35%). We found 102 spoligotypes distributed among seven genetic families (37.2% LAM, 15.8% Haarlem, 8.1% T, 3.4% U, 2.6% S, 2.1% X, and 0.9%, Beijing). The association analysis showed that the non-clustered isolates were related to prior treatment, relapse, orphan spoligotypes, and the Beijing family. The H family presented an association with the Arhuaco and Camëntŝá indigenous groups, the U family was associated with the Wounaan group, and the T family was associated with the Motilón Barí group. Conclusions: This is the first national study on M. tuberculosis characterization in indigenous groups. The study evidenced that diagnosis in indigenous people is late. We described 53% of orphan patterns that could be typical of the Colombian indigenous population. The high percentage of grouping by spoligotyping (62%) could indicate cases of active transmission, a situation that should be corroborated using a second genotyping marker. A new Beijing spoligotype (Beijing-like SIT 406) was identified in Colombia. RESUMEN Introducción. La tuberculosis es prioridad de salud pública. Los pueblos indígenas son vulnerables debido a los factores culturales determinantes que aumentan el riesgo de tuberculosis. Objetivo. Determinar la epidemiologia molecular y las características fenotípicas de los aislamientos de Mycobacterium tuberculosis de pueblos indígenas de Colombia entre 2009 y 2014. Materiales y métodos. Se hizo un estudio observacional analítico; se analizaron 234 aislamientos para determinar la sensibilidad a los fármacos antituberculosos y la estructura molecular usando spoligotyping. La información epidemiológica se recolectó utilizando el formato único de vigilancia de micobacterias. Resultados. Los aislamientos provenían de 41 grupos indígenas, principalmente los wayúu (13,10 %) y emberá chamí (11,35 %). Se encontraron 102 genotipos distribuidos en siete familias genéticas (37,2 %, LAM; 15,8 %, Haarlem; 8,1 %, T; 3,4 %, U; 2,6 %, S; 2,1 %, X, y 0,9%, Beijing). El análisis de asociación mostró que los aislamientos no agrupados se asociaron con el tratamiento previo, las recaídas, los genotipos huérfanos y la familia Beijing. La familia H presentó una asociación con los grupos indígenas arhuaco y camëntŝá, la familia U se asoció con el grupo wounaan y la familia T con el grupo motilón barí. Conclusiones. Este es el primer estudio nacional de caracterización de M. tuberculosis en grupos indígenas. Se evidenció que el diagnóstico en indígenas es tardío, y que 53 % de los patrones huérfanos podrían ser típicos de la población indígena colombiana. El alto porcentaje de agrupamiento por spoligotyping (62%) podría indicar casos de transmisión activa, una situación que debe ser corroborada usando un segundo marcador de genotipificación. Se identificó un nuevo genotipo (Beijing-like SIT 406) en Colombia.

Next generation sequencing provides new insights into the diversity and ecophysiology of bacteria communities throughout wastewater treatment plants (WWTP), as well as the fate of pathogens in wastewater treatment system. In the present study, we investigated the bacterial communities and human-associated Bacteroidales (HF183) marker in two WWTPs in North America that utilize Bardenpho treatment processes. Although, most pathogens were eliminated during wastewater treatment, some pathogenic bacteria were still observed in final effluents. The HF183 genetic marker demonstrated significant reductions between influent and post-Bardenpho treated samples in each WWTP, which coincided with changes in bacteria relative abundances and community compositions. Consistent with previous studies, the major phyla in wastewater samples were predominantly comprised by Proteobacteria (with Gammaproteobacteria and Alphaproteobacteria among the top two classes), Actinobacteria, Bacteroidetes, and Firmicutes. Dominant genera were often members of Proteobacteria and Firmicutes, including several pathogens of public health concern, such as Pseudomonas, Serratia, Streptococcus, Mycobacterium and Arcobacter. Pearson correlations were calculated to observe the seasonal variation of relative abundances of gene sequences at different levels based on the monthly average temperature. These findings profile how changes in bacterial communities can function as a robust method for monitoring wastewater treatment quality and performance for public and environmental health purposes.

Special Issue on Ecophysiology of Extremophiles., Microbial communities in hypersaline underground waters derive from ancient organisms trapped within the evaporitic salt crystals and are part of the poorly known subterranean biosphere. Here, we characterized the viral and prokaryotic assemblages present in the hypersaline springs that dissolve Triassic-Keuper evaporite rocks and feed the Añana Salt Valley (Araba/Alava, Basque Country, Spain). Four underground water samples (around 23% total salinity) with different levels of exposure to the open air were analysed by means of microscopy and metagenomics. Cells and viruses in the spring water had lower concentrations than what are normally found in hypersaline environments and seemed to be mostly inactive. Upon exposure to the open air, there was an increase in activity of both cells and viruses as well as a selection of phylotypes. The underground water was inhabited by a rich community harbouring a diverse set of genes coding for retinal binding proteins. A total of 35 viral contigs from 15 to 104 kb, representing partial or total viral genomes, were assembled and their evolutionary changes through the spring system were followed by SNP analysis and metagenomic island tracking. Overall, both the viral and the prokaryotic assemblages changed quickly upon exposure to the open air conditions., This work was funded by the Spanish Ministry of Science, Innovation and Universities grant MICROMATES (PGC2018-096956-B-C41 and C44, to J.A./F.S. and R.R.-M.), which was also supported with European Regional Development Fund (FEDER) funds, and by the Generalitat Valenciana grant PROMETEO/2017/129.

Background Optimal control of acute postoperative pain and prevention of chronic persistent pain in total hip arthroplasty (THA) remain a challenge. The main hypothesis was that peripheral nerve blocks improve postoperative analgesia. Methods Immediate postoperative pain (24 hours) was evaluated every hour in 510 patients using a verbal rating 11-point scale for patient self-reporting of pain (VRS-11). All patients received subarachnoid anesthesia (SA) and were randomly allocated in six groups: SA with morphine 0.1 (SA0.1) or 0.2 mg (SA0.2), fascia iliaca compartment block with dexamethasone 4 mg + levobupivacaine 0.375% 20 (FICB20) or 30 mL (FICB30), lateral femoral cutaneous nerve block with levobupivacaine 0.25% 5 ml (LFCNB) and FICB20+LFCNB. Standardized analgesia included intravenous metamizole magnesium, dexketoprofen and rescue with paracetamol and morphine, and/or regional rescue (FICB, LFCNB, femoral and sciatic nerve blocks). Results 37.5% of patients had at least one episode of pain, 31.3% of them needed rescue analgesia while the remaining 6.2% did not request analgesia. There were no significant differences between the groups in paracetamol, morphine and rescue nerve blocks requirements. There was pain only in 5.4% of the total PACU pain records: 3.1% mild pain, 1.7% moderate pain and 0.6% severe pain. Conclusions Combined with a multimodal analgesic approach, infra-inguinal FICB and LFCNB did not improve immediate postoperative analgesia for THA in our hospital. Other options and longer-term studies should be more extensively investigated to determine the role of peripheral blocks in postoperative pain treatment protocols.

