Whole-cortex in situ sequencing reveals input-dependent area identity.

The cerebral cortex is composed of neuronal types with diverse gene expression that are organized into specialized cortical areas. These areas, each with characteristic cytoarchitecture1,2, connectivity3,4 and neuronal activity5,6, are wired into modular networks3,4,7. However, it remains unclear whether these spatial organizations are reflected in neuronal transcriptomic signatures and how such signatures are established in development. Here we used BARseq, a high-throughput in situ sequencing technique, to interrogate the expression of 104 cell-type marker genes in 10.3 million cells, including 4,194,658 cortical neurons over nine mouse forebrain hemispheres, at cellular resolution. De novo clustering of gene expression in single neurons revealed transcriptomic types consistent with previous single-cell RNA sequencing studies8,9. The composition of transcriptomic types is highly predictive of cortical area identity. Moreover, areas with similar compositions of transcriptomic types, which we defined as cortical modules, overlap with areas that are highly connected, suggesting that the same modular organization is reflected in both transcriptomic signatures and connectivity. To explore how the transcriptomic profiles of cortical neurons depend on development, we assessed cell-type distributions after neonatal binocular enucleation. Notably, binocular enucleation caused the shifting of the cell-type compositional profiles of visual areas towards neighbouring cortical areas within the same module, suggesting that peripheral inputs sharpen the distinct transcriptomic identities of areas within cortical modules. Enabled by the high throughput, low cost and reproducibility of BARseq, our study provides a proof of principle for the use of large-scale in situ sequencing to both reveal brain-wide molecular architecture and understand its development.

Gating of homeostatic regulation of intrinsic excitability produces cryptic long-term storage of prior perturbations.

Neurons and neuronal circuits must maintain their function throughout the life of the organism despite changing environments. Previous theoretical and experimental work suggests that neurons monitor their activity using intracellular calcium concentrations to regulate their intrinsic excitability. Models with multiple sensors can distinguish among different patterns of activity, but previous work using models with multiple sensors produced instabilities that lead the models' conductances to oscillate and then to grow without bound and diverge. We now introduce a nonlinear degradation term that explicitly prevents the maximal conductances to grow beyond a bound. We combine the sensors' signals into a master feedback signal that can be used to modulate the timescale of conductance evolution. Effectively, this means that the negative feedback can be gated on and off according to how far the neuron is from its target. The modified model recovers from multiple perturbations. Interestingly, depolarizing the models to the same membrane potential with current injection or with simulated high extracellular K+ produces different changes in conductances, arguing that caution must be used in interpreting manipulations that serve as a proxy for increased neuronal activity. Finally, these models accrue traces of prior perturbations that are not visible in their control activity after perturbation but that shape their responses to subsequent perturbations. These cryptic or hidden changes may provide insight into disorders such as posttraumatic stress disorder that only become visible in response to specific perturbations.

Human epiblast lumenogenesis: From a cell aggregate to a lumenal cyst.

The formation of a central lumen in the human epiblast is a critical step for development. However, because the lumen forms in the epiblast coincident with implantation, the molecular and cellular events of this early lumenogenesis process cannot be studied in vivo. Recent developments using new model systems have revealed insight into the underpinnings of epiblast formation. To provide an up-to-date comprehensive review of human epiblast lumenogenesis, we highlight recent findings from human and mouse models with an emphasis on new molecular understanding of a newly described apicosome compartment, a novel 'formative' state of pluripotency that coordinates with epiblast polarization, and new evidence about the physical and polarized trafficking mechanisms contributing to lumenogenesis.

In-Depth Proteome Coverage of In Vitro-Cultured Treponema pallidum and Quantitative Comparison Analyses with In Vivo-Grown Treponemes.

