The drop plate method as an alternative for Azospirillum spp viable cell enumeration within the consensus protocol of the REDCAI network

Abstract Quality evaluation of commercial inoculants is essential to warrant an adequate cropresponse to inoculation within a biosecurity framework. In this sense, this work is aimed at standardizing and validating the drop plate method for the enumeration of Azospirillum viable cellsas an alternative to the spread plate technique, which is currently proposed in the consensusprotocol of the REDCAI network. Between 14 and 25 private and public laboratories partici-pated in three independent trials. We obtained consistent and robust results that allowed toconfirm that both techniques are equivalent, concluding that the drop plate method is an alternative enumeration technique that is adequate to be included in the abovementioned consensusprotocol.

Resumen La evaluación de la calidad de los inoculantes comerciales es fundamental para garantizar una adecuada respuesta de los cultivos a la inoculación dentro de un marco de bioseguridad. En este sentido, el objetivo de este trabajo fue la estandarización y validación de la técnica de la microgota para la cuantificación de Azospirillum como metodología alternativa a la técnica de siembra en superficie, propuesta actualmente en el protocolo consenso de la Red de Calidad de Inoculantes, REDCAI. Entre 14 y 25 laboratorios, tanto privados como públicos, participaron de tres ensayos independientes. A partir de ellos se obtuvieron resultados reproducibles y robustos que permiten confirmar que ambas técnicas son equivalentes y concluir que la técnica de recuento por la microgota es una alternativa adecuada para ser incluida dentro del mencionado protocolo consenso.

The drop plate method as an alternative for Azospirillum spp. viable cell enumeration within the consensus protocol of the REDCAI network.

Quality evaluation of commercial inoculants is essential to warrant an adequate crop response to inoculation within a biosecurity framework. In this sense, this work is aimed at standardizing and validating the drop plate method for the enumeration of Azospirillum viable cells as an alternative to the spread plate technique, which is currently proposed in the consensus protocol of the REDCAI network. Between 14 and 25 private and public laboratories participated in three independent trials. We obtained consistent and robust results that allowed to confirm that both techniques are equivalent, concluding that the drop plate method is an alternative enumeration technique that is adequate to be included in the abovementioned consensus protocol.

Occurrence and molecular characterization of cyst nematode species (Globodera spp.) associated with potato crops in Colombia.

Potato cyst nematodes (PCN) from the genus Globodera spp. cause major losses in the potato (Solanum tuberosum) industry worldwide. Despite their importance, at present little is known about the status of this plant pathogen in cultivated potatoes in Colombia. In this study, a total of 589 samples collected from 75 geographic localities in nine potato producing regions of Colombia (Cundinamarca, Boyacá, Antioquia, Nariño, Santander, Norte de Santander, Tolima, Caldas and Cauca) were assayed for the presence of potato cyst nematodes. Fifty-seven percent of samples tested positive for PCN. Based on phylogenetic analysis of the internal transcribed spacer region (ITS1-5.8S-ITS2) of the rRNA gene and D2-D3 expansion segments of the 28S rRNA gene, all populations but one were identified as Globodera pallida. Sequences of G. pallida from Colombia formed a monophyletic group closely related to Peruvian populations, with the lowest average number of nucleotide substitutions per site (Dxy = 0.002) and net nucleotide substitutions per site (Da = 0.001), when compared to G. pallida populations from Europe, South and North America. A single sample formed a well-supported subclade along with G. rostochiensis and G. tabacum from Japan, USA and Argentina. To our knowledge this is the first comprehensive survey of Globodera populations from Colombia that includes genetic data. Our findings on species diversity and phylogenetic relationships of Globodera populations from Colombia may help elucidate the status and distribution of Globodera species, and lead to the development of accurate management strategies for the potato cyst nematodes.

Wnt5a modulates dendritic spine dynamics through the regulation of Cofilin via small Rho GTPase activity in hippocampal neurons.

