The fate of Salmonella enterica and Listeria monocytogenes in the pulp of eight native Brazilian and exotic fruits.

Despite the wide variety of native and exotic fruits in Brazil, there is limited understanding of their ability to support pathogens during storage. This study aimed to evaluate the behavior of Salmonella enterica and Listeria monocytogenes inoculated into the pulp of eight fruits native and exotic to Brazil: Jenipapo (Genipa americana L.), Umbu (Spondias tuberosa Arruda), Maná (Solanum sessiliflorum), Cajá-manga (Spondias dulcis), Physalis (Physalis angulata L.), Feijoa (Acca sellowiana), Cupuaçu (Theobroma grandiflorum) (average pH < 3.3) and in a low acidy fruit: Abiu (Pouteria caimito) (pH 6.11). The pathogens were inoculated into the different fruits and stored at 10, 20, 30 and 37 °C for up to 12 h and 6 days, respectively. Among the fruits evaluated, Abiu was the only one that allowed Salmonella growth, showing higher δ-values at 20 and 30 °C (5.6 log CFU/g for both temperatures). For Physalis and Feijoa, there was a small reduction in the pathogen concentration (<1 log-cycle), mainly at 10 and 20 °C, indicating its ability to remain in the matrices. For the other fruits, notable negative δ-values were obtained, indicating a tendency towards microbial inactivation. The survival potential was significantly affected by temperature in Abiu, Maná, Cupuaçu, and Cajá-manga (p < 0.05). The same phenomena regarding δ-value were observed for L. monocytogenes population, with the greatest survival potential observed at 20 °C in Abiu (3.3 log CFU/g). Regarding the exponential growth rates in Abiu, the highest values were observed at 30 and 37 °C, both for Salmonella (4.6 and 4.9 log (CFU/g)/day, respectively) and for L. monocytogenes (2.8 and 2.7 log (CFU/g)/day, respectively), with no significant difference between both temperatures. Regarding microbial inactivation, L. monocytogenes showed greater resistance than Salmonella in practically all matrices. Jenipapo and Umbu were the pulps that, in general, had the greatest effect on reducing the population of pathogens. Furthermore, the increase in storage temperature seems to favor the increase on inactivation rates. In conclusion, Salmonella and L. monocytogenes can grow only in Abiu pulp, although they can survive in some acidic tropical fruits kept at refrigeration and abusive temperatures.

Investigating desiccation resistance, post-rehydration growth, and heat tolerance in desiccation-injured cells of Salmonella enterica isolated from the soybean production chain.

This study compared the resistance to different desiccation conditions of 190 Salmonella enterica strains previously isolated from the soybean meal production chain and belonging to 23 serovars. Additionally, the post-rehydration growth and heat tolerance of the strains previously exposed to desiccation were determined. Variability in desiccation resistance was observed both within and between serovars. Strains belonging to S. Havana and S. Schwarzengrund serovars were the most resistant, regardless of storage condition. The drying temperature (20 °C and 30 °C) did not influence the desiccation resistance of the Salmonella strains. On the other hand, increasing drying time from 1 to 7 days reduced Salmonella counts. The origin (isolation sources) also influenced the desiccation resistance of the Salmonella strains. The growth of the Salmonella strains after rehydration varied considerably depending on the drying conditions and incubation temperature during cultivation. An increase in the time and temperature of drying led to a reduction in population of most Salmonella strains after rehydration. Salmonella strains previously desiccated also showed differences in the heat tolerance in all temperature-time binomials tested. Some strains were highly resistant to heat tolerance conditions, presenting <1 log CFU/mL reduction from the initial population. The results obtained in this study suggest that the strategies to mitigate Salmonella in low-aw foods must consider the existence of high-stress resistant strains and their multiple-stress adaptability profiles, including effects of processing, food composition, and storage conditions.

Differential roles for ACBD4 and ACBD5 in peroxisome-ER interactions and lipid metabolism.

