Search

The study of microplastics (MPs) in soils is impeded by similarities between plastic and non-plastic particles and the misidentification of MP by current analytical methods such as visual microscopic examination. Soil MPs pose serious ecological and public health risks because of their abundance, persistence, and ubiquity. Thus, reliable identification methods are badly needed for scientific study. One possible solution is UV–Vis–NIR spectroscopy, which has the ability to rapidly identify and quantify concentrations of soil microplastics. In this study, a full-range, field portable spectrometer (350–2500 nm) with ultra-high spectral resolution (1.5 nm, 3.0 nm, and 3.8 nm) identified three types of common plastics: low-density polyethylene (LDPE), polyvinyl chloride (PVC), and polypropylene (PP). Three sets of artificially MP-treated vermiculite soil samples were prepared for model prediction testing and validation: 150 samples for model calibration and 50 samples for model validation. A partial least square regression model using the spectral signatures for quantification of soil and MP mixtures was built with all three plastic polymers. Prediction R2 values of all three polymers showed promising results: polypropylene R2 = 0.943, polyvinyl chloride R2 = 0.983, and polyethylene R2 = 0.957. Our study supports previous work showing that combining ultra-high-resolution UV–Vis–NIR spectrometry with quantitative modeling can improve the accuracy and speed of MP identification and quantification in soil.

Abstract Objectives Silicosis is an irreversible occupational lung disease resulting from crystalline silica inhalation. Previously, we discovered that Western diet (HFWD)-consumption increases susceptibility to silica-induced pulmonary inflammation and fibrosis. This study investigated the potential of HFWD to alter silica-induced effects on airway epithelial ion transport and smooth muscle reactivity. Methods Six-week-old male F344 rats were fed a HFWD or standard rat chow (STD) and exposed to silica (Min-U-Sil 5®, 15 mg/m3, 6 h/day, 5 days/week, for 39 d) or filtered air. Experimental endpoints were measured at 0, 4, and 8 weeks post-exposure. Transepithelial potential difference (Vt), short-circuit current (I SC) and transepithelial resistance (Rt) were measured in tracheal segments and ion transport inhibitors [amiloride, Na+ channel blocker; NPPB; Clˉ channel blocker; ouabain, Na+, K+-pump blocker] identified changes in ion transport pathways. Changes in airway smooth muscle reactivity to methacholine (MCh) were investigated in the isolated perfused trachea preparation. Results Silica reduced basal I SC at 4 weeks and HFWD reduced the I SC response to amiloride at 0 week compared to air control. HFWD + silica exposure induced changes in ion transport 0 and 4 weeks after treatment compared to silica or HFWD treatments alone. No effects on airway smooth muscle reactivity to MCh were observed.

Air sanitization is an important non-pharmaceutical intervention for mitigating the risk of indoor pathogen spreading. A dipropylene glycol-containing air sanitizer was tested against aerosolized Staphylococcus aureus and Klebsiella pneumoniae. The bacteria, suspended in a soil load, were aerosolized using a six-jet Collison nebulizer with pressurized air. The 25-m3 (~900 ft3) aerobiology chamber was maintained at 22 ± 2 °C and 50 ± 5% relative humidity per the U.S. Environmental Protection Agency’s 2012 Guidelines on air sanitizers. An initial 2-min air sample was collected from the chamber using a slit-to-agar sampler containing 150-mm Petri plates, with Trypticase soy agar (TSA) containing neutralizers to quench the microbicidal activity of the air sanitizer, to determine the initial bacterial challenge in the air. The air sanitizer was sprayed into the chamber from pressurized cans. Additional air samples were collected from the chamber over 10 min to detect surviving bacteria. The TSA plates were then incubated aerobically at 36 ± 1 °C for 90 ± 4 h and scored for bacterial colony-forming units. A 30-s spray of the air sanitizer reduced infectious S. aureus and K. pneumoniae titers by 3.0 log10 (99.9%) in 3.2 ± 0.3 min and 1.2 ± 0.0 min, respectively. Based on these findings, the EPA granted registration of the air sanitizer as the first product of its kind for indoor air sanitization.