Previous mass spectrometry (MS)-based global proteomics studies have detected a combined total of 86% of all Treponema pallidum proteins under infection conditions (in vivo-grown T. pallidum). Recently, a method was developed for the long-term culture of T. pallidum under in vitro conditions (in vitro-cultured T. pallidum). Herein, we used our previously reported optimized MS-based proteomics approach to characterize the T. pallidum global protein expression profile under in vitro culture conditions. These analyses provided a proteome coverage of 94%, which extends the combined T. pallidum proteome coverage from the previously reported 86% to a new combined total of 95%. This study provides a more complete understanding of the protein repertoire of T. pallidum. Further, comparison of the in vitro-expressed proteome with the previously determined in vivo-expressed proteome identifies only a few proteomic changes between the two growth conditions, reinforcing the suitability of in vitro-cultured T. pallidum as an alternative to rabbit-based treponemal growth. The MS proteomics data have been deposited in the MassIVE repository with the data set identifier MSV000093603 (ProteomeXchange identifier PXD047625).

Prevalence and risk of burnout among HIV service providers in South Africa and Zambia: findings from the HPTN 071 (PopART) trial.

BACKGROUND: In the high disease burden and resource-constrained contexts of sub-Saharan Africa (SSA), health workers experience a range of psychosocial stressors that leave them vulnerable to developing burnout, which can reduce service quality and negatively impact their own health and wellbeing. As universal testing and treatment (UTT) for HIV scales up across SSA, we sought to understand the implications of this human resource-intensive approach to HIV prevention to inform decision-making about health workforce staffing and support needs. METHODS: Using the Maslach Burnout Inventory-Human Services Survey (MBI-HSS), we assessed the prevalence of three domains of burnout-emotional exhaustion, depersonalization, and personal accomplishment-among three cadres of health workers delivering health services in areas receiving a UTT intervention in Zambia and South Africa. These cadres included health facility workers (n = 478), community health workers (n = 159), and a study-specific cadre of community HIV care providers (n = 529). We used linear regression to assess risk factors associated with emotional exhaustion, the only domain with sufficient variation in our sample. RESULTS: The MBI-HSS was completed by 1499/2153 eligible participants (69.6% response rate). Less than 1% of health workers met Maslach's definition for burnout. All groups of health workers reported lower levels of emotional exhaustion than found in previous studies of this type (mean score scores ranged from 10.7 to 15.4 out of 54 across health cadres). Higher emotional exhaustion was associated with higher educational attainment (ßadj = 2.24, 95% CI 0.76 to 3.72), greater years providing HIV services (ßadj = 0.20, 95% CI 0.03 to 0.36), and testing negative for HIV at last HIV test (ßadj = - 3.88 - 95% CI 5.69 to - 2.07). Working as a CHW was significantly associated with lower emotional exhaustion (ßadj = - 2.52, 95% CI - 4.69 to - 0.35). Among all health workers, irrespective of HIV status, witnessing stigmatizing behaviors towards people living with HIV among their co-workers was associated with significantly increased emotional exhaustion (ßadj = 3.38, 95% CI 1.99 to 4.76). CONCLUSIONS: The low level of burnout detected among health workers is reassuring. However, it remains important to assess how UTT may affect levels of emotional exhaustion among health workers over time, particularly in the context of emerging global pandemics, as burnout may impact the quality of HIV services they provide and their own mental health and wellbeing. Interventions to reduce HIV stigma in health facilities may protect against emotional exhaustion among health workers, as well as interventions to increase mindfulness and resilience among health workers at risk of burnout. Trial registration ClinicalTrials.gov number: NCT01900977.

The Pseudomonas aeruginosa homeostasis enzyme AlgL clears the periplasmic space of accumulated alginate during polymer biosynthesis.

Pseudomonas aeruginosa is an opportunistic human pathogen and a leading cause of chronic infection in the lungs of individuals with cystic fibrosis. After colonization, P. aeruginosa often undergoes a phenotypic conversion to mucoidy, characterized by overproduction of the alginate exopolysaccharide. This conversion is correlated with poorer patient prognoses. The majority of genes required for alginate synthesis, including the alginate lyase, algL, are located in a single operon. Previous investigations of AlgL have resulted in several divergent hypotheses regarding the protein's role in alginate production. To address these discrepancies, we determined the structure of AlgL and, using multiple sequence alignments, identified key active site residues involved in alginate binding and catalysis. In vitro enzymatic analysis of active site mutants highlights R249 and Y256 as key residues required for alginate lyase activity. In a genetically engineered P. aeruginosa strain where alginate biosynthesis is under arabinose control, we found that AlgL is required for cell viability and maintaining membrane integrity during alginate production. We demonstrate that AlgL functions as a homeostasis enzyme to clear the periplasmic space of accumulated polymer. Constitutive expression of the AlgU/T sigma factor mitigates the effects of an algL deletion during alginate production, suggesting that an AlgU/T-regulated protein or proteins can compensate for an algL deletion. Together, our study demonstrates the role of AlgL in alginate biosynthesis, explains the discrepancies observed previously across other P. aeruginosa ΔalgL genetic backgrounds, and clarifies the existing divergent data regarding the function of AlgL as an alginate degrading enzyme.