Dendritic spines are small, actin-rich protrusions that act as the receiving sites of most excitatory inputs in the central nervous system. The remodeling of the synapse architecture is mediated by actin cytoskeleton dynamics, a process precisely regulated by the small Rho GTPase family. Wnt ligands exert their presynaptic and postsynaptic effects during formation and consolidation of the synaptic structure. Specifically, Wnt5a has been identified as an indispensable synaptogenic factor for the regulation and organization of the postsynaptic side; however, the molecular mechanisms through which Wnt5a induces morphological changes resulting from actin cytoskeleton dynamics within dendritic spines remain unclear. In this work, we employ primary rat hippocampal cultures and HT22 murine hippocampal neuronal cell models, molecular and pharmacological tools, and fluorescence microscopy (laser confocal and epifluorescence) to define the Wnt5a-induced molecular signaling involved in postsynaptic remodeling mediated via the regulation of the small Rho GTPase family. We report that Wnt5a differentially regulates the phosphorylation of Cofilin in neurons through both Ras-related C3 botulinum toxin substrate 1 and cell division cycle 42 depending on the subcellular compartment and the extracellular calcium levels. Additionally, we demonstrate that Wnt5a increases the density of dendritic spines and promotes their maturation via Ras-related C3 botulinum toxin substrate 1. Accordingly, we find that Wnt5a requires the combined activation of small Rho GTPases to increase the levels of filamentous actin, thus promoting the stability of actin filaments. Altogether, these results provide evidence for a new mechanism by which Wnt5a may target actin dynamics, thereby regulating the subsequent morphological changes in dendritic spine architecture.

Prevalencia de Sífilis, Hepatitis B y Virus de Inmunodeficiencia Humana en población masculina privada de la libertad de un establecimiento carcelario en Bogotá-Colombia. Año 2019 / Prevalence of Syphilis, Hepatitis B and Human Immunodeficiency Virus in the male prison population in Bogotá, Colombia in 2019

Resumen Objetivo: Determinar la prevalencia de sífilis, hepatitis B y virus de la inmunodeficiencia humana en una población privada de la libertad de un establecimiento carcelario masculino de Bogotá D.C.-Colombia en 2019. Materiales y métodos: Se realizó un estudio de corte transversal en un establecimiento carcelario masculino de Bogotá, se incluyeron personas privadas de la libertad, mayores de 18 años. Los sujetos fueron sometidos a pruebas de detección de anticuerpos contra el Treponema pallidum, Antígenos de Superficie contra hepatitis B (HBsAg) y Virus de Inmunodeficiencia Humana (VIH) y respondieron un cuestionario estructurado para la descripción de conductas de riesgo. Resultados: Participaron 447 sujetos, ubicados en 7 pabellones del establecimiento carcelario. La prevalencia de sífilis fue del 5.8% (IC95% 3.8 - 8.4), del 1.1% para VIH (IC95% 0.4 - 2.6), y del 0.45% para hepatitis B crónica (IC95% 0.05 - 1.6). Discusión: A pesar de que la prevalencia documentada para estas patologías es más alta que en la población general, los resultados son más bajos que los reporta dos en instituciones de condiciones similares en otras latitudes. Se recomienda que el establecimiento continúe desarrollando políticas de promoción y prevención de estas patologías dentro de su población.

Abstract Objective: To determine the prevalence of syphilis, hepatitis B and the human immunodeficiency virus (HIV) in the male prison population in Bogotá, Colombia in 2019. Materials and methods: A cross-sectional study was carried out in a male prison center in Bogotá, in which sequential sampling, stratified by ward, included people deprived of liberty, over 18 years of age and who voluntarily agreed to participate in the investigation. Subjects underwent tests for antibodies to Treponema pallidum, Surface Antigens against hepatitis B (HBsAg) and Human Immunodeficiency Virus (HIV) and they answered a structured questionnaire for the description of risk behaviors. Results: A total of 447 subjects were included, belonging to 7 prison wards. The prevalence of syphilis was 5.8% (95% CI 3.8 - 8.4), 0.5% for chronic hepatitis B (95% CI 0.05 - 1.6) and 1.1% for HIV (95% CI 0.4 - 2.6). Discussion: Although the documented prevalence for these pathologies is higher than in the general population, the results are lower than those reported in other institutions with similar conditions in other latitudes. It is recommended that the institution continue to strengthen its policies for the promotion and prevention of these pathologies within its population.