Abstract Despite the longstanding use of nitrous oxide and descriptions of its psychological effects more than a century ago, there is a paucity of neurobiological investigation of associated psychedelic experiences. We measure the brain’s functional geometry (through analysis of cortical gradients) and temporal dynamics (through analysis of co-activation patterns) using human resting-state functional magnetic resonance imaging data acquired before and during administration of 35% nitrous oxide. Both analyses demonstrate that nitrous oxide reduces functional differentiation in frontoparietal and somatomotor networks. Importantly, the subjective psychedelic experience induced by nitrous oxide is inversely correlated with the degree of functional differentiation. Thus, like classical psychedelics acting on serotonin receptors, nitrous oxide flattens the functional geometry of the cortex and disrupts temporal dynamics in association with psychoactive effects.

Background: Fibromyalgia (FM) is a chronic pain disorder and is associated with disability, and high levels of pain and suffering. FM is known to co-occur with obesity and obstructive sleep apnea (OSA). Individuals with FM often experience symptoms of pain, depression and anxiety, sleep disturbances, and fatigue. These symptoms may be exacerbated by OSA and contribute to the symptoms’ severity in FM. Obesity is a common comorbidity in OSA patients, and as FM and OSA are related in some patients, obesity also may contribute to FM symptom severity. For healthcare providers to effectively manage FM patients, a better understanding of the co-occurrence between these FM comorbidities and psychological factors is needed. Methods: This study was approved by IRB and conducted using a retrospective EPIC chart review. To identify FM, the following ICD-9 codes were used: (729.1) and ICD-10 (M79.7) codes. To identify patients with OSA, the following ICD-9 codes were used: (327.23) and ICD-10 (G47.33). Body Mass Index (BMI), the total number of medical diagnoses, and psychiatric conditions were documented for each patient. The prevalence of psychiatric conditions including depression and anxiety was compared between patients with and without obesity (BMI > 30), and patients with fewer than 25 medical diagnoses and those with 25 or more diagnoses. A chart review was conducted to identify patients with fibromyalgia with prior serum cortisol testing within the last ten years. Cortisol levels were compared and patients were divided into six groups: 1. FM without identified psychiatric conditions; 2. FM with psychiatric diagnosis of adjustment disorders and insomnia; 3. FM with psychiatric diagnosis of depressive disorders; 4. FM with psychiatric diagnosis of bipolar disorders; 5. FM with psychiatric diagnosis of mixed anxiety and depression; 6. FM with psychiatric diagnosis of anxiety disorders. Available C-reactive protein (CRP) values were gathered. Results: The total FM and OSA population was N = 331. The mean age of the patient population was 63.49 years old, with 297 being female. The diagnoses mean was 31.79 ± 17.25 and the mean total psychiatric diagnoses was 2.80 ± 1.66. The mean BMI was 36.69 ± 8.86, with obesity present in 77.95% of the patients. A total of 66.99% of patients had comorbid anxiety and depression with 25 or more medical problems vs. 33.01% of patients who had fewer than 25 medical problems (odds ratio = 1.50). Patients with a BMI < 30 (N = 71) had rates of anxiety and depression at 64.79% and a mean total of 2.79 ± 1.66 psychiatric diagnoses, whereas patients with a BMI > 30 (N = 258) had rates of anxiety and depression at 61.63% (odds ratio = 1.28) and a mean total of 2.80 ± 1.66 psychiatric diagnoses. The most common other psychiatric conditions among FM/OSA patients included hypersomnia and substance use disorders. Cortisol data: Available cortisol results: FM n = 64, female: 59, male: 5, mean age: 63, average BMI: 38.8. The averages for serum cortisol alone for groups 1–6, respectively, are 9.06, 5.49, 13.00, 14.17, 12.25, and 16.03 μg/dL. These results indicate a relatively upward cortisol serum value by the addition of several psychiatric conditions, with the most notable being anxiety for patients with FM. CRP values were available for 53 patients with an average CRP of 4.14. Discussion: Higher rates of anxiety and depression were present in FM patients with 25 or more diagnoses. The odds ratios indicate that a patient with 25 or more medical problems was 1.5 times more likely to have anxiety and depression than those with fewer diagnoses. Additionally, those with a BMI > 30 were 1.3 times more likely to have anxiety and depression than those with a normal BMI. Conclusion: addressing psychological factors in FM and OSA is important as high healthcare utilization is common in patients with FM and OSA.

