Database selection for shotgun metaproteomic of low-diversity dairy microbiomes.

The metaproteomics field has recently gained more and more interest as a valuable tool for studying both the taxonomy and function of microbiomes, including those used in food fermentations. One crucial step in the metaproteomics pipeline is selecting a database to obtain high-quality taxonomical and functional information from microbial communities. One of the best strategies described for building protein databases is using sample-specific or study-specific protein databases obtained from metagenomic sequencing. While this is true for high-diversity microbiomes (such as gut and soil), there is still a lack of validation for different database construction strategies in low-diversity microbiomes, such as those found in fermented dairy products where starter cultures containing few species are used. In this study, we assessed the performance of various database construction strategies applied to metaproteomics on two low-diversity microbiomes obtained from cheese production using commercial starter cultures and analyzed by LC-MS/MS. Substantial differences were detected between the strategies, and the best performance in terms of the number of peptides and proteins identified from the spectra was achieved by metagenomic-derived databases. However, extensive databases constructed from a high number of available online genomes obtained a similar taxonomical and functional annotation of the metaproteome compared to the metagenomic-derived databases. Our results indicate that, in the case of low-diversity dairy microbiomes, the use of publically available genomes to construct protein databases can be considered as an alternative to metagenome-derived databases.

Host DNA depletion methods and genome-centric metagenomics of bovine hindmilk microbiome.

Bovine mastitis is a multi-etiological and complex disease, resulting in serious economic consequences for dairy farmers and industry. In recent years, the microbiological evaluation of raw milk has been investigated in-depth using next-generation sequencing approaches such as metataxonomic analysis. Despite this, host DNA is a major concern in the shotgun metagenomic sequencing of microbial communities in milk samples, and it represents a big challenge. In this study, we aimed to evaluate different methods for host DNA depletion and/or microbial DNA enrichment and assess the use of PCR-based whole genome amplification in milk samples with high somatic cell count (SCC) by using short- and long-read sequencing technologies. Our results evidenced that DNA extraction performed differently in terms of host DNA removal, impacting metagenome composition and functional profiles.. Moreover, the ratio of SCC/bacteria ultimately impacts microbial DNA yield, and samples with low SCC (SCC below 100,000 cells/mL) are the most problematic. When milk samples with high SCC (SCC above 200,000 cells/mL) underwent multiple-displacement amplification (MDA), we successfully recovered high-quality metagenome-assembled genomes (MAGs), and long-read sequencing was feasible even for samples with low DNA concentration. By associating MDA and short-read sequencing, we recovered two times more MAGs than in untreated samples, and an ongoing co-infection not reported by traditional methods was detected for mastitis pathogen. Overall, this new approach will improve the detection of mastitis-associated microorganisms and make it possible to examine host-microbiome interactions in bovine mastitis.IMPORTANCENext-generation sequencing technologies have been widely used to gain new insights into the diversity of the microbial community of milk samples and dairy products for different purposes such as microbial safety, profiling of starter cultures, and host-microbiome interactions. Milk is a complex food matrix, and additionally, the presence of host nucleic acid sequences is considered a contaminant in untargeted high-throughput sequencing studies. Therefore, genomic-centric metagenomic studies of milk samples focusing on the health-disease status in dairy cattle are still scarce, which makes it difficult to evaluate the microbial ecophysiology of bovine hindmilk. This study provides an alternative method for genome-centric metagenome studies applied to hindmilk samples with high somatic cell content, which is indispensable to examining host-microbiome interactions in bovine mastitis.

Interactions between Starmerella bacillaris and Saccharomyces cerevisiae during sequential fermentations influence the release of yeast mannoproteins and impact the protein stability of an unstable wine.

Wine protein haze formation is a problem due to grape proteins aggregation during wine storage. The cell wall components of wine yeasts, particularly high molecular weight mannoproteins, have a protective effect against haze formation, although their involvement remains poorly understood. This study aimed at characterizing glycosylated proteins released by Starmerella bacillaris and Saccharomyces cerevisiae during single and sequential fermentations in a synthetic must, and testing their impact on wine protein stability. Mannoproteins-rich extracts from sequential fermentations showed an increase in the low MW polysaccharide fraction and, when added to an unstable wine, had a greater effect on protein stability than S. cerevisiae extracts. Shotgun proteomics approaches revealed that the identified cell wall proteins exclusively found in sequential fermentations were produced by both S. bacillaris (MKC7, ENG1) and S. cerevisiae (Bgl2p). Moreover, sequential fermentations significantly increased the expression of Scw4p and 1,3-beta-glucanosyltransferase (GAS5), produced by S. cerevisiae. Finally, some of the key proteins identified might play a positive role in increasing wine protein stability.