Here, we present the draft genome sequence of strain UYCP14C, a rhizobium isolated from Calliandra parvifolia nodules. The assembled genome size was around 9.8 million bp, containing 9,031 predicted protein-coding sequences, including several symbiotic and nitrogen fixation genes., Here, we present the draft genome sequence of strain UYCP14C, a rhizobium isolated from Calliandra parvifolia nodules. The assembled genome size was around 9.8 million bp, containing 9,031 predicted protein-coding sequences, including several symbiotic and nitrogen fixation genes. UYCP14C appears to be a novel species of the plant growth-promoting Paraburkholderia genus.

Modern high-throughput ‘omics’ science tools (including genomics, transcriptomics, proteomics, metabolomics and microbiomics) are currently being applied to nutritional sciences to unravel the fundamental processes of health effects ascribed to particular nutrients in humans and to contribute to more precise nutritional advice. Diet and food components are key environmental factors that interact with the genome, transcriptome, proteome, metabolome and the microbiota, and this life-long interplay defines health and diseases state of the individual. Rheumatoid arthritis (RA) is a chronic autoimmune disease featured by a systemic immune-inflammatory response, in genetically susceptible individuals exposed to environmental triggers, including diet. In recent years increasing evidences suggested that nutritional factors and gut microbiome have a central role in RA risk and progression. The aim of this review is to summarize the main and most recent applications of ‘omics’ technologies in human nutrition and in RA research, examining the possible influences of some nutrients and nutritional patterns on RA pathogenesis, following a nutrigenomics approach. The opportunities and challenges of novel ‘omics technologies’ in the exploration of new avenues in RA and nutritional research to prevent and manage RA will be also discussed.

Diatoms are unicellular organisms containing red algal-derived plastids that probably originated as result of serial endosymbioses between an ancestral heterotrophic organism and a red alga or cryptophyta algae from which has only the chloroplast left. Diatom mitochondria are thus believed to derive from the exosymbiont. Unlike animals and fungi, diatoms seem to contain ancestral respiratory chains. In support of this, genes encoding gamma type carbonic anhydrases (CAs) whose products were shown to be intrinsic complex I subunits in plants, Euglena and Acanthamoeba were found in diatoms, a representative of Stramenopiles. In this work, we experimentally show that mitochondrial complex I in diatoms is a large complex containing gamma type CA subunits, supporting an ancestral origin. By using a bioinformatic approach, a complex I integrated CA domain with heterotrimeric subunit composition is proposed.

Objectives A high incidence of pulmonary embolism (PE) is reported in patients with critical coronavirus disease 2019 (COVID‐19). Neutrophils may contribute to this through a process referred to as immunothrombosis. The aim of this study was to investigate the occurrence of neutrophil subpopulations in blood preceding the development of COVID‐19 associated PE. Methods We studied COVID‐19 patients admitted to the ICU of our tertiary hospital between 19‐03‐2020 and 17‐05‐2020. Point‐of‐care fully automated flow cytometry was performed prior to ICU admission, measuring the neutrophil activation/maturation markers CD10, CD11b, CD16 and CD62L. Neutrophil receptor expression was compared between patients who did or did not develop PE (as diagnosed on CT angiography) during or after their ICU stay. Results Among 25 eligible ICU patients, 22 subjects were included for analysis, of whom nine developed PE. The median (IQR) time between neutrophil phenotyping and PE occurrence was 9 (7‐12) days. A significant increase in the immune‐suppressive neutrophil phenotype CD16bright/CD62Ldim was observed on the day of ICU admission (P = 0.014) in patients developing PE compared to patients who did not. Conclusion The increase in this neutrophil phenotype indicates that the increased number of CD16bright/CD62Ldim neutrophils might be used as prognostic marker to predict those patients that will develop PE in critical COVID‐19 patients.

SummaryThe SARS-CoV-2 pandemic has affected more than 70 million people worldwide and resulted in over 1.5 million deaths. A broad deployment of effective immunization campaigns to achieve population immunity at global scale will depend on the biological and logistical attributes of the vaccine. Here, two adeno-associated viral (AAV)-based vaccine candidates demonstrate potent immunogenicity in mouse and nonhuman primates following a single injection. Peak neutralizing antibody titers remain sustained at 5 months and are complemented by functional memory T-cells responses. The AAVrh32.33 capsid of the AAVCOVID vaccine is an engineered AAV to which no relevant pre-existing immunity exists in humans. Moreover, the vaccine is stable at room temperature for at least one month and is produced at high yields using established commercial manufacturing processes in the gene therapy industry. Thus, this methodology holds as a very promising single dose, thermostable vaccine platform well-suited to address emerging pathogens on a global scale.

Inflammation is a key mechanism of the immune system that can be elicited by several factors, among them several chemical, physical and biological agents. Once stimulated, the inflammatory response activates a series of signaling pathways and a number of immune cells which promote, in a very coordinated manner, the neutralization of the infectious agent. However, if uncontrolled, the inflammatory status may become chronic leading, potentially, to tissue damage and disease onset. Several risk factors are associated with the development of chronic inflammation and, among these factors, diet plays an essential role. In this chapter the effects of some dietary bioactive compounds, including micronutrients, omega-3 fatty acids, nucleotides and polyphenols, on the immunoinflammatory responses in different cellular, animal and human studies have been summarized.