MicroRNA-Transcription factor regulatory networks in the early strobilar development of Echinococcus granulosus protoscoleces.

BACKGROUND: Echinococcus granulosus sensu lato has a complex developmental biology with a variety of factors relating to both intermediate and final hosts. To achieve maximum parasite adaptability, the development of the cestode is dependent on essential changes in transcript regulation. Transcription factors (TFs) and miRNAs are known as master regulators that affect the expression of downstream genes through a wide range of metabolic and signaling pathways. In this study, we aimed to develop a regulatory miRNA-Transcription factor (miRNA-TF) network across early developmental stages of E. granulosus protoscoleces by performing in silico analysis, and to experimentally validate TFs expression in protoscoleces obtained from in vitro culture, and from in vivo experiments. RESULTS: We obtained list of 394 unique E. granulosus TFs and matched them with 818 differentially expressed genes which identified 41 predicted TFs with differential expression. These TFs were used to predict the potential targets of 31 differentially expressed miRNAs. As a result, eight miRNAs and eight TFs were found, and the predicted network was constructed using Cytoscape. At least four miRNAs (egr-miR-124a, egr-miR-124b-3p, egr-miR-745-3p, and egr-miR-87-3p) and their corresponding differentially expressed TFs (Zinc finger protein 45, Early growth response protein 3, Ecdysone induced protein 78c and ETS transcription factor elf 2) were highlighted in this investigation. The expression of predicted differentially expressed TFs obtained from in vitro and in vivo experiments, were experimentally validated by quantitative polymerase chain reaction. This confirmed findings of RNA-seq data. CONCLUSION: miRNA-TF networks presented in this study control some of the most important metabolic and signaling pathways in the development and life cycle of E. granulosus, providing a potential approach for disrupting the early hours of dog infection and preventing the development of the helminth in the final host.

A pathogenic role for cystic fibrosis transmembrane conductance regulator in celiac disease.

Intestinal handling of dietary proteins usually prevents local inflammatory and immune responses and promotes oral tolerance. However, in ~ 1% of the world population, gluten proteins from wheat and related cereals trigger an HLA DQ2/8-restricted TH1 immune and antibody response leading to celiac disease. Prior epithelial stress and innate immune activation are essential for breaking oral tolerance to the gluten component gliadin. How gliadin subverts host intestinal mucosal defenses remains elusive. Here, we show that the α-gliadin-derived LGQQQPFPPQQPY peptide (P31-43) inhibits the function of cystic fibrosis transmembrane conductance regulator (CFTR), an anion channel pivotal for epithelial adaptation to cell-autonomous or environmental stress. P31-43 binds to, and reduces ATPase activity of, the nucleotide-binding domain-1 (NBD1) of CFTR, thus impairing CFTR function. This generates epithelial stress, tissue transglutaminase and inflammasome activation, NF-κB nuclear translocation and IL-15 production, that all can be prevented by potentiators of CFTR channel gating. The CFTR potentiator VX-770 attenuates gliadin-induced inflammation and promotes a tolerogenic response in gluten-sensitive mice and cells from celiac patients. Our results unveil a primordial role for CFTR as a central hub orchestrating gliadin activities and identify a novel therapeutic option for celiac disease.

From the Neuroscience of Individual Variability to Climate Change.

Years of basic neuroscience on the modulation of the small circuits found in the crustacean stomatogastric ganglion have led us to study the effects of temperature on the motor patterns produced by the stomatogastric ganglion. While the impetus for this work was the study of individual variability in the parameters determining intrinsic and synaptic conductances, we are confronting substantial fluctuations in the stability of the networks to extreme temperature; these may correlate with changes in ocean temperature. Interestingly, when studied under control conditions, these wild-caught animals appear to be unchanged, but it is only when challenged by extreme temperatures that we reveal the consequences of warming oceans.