Peroxisomes and the endoplasmic reticulum (ER) are intimately linked subcellular organelles, physically connected at membrane contact sites. While collaborating in lipid metabolism, for example, of very long-chain fatty acids (VLCFAs) and plasmalogens, the ER also plays a role in peroxisome biogenesis. Recent work identified tethering complexes on the ER and peroxisome membranes that connect the organelles. These include membrane contacts formed via interactions between the ER protein VAPB (vesicle-associated membrane protein-associated protein B) and the peroxisomal proteins ACBD4 and ACBD5 (acyl-coenzyme A-binding domain protein). Loss of ACBD5 has been shown to cause a significant reduction in peroxisome-ER contacts and accumulation of VLCFAs. However, the role of ACBD4 and the relative contribution these two proteins make to contact site formation and recruitment of VLCFAs to peroxisomes remain unclear. Here, we address these questions using a combination of molecular cell biology, biochemical, and lipidomics analyses following loss of ACBD4 or ACBD5 in HEK293 cells. We show that the tethering function of ACBD5 is not absolutely required for efficient peroxisomal ß-oxidation of VLCFAs. We demonstrate that loss of ACBD4 does not reduce peroxisome-ER connections or result in the accumulation of VLCFAs. Instead, the loss of ACBD4 resulted in an increase in the rate of ß-oxidation of VLCFAs. Finally, we observe an interaction between ACBD5 and ACBD4, independent of VAPB binding. Overall, our findings suggest that ACBD5 may act as a primary tether and VLCFA recruitment factor, whereas ACBD4 may have regulatory functions in peroxisomal lipid metabolism at the peroxisome-ER interface.

Lateral septum adenosine A2A receptors control stress-induced depressive-like behaviors via signaling to the hypothalamus and habenula.

Major depressive disorder ranks as a major burden of disease worldwide, yet the current antidepressant medications are limited by frequent non-responsiveness and significant side effects. The lateral septum (LS) is thought to control of depression, however, the cellular and circuit substrates are largely unknown. Here, we identified a subpopulation of LS GABAergic adenosine A2A receptors (A2AR)-positive neurons mediating depressive symptoms via direct projects to the lateral habenula (LHb) and the dorsomedial hypothalamus (DMH). Activation of A2AR in the LS augmented the spiking frequency of A2AR-positive neurons leading to a decreased activation of surrounding neurons and the bi-directional manipulation of LS-A2AR activity demonstrated that LS-A2ARs are necessary and sufficient to trigger depressive phenotypes. Thus, the optogenetic modulation (stimulation or inhibition) of LS-A2AR-positive neuronal activity or LS-A2AR-positive neurons projection terminals to the LHb or DMH, phenocopied depressive behaviors. Moreover, A2AR are upregulated in the LS in two male mouse models of repeated stress-induced depression. This identification that aberrantly increased A2AR signaling in the LS is a critical upstream regulator of repeated stress-induced depressive-like behaviors provides a neurophysiological and circuit-based justification of the antidepressant potential of A2AR antagonists, prompting their clinical translation.

Generation of Reporter Cell Lines for Endogenous Expression Analysis of Peroxisomal Proteins.

Peroxisomes are multifunctional, ubiquitous, and dynamic organelles. They are responsible for diverse metabolic and physiological functions and communicate with other organelles, including the ER, mitochondria, lipid droplets, and lysosomes, through membrane contact sites. However, despite their importance for healthy cell function, remarkably, little is known about how peroxisomes and peroxisomal proteins are regulated under physiological conditions in human cells. Here, we present a method to generate reporter cell lines to measure endogenous expression of peroxisomal proteins of interest. By CRISPR-mediated knock-in of an easily detectable protein-coding tag in-frame into the relevant genomic loci, endogenous levels of the protein of interest in a cell population can be quantified in a high-throughput manner under different conditions. This has important implications for the fundamental understanding of how peroxisomal proteins are regulated and may reveal the therapeutic potential of modulating peroxisomal protein expression to improve cell performance.

The model of litter size reduction induces long-term disruption of the gut-brain axis: An explanation for the hyperphagia of Wistar rats of both sexes.

The gut microbiota affects the host's metabolic phenotype, impacting health and disease. The gut-brain axis unites the intestine with the centers of hunger and satiety, affecting the eating behavior. Deregulation of this axis can lead to obesity onset. Litter size reduction is a well-studied model for infant obesity because it causes overnutrition and programs for obesity. We hypothesize that animals raised in small litters (SL) have altered circuitry between the intestine and brain, causing hyperphagia. We investigated vagus nerve activity, the expression of c-Fos, brain-derived neurotrophic factor (BDNF), gastrointestinal (GI) hormone receptors, and content of bacterial phyla and short-chain fatty acids (SCFAs) in the feces of adult male and female Wistar rats overfed during lactation. On the 3rd day after birth, litter size was reduced to 3 pups/litter (SL males or SL females) until weaning. Controls had normal litter size (10 pups/litter: 5 males and 5 females). The rats were killed at 5 months of age. The male and female offspring were analyzed separately. The SL group of both sexes showed higher food consumption and body adiposity than the respective controls. SL animals presented dysbiosis (increased Firmicutes, decreased Bacteroidetes) and had increased vagus nerve activity. Only the SL males had decreased hypothalamic GLP-1 receptor expression, while only the SL females had lower acetate and propionate in the feces and higher CCK receptor expression in the hypothalamus. Thus, overfeeding during lactation differentially changes the gut-brain axis, contributing to hyperphagia of the offspring of both sexes.