Emerging Synaptic Molecules as Candidates in the Etiology of Neurological Disorders.

Synapses are complex structures that allow communication between neurons in the central nervous system. Studies conducted in vertebrate and invertebrate models have contributed to the knowledge of the function of synaptic proteins. The functional synapse requires numerous protein complexes with specialized functions that are regulated in space and time to allow synaptic plasticity. However, their interplay during neuronal development, learning, and memory is poorly understood. Accumulating evidence links synapse proteins to neurodevelopmental, neuropsychiatric, and neurodegenerative diseases. In this review, we describe the way in which several proteins that participate in cell adhesion, scaffolding, exocytosis, and neurotransmitter reception from presynaptic and postsynaptic compartments, mainly from excitatory synapses, have been associated with several synaptopathies, and we relate their functions to the disease phenotype.

Posttranslational Modifications Regulate the Postsynaptic Localization of PSD-95.

The postsynaptic density (PSD) consists of a lattice-like array of interacting proteins that organizes and stabilizes synaptic receptors, ion channels, structural proteins, and signaling molecules required for normal synaptic transmission and synaptic function. The scaffolding and hub protein postsynaptic density protein-95 (PSD-95) is a major element of central chemical synapses and interacts with glutamate receptors, cell adhesion molecules, and cytoskeletal elements. In fact, PSD-95 can regulate basal synaptic stability as well as the activity-dependent structural plasticity of the PSD and, therefore, of the excitatory chemical synapse. Several studies have shown that PSD-95 is highly enriched at excitatory synapses and have identified multiple protein structural domains and protein-protein interactions that mediate PSD-95 function and trafficking to the postsynaptic region. PSD-95 is also a target of several signaling pathways that induce posttranslational modifications, including palmitoylation, phosphorylation, ubiquitination, nitrosylation, and neddylation; these modifications determine the synaptic stability and function of PSD-95 and thus regulate the fates of individual dendritic spines in the nervous system. In the present work, we review the posttranslational modifications that regulate the synaptic localization of PSD-95 and describe their functional consequences. We also explore the signaling pathways that induce such changes.

Effects of dietary supplementation with lemon verbena extracts on serum inflammatory markers of multiple sclerosis patients / Efecto de la suplementación dietética con extractos de hierbaluisa en los marcadores de inflamación en suero de personas con esclerosis múltiple

Introduction: Inflammation is one of the main contributory factors to the etiopathogenesis of multiple sclerosis (MS). Dietary interventions with Lipia citriadora (lemon verbena) extracts have been proved to be effective in the prevention of inflammatory diseases. Objectives: The aim of this study is to evaluate the effect of lemon verbena supplementation in pro- and anti- inflammatory serum biomarkers of patients with different clinical subtypes of multiple sclerosis. Methods: The effect of lemon verbena supplementation (10% w/w verbascoside) was evaluated in a randomized, double-blinded placebo-controlled study with 30 participants classified in relapsing-remitting (n=10), primary progressive (n=5) and secondary progressive (n=15) MS presentations. Serum cytokine and C reactive protein levels were assessed in intervention and control groups for each MS clinical subtype after 28 days of dietary supplementation. Results: Serum levels of C reactive protein and 8 cytokines/ inflammatory (IFN-γ, IL-12, IL-23, IL-6, TNF-α, TGF-β, IL-4 and IL-10) markers were studied. Secondary progressive MS- supplemented patients showed C reactive protein concentrations significantly lower compared to the placebo group (p<0.005). IFN-γ levels decreased for all MS-treated groups whereas IL-12 diminished levels were observed for relapsing-remitting type (p<0.05). Anti-inflammatory cytokine concentrations of IL-4 (difference 2.98 ± 2.99 pg/mL) and IL-10 (difference 1.78 ± 5.54 pg/mL) increased in secondary progressive MS patients (p<0.05). Conclusion: The variation of several pro- and anti-inflammatory markers after supplementation suggests that lemon verbena extracts may affect cytokine profiles in multiple sclerosis. Further investigation on dietary components with antioxidant and anti-inflammatory properties may contribute to understand MS pathogenesis and ameliorate MS symptoms (AU)