Genomic and proteomic characterization of vB_SauM-UFV_DC4, a novel Staphylococcus jumbo phage.

Staphylococcus aureus is one of the most relevant mastitis pathogens in dairy cattle, and the acquisition of antimicrobial resistance genes presents a significant health issue in both veterinary and human fields. Among the different strategies to tackle S. aureus infection in livestock, bacteriophages have been thoroughly investigated in the last decades; however, few specimens of the so-called jumbo phages capable of infecting S. aureus have been described. Herein, we report the biological, genomic, and structural proteomic features of the jumbo phage vB_SauM-UFV_DC4 (DC4). DC4 exhibited a remarkable killing activity against S. aureus isolated from the veterinary environment and stability at alkaline conditions (pH 4 to 12). The complete genome of DC4 is 263,185 bp (GC content: 25%), encodes 263 predicted CDSs (80% without an assigned function), 1 tRNA (Phe-tRNA), multisubunit RNA polymerase, and an RNA-dependent DNA polymerase. Moreover, comparative analysis revealed that DC4 can be considered a new viral species belonging to a new genus DC4 and showed a similar set of lytic proteins and depolymerase activity with closely related jumbo phages. The characterization of a new S. aureus jumbo phage increases our understanding of the diversity of this group and provides insights into the biotechnological potential of these viruses. KEY POINTS: • vB_SauM-UFV_DC4 is a new viral species belonging to a new genus within the class Caudoviricetes. • vB_SauM-UFV_DC4 carries a set of RNA polymerase subunits and an RNA-directed DNA polymerase. • vB_SauM-UFV_DC4 and closely related jumbo phages showed a similar set of lytic proteins.

Composition and fate of heat-resistant anaerobic spore-formers in the milk powder production line.

Anaerobic spore-forming bacteria are a continuous threat to the dairy industry due to their ability to withstand processing conditions, such as those during heat treatment. These ubiquitous microorganisms have ample opportunity for multiple entry points into the milk chain, creating food quality and safety issues. Certain spore-formers, namely bacilli and clostridia, are more problematic due to their ability to spoil dairy products and pathogenicity. In this study, we investigated how milk treatment and milk powder production influenced the composition and survival of anaerobic spore-formers. Samples were obtained on three different days (replicate blocks) during the production of dairy powders and examined in a culture-dependent manner using the most probable number method coupled with 16S rRNA amplicon sequencing and metagenomic analysis of the enriched samples. Results revealed that the milk separation greatly affected the spore-former presence and composition which were detected along the entire production line from raw material to milk powders. Throughout the various points of the production line, the occurrence of species belonging to the Bacillus cereus sensu lato was higher than that of clostridia. Sequence variants (SVs) belonging to the anaerobic spore-forming genus Clostridium were taxonomically assigned to two SVs groups and were detected in all three replicate blocks. A total of 19 metagenome-assembled genomes were recovered from nine enrichments. Four near-complete and two medium-quality genomes were found in raw milk/milk powder samples and further assigned as Clostridium tyrobutyricum and Clostridium diolis, which may constitute a problem in the finished dairy product. In conclusion, our findings highlight spore-formers' importance on dairy quality and may aid in their intervention and control in the dairy production line.

Contrasting leaf intercellular space development in sorghum and maize modulates different tolerance capacity to water limitation.

The objective of this study was to evaluate the relationship between intercellular spaces and leaf gas exchange and the effect of total intercellular space on the growth of maize and sorghum under water restriction. The experiments were conducted in a greenhouse in a 2 × 3 factorial arrangement (two plant types and three water conditions: field capacity (FC = 100%), 75%FC, and 50%FC) with 10 replicates. The lack of water was a limiting factor for maize because it showed reductions in leaf area, leaf thickness, biomass, and gas exchange parameters, while sorghum remained unchanged, maintaining its water-use efficiency. This maintenance was correlated with the growth of intercellular spaces in sorghum leaves because the increased internal volume led to better CO2 control and prevented excessive water loss under drought stress. In addition, sorghum had more stomata than maize. These characteristics contributed to the drought tolerance of sorghum, while maize could not make the same adjustments. Therefore, changes in intercellular spaces promoted adjustments to avoid water loss and may have improved CO2 diffusion, characteristics that are important for drought-tolerant plants.