Quantification of mitophagy using mKeima-mito in cultured human primary retinal pigment epithelial cells.

The retinal pigment epithelium is a pigmented monolayer of cells that help maintain a healthy retina. Loss of this essential cell layer is implicated in a number of visual disorders, including age-related macular degeneration (AMD). Utilizing primary RPE cultures to investigate disease is an important step in understanding disease mechanisms. However, the use of primary RPE cultures presents a number of challenges, including the limited number of cells available and the presence of auto-fluorescent pigment that interferes with quantifying fluorescent probes. Additionally, primary RPE are difficult to transfect with exogenous nucleic acids traditionally used for fluorescent imaging. To overcome these challenges, we used an adeno-associated viral (AAV) vector to express a pH sensitive fluorescent protein, mKeima, fused to the mitochondrial targeting sequence of cytochrome oxidase subunit 8A (mKeima-mito). mKeima-mito allows for quantification of mitochondrial autophagy (mitophagy) in live-cell time-lapse imaging experiments. We also developed an image analysis pipeline to selectively quantify mKeima-mito while removing the signal of auto-fluorescent pigment from the dataset by utilizing information from the mKeima fluorescent channels. These techniques are demonstrated in primary RPE cultures expressing mKeima-mito treated with 2-[2-[4-(trifluoromethoxy)phenyl]hydrazinylidene]-propanedinitrile (FCCP), an uncoupler that depolarizes the mitochondrial membrane and leads to mitochondrial fragmentation and mitophagy. The techniques outlined provide a roadmap for investigating disease mechanisms or the effect of treatments utilizing fluorescent probes in an important cell culture model.

Exomer complex regulates protein traffic at the TGN through differential interactions with cargos and clathrin adaptor complexes.

Protein sorting at the trans-Golgi network (TGN) usually requires the assistance of cargo adaptors. However, it remains to be examined how the same complex can mediate both the export and retention of different proteins or how sorting complexes interact among themselves. In Saccharomyces cerevisiae, the exomer complex is involved in the polarized transport of some proteins from the TGN to the plasma membrane (PM). Intriguingly, exomer and its cargos also show a sort of functional relationship with TGN clathrin adaptors that is still unsolved. Here, using a wide range of techniques, including time-lapse and BIFC microscopy, we describe new molecular implications of the exomer complex in protein sorting and address its different layers of functional interaction with clathrin adaptor complexes. Exomer mutants show impaired amino acid uptake because it facilitates not only the polarized delivery of amino acid permeases to the PM but also participates in their endosomal traffic. We propose a model for exomer where it modulates the recruitment of TGN clathrin adaptors directly or indirectly through the Arf1 function. Moreover, we describe an in vivo competitive relationship between the exomer and AP-1 complexes for the model cargo Chs3. These results highlight a broad role for exomer in regulating protein sorting at the TGN that is complementary to its role as cargo adaptor and present a model to understand the complexity of TGN protein sorting.

Depressive Symptoms Associated With Musculoskeletal Pain in Inactive Adults During COVID-19 Quarantine.

BACKGROUND: Depression has been associated with episodes of musculoskeletal pain. However, it is not clear whether such relationships could be mitigated according to the physical activity level. AIM: To describe, during the COVID-19 pandemic, the relationship between depression and musculoskeletal pain according to the physical activity levels. DESIGN: Cross-sectional study. METHODS: This research was conducted in Brazil between May 5 and March 17, 2020. Participants (N = 1872; 58% women) were invited through social media to answer a structured online questionnaire. Depressive symptoms were assessed through self-report of perception of depression during quarantine. Musculoskeletal pain was assessed based on the Nordic questionnaire identifying nine possible pain points in the body. Physical activity was assessed based on the weekly frequency, intensity, and duration of each session of physical activity the participants engaged in during COVID-19. The logistic binary regression analyzed the associations between depressive symptoms and musculoskeletal pain according to the participants' level of physical activity. RESULTS: Depressive symptoms were associated with pain in six different regions of the body in physically inactive participants. In physically inactive participants, those with depressive symptoms 1.51 (95% CI = 1.04-2.19) and 2.78 (95% CI = 1.81-4.26) times more likely to have pain in one or two and ≥three regions body regions, respectively. In active participants, depressive symptoms were not associated with pain. CONCLUSION: During the COVID-19 pandemic, depression was associated with musculoskeletal pain in physically inactive participants.