Introducción: La inflamación es uno de los principales factores que contribuyen en la etiopatogénesis de la esclerosis múltiple (EM). Se ha demostrado que las intervenciones en la dieta con extractos de Lipia citriadora (hierbaluisa) son efectivas en la prevención de las enfermedades inflamatorias. Objectivos: El objetivo de este estudio es evaluar el efecto de la suplementación con extractos de hierbaluisa en los biomarcadores de inflamación en suero de pacientes con diferentes subtipos clínicos de esclerosis múltiple. Métodos: El efecto de la suplementación con hierbaluisa (10 % p/p verbascósido) se evaluó mediante un estudio aleatorizado de doble ciego controlado con grupo placebo, constituido por 30 participantes clasificados según la forma de presentación de EM en: remitentes-recaídas (n=10), primaria progresiva (n=5) y secundaria progresiva (n=15). Los niveles de citoquinas y proteína C reactiva en suero se valoraron en los grupos intervención y control de cada uno de los subtipos clínicos de EM después de 28 días de suplementación en la dieta. Resultados: Se estudiaron los niveles en suero de proteína C reactiva y de 8 citoquinas como biomarcadores deinflamación (IFN-γ, IL-12, IL-23, IL-6, TNF-α, TGF-β, IL-4 e IL-10). Los pacientes del grupo de intervención con EM secundaria progresiva presentaron concentraciones de proteína C reactiva significativamente más bajos comparados con el grupo placebo (p<0.005). Los niveles de IFN-γ disminuyeron en todos los grupos tratados a la vez que se detectaron niveles inferiores de IL-12 en las formas secundaria progresiva y remitente-recaídas (p<0.05). Las concentraciones de las citoquinas anti-inflamatorias: IL-4 (diferencia 2,98 ± 2,99 pg/mL) y IL-10 (diferencia 1,78 ± 5,54 pg/mL) aumentaron en los pacientes con EM secundaria progresiva (p<0.05). Conclusión: La variación en la concentración de varias citoquinas pro- y anti-inflamatorias después de la suplementación con los extractos de hierbaluisa puede afectar al perfil de las citoquinas en la esclerosis múltiple. La investigación futura de los componentes de la dieta con propiedades anti-inflamatorias y antioxidantes puede contribuir a entender la patógenesis de la esclerosis múltiple así como a disminuir sus síntomas (AU)

Autophagic response in the Rabbit Hemorrhagic Disease, an animal model of virally-induced fulminant hepatic failure.

The Rabbit Hemorrhagic Disease Virus (RHDV) induces a severe disease that fulfils many requirements of an animal model of fulminant hepatic failure. However, a better knowledge of molecular mechanisms contributing to liver damage is required, and it is unknown whether the RHDV induces liver autophagy and how it relates to apoptosis. In this study, we attempted to explore which signalling pathways were involved in the autophagic response induced by the RHDV and to characterize their role in the context of RHDV pathogenesis. Rabbits were infected with 2 × 104 hemmaglutination units of a RHDV isolate. The autophagic response was measured as presence of autophagic vesicles, LC3 staining, conversion of LC3-I to autophagosome-associated LC3-II and changes in expression of beclin-1, UVRAG, Atg5, Atg12, Atg16L1 and p62/SQSTM1. RHDV-triggered autophagy reached a maximum at 24 hours post-infection (hpi) and declined at 30 and 36 hpi. Phosphorylation of mTOR also augmented in early periods of infection and there was an increase in the expression of the endoplasmic reticulum chaperones BiP/GRP78, CHOP and GRP94. Apoptosis, measured as caspase-3 activity and expression of PARP-1, increased significantly at 30 and 36 hpi in parallel to the maximal expression of the RHDV capsid protein VP60. These data indicate that RHDV infection initiates a rapid autophagic response, perhaps in an attempt to protect liver, which associates to ER stress development and is independent from downregulation of the major autophagy suppressor mTOR. As the infection continues and the autophagic response declines, cells begin to exhibit apoptosis.