Metataxonomic analysis and host proteome response in dairy cows with high and low somatic cell count: a quarter level investigation.

Host response to invasive microbes in the bovine udder has an important role on the animal health and is essential to the dairy industry to ensure production of high-quality milk and reduce the mastitis incidence. To better understand the biology behind these host-microbiome interactions, we investigated the somatic cell proteomes at quarter level for four cows (collected before and after milking) using a shotgun proteomics approach. Simultaneously, we identified the quarter microbiota by amplicon sequencing to detect presence of mastitis pathogens or other commensal taxa. In total, 32 quarter milk samples were analyzed divided in two groups depending on the somatic cell count (SCC). The high SCC group (>100,000 cell/mL) included 10 samples and significant different proteome profiles were detected. Differential abundance analysis uncovers a specific expression pattern in high SCC samples revealing pathways involved in immune responses such as inflammation, activation of the complement system, migration of immune cells, and tight junctions. Interestingly, different proteome profiles were also identified in quarter samples containing one of the two mastitis pathogens, Staphylococcus aureus and Streptococcus uberis, indicating a different response of the host depending on the pathogen. Weighted correlation network analysis identified three modules of co-expressed proteins which were correlated with the SCC in the quarters. These modules contained proteins assigned to different aspects of the immune response, but also amino sugar and nucleotide sugar metabolism, and biosynthesis of amino acids. The results of this study provide deeper insights on how the proteome expression changes at quarter level in naturally infected cows and pinpoint potential interactions and important biological functions during host-microbe interaction.

Lactobacillus gasseri LG-G12 Restores Gut Microbiota and Intestinal Health in Obesity Mice on Ceftriaxone Therapy.

Gut microbiota imbalance is associated with the occurrence of metabolic diseases such as obesity. Thus, its modulation is a promising strategy to restore gut microbiota and improve intestinal health in the obese. This paper examines the role of probiotics, antimicrobials, and diet in modulating gut microbiota and improving intestinal health. Accordingly, obesity was induced in C57BL/6J mice, after which they were redistributed and fed with an obesogenic diet (intervention A) or standard AIN-93 diet (intervention B). Concomitantly, all the groups underwent a treatment phase with Lactobacillus gasseri LG-G12, ceftriaxone, or ceftriaxone followed by L. gasseri LG-G12. At the end of the experimental period, the following analysis was conducted: metataxonomic analysis, functional profiling of gut microbiota, intestinal permeability, and caecal concentration of short-chain fatty acids. High-fat diet impaired bacterial diversity/richness, which was counteracted in association with L. gasseri LG-G12 and the AIN-93 diet. Additionally, SCFA-producing bacteria were negatively correlated with high intestinal permeability parameters, which was further confirmed via functional profile prediction of the gut microbiota. A novel perspective on anti-obesity probiotics is presented by these findings based on the improvement of intestinal health irrespective of undergoing antimicrobial therapy or not.

Kombuchas from Green and Black Tea Modulate the Gut Microbiota and Improve the Intestinal Health of Wistar Rats Fed a High-Fat High-Fructose Diet.

The Western diet can negatively affect the gut microbiota and is associated with metabolic disorders. Kombucha, a tea fermented by a symbiotic culture of bacteria and yeast (SCOBY), is known for its bioactive properties and has become popular in the last years. In this study, we evaluated the effects of regular kombucha consumption on the gut microbiota and on outcomes related to the intestinal health of Wistar rats fed a high-fat high-fructose diet. After eight weeks receiving a standard diet (AIN-93M) (n = 10) or a high-fat and high-fructose diet (HFHF) (n = 30) to induce metabolic disorders, the animals were subdivided into four groups: AIN-93M (n = 10); HFHF (n = 10); GTK (HFHF + green tea kombucha (n = 10); and BTK (HFHF + black tea kombucha; n = 10) for 10 weeks. Although body composition did not differ among the groups, the HFHF diet was associated with metabolic alterations, and stimulated the growth of gram-negative bacteria such as Proteobacteria and Bacteroides. Kombucha ingestion could somewhat modulate the gut microbiota, attenuating the effects of a Western diet by increasing propionate production and favoring the growth of beneficial bacteria, such as Adlercreutzia in the GTK group. Our results suggest that regular kombucha consumption may be beneficial to intestinal health, which can be mostly attributed to its high content and diversity of phenolic compounds.