During the recent pandemic of COVID-19 (SARS-CoV-2), influential public health agencies such as the World Health Organization (WHO) and the U.S. Centers for Disease Control and Prevention (CDC) have favored the view that SARS CoV-2 spreads predominantly via droplets. Many experts in aerobiology have openly opposed that stance, forcing a vigorous debate on the topic. In this review, we discuss the various proposed modes of viral transmission, stressing the interdependencies between droplet, aerosol, and fomite spread. Relative humidity and temperature prevailing determine the rates at which respiratory aerosols and droplets emitted from an expiratory event (sneezing, coughing, etc.) evaporate to form smaller droplets or aerosols, or experience hygroscopic growth. Gravitational settling of droplets may result in contamination of environmental surfaces (fomites). Depending upon human, animal and mechanical activities in the occupied space indoors, viruses deposited on environmental surfaces may be re-aerosolized (re-suspended) to contribute to aerosols, and can be conveyed on aerial particulate matter such as dust and allergens. The transmission of respiratory viruses may then best be viewed as resulting from dynamic virus spread from infected individuals to susceptible individuals by various physical states of active respiratory emissions, instead of the current paradigm that emphasizes separate dissemination by respiratory droplets, aerosols or by contaminated fomites. To achieve the optimum outcome in terms of risk mitigation and infection prevention and control (IPAC) during seasonal infection peaks, outbreaks, and pandemics, this holistic view emphasizes the importance of dealing with all interdependent transmission modalities, rather than focusing on one modality.

Summary: Ovarian high-grade serous carcinoma (HGSC) is the most common subtype of ovarian cancer with limited therapeutic options and a poor prognosis. In recent years, poly-ADP ribose polymerase (PARP) inhibitors have demonstrated significant clinical benefits, especially in patients with BRCA1/2 mutations. However, acquired drug resistance and relapse is a major challenge. Indisulam (E7070) has been identified as a molecular glue that brings together splicing factor RBM39 and DCAF15 E3 ubiquitin ligase resulting in polyubiquitination, degradation, and subsequent RNA splicing defects. In this work, we demonstrate that the loss of RBM39 induces splicing defects in key DNA damage repair genes in ovarian cancer, leading to increased sensitivity to cisplatin and various PARP inhibitors. The addition of indisulam also improved olaparib response in mice bearing PARP inhibitor-resistant tumors. These findings demonstrate that combining RBM39 degraders and PARP inhibitors is a promising therapeutic approach to improve PARP inhibitor response in ovarian HGSC.

A DNA damage-inducible mutagenic gene cassette has been implicated in the emergence of drug resistance in Mycobacterium tuberculosis during anti-tuberculosis (TB) chemotherapy. However, the molecular composition and operation of the encoded ‘mycobacterial mutasome’ – minimally comprising DnaE2 polymerase and ImuA′ and ImuB accessory proteins – remain elusive. Following exposure of mycobacteria to DNA damaging agents, we observe that DnaE2 and ImuB co-localize with the DNA polymerase III β subunit (β clamp) in distinct intracellular foci. Notably, genetic inactivation of the mutasome in an imuBAAAAGG mutant containing a disrupted β clamp-binding motif abolishes ImuB–β clamp focus formation, a phenotype recapitulated pharmacologically by treating bacilli with griselimycin and in biochemical assays in which this β clamp-binding antibiotic collapses pre-formed ImuB–β clamp complexes. These observations establish the essentiality of the ImuB–β clamp interaction for mutagenic DNA repair in mycobacteria, identifying the mutasome as target for adjunctive therapeutics designed to protect anti-TB drugs against emerging resistance.

Collective cell migration plays an essential role in vertebrate development, yet the extent to which dynamically changing microenvironments influence this phenomenon remains unclear. Observations of the distribution of the extracellular matrix (ECM) component fibronectin during the migration of loosely connected neural crest cells (NCCs) lead us to hypothesize that NCC remodeling of an initially punctate ECM creates a scaffold for trailing cells, enabling them to form robust and coherent stream patterns. We evaluate this idea in a theoretical setting by developing an individual-based computational model that incorporates reciprocal interactions between NCCs and their ECM. ECM remodeling, haptotaxis, contact guidance, and cell-cell repulsion are sufficient for cells to establish streams in silico, however, additional mechanisms, such as chemotaxis, are required to consistently guide cells along the correct target corridor. Further model investigations imply that contact guidance and differential cell-cell repulsion between leader and follower cells are key contributors to robust collective cell migration by preventing stream breakage. Global sensitivity analysis and simulated gain- and loss-of-function experiments suggest that long-distance migration without jamming is most likely to occur when leading cells specialize in creating ECM fibers, and trailing cells specialize in responding to environmental cues by upregulating mechanisms such as contact guidance.