Objetivo. Evaluar la capacidad operativa de nueve pruebas serológicas rápidas para detectar la respuesta de anticuerpos IgM/IgG en el suero de pacientes con SARS-CoV-2 en diferentes etapas clínicas. Métodos. Se diseñó un estudio transversal de las pruebas serológicas rápidas para comparar su rendimiento respecto del diagnóstico del SARS-CoV-2. Se utilizaron en total 293 muestras, inclusive muestras de suero de pacientes negativos, asintomáticos y sintomáticos. Resultados. La sensibilidad de las pruebas evaluadas fue baja y moderada en las muestras de suero del grupo de pacientes asintomáticos y en el grupo de pacientes con menos de 11 días desde el inicio de los síntomas. La especificidad de las pruebas de anticuerpos anti-SARS-CoV-2 varió entre 86,5%-99% para la IgM y 86,5%-99,5% para la IgG. La sensibilidad y la razón de verosimilitud (likelihood ratio) fueron diferentes según los grupos de estudio. La utilidad de estas pruebas se limita a los pacientes sintomáticos y su sensibilidad es superior al 85% después de 11 días de la aparición de los síntomas. Conclusiones. Las pruebas serológicas no son una estrategia adecuada para la identificación de los pacientes asintomáticos y presintomáticos. Las pruebas serológicas rápidas para la detección de anticuerpos específicos anti-SARS-CoV-2 pueden ser utilizadas como ayuda diagnóstica, pero el diagnóstico debe ser confirmado por RT-PCR. Las pruebas rápidas deben reservarse para los pacientes con síntomas que duren más de 11 días. Objective. To evaluate the operative capacity of nine serological rapid tests to detect the IgM/IgG antibodies response in serum from patients with SARS-CoV-2 in different clinical stages. Methods. A cross-sectional study of serological rapid tests was designed to compare the performance of the evaluated immunochromatographic tests for the diagnosis of SARS-CoV-2. A total of 293 samples was used, including negatives, asymptomatic, and symptomatic serum samples. Results. The sensitivity of the evaluated tests was low and moderate in the groups of asymptomatic serum samples and the group of serums coming from patients with less than 11 days since the onset of the symptoms. The specificity for the anti-SARS-CoV-2 antibodies tests ranged between 86.5%-99% for IgM and 86.5%-99.5% for IgG. The sensitivity and the likelihood ratio were different according to the study groups. The usefulness of these tests is restricted to symptomatic patients and their sensitivity is greater than 85% after 11 days from the appearance of symptoms. Conclusions. Serological tests are not an adequate strategy for the identification of asymptomatic and pre-symptomatic patients. Serological rapid tests for the detection of specific anti-SARS-CoV-2 antibodies can be used as a diagnostic aid, but diagnosis must be confirmed by RT-PCR. Rapid tests should be reserved for patients with symptoms lasting more than 11 days.

In flowering plants, pollen tubes undergo a journey that starts in the stigma and ends in the ovule with the delivery of the sperm cells to achieve double fertilization. The pollen cell wall plays an essential role to accomplish all the steps required for the successful delivery of the male gametes. This extended path involves female tissue recognition, rapid hydration and germination, polar growth, and a tight regulation of cell wall synthesis and modification, as its properties change not only along the pollen tube but also in response to guidance cues inside the pistil. In this review, we focus on the most recent advances in elucidating the molecular mechanisms involved in the regulation of cell wall synthesis and modification during pollen germination, pollen tube growth, and rupture. Fil: Cascallares, María Milagros. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina Fil: Setzes, Nicolás. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina Fil: Marchetti, Maria Fernanda. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina Fil: Lopez, Gabriel Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina Fil: Distefano, Ayelen Mariana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina Fil: Cainzos, Maximiliano. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina Fil: Zabaleta, Eduardo Julian. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina Fil: Pagnussat, Gabriela Carolina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina

Intensive land use favors eutrophication processes and algae bloom proliferation in freshwaters, which is considered to be one of the main environmental issues worldwide. In general, and particularly in South America, inland water monitoring only covers the main water bodies due to the high costs and efforts involved. In order to improve the coverage of spatial and temporal of algae bloom monitoring, remote sensing serves as an alternative tool. Thereby, the analysis of significant spatial clusters of high values (hotspots) and low values (coldspots) of chlorophyll-a has been applied in coastal studies; however, at present, there are no studies in freshwaters. In this study, Getis-Ord Gi* hotspot analysis was applied to detect spatial distribution patterns of algae bloom dynamics in small- and medium-sized freshwater bodies. Four in situ samplings were carried out in five suburban lakes of Uruguay, in agreement with the satellite capture. Total and cyanobacterial chlorophyll-a concentration, and suspended solids were evaluated. Linear models were developed by combining pre-established indexes with additional Sentinel-2 spectral bands and in situ data. The relationship between red and red edge regions allowed mapping the chlorophyll-a in the study lakes with an adjustment of R2 = 0.83. Hotspot analysis was performed with the selected linear model, and significant chlorophyll-a variability within each lake was successfully detected. The novel application of hotspots analyses presented in this work represents a contribution to advance knowledge in the remote detection of algae bloom dynamics and improve monitoring capabilities of inland water bodies.

The cytochrome P450 superfamily is a large enzymatic protein family that is widely distributed along diverse kingdoms. In plants, cytochrome P450 monooxygenases (CYPs) participate in a vast array of pathways leading to the synthesis and modification of multiple metabolites with variable and important functions during different stages of plant development. This includes the biosynthesis and degradation of a great assortment of compounds implicated in a variety of physiological responses, such as signaling and defense, organ patterning and the biosynthesis of structural polymers, among others. In this review, we summarize the characteristics of the different families of plant CYPs, focusing on the most recent advances in elucidating the roles of CYPs in plant growth and development and more specifically, during plant gametogenesis, fertilization and embryogenesis.

Several dinucleotide cyclases, including cyclic GMP–AMP synthase, and their involvement in STING-mediated immunity have been extensively studied. In this study, we tested five bacterial diguanylate cyclases from the Gram-negative bacterium Salmonella Enteritidis, identifying AdrA as the most potent inducer of a STING-mediated IFN response. AdrA wild-type (wt) or its inactive version AdrA mutant (mut) were delivered by an adenovirus (Ad) vector. Dendritic cells obtained from wt mice and infected in vitro with Ad vector containing AdrA wt, but not mut, had increased activation markers and produced large amounts of several immunostimulatory cytokines. For dendritic cells derived from STING-deficient mice, no activation was detected. The potential antiviral activity of AdrA was addressed in hepatitis B virus (HBV)-transgenic and adenovirus-associated virus (AAV)-HBV mouse models. Viremia in serum of Ad AdrA wt–treated mice was reduced significantly compared with that in Ad AdrA mut–injected mice. The viral load in the liver at sacrifice was in line with this finding. To further elucidate the molecular mechanism(s) by which AdrA confers its antiviral function, the response in mice deficient in STING or its downstream effector molecules was analyzed. wt and IFN-αR (IFNAR)−/− animals were additionally treated with anti–TNF-α (Enbrel). Interestingly, albeit less pronounced than in wt mice, in IFNAR−/− and Enbrel-treated wt mice, a reduction of serum viremia was achieved—an observation that was lost in anti–TNF-α–treated IFNAR−/− animals. No effect of AdrA wt was seen in STING-deficient animals. Thus, although STING is indispensable for the antiviral activity of AdrA, type I IFN and TNF-α are both required and act synergistically.