BKC-2, a New BKC Variant Detected in MCR-9.1-Producing Enterobacter hormaechei subsp. xiangfangensis.

We characterized a multidrug-resistant (MDR) Enterobacter spp. isolate highlighting the genetic aspects of the antimicrobial resistance genes. An Enterobacter spp. isolate (Ec61) was recovered in 2014 from a transtracheal aspirate sample from a patient admitted to a Brazilian tertiary hospital and submitted to further microbiological and genomic characterization. Ec61 was identified as Enterobacter hormaechei subsp. xiangfangensis strain ST451, showing an MDR profile and the presence of genes codifying the new ß-lactamase variants BKC-2 and ACT-84 and the mobile colistin resistance gene mcr-9.1.

Evaluation of kinase inhibitors as potential therapeutics for flavivirus infections.

The recent introduction of Zika virus (ZIKV), the recurrence of dengue virus (DENV), and the lethality of yellow fever virus (YFV) have had a significant impact on Brazilian society and public health. Here, we targeted two cellular kinases implicated in cell proliferation and cancer that are also important for viral replication: mitogen-activated protein kinase kinase (MEK) and Src. We used two MEK inhibitors - trametinib and selumetinib - and two Src inhibitors - saracatinib and bosutinib - to inhibit ZIKV, DENV, and YFV replication in cell culture. The cytotoxicity of the four inhibitors was determined by the observation of abnormal morphology and quantification of adherent cells by crystal violet staining. The antiviral activity of these drugs was assessed based on the reduction of plaque-forming units in cell culture as evidence of the inhibition of the replication of the selected flaviviruses. All four inhibitors showed antiviral activity, but among them, trametinib was the safest and most efficacious against all of the viruses, inhibiting the replication of ZIKV and YFV by 1000-fold, and DENV2/3 by nearly 100-fold. This pan-antiviral effect shows that trametinib could be repurposed for the treatment of flaviviral infections.

Plasma membrane to vacuole traffic induced by glucose starvation requires Gga2-dependent sorting at the trans-Golgi network.

BACKGROUND INFORMATION: In the yeast Saccharomyces cerevisiae, acute glucose starvation induces rapid endocytosis followed by vacuolar degradation of many plasma membrane proteins. This process is essential for cell viability, but the regulatory mechanisms that control it remain poorly understood. Under normal growth conditions, a major regulatory decision for endocytic cargo occurs at the trans-Golgi network (TGN) where proteins can recycle back to the plasma membrane or can be recognized by TGN-localised clathrin adaptors that direct them towards the vacuole. However, glucose starvation reduces recycling and alters the localization and post-translational modification of TGN-localised clathrin adaptors. This raises the possibility that during glucose starvation endocytosed proteins are routed to the vacuole by a novel mechanism that bypasses the TGN or does not require TGN-localised clathrin adaptors. RESULTS: Here, we investigate the role of TGN-localised clathrin adaptors in the traffic of several amino acid permeases, including Can1, during glucose starvation. We find that Can1 transits through the TGN after endocytosis in both starved and normal conditions. Can1 and other amino acid permeases require TGN-localised clathrin adaptors for maximal delivery to the vacuole. Furthermore, these permeases are actively sorted to the vacuole, because ectopically forced de-ubiquitination at the TGN results in the recycling of the Tat1 permase in starved cells. Finally, we report that the Mup1 permease requires the clathrin adaptor Gga2 for vacuolar delivery. In contrast, the clathrin adaptor protein complex AP-1 plays a minor role, potentially in retaining permeases in the TGN, but it is otherwise dispensable for vacuolar delivery. CONCLUSIONS AND SIGNIFICANCE: This work elucidates one membrane trafficking pathway needed for yeast to respond to acute glucose starvation. It also reveals the functions of TGNlocalised clathrin adaptors in this process. Our results indicate that the same machinery is needed for vacuolar protein sorting at the GN in glucose starved cells as is needed in the presence of glucose. In addition, our findings provide further support for the model that the TGN is a transit point for many endocytosed proteins, and that Gga2 and AP-1 function in distinct pathways at the TGN.