Maintaining social contacts: The physiological relevance of organelle interactions.

Membrane-bound organelles in eukaryotic cells form an interactive network to coordinate and facilitate cellular functions. The formation of close contacts, termed "membrane contact sites" (MCSs), represents an intriguing strategy for organelle interaction and coordinated interplay. Emerging research is rapidly revealing new details of MCSs. They represent ubiquitous and diverse structures, which are important for many aspects of cell physiology and homeostasis. Here, we provide a comprehensive overview of the physiological relevance of organelle contacts. We focus on mitochondria, peroxisomes, the Golgi complex and the plasma membrane, and discuss the most recent findings on their interactions with other subcellular organelles and their multiple functions, including membrane contacts with the ER, lipid droplets and the endosomal/lysosomal compartment.

Antioxidant activity and bioaccessibility of phenolic compounds in white, red, blue, purple, yellow and orange edible flowers through a simulated intestinal barrier.

This study assessed the phenolics and their bioaccessibility through an in vitro digestion system coupled to a simulated intestinal barrier in eight edible flowers of distinct colors, namely mini rose, torenia, mini daisy, clitoria, cosmos, cravine, begonia and tagete. The antioxidant activity of the flowers before in vitro digestion, in their derived dialyzed and non-dialyzed fractions was evaluated using distinct approaches. All flowers presented in their composition phenolic acids, stilbenes, flavanol, anthocyanin, flavonol and flavanone, however distinct compounds and contents were found in each flower. The bioaccessibility varied among the phenolics and within the flower source (p < 0.05). Cosmos presented the highest (p < 0.05) content of phenolics and activity in ORAC assay before in vitro digestion and in dialyzed and non-dialyzed fraction; the observed activity was correlated (r = 0.9) to its major compounds, hesperidin and rutin, as well as to caftaric acid and procyanidin B2. Mini rose displayed the highest antioxidant activity in FRAP and DPPH assays before in vitro digestion; its dialyzed and non-dialyzed fraction showed the highest activity in FRAP, correlated to pelargonidin 3,5-diglucoside, catechin, epicatechin galate, epicagocatechin galate, procyanidin A2, quercitin 3-glucoside and trans-resveratrol (r = 0.9). In DPPH assay, mini rose showed the highest activity in the non-dialyzed fraction, while cravine showed the highest activity in the dialyzed fraction, which was mainly correlated to syringic acid (r = 1.0), pelargonidin 3,5-diglucoside and epicatechin (r = 0.9). Results show great variability in the phenolic composition and their bioaccessibility among the edible flowers studied. Our findings indicate cosmos and mini rose as sources of bioaccessible phenolics with great antioxidant activity.

Ecogenomics of the Marine Benthic Filamentous Cyanobacterium Adonisia.

Turfs are among the major benthic components of reef systems worldwide. The nearly complete genome sequences, basic physiological characteristics, and phylogenomic reconstruction of two phycobiliprotein-rich filamentous cyanobacteria strains isolated from turf assemblages from the Abrolhos Bank (Brazil) are investigated. Both Adonisia turfae CCMR0081T (= CBAS 745T) and CCMR0082 contain approximately 8 Mbp in genome size and experiments identified that both strains exhibit chromatic acclimation. Whereas CCMR0081T exhibits chromatic acclimation type 3 (CA3) regulating both phycocyanin (PC) and phycoerythrin (PE), CCMR0082 strain exhibits chromatic acclimation type 2 (CA2), in correspondence with genes encoding specific photosensors and regulators for PC and PE. Furthermore, a high number and diversity of secondary metabolite synthesis gene clusters were identified in both genomes, and they were able to grow at high temperatures (28 °C, with scant growth at 30 °C). These characteristics provide insights into their widespread distribution in reef systems.

The presence of ochratoxin A does not influence Saccharomyces cerevisiae growth kinetics but leads to the formation of modified ochratoxins.