Production of polyhydroxyalkanoates (PHAs) as bio-alternative to petroleum-based plastics is an important field in the biorefinery to move forward in the development of the circular economy. PHAs are bioplastics stored inside microbial cells as carbon reservoirs and can be produced from a broad range of renewable resources such as waste streams. One important waste stream is food waste that can be converted into volatile fatty acids (VFAs) by anaerobic digestion. The produced effluent from food waste is not only rich in VFAs but also, other nutrients such as nitrogen and phosphorus that can be used by the microorganisms to produce PHAs. The aim of this research is to convert VFAs produced from food waste into PHAs, in which two approaches have been studied. The first approach was to use microbial mixed cultures (MMCs) while the second used microbial pure cultures. The MMCs were enriched in sequencing batch bioreactor cultivations, where nitrogen and carbon starvation were combined to enhance the selection phase. PHA accumulation of the selected cultures was studied in nitrogen-limited fed-batch cultivations. The second approach studied five different PHA producing bacteria: Cupriavidus necator, Burkholderia cepacea, Bacillus megaterium, Bacillus cereus and Bacillus cereus. To select the most promising bacteria, synthetic medium with the same VFAs composition as in MMCs study was used for pre-screening experiments. Both, pure and mixed culture studies, resulted in the production of PHAs containing (R)-3-hydroxybutyrate, (R)-3-hydroxyvalerate and (R)-3-hydroxyhexanoate as monomers and VFAs were consumed with a high rate by the microorganisms.

Regulated cell death plays key roles during essential processes along the plant life cycle. It takes part of specific developmental programs and maintains the organism homeostasis in response to unfavorable environments. Ferroptosis is a recently discovered iron-dependent cell death pathway characterized by the accumulation of lipid reactive oxygen species. Ferroptosis in plants shares all the main hallmarks described for ferroptosis in other systems. Those specific features include biochemical and morphological signatures that seem conserved among species. However, plant cells have specific metabolic pathways and a high degree of metabolic compartmentalization. Together with their particular morphology, these features add more complexity to the plant ferroptosis pathway. In this review, we summarize the most recent advances in elucidating the roles of ferroptosis in plants, focusing on specific triggers, main players and underlying pathways. Fil: Distefano, Ayelen Mariana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina Fil: Lopez, Gabriel Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina Fil: Setzes, Nicolás. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina Fil: Marchetti, Maria Fernanda. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina Fil: Cainzos Gelabert, Maximiliano Guillermo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina Fil: Cascallares, María Milagros. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina Fil: Zabaleta, Eduardo Julian. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina Fil: Pagnussat, Gabriela Carolina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina

Development of advanced gene and cell therapies for the treatment of genetic diseases requires confident animal and cellular models to test their efficacy and is crucial in the cases where no primary samples from patients are available. CRISPR/Cas9 technology, has become one of the most spread endonuclease tools for editing the genome at will. Moreover, it is known that the use of two guides tends to produce the precise deletion between the guides via NHEJ. Different distances between guides were tested (from 8 to 500 base pairs). We found that distances between the two cutting sites and orientation of Cas9 protein-DNA interaction are important for the efficiency within the optimal range (30-60 bp), we could obtain new genetically reproducible models for two rare disease, a Pyruvate Kinase Deficiency model, using human primary cells, and a (forin vivoprimary hyperoxaluria therapeutic mouse model. We demonstrate that the use of a 2-guideRNAs at the optimal distance and orientation is a powerful strategy for disease modelling in those diseases where the availability of primary cells is limited.

Funder: Instituto Nacional de Salud, Bogotá, Colombia, Salmonella Typhi (S. Typhi) is the causative agent of typhoid fever; a systemic disease affecting ~20 million people per year globally. There are little data regarding the contemporary epidemiology of typhoid in Latin America. Consequently, we aimed to describe some recent epidemiological aspects of typhoid in Colombia using cases reported to the National Public Health Surveillance System (Sivigila) between 2012 and 2015. Over the four-year reporting period there were 836 culture confirmed cases of typhoid in Colombia, with the majority (676/836; 80.1%) of reported cases originated from only seven departments. We further characterized 402 S. Typhi isolates with available corresponding data recovered from various departments of Colombia through antimicrobial susceptibility testing and molecular subtyping. The majority (235/402; 58.5%) of these typhoid cases occurred in males and were most commonly reported in those aged between 10 and 29 years (218/402; 54.2%); there were three (0.74%) reported fatalities. The overwhelming preponderance (339/402; 84.3%) of S. Typhi were susceptible to all tested antimicrobials. The most common antimicrobial to which the organisms exhibited non-susceptibility was ampicillin (30/402;7.5%), followed by nalidixic acid (23/402, 5.7%). Molecular subtyping identified substantial genetic diversity, which was well distributed across the country. Despite the diffuse pattern of S. Typhi genotypes, we identified various geographical hotspots of disease associated with local dominant genotypes. Notably, we found limited overlap of Colombian genotypes with organisms reported in other Latin American countries. Our work highlights a substantial burden of typhoid in Colombia, characterized by sustained transmission in some regions and limited epidemics in other departments. The disease is widely distributed across the country and associated with multiple antimicrobial susceptible genotypes that appear to be restricted to Colombia. This study provides a current perspective for typhoid in Latin America and highlights the importance of pathogen-specific surveillance to add insight into the limited epidemiology of typhoid in this region.

OBJECTIVES To assess the impact of a history of liver metastases on survival in patients undergoing surgery for lung metastases from colorectal carcinoma. METHODS We reviewed recent studies identified by searching MEDLINE and EMBASE using the Ovid interface, with the following search terms: lung metastasectomy, pulmonary metastasectomy, lung metastases and lung metastasis, supplemented by manual searching. Inclusion criteria were that the research concerned patients with lung metastases from colorectal cancer undergoing surgery with curative intent, and had been published between 2007 and 2014. Exclusion criteria were that the paper was a review, concerned surgical techniques themselves (without follow-up), and included patients treated non-surgically. Using Stata 14, we performed aggregate data and individual data meta-analysis using random-effect and Cox multilevel models respectively. RESULTS We collected data on 3501 patients from 17 studies. The overall median survival was 43 months. In aggregate data meta-analysis, the hazard ratio for patients with previous liver metastases was 1.19 (95% CI 0.90-1.47), with low heterogeneity (I2 4.3%). In individual data meta-analysis, the hazard ratio for these patients was 1.37 (95% CI 1.14-1.64; p

N-ethyl perfluorooctane sulfonamide (EtFOSA) is the active ingredient of Sulfluramid, a pesticide which is used extensively in South America for control of leaf-cutting ants. Despite being a known precursor to perfluorooctanesulfonate (PFOS), the importance of EtFOSA as a source of environmental PFOS remains unclear. In the present work, uptake, leaching, and biodegradation of EtFOSA and its transformation products were assessed over 81 days in soil-carrot ( Daucus carota ssp sativus) mesocosms for the first time. Experiments performed in the presence of carrot produced PFOS yields of up to 34% using a technical EtFOSA standard and up to 277% using Grão Forte, a commercial Sulfluramid bait formulation containing 0.0024% EtFOSA. Perfluorooctane sulfonamido acetate (FOSAA), perfluorooctane sulfonamide (FOSA), and perfluorooctanoic acid (PFOA) also formed over the course of the experiments, with the latter substance attributed to the presence of perfluorooctanamide impurities. The leachate contained low levels of transformation products and a high FOSA:PFOS ratio, consistent with recent observations in Brazilian surface water. In carrots, the more hydrophilic transformation products (e.g., PFOS) occurred primarily in the leaves, while the more hydrophobic products (e.g., FOSA, FOSAA, and EtFOSA) occurred in the peel and core. Remarkably, isomer-specific analysis revealed that the linear EtFOSA isomer biodegraded significantly faster than branched isomers. These data collectively show that the application of Sulfluramid baits can lead to the occurrence of PFOS in crops and in the surrounding environment, in considerably higher yields than previously thought.