Synbiotic modulates intestinal microbiota metabolic pathways and inhibits DMH-induced colon tumorigenesis through c-myc and PCNA suppression.

The use of probiotic and synbiotic is a promising strategy to modulate the intestinal microbiota, and thereby modify the risk of diseases. In this study, the effect of probiotic VSL#3, isolated or associated with a yacon-based product (PBY), on the functional metabolic pathways of the microbiota, in a colorectal carcinogenesis model, was evaluated. For this, mice induced to carcinogenesis were fed with standard diet AIN-93 M (CON), diet AIN-93 M and VSL#3 (PRO) or diet AIN-93 M with yacon and VSL#3 (SYN). The SYN group showed a highly differentiated intestinal community based on the MetaCyc pathways. Of the 351 predicted functional pathways, 222 differed between groups. Most of them were enriched in the SYN group, namely: amino acid biosynthesis pathways, small molecule biosynthesis pathways (cofactors, prosthetic groups, electron carriers and vitamins) carbohydrate degradation pathways and fermentation pathways. In addition, the synbiotic was able to stimulate the anti-inflammatory immune response and reduce the gene expression of PCNA and c-myc. Thus, we conclude that the synbiotic impacted more significantly the metabolic functions of the microbiota compared to the isolated use of probiotic. We believe that the enrichment of these pathways can exert antiproliferative action, reducing colorectal carcinogenesis. The prediction of the functional activity of the microbiota is a promising tool for understanding the influence of the microbiome on tumor development.

Longitudinal dynamics of the bovine udder microbiota.

BACKGROUND: In recent years, the number of studies concerning microbiota of the intramammary environment has increased rapidly due to the development of high-throughput sequencing technologies that allow mapping of microbiota without culturing. This has revealed that an environment previously thought to be sterile in fact harbours a microbial community. Since this discovery, many studies have investigated the microbiota of different parts of the udder in various conditions. However, few studies have followed the changes that occur in the udder microbiota over time. In this study, the temporal dynamics of the udder microbiota of 10 cows, five with a low somatic cell count (SCC, SCC < 100,000 cells/mL) and five with a high SCC (SCC > 100,000 cells/mL), were followed over 5 months to gather insights into this knowledge gap. RESULTS: Analysis of the temporal changes in the microbial composition of milk from udders with a low SCC revealed a dynamic and diverse microbiota. When an imbalance due to one dominating genus was recorded, the dominant genus quickly vanished, and the high diversity was restored. The genera dominating in the samples with a high SCC remained the dominant genera throughout the whole sampling period. These cows generally displayed a heightened SCC or an intramammary infection in at least one quarter though-out the sampling period. CONCLUSION: Our results show that the bovine udder has a diverse microbiota, and that the composition and diversity of this community affects udder health with regards to SCC. Understanding what influences the composition and stability of this community has important implications for the understanding, control, and treatment of mastitis.

Chia (Salvia hispanica L.) Flour and Oil Ameliorate Metabolic Disorders in the Liver of Rats Fed a High-Fat and High Fructose Diet.

We hypothesized that the consumption of chia (Salvia hispanica L.) flour (CF) and chia oil (CO) improves metabolic disorders in the liver of Wistar rats (Rattus norvegicus domestica) fed a high-fat and high-fructose (HFHF) diet. The animals were fed a HFHF diet (n = 30) or AIN93-M standard diet (n = 10) for eight weeks. After this period, the animals fed HFHF were divided into three groups (n = 10): HFHF diet, HFHF plus 14.7% of CF, and HFHF plus 4% of CO. Histological and biochemical analyses, gene expression, protein levels related to inflammation, and oxidative stress were evaluated in the liver. The HFHF diet caused lipogenesis, liver steatosis, oxidative stress, and inflammation in the animals. The CF and CO intake increased the liver total antioxidant capacity and superoxide dismutase, decreased nitric oxide levels and liver steatosis. Furthermore, the CF and CO led to the upregulation of Cpt1a and Adipor2, respectively, whereas CF downregulated Srebf1. CO intake decreased blood glucose, triglycerides, and the animals' body weight. Chia did not show effects on mitigating liver pro-inflammatory status, which it may indicate occurs later. The addition of chia into an unbalanced diet is a good and relevant strategy to reduce liver metabolic disorders caused by the high consumption of fructose and saturated fat.