The implementation of routine procedural transvaginal sonography to decrease retained products of conception: a quality improvement initiative.

BACKGROUND: Retained products of conception (POC) following uterine evacuation can lead to adverse sequelae, including hemorrhage, endometritis, intrauterine adhesions, and reoperation. Use of procedural transvaginal sonography (TVUS) in the operating room has been proposed to help decrease retained POC. METHODS: A retrospective review of all first trimester uterine evacuation procedures from 1/2015 to 2/2017 was performed, noting use of transabdominal ultrasonography, retained products of conception, and complications. A practice change was implemented in May 2018, in which routine intra-procedural TVUS use was initiated. A second retrospective chart review was conducted to assess for post-implementation incidence of retained POC, re-operation, and associated complications. RESULTS: Prior to intra-procedural TVUS implementation, 130 eligible procedures were performed during the specified timeframe, with 9/130 (6.9%) incidence of retained products of conception. TAUS was performed in 59/130 (45.4%) of procedures, and 4/9 (44.4%) of those with retained products. There were eight re-operative procedures in seven patients, and two patients were treated with misoprostol. Complications included hemorrhage, Asherman's syndrome and endometritis. Following implementation, 95 first trimester procedures were performed with transvaginal sonography, with 0 (0%) cases of retained POC (p = 0.01), no incidences of re-operation (p = 0.02), and one case of Asherman's syndrome. TVUS findings led to additional focused suction curettage in 20/95 (21.0%) of procedures. The endometrium was measured on procedure completion in 64 procedures, with a mean thickness of 5.5 mm (1-12 mm). CONCLUSION: Implementation of routine TVUS during uterine evacuation may reduce the incidence of retained POC and associated reoperation rates. Further multi-center trials are needed to confirm this finding.

MicroRNA profile of the strobilated worms of Echinococcus granulosus derived from in vivo and in vitro systems by using high-throughput approach.

MicroRNAs are critical gene regulators at the post-transcriptional level and play essential roles in numerous developmental processes in metazoan parasites including the causative agent of cystic echinococcosis, Echinococcus granulosus. The molecular basis of different patterns of E. granulosus development in the canine definitive host and in in vitro culture systems is poorly understood. In the present study, miRNA transcriptomes of the strobilated worms derived from experimental infection in the definitive host were compared with those from diphasic culture system after 60-day protoscoleces cultivation. Total RNA was extracted from in vivo- and in vitro-derived strobilated worms. Small RNA libraries were constructed, and deep sequencing was performed. Subsequently, differential miRNA expressions and target predictions were obtained, and pathway analysis was performed by gene ontology and KEGG. Seven miRNAs were differentially expressed between the in vivo- and in vitro-derived worms. In addition, we reported 13 novel miRNA candidates and 42 conserved miRNAs. Four out of five top miRNAs with the highest read counts were shared between the in vivo and in vitro-derived worms, i.e., egr-miR-10a-5p, egr-let-7-5p, egr-bantam-3p, and egr-miR-71-5p. Target prediction of the differential miRNAs between the two systems showed significant differences in the membrane-enclosed lumen, membrane part, and an intrinsic component of the membrane. Findings of KEGG analysis indicated that differentially expressed miRNAs were involved in hippo, MAPK, and WNT signaling pathways. The study demonstrated a significant difference in miRNA transcriptomes and related signaling pathways between the two systems, suggesting the importance of host-parasite interplay in the fate of protoscoleces development in in vivo and in vitro systems.

Spatiotemporal distribution and human health risk assessment of potential toxic species in soils of urban and surrounding crop fields from an agricultural area, Córdoba, Argentina.