In the gene therapy field, re-administration of adeno-associated virus (AAV) is an important topic because a decrease in therapeutic protein expression might occur over time. However, an efficient re-administration with the same AAV serotype is impossible due to serotype-specific, anti-AAV neutralizing antibodies (NABs) that are produced after initial AAV treatment. To address this issue, we explored the feasibility of using chimeric AAV serotype 5 (AAV5ch) and AAV1 for repeated liver-targeted gene delivery. To develop a relevant model, we immunized animals with a high dose of AAV5ch-human secreted embryonic alkaline phosphatase (hSEAP) that generates high levels of anti-AAV5ch NAB. Secondary liver transduction with the same dose of AAV1-human factor IX (hFIX) in the presence of high levels of anti-AAV5ch NAB proved to be successful because expression/activity of both reporter transgenes was observed. This is the first time that two different transgenes are shown to be produced by non-human primate (NHP) liver after sequential administration of clinically relevant doses of both AAV5ch and AAV1. The levels of transgene proteins achieved after delivery with AAV5ch and AAV1 illustrate the possibility of both serotypes for liver targeting. Furthermore, transgene DNA and RNA biodistribution patterns provided insight into the potential cause of decrease or loss of transgene protein expression over time in NHPs.

// Jacob M. Loupe 3, * , Patrick J. Miller 4, * , Judy S. Crabtree 1 , Jovanny Zabaleta 2 and Andrew D. Hollenbach 1 1 Louisiana State University Health Sciences Center, Department of Genetics, New Orleans, LA 70112, USA 2 Louisiana State University Health Sciences Center, Department of Pediatrics and Stanley S. Scott Cancer Center, New Orleans, LA 70112, USA 3 Current/Present address: Center for Human Genetic Research, Massachusetts General Hospital, Richard B. Simches Research Center, Boston, MA 02114, USA 4 Current/Present address: Tulane University, New Orleans, LA 70112, USA * These authors have contributed equally to this work Correspondence to: Andrew D. Hollenbach, email: aholle@lsuhsc.edu Keywords: Alveolar Rhabdomyosarcoma, myogenesis, PAX3-FOXO1 Received: April 28, 2017 Accepted: July 03, 2017 Published: July 29, 2017 ABSTRACT The differentiation status of tumors is used as a prognostic indicator, with tumors comprised of less differentiated cells exhibiting higher levels of aggressiveness that correlate with a poor prognosis. Although oncogenes contribute to blocking differentiation, it is not clear how they globally alter miRNA expression during differentiation to achieve this result. The pediatric sarcoma Alveolar Rhabdomyosarcoma, which is primarily characterized by the expression of the PAX3-FOXO1 oncogenic fusion protein, consists of undifferentiated muscle cells. However, it is unclear what role PAX3-FOXO1 plays in promoting the undifferentiated state. We demonstrate that expression of PAX3-FOXO1 globally alters the expression of over 80 individual miRNA during early myogenic differentiation, resulting in three primary effects: 1) inhibition of the expression of 51 miRNA essential for promoting myogenesis, 2) promoting the aberrant expression of 43 miRNA not normally expressed during myogenesis, and 3) altering the expression pattern of 39 additional miRNA. Combined, these changes are predicted to have an overall negative effect on myogenic differentiation. This is one of the first studies describing how an oncogene globally alters miRNA expression to block differentiation and has clinical implications for the development of much needed multi-faceted tumor-specific therapeutic regimens.

The interaction between nutrition and human infectious diseases has always been recognized. With the emergence of molecular tools and post-genomics, high-resolution sequencing technologies, the gut microbiota has been emerging as a key moderator in the complex interplay between nutrients, human body, and infections. Much of the host–microbial and nutrition research is currently based on animals or simplistic in vitro models. Although traditional in vivo and in vitro models have helped to develop mechanistic hypotheses and assess the causality of the host–microbiota interactions, they often fail to faithfully recapitulate the complexity of the human nutrient–microbiome axis in gastrointestinal homeostasis and infections. Over the last decade, remarkable progress in tissue engineering, stem cell biology, microfluidics, sequencing technologies, and computing power has taken place, which has produced a new generation of human-focused, relevant, and predictive tools. These tools, which include patient-derived organoids, organs-on-a-chip, computational analyses, and models, together with multi-omics readouts, represent novel and exciting equipment to advance the research into microbiota, infectious diseases, and nutrition from a human-biology-based perspective. After considering some limitations of the conventional in vivo and in vitro approaches, in this review, we present the main novel available and emerging tools that are suitable for designing human-oriented research.

Metaplastic breast carcinoma (MBC) is a clinically aggressive and rare subtype of breast cancer, with similar features to basal-like breast cancers. Due rapid growth rates and characteristic heterogeneity, MBC is often unresponsive to standard chemotherapies; and novel targeted therapeutic discovery is urgently needed. Histone deacetylase inhibitors (DACi) suppress tumor growth and metastasis through regulation of the epithelial-to-mesenchymal transition axis in various cancers, including basal-like breast cancers.We utilized a new MBC patient-derived xenograft (PDX) to examine the effect of DACi therapy on MBC. Cell morphology, cell cycle-associated gene expressions, transwell migration, and metastasis were evaluated in patient-derived cells and tumors after treatment with romidepsin and panobinostat. Derivations of our PDX model, including cells, spheres, organoids, explants, andin vivoimplanted tumors were treated. Finally, we tested the effects of combining DACi with approved chemotherapeutics on relative cell biomass.DACi significantly suppressed the total number of lung metastasisin vivousing our PDX model, suggesting a role for DACi in preventing circulating tumor cells from seeding distal tissue sites. These data were supported by our findings that DACi reduced cell migration, populations, and expression of mesenchymal-associated genes. While DACi treatment did affect cell cycle-regulating genesin vitro,tumor growth was not affected compared to controls. Importantly, gene expression results varied depending on the cellular or tumor system used, emphasizing the importance of using multiple derivations of cancer models in preclinical therapeutic discovery research. Furthermore, DACi sensitized and produced a synergistic effect with approved oncology therapeutics on inherently resistant MBC.This study introduced a role for DACi in suppressing the migratory and mesenchymal phenotype of MBC cells through regulation of the epithelial-mesenchymal transition axis and suppression of the CTC population. Preliminary evidence that DACi treatment in combination with MEK1/2 inhibitors exerts a synergistic effect on MBC cells was also demonstrated.