Assessment of the microbiological origin of blowing defects in Grana Padano Protected Designation of Origin cheese.

Recognized worldwide for its history, flavor, and high nutritional quality, Grana Padano (GP) is one of the most traditional Italian raw-milk, hard-cooked, long-ripened cheese. Throughout GP manufacturing, some well-known and undesired bacterial species, such as clostridia, can proliferate and lead to spoilage defects that mischaracterize the final product; however, little is known about the development of late-blowing defects in hard cheese samples without clostridia. Therefore, in this study we aimed to use metataxonomic analysis to identify bacterial taxa associated with the development of late-blowing defect in GP samples. Furthermore, the presence of several heterofermentative lactobacilli species in defective zones were verified by primer-specific PCR assay. Considering α- and ß-diversity analyses, no statistically significant differences were detected between cheese samples with and without blowing defect. Following taxonomic assignment, Lactobacillus and Streptococcus were the dominant genera, whereas clostridia-related taxa were not detected in any of the 20 analyzed samples. Using EdgeR, the genera Propionibacterium and Acinetobacter were found to be prevalently more abundant in samples categorized as having "big regular holes." In samples with "small regular holes," multiplex PCR amplification revealed differences in terms of Lactobacillus population composition, mainly obligate homofermentative lactobacilli, between defective and non-defective zones of the same cheese wheel. This study demonstrated that GP samples with blowing defects not caused by clostridial development share similar biodiversity indices with GP collected from control zones, but an imbalance of obligate homofermentative lactobacilli was noticed between samples, which requires further analysis to better comprehend the exact mechanism involved in this process.

Root anatomy, growth, and development of Typha domingensis Pers. (Typhaceae) and their relationship with cadmium absorption, accumulation, and tolerance.

Typha domingensis Pers. is a plant that grows in marshy environments, where cadmium (Cd) accumulates. The root is the first organ that comes into contact with the metal. The aim of this study was to evaluate the effect of Cd on the roots of T. domingensis. The experiment was conducted in a greenhouse using different Cd concentrations: (1) 0 µM (control), (2) 10 µM, and (3) 50 µM, with 10 replicates for 90 days. The plants were placed in plastic containers containing 5 L of nutrient solution modified with the different Cd concentrations. At the end of the experiment, the roots were measured, sampled, fixed, and subjected to usual plant microtechniques. The slides were observed and photographed under light microscopy and analyzed in ImageJ software. To measure Cd absorption, atomic-absorption spectrometry was used. The data were subjected to analysis of variance and comparison of means by the Scott-Knott test at P < 0.05. When exposed to 50 µM of Cd, the roots accumulated 99.35% of the Cd. At this concentration, there was a reduction in the exodermis but there was an increase in the diameter of the cortical cells and in the proportion of aerenchyma in the cortex. There was an increase in the root cap, which guaranteed the protection of the primary meristems. Therefore, T. domingensis adjusts its root anatomy improving the Cd tolerance and shows potential for phytoremediation purposes.

Starmerella bacillaris Released in Vineyards at Different Concentrations Influences Wine Glycerol Content Depending on the Vinification Protocols.

Starmerella bacillaris is a non-Saccharomyces yeast proposed for must fermentation together with Saccharomyces cerevisiae because of its high glycerol and moderate volatile acidity production. Furthermore, it was demonstrated that the same S. bacillaris strains that possess interesting technological properties exhibited antifungal activity against Botrytis cinerea, suggesting the release of this yeast in the vineyard. To obtain a positive effect during the following winemaking process, the maintenance of suitable concentrations of S. bacillaris is essential. Therefore, to obtain information on the survival of S. bacillaris, a small-scale field trial was performed. One week before the harvest, two different concentrations of S. bacillaris (106 and 107 cells/mL) were sprayed on Pinot grigio bunches, and the strain concentration was monitored by means of qPCR during the subsequent fermentation process. In addition, the combined effect of different winemaking techniques was evaluated, i.e., the vinification of juice, juice with marc and cryomaceration treatment. Results demonstrated that, under the tested conditions, S. bacillaris released in the vineyard remained viable for one week on grape bunches and increased glycerol content during the subsequent fermentation process. Different vinification protocols influenced cell concentrations. In particular, the cryomaceration treatment, due to the use of low temperature, supported S. bacillaris growth due to its cryotolerant aptitude. The collected data open new perspectives on the control of alcoholic fermentation, involving both vineyard and cellar management.