Yeasts are able to reduce the levels of ochratoxin A in fermentative processes; and, through their enzymatic complex, these micro-organisms are also capable of forming modified mycotoxins. These mycotoxins are often underreported, and may increase health risks after ingestion of contaminated food. In this sense, this study aims to evaluate whether the presence of ochratoxin A influences yeast growth kinetic parameters and to elucidate the formation of modified ochratoxin by Saccharomyces cerevisiae strains during fermentation. Three S. cerevisiae strains (12 M, 01 PP, 41 PP) were exposed to OTA at the concentrations of 10, 20 and 30 µg/L. The Baranyi model was fitted to the growth data (Log CFU/mL), and the identification of modified ochratoxins was performed through High Resolution Mass Spectrometry. The presence of ochratoxin A did not influence the growth of S. cerevisiae strains. Four pathways were proposed for the metabolization of OTA: dechlorination, hydrolysis, hydroxylation, and conjugation. Among the elected targets, the following were identified: ochratoxin α, ochratoxin ß, ochratoxin α methyl ester, ochratoxin B methyl ester, ethylamide ochratoxin A, ochratoxin C, hydroxy-ochratoxin A, hydroxy-ochratoxin A methyl ester, and ochratoxin A cellobiose ester. These derivatives formed from yeast metabolism may contribute to the occurrence of underreporting levels of total mycotoxin in fermented products.

Epidemiology and forensic aspects of carbon monoxide intoxication in Portugal: A three years' analysis.

The presented manuscript describes the carbon monoxide (CO) related deaths in Portugal over a period of 3 years, based on autopsies carried out at the National Institute of Legal Medicine and Forensic Sciences, from January 2012 to December 2014. Three hundred and forty-seven forensic autopsy reports with carboxyhaemoglobin (COHb) analysis requests were analysed and subdivided into three main groups: (1) improbable CO intoxication; (2) possible CO intoxication; (3) highly probable CO intoxication. In group 1, COHb analysis was negative, and the death circumstances, as well as the post mortem findings, didn't corroborate an exposition to CO. In group 2, with COHb positive in 1/3 of the cases, the death circumstances corroborated an exposition to CO, but the post mortem findings weren't enough to confirm an exposition to this substance. In group 3, the results of COHb were positive, and both circumstances of death and post mortem findings corroborated an exposition to CO. The first group (113 cases) had no specific suspicion of a CO intoxication and, thus, the request of a COHb analysis had no particular basis, reflected in the low COHb achieved percentage (between 0 and 12). In the second group (164 cases), 29% of the cases were directly or indirectly related to CO exposure (between 0% and 94%). In the third group (70 cases), 56 deaths were due to CO intoxication and 14 due to burns after CO inhalation (between 18% and 91%). This study intended to do, not only a 3-year assessment of CO poisoning, but also to enhance the fact that circumstantial information, as well as a correct evaluation at the forensic autopsy data are crucial, and allow an enhanced diagnosis of possible intoxication, as well as a better guidance for the consequent toxicological analysis requests.

Caffeine Controls Glutamatergic Synaptic Transmission and Pyramidal Neuron Excitability in Human Neocortex.

Caffeine is the most widely used psychoactive drug, bolstering attention and normalizing mood and cognition, all functions involving cerebral cortical circuits. Whereas studies in rodents showed that caffeine acts through the antagonism of inhibitory A1 adenosine receptors (A1R), neither the role of A1R nor the impact of caffeine on human cortical neurons is known. We here provide the first characterization of the impact of realistic concentrations of caffeine experienced by moderate coffee drinkers (50 µM) on excitability of pyramidal neurons and excitatory synaptic transmission in the human temporal cortex. Moderate concentrations of caffeine disinhibited several of the inhibitory A1R-mediated effects of adenosine, similar to previous observations in the rodent brain. Thus, caffeine restored the adenosine-induced decrease of both intrinsic membrane excitability and excitatory synaptic transmission in the human pyramidal neurons through antagonism of post-synaptic A1R. Indeed, the A1R-mediated effects of endogenous adenosine were more efficient to inhibit synaptic transmission than neuronal excitability. This was associated with a distinct affinity of caffeine for synaptic versus extra-synaptic human cortical A1R, probably resulting from a different molecular organization of A1R in human cortical synapses. These findings constitute the first neurophysiological description of the impact of caffeine on pyramidal neuron excitability and excitatory synaptic transmission in the human temporal cortex, providing adequate ground for the effects of caffeine on cognition in humans.