The Arabidopsis genome encodes >450 proteins containing the pentatricopeptide repeat (PPR) motif. The PPR proteins are classified into two groups, termed as P and P Long-Short (PLS) classes. Typically, the PLS subclass proteins are mainly involved in the RNA editing of mitochondrial and chloroplast transcripts, whereas most of the analyzed P subclass proteins have been mainly implicated in RNA metabolism, such as 5′ or 3′ transcript stabilization and processing, splicing and translation. Mutations of PPR genes often result in embryogenesis and altered seedling developmental defect phenotypes, but only a limited number of ppr mutants have been characterized in detail. In this report, we show that null mutations in the EMB2794 gene result in embryo arrest, due to altered splicing of nad2 transcripts in the Arabidopsis mitochondria. In angiosperms, nad2 has five exons that are transcribed individually from two mitochondrial DNA regions. Biochemical and in vivo analyses further indicate that recombinant or transgenic EMB2794 proteins bind to the nad2 pre-mRNAs in vitro as well as in vivo, suggesting a role for this protein in trans-splicing of nad2 intron 2 and possibly in the stability of the second pre-mRNA of nad2. Homozygous emb2794 lines, showing embryo-defective phenotypes, can be partially rescued by the addition of sucrose to the growth medium. Mitochondria of rescued homozygous mutant plants contain only traces of respiratory complex I, which lack the NADH-dehydrogenase activity. Fil: Marchetti, Maria Fernanda. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina Fil: Cainzos, Maximiliano. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina Fil: Shevtsov, Sofia. The Hebrew University of Jerusalem; Israel Fil: Cordoba, Juan Pablo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; Argentina Fil: Sultan, Laure Dora. The Hebrew University of Jerusalem; Israel Fil: Brennicke, Axel. Universitat Ulm; Alemania Fil: Takenaka, Mizuki. Kyoto University; Japón Fil: Pagnussat, Gabriela Carolina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina Fil: Ostersetzer Biran, Oren. The Hebrew University of Jerusalem; Israel Fil: Zabaleta, Eduardo Julian. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina

Gastric cancer is one of the most common and deadliest cancers worldwide. There is a significant diversity in risk and aggressiveness associated with the disease in terms of factors such as ethnicity, age, and diet. However, one thing that is common is the fact that infection with Helicobacter pylori (H. pylori) is often associated with the development of gastric adenocarcinoma. Infection with H. pylori induces a series of events that lead to non-atrophic gastritis (NAG), multifocal atrophic gastritis (MAG), intestinal metaplasia (IM), dysplasia, and cancer. All the premalignant, proinflammatory stages are treatable, but it has been shown that the rate of progression to more advanced lesions is higher than the rate of regression. Most of the research efforts have focused on understanding the cancer stage, but there is still a large gap in the knowledge of biomarkers of the inflammatory stages and those of the progression/regression. Using samples from African American and Caucasian individuals with gastritis, we have been able to identify specific sets of single-nucleotide polymorphisms (SNPs) and haplotypes in cytokine genes associated with ethnicity. Interestingly, those same SNPs are also associated with advanced gastric lesions. On the other hand, using a cohort of Hispanic/Latino individuals with gastritis, we identified CD44 as a marker of disease progression and DMBT1 as a marker of disease aggressiveness. Both markers were validated in in vitro and in vivo systems.

Breast cancer (BC) is the most common malignancy in women worldwide. Different risk factors are associated with the disease which is classified into several intrinsic subtypes according to expression of hormone receptors (estrogen and progesterone) and human epidermal growth factor receptor 2 (HER2). The advent of high throughput techniques has allowed the molecular classification of BC and development of diagnostic/prognostic tools to predict disease outcome and progression. However, most molecular classifications and diagnostic/prognostic tools were developed using samples from European/Caucasian women. Hispanic/Latino describes an admixed group of individuals with different fractions of European, African, and Indigenous American ancestries. We determined the frequency of the different intrinsic subtypes of BC in Colombia and their association with genetic ancestry and found that luminal B subtype was the most prevalent in Colombian women with BC. We found that African ancestry was associated with more aggressive forms of BC. We then used next-generation sequencing techniques to determine the underlying markers of luminal B subtype of BC in Colombia. We found 67 genes differentially expressed between luminal A and luminal B subtypes, and of those, six were common between patients with high European/low Indigenous American ancestries. Interestingly, three genes (ERBB2, MIEN1, and GRB7) form a genomic “block” in chromosome 17 from which ERBB2 and GRB7 are co-expressed. The expression of ERBB2 was independent of HER-2 labeling. Our results suggest a major role of ethnicity in the modulation of these genes in BC in Hispanic/Latina women and may be the basis for future larger studies to investigate the role of ethnicity in BC susceptibility in minority groups.

Background: Colorectal cancer is the third most deadly cancer among African Americans (AA). When compared to Caucasian Americans (CA), AA present with more advanced disease and lower survival rates. Here, we investigated if differences in tumor immunology could be contributive to disparities observed between these populations.Methods: We examined gene expression of tumor and non-tumor adjacent tissues from AA and CA by whole transcriptome sequencing, and generated scores for immune cell populations by NanoString. In addition, we utilized “The Cancer Genome Atlas” (TCGA) database from AA and CA as a validation cohort. Finally, we measured the secretion of cytokines characteristic of effector T helper cell (Th) subsets by ELISA using plasma from each AA and CA participant.Results: Colon tumors from AA patients showed significant fold-change increase in gene expression when compared to CA for FOXP3 (6.22 vs. 3.22), IL1B (103 vs. 11.4) and IL8 (220 vs. 28.9) (p < 0.05). In contrast, among CA we observed statistically higher gene expression of markers associated with antitumor activity such as GZMB (Granzyme B), IFNG and the immunotherapy targets PDL1 (CD274) and CTLA4 (p < 0.05). TCGA data validated our observed higher gene expression of GZMB and PDL1 in CA patients when compared to AA. Notably, our observations on immune cell populations show that AA tumors have significantly higher number of exhausted CD8+ cells (p < 0.01), mast cells (p < 0.02) and increased T regulatory cells when compared to CA. AA colon cancer patients differed from CA in cytokine production patterns in plasma (i.e., reduced IL-12).Conclusions: Our study demonstrates significant differences of the immunological profiles of colon tumors from AA compared to CA that suggest a deficiency of appropriate immune defense mechanisms in terms of gene expression, recruitment of immune cells and systemic secretion of cytokines. As such, these immune differences could be mitigated through population-specific therapeutic approaches.