Stomatal cavity modulates the gas exchange of Sorghum bicolor (L.) Moench. grown under different water levels.

This work aimed to evaluate the effects of lower water levels on leaf intercellular spaces and to assess their relations with the gas exchange, anatomy, and growth of Sorghum bicolor. Experiments were conducted in a greenhouse, in which plants were subjected to three water conditions (ten replicates, n = 30): well-irrigated, decreased irrigation, and limited irrigation. Lower water levels had no significant effect on the growth of S. bicolor but increased the biomass of the roots. Moreover, the number of leaves, leaf area, and leaf size as well as the chlorophyll content were not affected by lower water levels, and no significant changes were detected for whole plant photosynthesis, transpiration, or stomatal conductance. The water content of the plants and the water potential remained unchanged. However, compared with other treatments, the decreased irrigation decreased water loss and increased the water retention. Lower water levels increased the intercellular CO2 percentage, mesophyll area, and proportion of stomatal cavities and promoted minor changes in leaf tissue and stomatal traits. The increased stomatal cavities provided higher CO2 uptake and prevented excessive water loss. Thus, modifications to the intercellular spaces promoted conditions to avoid excessive water loss while concurrently improving CO2 uptake, which are important traits for drought-tolerant plants.

Expression of Human Immunodeficiency Virus Transactivator of Transcription (HIV-Tat1-86) Protein Alters Nociceptive Processing that is Sensitive to Anti-Oxidant and Anti-Inflammatory Interventions.

Despite the success of combined antiretroviral therapy (cART) in reducing viral load, a substantial portion of Human Immunodeficiency Virus (HIV)+ patients report chronic pain. The exact mechanism underlying this co-morbidity even with undetectable viral load remains unknown, but the transactivator of transcription (HIV-Tat) protein is of particular interest. Functional HIV-Tat protein is observed even in cerebrospinal fluid of patients who have an undetectable viral load. It is hypothesized that Tat protein exposure is sufficient to induce neuropathic pain-like manifestations via both activation of microglia and generation of oxidative stress. iTat mice conditionally expressed Tat(1-86) protein in the central nervous system upon daily administration of doxycycline (100 mg/kg/d, i.p., up to 14 days). The effect of HIV-Tat protein exposure on the well-being of the animal was assessed using sucrose-evoked grooming and acute nesting behavior for pain-depressed behaviors, and the development of hyperalgesia assessed with warm-water tail-withdrawal and von Frey assays for thermal hyperalgesia and mechanical allodynia, respectively. Tissue harvested at select time points was used to assess ex vivo alterations in oxidative stress, astrocytosis and microgliosis, and blood-brain barrier integrity with assays utilizing fluorescence-based indicators. Tat protein induced mild thermal hyperalgesia but robust mechanical allodynia starting after 4 days of exposure, reaching a nadir after 7 days. Changes in nociceptive processing were associated with reduced sucrose-evoked grooming behavior without altering acute nesting behavior, and in spinal cord dysregulated free radical generation as measured by DCF fluorescence intensity, altered immunohistochemical expression of the gliotic markers, Iba-1 and GFAP, and increased permeability of the blood-brain barrier to the small molecule fluorescent tracer, sodium fluorescein, in a time-dependent manner. Pretreatment with the anti-inflammatory, indomethacin (1 mg/kg/d, i.p.), the antioxidant, methylsulfonylmethane (100 mg/kg/d i.p.), or the immunomodulatory agent, dimethylfumarate (100 mg/kg/d p.o.) thirty minutes prior to daily injections of doxycycline (100 mg/kg/d i.p.) over 7 days significantly attenuated the development of Tat-induced mechanical allodynia. Collectively, the data suggests that even acute exposure to HIV-1 Tat protein at pathologically relevant levels is sufficient to produce select neurophysiological and behavioral manifestations of chronic pain consistent with that reported by HIV-positive patients.