The spatiotemporal distribution of potentially toxic species was studied in the topsoil of parks, playgrounds, and surrounding crop fields of Marcos Juarez City in Córdoba province, Argentina. The content of available metals and of some pesticides used in the region was determined. The mean values of available metal concentrations in all samples, expressed in mg kg-1, were 7.99 ± 6.58, 0.89 ± 0.71, 0.35 ± 0.26, and 1.50 ± 1.40 for Pb, Cr, Cd, and Ni, respectively. Pearson's correlation coefficients, coefficient of variation (%), and principal component analysis were used to explore whether variations in metal content were associated with anthropogenic factors. Agrochemicals such as lindane, chlorobenzilate, endosulfan, endrin, permethrin, and chlorpyrifos were found in crop field soil samples. In turn, chlorothalonil, chlordanes, methoxychlor, DDT, permethrin, and chlorpyrifos were detected in park and playground soil samples. The degree of pollution with possible effects on environmental health was evaluated using the Nemerow integrated pollution index (NIPI) and the modified degree of contamination (mCd). In some campaigns, the values obtained from crop fields ranged from low to high pollution levels during periods of agrochemical application. Noteworthy, in periods of low agrochemical application in crop fields, a high level of pollution was observed in parks and playgrounds. For children, the hazard index (HI) values were higher than the threshold value of 1, suggesting a potential health risk. This study provides valuable information regarding land management practices and highlights the importance of monitoring and implementing policies to reduce human health risks.

Effects of freeze-dried pulp of Eugenia uniflora L. and Opuntia ficus-indica fruits on quality attributes of beef patties enriched with n-3 fatty acids.

The objective of this work was to evaluate the effect of freeze-dried pulps of Eugenia uniflora L. and Opuntia ficus-indica fruits on oxidative stability of n-3-enriched beef patties. Besides, the effect of freeze-dried pulps on cooking yield, color, texture parameters and sensory acceptance of patties was also evaluated. Eugenia uniflora L. (orange-skinned variety) and Opuntia ficus-indica (orange-skinned and red-skinned varieties) fruits were collected, their pulp was lyophilized and incorporated, as a natural source of antioxidant, in beef patties enriched with canola oil as a source of polyunsaturated fatty acids. Oxidation stability of samples was monitored by measuring thiobarbituric acid-reactive substances during refrigerated storage. Results show that all freeze-dried pulps limited lipid oxidation to an acceptable level during 15 days refrigerated storage, having Opuntia ficus-indica (red-skinned variety) the highest inhibitory activity. Besides, the addition of freeze-dried pulps increased the cooking yield and decreased the hardness of beef patties. Respect to sensorial parameters, patties elaborated with no added freeze-dried fruit pulp and patties elaborated with freeze-dried pulp of Opuntia ficus-indica (red-skinned variety) presented the highest overall acceptance of consumers, remaining as a future task to improve the color and taste of patties elaborated with freeze-dried pulps of Eugenia uniflora L. and Opuntia ficus-indica (orange-skinned variety). This research reveals the effectiveness of studied fruits to reduce lipid oxidation in beef patties. In addition, preliminary information about which sensorial parameters of these products should be improved in futures incorporation of pulps in the manufacture of healthy meat products.

Adaptor protein complex-1 (AP-1) is recruited by the HEATR5 protein Laa1 and its co-factor Laa2 in yeast.

Cellular membrane trafficking mediated by the clathrin adaptor protein complex-1 (AP-1) is important for the proper composition and function of organelles of the endolysosomal system. Normal AP-1 function requires proteins of the HEAT repeat-containing 5 (HEATR5) family. Although HEATR5 proteins were first identified based on their ability to interact with AP-1, the functional significance of this interaction was unknown. We used bioinformatics-based phenotypic profiling and information from genome-wide fluorescence microscopy studies in the budding yeast Saccharomyces cerevisiae to identify a protein, Laa2, that mediates the interaction between AP-1 and the yeast HEATR5 protein Laa1. Further characterization of Laa2 revealed that it binds to both Laa1 and AP-1. Laa2 contains a motif similar to the characterized γ-ear-binding sites found in other AP-1-binding proteins. This motif in Laa2 is essential for the Laa1-AP-1 interaction. Moreover, mutation of this motif disrupted AP-1 localization and function and caused effects similar to mutations that remove the γ-ear of AP-1. These results indicate that Laa2 mediates the interaction between Laa1 and AP-1 and reveal that this interaction promotes the stable association of AP-1 with membranes in yeast.