Aspergillus versicolor is ubiquitous in the environment and is particularly abundant in damp indoor spaces. Exposure to Aspergillus species, as well as other environmental fungi, has been linked to respiratory health outcomes, including asthma, allergy, and even local or disseminated infection. However, the pulmonary immunological mechanisms associated with repeated exposure to A. versicolor have remained relatively uncharacterized. Here, A. versicolor was cultured and desiccated on rice then placed in an acoustical generator system to achieve aerosolization. Mice were challenged with titrated doses of aerosolized conidia to examine deposition, lymphoproliferative properties, and immunotoxicological response to repeated inhalation exposures. The necessary dose to induce lymphoproliferation was identified, but not infection-like pathology. Further, it was determined that the dose was able to initiate localized immune responses. The data presented in this study demonstrate an optimized and reproducible method for delivering A. versicolor conidia to rodents via nose-only inhalation. Additionally, the feasibility of a long-term repeated exposure study was established. This experimental protocol can be used in future studies to investigate the physiological effects of repeated pulmonary exposure to fungal conidia utilizing a practical and relevant mode of delivery. In total, these data constitute an important foundation for subsequent research in the field.

Abstract Background The functional understanding of genetic interaction networks and cellular mechanisms governing health and disease requires the dissection, and multifaceted study, of discrete cell subtypes in developing and adult animal models. Recombinase-driven expression of transgenic effector alleles represents a significant and powerful approach to delineate cell populations for functional, molecular, and anatomical studies. In addition to single recombinase systems, the expression of two recombinases in distinct, but partially overlapping, populations allows for more defined target expression. Although the application of this method is becoming increasingly popular, its experimental implementation has been broadly restricted to manipulations of a limited set of common alleles that are often commercially produced at great expense, with costs and technical challenges associated with production of intersectional mouse lines hindering customized approaches to many researchers. Here, we present a simplified CRISPR toolkit for rapid, inexpensive, and facile intersectional allele production. Results Briefly, we produced 7 intersectional mouse lines using a dual recombinase system, one mouse line with a single recombinase system, and three embryonic stem (ES) cell lines that are designed to study the way functional, molecular, and anatomical features relate to each other in building circuits that underlie physiology and behavior. As a proof-of-principle, we applied three of these lines to different neuronal populations for anatomical mapping and functional in vivo investigation of respiratory control. We also generated a mouse line with a single recombinase-responsive allele that controls the expression of the calcium sensor Twitch-2B. This mouse line was applied globally to study the effects of follicle-stimulating hormone (FSH) and luteinizing hormone (LH) on calcium release in the ovarian follicle. Conclusions The lines presented here are representative examples of outcomes possible with the successful application of our genetic toolkit for the facile development of diverse, modifiable animal models. This toolkit will allow labs to create single or dual recombinase effector lines easily for any cell population or subpopulation of interest when paired with the appropriate Cre and FLP recombinase mouse lines or viral vectors. We have made our tools and derivative intersectional mouse and ES cell lines openly available for non-commercial use through publicly curated repositories for plasmid DNA, ES cells, and transgenic mouse lines.

The locus coeruleus (LC) houses the vast majority of noradrenergic neurons in the brain and regulates many fundamental functions, including fight and flight response, attention control, and sleep/wake cycles. While efferent projections of the LC have been extensively investigated, little is known about its local circuit organization. Here, we performed large-scale multipatch recordings of noradrenergic neurons in adult mouse LC to profile their morpho-electric properties while simultaneously examining their interactions. LC noradrenergic neurons are diverse and could be classified into two major morpho-electric types. While fast excitatory synaptic transmission among LC noradrenergic neurons was not observed in our preparation, these mature LC neurons connected via gap junction at a rate similar to their early developmental stage and comparable to other brain regions. Most electrical connections form between dendrites and are restricted to narrowly spaced pairs or small clusters of neurons of the same type. In addition, more than two electrically coupled cell pairs were often identified across a cohort of neurons from individual multicell recording sets that followed a chain-like organizational pattern. The assembly of LC noradrenergic neurons thus follows a spatial and cell-type-specific wiring principle that may be imposed by a unique chain-like rule.