Summary In vertebrates, GATA2 is a master regulator of hematopoiesis and is expressed throughout embryo development and in adult life. Although the essential role of GATA2 in mouse hematopoiesis is well established, its involvement during early human hematopoietic development is not clear. By combining time-controlled overexpression of GATA2 with genetic knockout experiments, we found that GATA2, at the mesoderm specification stage, promotes the generation of hemogenic endothelial progenitors and their further differentiation to hematopoietic progenitor cells, and negatively regulates cardiac differentiation. Surprisingly, genome-wide transcriptional and chromatin immunoprecipitation analysis showed that GATA2 bound to regulatory regions, and repressed the expression of cardiac development-related genes. Moreover, genes important for hematopoietic differentiation were upregulated by GATA2 in a mostly indirect manner. Collectively, our data reveal a hitherto unrecognized role of GATA2 as a repressor of cardiac fates, and highlight the importance of coordinating the specification and repression of alternative cell fates., Graphical Abstract, Highlights • GATA2 promotes hemogenic emergence during human iPSC differentiation • GATA2 enhances hematopoietic differentiation of human iPSCs • GATA2 acts as a direct repressor of cardiac fates during mesoderm specification, Giorgetti A. and colleagues demonstrate that GATA2 induction in early mesodermal cells leads to the robust generation of hemogenic endothelial and hematopoietic cells from human iPSCs. They show the ability of GATA2 to instruct mesodermal precursors toward a hematopoietic cell fate and concurrently inhibit cardiac fates. This study expands our understanding of the regulatory networks that control early human hematopoiesis.

BRCA1 and BRCA2 genes are involved in DNA double-strand break repair and related to breast cancer. Shift work is associated with biological clock alterations and with a higher risk of breast cancer. The aim of this study was to investigate the variability of expression of BRCA genes through the day in healthy subjects and to measure BRCA expression levels in shift workers. The study was approached in two ways. First, we examined diurnal variation of BRCA1 and BRCA2 genes in lymphocytes of 15 volunteers over a 24-hour period. Second, we measured the expression of these genes in lymphocytes from a group of shift and daytime workers. The change in 24-hour expression levels of BRCA1 and BRCA2 genes was statistically significant, decreasing from the peak at midday to the lowest level at midnight. Lower levels for both genes were found in shift workers compared to daytime workers. Diurnal variability of BRCA1 and BRCA2 expression suggests a relation of DNA double-strand break repair system with biological clock. Lower levels of BRCA1 and BRCA2 found in shift workers may be one of the potential factors related to the higher risk of breast cancer.

Background: Epstein–Barr virus (EBV)-positive gastric cancers have clinicopathologic differences from EBV-negative tumors and lack TP53 mutation. Serologic profiles may inform viral contribution to carcinogenesis. Methods: We compared humoral responses of EBV-positive (n = 67) and EBV-negative (n = 137) patients with gastric cancer from the International EBV-Gastric Cancer Consortium. Serum antibodies against four EBV proteins, nuclear (EBNA), viral capsid (VCA), early-diffuse (EA-D), and Zta replication activator (ZEBRA), and to p53 were assessed by multiplex assays. OR of antibody level tertiles (T1–T3) were adjusted by logistic regression. We also conducted a meta-analysis of reported anti-p53 seropositivity in gastric cancer. Results: Consistent with EBV's ubiquity, 99% of patients were seropositive for anti-EBNA and 98% for anti-VCA, without difference by tumor EBV status. Seropositivity varied between patients with EBV-positive and EBV-negative tumors for anti-EA-D (97% vs. 67%, respectively, P < 0.001) and anti-ZEBRA (97% vs. 85%, respectively, P = 0.009). Adjusted ORs (vs. T1) for patients with EBV-positive versus EBV-negative tumors were significantly elevated for higher antibodies against EBNA (2.6 for T2 and 13 for T3), VCA (1.8 for T2 and 2.4 for T3), EA-D (6.0 for T2 and 44 for T3), and ZEBRA (4.6 for T2 and 12 for T3). Antibodies to p53 were inversely associated with EBV positivity (3% vs. 15%; adjusted OR = 0.16, P = 0.021). Anti-p53 prevalence from the literature was 15%. Conclusions: These serologic patterns suggest viral reactivation in EBV-positive cancers and identify variation of p53 seropositivity by subtype. Impact: Anti-EBV and anti-p53 antibodies are differentially associated with tumor EBV positivity. Serology may identify EBV-positive gastric cancer for targeted therapies.

Adeno-associated virus (AAV)–based liver gene therapy has been shown to be clinically successful. However, the presence of circulating neutralizing antibodies (NABs) against AAV vector capsids remains a major challenge as it may prevent successful transduction of the target cells. Therefore, there is a need to develop strategies that would enable AAV-mediated gene delivery to patients with preexisting anti-AAV NABs. In the current study, the feasibility of using an immunoadsorption (IA) procedure for repeated, liver-targeted gene delivery in nonhuman primates was explored. The animals were administered IV with recombinant AAV5 (rAAV5) carrying the reporter gene human secreted embryonic alkaline phosphatase (hSEAP). Seven weeks after the first rAAV treatment, all of the animals were readministered with rAAV5 carrying the therapeutic hemophilia B gene human factor IX (hFIX). Half of the animals administered with rAAV5-hSEAP underwent IA prior to the second rAAV5 exposure. The transduction efficacies of rAAV5-hSEAP and rAAV5-hFIX were assessed by measuring the levels of hSEAP and hFIX proteins. Although no hFIX was detected after rAAV5-hFIX readministration without prior IA, all animals submitted to IA showed therapeutic levels of hFIX expression, and a threshold of anti-AAV5 NAB levels compatible with successful readministration was demonstrated. In summary, our data demonstrate that the use of a clinically applicable IA procedure enables successful readministration of an rAAV5-based gene transfer in a clinically relevant animal model. Finally, the analysis of anti-AAV NAB levels in human subjects submitted to IA confirmed the safety and efficacy of the procedure to reduce anti-AAV NABs. Furthermore, clinical translation was assessed using an immunoglobulin G assay as surrogate.