Melatonin modulates the autophagic response in acute liver failure induced by the rabbit hemorrhagic disease virus.

Autophagy is an important survival pathway and participates in the host response to infection. Beneficial effects of melatonin have been previously reported in an animal model of acute liver failure (ALF) induced by the rabbit hemorrhagic disease virus (RHDV). This study was aimed to investigate whether melatonin protection against liver injury induced by the RHDV associates to modulation of autophagy. Rabbits were infected with 2 × 10(4) hemagglutination units of a RHDV isolate and received 20 mg/kg melatonin at 0, 12, and 24 hr postinfection. RHDV induced autophagy, with increased expression of beclin-1, ubiquitin-like autophagy-related (Atg)5, Atg12, Atg16L1 and sequestrosome 1 (p62/SQSTM1), protein 1 light chain 3 (LC3) staining, and conversion of LC3-I to autophagosome-associated LC3-II. These effects reached a maximum at 24 hr postinfection, in parallel to extensive colocalization of LC3 and lysosome-associated membrane protein (LAMP)-1. The autophagic response induced by RHDV infection was significantly inhibited by melatonin administration. Melatonin treatment also resulted in decreased immunoreactivity for RHDV viral VP60 antigen and a significantly reduction in RHDV VP60 mRNA levels, oxidized to reduced glutathione ratio (GSSG/GSH), caspase-3 activity, and immunoglobulin-heavy-chain-binding protein (BiP) and CCAAT/enhancer-binding protein homologous protein (CHOP) expression. Results indicate that, in addition to its antioxidant and antiapoptotic effects, and the suppression of ER stress, melatonin induces a decrease in autophagy associated with RHDV infection and inhibits RHDV RNA replication. Results obtained reveal novel molecular pathways accounting for the protective effect of melatonin in this animal model of ALF.

Effects of dietary supplementation with lemon verbena extracts on serum inflammatory markers of multiple sclerosis patients.

INTRODUCTION: Inflammation is one of the main contributory factors to the etiopathogenesis of multiple sclerosis (MS). Dietary interventions with Lipia citriadora (lemon verbena) extracts have been proved to be effective in the prevention of inflammatory diseases. OBJECTIVES: The aim of this study is to evaluate the effect of lemon verbena supplementation in pro- and anti- inflammatory serum biomarkers of patients with different clinical subtypes of multiple sclerosis. METHODS: The effect of lemon verbena supplementation (10% w/w verbascoside) was evaluated in a randomized, double-blinded placebo-controlled study with 30 participants classified in relapsing-remitting (n=10), primary progressive (n=5) and secondary progressive (n=15) MS presentations. Serum cytokine and C reactive protein levels were assessed in intervention and control groups for each MS clinical subtype after 28 days of dietary supplementation. RESULTS: Serum levels of C reactive protein and 8 cytokines/ inflammatory (IFN-γ, IL-12, IL-23, IL-6, TNF-α, TGF-ß, IL-4 and IL-10) markers were studied. Secondary progressive MS- supplemented patients showed C reactive protein concentrations significantly lower compared to the placebo group (p.

Introducción: La inflamación es uno de los principales factores que contribuyen en la etiopatogénesis de la esclerosis múltiple (EM). Se ha demostrado que las intervenciones en la dieta con extractos de Lipia citriadora (hierbaluisa) son efectivas en la prevención de las enfermedades inflamatorias. Objectivos: El objetivo de este estudio es evaluar el efecto de la suplementación con extractos de hierbaluisa en los biomarcadores de inflamación en suero de pacientes con diferentes subtipos clínicos de esclerosis múltiple. Métodos: El efecto de la suplementación con hierbaluisa (10 % p/p verbascósido) se evaluó mediante un estudio aleatorizado de doble ciego controlado con grupo placebo, constituido por 30 participantes clasificados según la forma de presentación de EM en: remitentes-recaídas (n=10), primaria progresiva (n=5) y secundaria progresiva (n=15). Los niveles de citoquinas y proteína C reactiva en suero se valoraron en los grupos intervención y control de cada uno de los subtipos clínicos de EM después de 28 días de suplementación en la dieta. Resultados: Se estudiaron los niveles en suero de proteína C reactiva y de 8 citoquinas como biomarcadores de inflamación (IFN-, IL-12, IL-23, IL-6, TNF-, TGF-, IL-4 e IL-10). Los pacientes del grupo de intervención con EM secundaria progresiva presentaron concentraciones de proteína C reactiva significativamente más bajos comparados con el grupo placebo (p.