The contiguous United States (CONUS) harbor a significant non-native species diversity. However, spatiotemporal trends of some groups such as terrestrial gastropods (i.e., land snails and slugs) have not been comprehensively considered, and therefore management has been hindered. Here, our aims were to 1.) compile a dataset of all non-native terrestrial gastropod species with CONUS occurrence records, 2.) assess overarching spatiotemporal patterns associated with these records, 3.) describe the continental origin of each species, and 4.) compare climatic associations of each species in their indigenous and introduced CONUS ranges. We compiled a georeferenced dataset of 10,097 records for 22 families, 48 genera, and 69 species, with > 70% of records sourced from the citizen science database iNaturalist. The species Cornu aspersum Müller, 1774 was most prevalent with 3,672 records. The majority (> 92%) of records exhibit an indigenous Western European and Mediterranean distribution, with overlap in broad-scale climatic associations between indigenous and CONUS ranges. Records are most dense in urban metropolitan areas, with the highest proportion of records and species richness in the state of California. We show increased prevalence of non-native species through time, largely associated with urbanized areas with high human population density. Moreover, we show strong evidence for a role for analogous climates in dictating geographic fate and pervasiveness between indigenous and CONUS ranges for non-native species.

Heterogeneity of glucose-stimulated insulin secretion (GSIS) in pancreatic islets is physiologically important but poorly understood. Here, we utilize mouse islets to determine how microtubules (MTs) affect secretion toward the vascular extracellular matrix at single cell and subcellular levels. Our data indicate that MT stability in the β-cell population is heterogenous, and that GSIS is suppressed in cells with highly stable MTs. Consistently, MT hyper-stabilization prevents, and MT depolymerization promotes the capacity of single β-cell for GSIS. Analysis of spatiotemporal patterns of secretion events shows that MT depolymerization activates otherwise dormant β-cells via initiation of secretion clusters (hot spots). MT depolymerization also enhances secretion from individual cells, introducing both additional clusters and scattered events. Interestingly, without MTs, the timing of clustered secretion is dysregulated, extending the first phase of GSIS and causing oversecretion. In contrast, glucose-induced Ca2+ influx was not affected by MT depolymerization yet required for secretion under these conditions, indicating that MT-dependent regulation of secretion hot spots acts in parallel with Ca2+ signaling. Our findings uncover a novel MT function in tuning insulin secretion hot spots, which leads to accurately measured and timed response to glucose stimuli and promotes functional β-cell heterogeneity.

Uropathogenic Escherichia coli (UPEC) proliferate within superficial bladder umbrella cells to form intracellular bacterial communities (IBCs) during early stages of urinary tract infections. However, the dynamic responses of IBCs to host stresses and antibiotic therapy are difficult to assess in situ. We develop a human bladder-chip model wherein umbrella cells and bladder microvascular endothelial cells are co-cultured under flow in urine and nutritive media respectively, and bladder filling and voiding mimicked mechanically by application and release of linear strain. Using time-lapse microscopy, we show that rapid recruitment of neutrophils from the vascular channel to sites of infection leads to swarm and neutrophil extracellular trap formation but does not prevent IBC formation. Subsequently, we tracked bacterial growth dynamics in individual IBCs through two cycles of antibiotic administration interspersed with recovery periods which revealed that the elimination of bacteria within IBCs by the antibiotic was delayed, and in some instances, did not occur at all. During the recovery period, rapid proliferation in a significant fraction of IBCs reseeded new foci of infection through bacterial shedding and host cell exfoliation. These insights reinforce a dynamic role for IBCs as harbors of bacterial persistence, with significant consequences for non-compliance with antibiotic regimens.

Molecular studies have recently led to the detection of many cryptic species complexes within morphologically ambiguous species formerly undescribed by the scientific community. Organisms such as land snails are at a particularly higher risk of species misidentification and misinterpretation, in that gastropod systematics are based almost entirely on external shell morphology. Subterranean ecosystems are associated with especially high degrees of cryptic speciation, largely owing to the abiotic similarities of these systems. In this study, we attempt to diagnose the potential cryptic diversity in the troglobitic land snail Helicodiscus barri. Land snails are generally associated with having low vagility, and as such this species’ broad, mosaic distribution indicates the misdiagnosis of this organism as a single species. We analyze both mitochondrial (16S, CO1) and nuclear (28S, H3) genetic data for 23 populations. Phylogeny for H. barri was reconstructed using both maximum-likelihood and Bayesian approaches to assess relationships among populations, and two species delimitation methods (mPTP and ABGD) were used to detect the presence of unique molecular operational taxonomic units (MOTUs). Species delimitation results revealed seven and sixteen MOTUs respectively, suggesting the presence of several cryptic lineages within H. barri. To assess how external shell morphology corresponds with patterns of genetic and environmental variation, two morphometric approaches were used incorporating 115 shells from 31 populations. Both morphometric approaches reveal a significant environmental influence on shell morphology, and one approach showed the significance of MOTU groups. We discuss the delimitation and morphometric results and additionally provide discussion on the taxonomic and conservation implications of this study.