Background: Hereditary hemorrhagic telangiectasia is an autosomal dominant vascular disorder caused by heterozygous, loss-of-function mutations in 4 transforming growth factor beta (TGFβ) pathway members, including the central transcriptional mediator of the TGFβ pathway, Smad4. Loss of Smad4 causes the formation of inappropriate, fragile connections between arteries and veins called arteriovenous malformations (AVMs), which can hemorrhage leading to stroke, aneurysm, or death. Unfortunately, the molecular mechanisms underlying AVM pathogenesis remain poorly understood, and the TGFβ downstream effectors responsible for hereditary hemorrhagic telangiectasia–associated AVM formation are currently unknown. Methods: To identify potential biological targets of the TGFβ pathway involved in AVM formation, we performed RNA- and chromatin immunoprecipitation–sequencing experiments on BMP9 (bone morphogenetic protein 9)-stimulated endothelial cells (ECs) and isolated ECs from a Smad4-inducible, EC-specific knockout ( Smad4 -iECKO) mouse model that develops retinal AVMs. These sequencing studies identified the angiopoietin-Tek signaling pathway as a downstream target of SMAD4. We used monoclonal blocking antibodies to target a specific component in this pathway and assess its effects on AVM development. Results: Sequencing studies uncovered 212 potential biological targets involved in AVM formation, including the EC surface receptor, TEK (TEK receptor tyrosine kinase) and its antagonistic ligand, ANGPT2 (angiopoietin-2). In Smad4 -iECKO mice, Angpt2 expression is robustly increased, whereas Tek levels are decreased, resulting in an overall reduction in angiopoietin-Tek signaling. We provide evidence that SMAD4 directly represses Angpt2 transcription in ECs. Inhibition of ANGPT2 function in Smad4 -deficient mice, either before or after AVMs form, prevents and alleviates AVM formation and normalizes vessel diameters. These rescue effects are attributed to a reversion in EC morphological changes, such as cell size and shape that are altered in the absence of Smad4. Conclusions: Our studies provide a novel mechanism whereby the loss of Smad4 causes increased Angpt2 transcription in ECs leading to AVM formation, increased blood vessel calibers, and changes in EC morphology in the retina. Blockade of ANGPT2 function in an in vivo Smad4 model of hereditary hemorrhagic telangiectasia alleviated these vascular phenotypes, further implicating ANGPT2 as an important TGFβ downstream mediator of AVM formation. Therefore, alternative approaches that target ANGPT2 function may have therapeutic value for the alleviation of hereditary hemorrhagic telangiectasia symptoms, such as AVMs.

We have generated two human induced pluripotent stem cell (iPSC) lines from CD133+ cells isolated from umbilical cord blood (CB) of a female child using non-integrative Sendai virus. Here we describe the complete characterization of these iPSC lines: PRYDi-CB5 and PRYDi-CB40.

Presently, biopreservation through protective bacterial cultures and their antimicrobial products or using antibacterial compounds derived from plants are proposed as feasible strategies to maintain the long shelf-life of products. Another emerging category of food biopreservatives are bacteriophages or their antibacterial enzymes called “phage lysins” or “enzybiotics”, which can be used directly as antibacterial agents due to their ability to act on the membranes of bacteria and destroy them. Bacteriophages are an alternative to antimicrobials in the fight against bacteria, mainly because they have a practically unique host range that gives them great specificity. In addition to their potential ability to specifically control strains of pathogenic bacteria, their use does not generate a negative environmental impact as in the case of antibiotics. Both phages and their enzymes can favor a reduction in antibiotic use, which is desirable given the alarming increase in resistance to antibiotics used not only in human medicine but also in veterinary medicine, agriculture, and in general all processes of manufacturing, preservation, and distribution of food. We present here an overview of the scientific background of phages and enzybiotics in the food industry, as well as food applications of these biopreservatives. SODERCAN | Ref. RH20-XX-032, FAGOFOOD

Production of polyhydroxyalkanoates is an important field in the biorefinery as bio-alternative to conventional plastics. However, its commercialization is still limited by high production cost. In this study, a process with the potential to reduce the production cost of polyhydroxyalkanoates was proposed. Mixed cultures accumulated polyhydroxyalkanoates using volatile fatty acid-rich effluents from waste streams, without pH and temperature control. In addition, the impact of two types of carbon sources was investigated by analyzing the microbial community as well as the polyhydroxyalkanoate accumulation capacity. Mixed cultures successfully adapted to different substrates, consuming the volatile fatty acids in their totality. The phyla Proteobacteria, Bacteroidetes and Firmicutes dominated the bacterial community. The highest polyhydroxyalkanoate content was 43.5% w/w, which is comparable to contents reported from mixed cultures using synthetic carbon sources. The biopolymer consisted of (R)-3-hydroxybutyrate 94.8 ± 1.7% w/w and (R)-3-hydroxyvaletare 5.2 ± 1.7% w/w.

// Jacob M. Loupe 1,4,* , Patrick J. Miller 1,5,* , Benjamin P. Bonner 1,* , Elaine C. Maggi 1 , Jyothi Vijayaraghavan 1 , Jovanny Zabaleta 2 , Christopher M. Taylor 3 , Fern Tsien 1 , Judy S. Crabtree 1 and Andrew D. Hollenbach 1 1 Department of Genetics, Louisiana State University Health Sciences Center, New Orleans, LA, USA 2 Department of Pediatrics and Stanley S. Scott Cancer Center, Louisiana State University Health Sciences Center, New Orleans, LA, USA 3 Department of Microbiology, Immunology, and Parasitology, Louisiana State University Health Sciences Center, New Orleans, LA, USA 4 Center for Human Genetic Research, Massachusetts General Hospital, Richard B. Simches Research Center, Boston, MA, USA 5 Tulane University, New Orleans, LA, USA * These authors have contributed equally to this work Correspondence to: Andrew D. Hollenbach, email: // Keywords : aneuploidy, Pax3-FOXO1, myogenesis, phosphorylation, alveolar rhabdomyosarcoma Received : April 07, 2016 Accepted : August 10, 2016 Published : August 30, 2016 Abstract While many solid tumors are defined by the presence of a particular oncogene, the role that this oncogene plays in driving transformation through the acquisition of aneuploidy and overcoming growth arrest are often not known. Further, although aneuploidy is present in many solid tumors, it is not clear whether it is the cause or effect of malignant transformation. The childhood sarcoma, Alveolar Rhabdomyosarcoma (ARMS), is primarily defined by the t(2;13)(q35;q14) translocation, creating the PAX3-FOXO1 fusion protein. It is unclear what role PAX3-FOXO1 plays in the initial stages of tumor development through the acquisition and persistence of aneuploidy. In this study we demonstrate that PAX3-FOXO1 serves as a driver mutation to initiate a cascade of mRNA and miRNA changes that ultimately reprogram proliferating myoblasts to induce the formation of ARMS. We present evidence that cells containing PAX3-FOXO1 have changes in the expression of mRNA and miRNA essential for maintaining proper chromosome number and structure thereby promoting aneuploidy. Further, we demonstrate that the presence of PAX3-FOXO1 alters the expression of growth factor related mRNA and miRNA, thereby overriding aneuploid-dependent growth arrest. Finally, we present evidence that phosphorylation of PAX3-FOXO1 contributes to these changes. This is one of the first studies describing how an oncogene and post-translational modifications drive the development of a tumor through the acquisition and persistence of aneuploidy. This mechanism has implications for other solid tumors where large-scale genomics studies may elucidate how global alterations contribute to tumor phenotypes allowing the development of much needed multi-faceted tumor-specific therapeutic regimens.