The role of reactive oxygen species and nitric oxide in the formation of root cortical aerenchyma under cadmium contamination.

The present study aimed to evaluate root cortical aerenchyma formation in response to Cd-driven hydrogen peroxide (H2 O2 ) production and the role of nitric oxide (NO) in the alleviation of Cd oxidative stress in maize roots and its effects on aerenchyma development. Maize plants were subjected to continuous flooding for 30 days, and the following treatments were applied weekly: Cd(NO3 )2 at 0, 10, and 50 µM and Na2 [Fe(CN)5 NO]·2H2 O (an NO donor) at 0.5, 0.1, and 0.2 µM. The root biometrics; oxidative stress indicators H2 O2 and malondialdehyde (MDA); and activities of catalase (CAT), superoxide dismutase (SOD), and ascorbate peroxidase (APX) were analyzed. The root dry and fresh masses decreased at higher concentrations of NO and Cd. H2 O2 also decreased at higher NO concentrations; however, MDA increased only at higher Cd levels. SOD activity decreased at higher concentrations of NO, but CAT activity increased. Aerenchyma development decreased in response to NO. Consequently, NO acts as an antagonist to Cd, decreasing the concentration of H2 O2 by reducing SOD activity and increasing CAT activity. Although H2 O2 is directly linked to aerenchyma formation, increased H2 O2 concentrations are necessary for root cortical aerenchyma development.

Impact of deoxynivalenol on intestinal explants of broiler chickens: An ex vivo model to assess antimycotoxins additives.

The impact of deoxynivalenol (DON) upon intestinal tissue of broilers was assessed by using jejunal explants in Ussing chambers and analyzing histopathological and immunohistochemical parameters; this system was also applied to evaluate the efficacy of an antimycotoxins additive (AMA). The explants were subjected to the following treatments within each experiment for 120 min: Experiment 1) T1 (control) - buffer solution, and T2 - 10 mg/L DON; and Experiment 2) T1 (control) - buffer solution, T2 - 10 mg/L DON, T3 - AMA (0.5%), and T4 - 10 mg/L DON + 0.5% AMA. In Experiment 1, DON triggered a reduction in the size of enterocytes as well as of their nuclei, an increase in cytoplasmic vacuolization and apical denudation of villi. Apoptotic cells count was also greater in DON-exposed explants. In Experiment 2, the AMA mitigated DON harmful effects; cytoplasmic vacuolization of enterocytes was reduced and the size of their nuclei was preserved. The additive also promoted a partial decrease in microvillus integrity, in size of enterocytes and in apoptotic cells count. The tested ex vivo model demonstrated the impact of DON upon the intestine as well as the efficacy of the AMA against its damaging effects.

Aerenchyma, gas diffusion, and catalase activity in Typha domingensis: a complementary model for radial oxygen loss.

Radial oxygen loss is a physical phenomenon that occurs naturally in aquatic plants. Typha domingensis was chosen as a model plant because it possesses basic morphological characteristics, such as a stem (rhizome) that produces leaves and adventitious roots, which are present in many aquatic plants. This study aimed to evaluate the following: the relevance of the anatomy of T. domingensis on gas diffusion among organs; the influence of plant parts on radial oxygen loss; the role of catalase in radial oxygen loss; and the proposition of a novel explanation for the downward diffusion of oxygen through the organs of this aquatic macrophyte and into the environment. Typha domingensis plants were cultivated in a greenhouse under different conditions: plants with intact leaves, plants with leaves cut in half, and plants without leaves. Furthermore, we evaluated the percentage of aerenchyma in different vegetative organs, the minimum pressure required for radial oxygen loss, the daily variations of dissolved oxygen, and the roots' catalase activity. The results demonstrated that certain cellular features contributed to decreased oxygen diffusion among the organs, specifically, those found in the leaf-rhizome and root-rhizome interfaces as well as the suberin and lignin layers in these regions. Additionally, our experiments with a catalase activator and inhibitor validated that a significant amount of the oxygen released in radial oxygen loss could not, in fact, be exclusively supplied by the atmosphere. Thus, a complementary model is proposed in which catalase activity is an important component of radial oxygen loss.