Melatonin treatment reduces endoplasmic reticulum stress and modulates the unfolded protein response in rabbits with lethal fulminant hepatitis of viral origin.

Hepatocyte apoptosis plays an important role in the development of fulminant hepatic failure (FHF). The objective of this study was to investigate whether endoplasmic reticulum (ER) stress and unfolded protein response (UPR) inhibition is an underlying mechanism of melatonin anti-apoptotic effects in an animal model of FHF of viral origin induced by the rabbit hemorrhagic disease virus (RHDV). Rabbits were experimentally infected with 2 × 10(4) hemagglutination units of a RHDV isolate and received melatonin at two concentrations of 10 mg/kg and 20 mg/kg at 0 hr, 12 hr and 24 hr postinfection. RHDV infection induced increased expression of CCAAT/enhancer-binding protein homologous protein (CHOP), immunoglobulin heavy chain binding protein (BiP/GRP78), glucose-regulated protein 94 (GRP94), phospho-c-Jun N-terminal kinase (JNK) and caspase-12. These effects were attenuated by melatonin. Double immunofluorescence staining showed colocalization of CHOP and cleaved caspase-3 in liver sections of RHDV-infected rabbits, while immunostaining decreased markedly with melatonin treatment. RHDV infection resulted in significant increases in the mRNA levels of activating transcription factor 6 (ATF6), ATF4, inositol-requiring enzyme 1 (IRE1), spliced X-box binding protein-1 (XBP1s) and tumor necrosis factor receptor-associated factor 2 (TRAF2). Melatonin attenuated the extent of the changes. Data obtained provide evidence that in rabbits with experimental infection by RHDV, reduction in apoptotic liver damage by melatonin is associated with attenuation of ER stress through a modulation of the three arms of UPR signaling and further support a potential hepatoprotective role of melatonin in FHF.

Quercetin treatment ameliorates inflammation and fibrosis in mice with nonalcoholic steatohepatitis.

We investigated whether quercetin protects from steatosis and limits the expression of proinflammatory and fibrogenic genes in C57BL/6J mice with nonalcoholic steatohepatitis (NASH) induced by feeding a methionine-choline-deficient (MCD) diet. Quercetin (50 mg/kg) was given by oral route daily. Mice were randomly divided into 4 groups that received for 2 or 4 wk: the control diet plus vehicle, control diet plus quercetin, MCD diet plus vehicle, and MCD diet plus quercetin. At both 2 and 4 wk, feeding the MCD diet resulted in liver steatosis, inflammatory cell accumulation, oxidative stress evaluated by the concentration of TBARS, and fibrosis evidenced by the staining of α-smooth muscle actin-positive cells in the liver. At both 2 and 4 wk, the MCD diet induced an increase in the mRNA levels of Il6, Tnf, Ptgs2, and Hmgb1 and increased the protein concentrations of Toll-like receptor-4, c-Jun terminal kinase, and p65 NFκB subunit compared with control rats. Feeding the mice the MCD diet also triggered an increase of Col1a1, Col3a1, Plod3, Tgfb1, Smad3, Smad7, Pdgfb, Ctgf, Areg, Mmp9, and Timp1 mRNA levels. These effects were totally or partially prevented by treatment with quercetin. The data obtained suggest that attenuation of multiple profibrotic and proinflammatory gene pathways contributes to the beneficial effects of quercetin in mice with MCD diet-induced steatohepatitis.