Incidence and growth of Salmonella enterica on the peel and pulp of avocado (Persea americana) and custard apple (Annona squamosa).

The aim of this study was to assess the incidence and to estimate the growth kinetic parameters (maximum growth rate, µ; lag time, λ; and maximum population, κ) of Salmonella on the peel and pulp of avocado (Perseaamericana var. americana) and custard apple (Annona squamosa L.) as affected by temperature (10-30°C). The incidence of Salmonella was assessed on the peel and pulp of the fruits (n=200 of each fruit), separately, totalizing 800 analyses. Only three samples of custard apple pulp were positive for Salmonella enterica and the three isolates recovered belonged to serotype S. Typhimurium. Salmonella was not recovered from avocado and custard apple peels and from avocado pulp. Generally, the substrate (pulp or peel) of growth did not affect µ values of S. enterica (p>0.05). Very similar µ values were found for S. enterica inoculated in custard apple and avocado. S. enterica presented the highest λ in the peel of the fruits. The growth of S. enterica resulted in larger λ in custard apple in comparison to avocado. For example, the λ of S. enterica in the pulp of custard apple and avocado were 47.0±0.78h and 10.0±3.78h, respectively. The lowest values of κ were obtained at the lower storage temperature conditions (10°C). For instance, κ values of 3.7±0.06log CFU/g and 2.9±0.03log CFU/g were obtained from the growth of S. enterica in avocado and custard apple pulps at 10°C (p<0.05), respectively. On the other hand, at 30°C, κ values were 6.5±0.25log CFU/g and 6.5±0.05log CFU/g, respectively. Significantly higher κ were obtained from the growth of S. enterica in the pulp than in the peel of the fruits (p<0.05). For instance, the growth of S. enterica in the pulp of avocado led to a κ value of 6.5±0.25log CFU/g, while in the peel led to a κ value of 4.6±0.23log CFU/g (p<0.05). In general, growth kinetic parameters indicated that avocado comprises a better substrate than custard apple for the growth of S. enterica. The square root model fitted to the data obtained in this study and to the growth data available in the literature for other tropical low acid fruits indicated high variability in µ and λ of Salmonella. The results obtained in this study show that whole low acid tropical fruits can harbor Salmonella, and that this foodborne pathogen can not only survive but also grow both on the peel and pulp of low acid tropical fruits, such as avocado and custard apple.

Age-Related Changes in the Synaptic Density of Amyloid-ß Protein Precursor and Secretases in the Human Cerebral Cortex.

Amyloid-ß protein precursor (AßPP) is involved in synaptic formation and function. In the human cingulate cortex, AßPP was preferentially located in the presynaptic active zone as in rodents, indicating a preserved subsynaptic AßPP distribution across species and brain regions. Synaptic AßPP immunoreactivity was decreased with aging in cortical samples collected from autopsies of males (20-80 years), whereas the synaptic levels of α-secretase (ADAM10) and ß-secretase (BACE1) did not significantly change. Decreased AßPP levels may be related to lower allostasis of synapses in the aged brain and their greater susceptibility to dysfunction characteristic of the onset of neurodegenerative disorders.

Incidence, populations and diversity of fungi from raw materials, final products and air of processing environment of multigrain whole meal bread.

This study aimed to assess the incidence, to quantify and to assess the diversity of fungi in a multigrain whole meal bread processing plant. Two hundred and eight one (n=281) samples were analyzed, including raw materials (n=120), air samples (n=136) and multigrain breads (n=25). Among the raw materials, the whole corn flour showed the highest counts of fungi (4.8logCFU/g), followed by whole-wheat flour (3.1logCFU/g). The counts of fungi in the air of processing environment were higher in post-baking steps (oven output, cooling, slicing, packaging) than in pre-baking steps (weighing and mixer) (p<0.05). Species of fungi isolated from spoiled bread samples stored at 5, 20, 25 and 30, and 40°C corresponded mostly to Penicillium paneum and Penicillium polonicum isolated from 20 and 24% of samples, respectively. These species were also isolated from raw materials (P. paneum and P. polonicum) and air collected at different processing sampling points (P. polonicum). The high counts of filamentous fungi in raw materials and air samples in processing steps such as cooling, slicing, and packaging, suggest that contamination that may occur in these steps can be critical for the shelf life of breads. The results of this study highlight that the prevention of contamination of breads by fungal spores is still a challenge for bakery industries and that other strategies such as control of germination and growth of spoilage fungi through the development of more stable formulations have to be developed.