Summary: Conscious access to sensory information is likely gated at an intermediate site between primary sensory and transmodal association cortices, but the structure responsible remains unknown. We perform functional neuroimaging to determine the neural correlates of conscious access using a volitional mental imagery task, a report paradigm not confounded by motor behavior. Titrating propofol to loss of behavioral responsiveness in healthy volunteers creates dysfunction of the anterior insular cortex (AIC) in association with an impairment of dynamic transitions of default-mode and dorsal attention networks. Candidate subcortical regions mediating sensory gating or arousal (thalamus, basal forebrain) fail to show this association. The gating role of the AIC is consistent with findings in awake participants, whose conscious access is predicted by pre-stimulus AIC activity near perceptual threshold. These data support the hypothesis that AIC, situated at an intermediate position of the cortical hierarchy, regulates brain network transitions that gate conscious access.

Given the importance of DNA methylation in protection of the genome against transposable elements and transcriptional regulation in other taxonomic groups, the diversity in both levels and patterns of DNA methylation in the insects raises questions about its function and evolution. We show that the maintenance DNA methyltransferase, DNMT1, affects meiosis and is essential to fertility in milkweed bugs, Oncopeltus fasciatus, while DNA methylation is not required in somatic cells. Our results support the hypothesis that Dnmt1 is required for the transition of germ cells to gametes in O. fasciatus and that this function is conserved in male and female gametogenesis. They further suggest that DNMT1 has a function independent of DNA methylation in germ cells. Our results raise thequestion as to how a gene that is so critical to fitness across multiple insect species is able to diverge widely across the insect tree of life.

We establish a murine lung-on-chip infection model and use time-lapse imaging to reveal the dynamics of host-Mycobacterium tuberculosis interactions at an air-liquid interface with a spatiotemporal resolution unattainable in animal models and to probe the direct role of pulmonary surfactant in early infection. Surfactant deficiency results in rapid and uncontrolled bacterial growth in both macrophages and alveolar epithelial cells. In contrast, under normal surfactant levels, a significant fraction of intracellular bacteria are non-growing. The surfactant-deficient phenotype is rescued by exogenous addition of surfactant replacement formulations, which have no effect on bacterial viability in the absence of host cells. Surfactant partially removes virulence-associated lipids and proteins from the bacterial cell surface. Consistent with this mechanism, the attenuation of bacteria lacking the ESX-1 secretion system is independent of surfactant levels. These findings may partly explain why smokers and elderly persons with compromised surfactant function are at increased risk of developing active tuberculosis.

Meiotic drivers are parasitic loci that force their own transmission into greater than half of the offspring of a heterozygote. Many drivers have been identified, but their molecular mechanisms are largely unknown. The wtf4 gene is a meiotic driver in Schizosaccharomyces pombe that uses a poison-antidote mechanism to selectively kill meiotic products (spores) that do not inherit wtf4. Here, we show that the Wtf4 proteins can function outside of gametogenesis and in a distantly related species, Saccharomyces cerevisiae. The Wtf4poison protein forms dispersed, toxic aggregates. The Wtf4antidote can co-assemble with the Wtf4poison and promote its trafficking to vacuoles. We show that neutralization of the Wtf4poison requires both co-assembly with the Wtf4antidote and aggregate trafficking, as mutations that disrupt either of these processes result in cell death in the presence of the Wtf4 proteins. This work reveals that wtf parasites can exploit protein aggregate management pathways to selectively destroy spores.

Two distinct mechanisms for primordial germ cell (PGC) specification are observed within Bilatera: early determination by maternal factors or late induction by zygotic cues. Here we investigate the molecular basis for PGC specification in Nematostella, a representative pre-bilaterian animal where PGCs arise as paired endomesodermal cell clusters during early development. We first present evidence that the putative PGCs delaminate from the endomesoderm upon feeding, migrate into the gonad primordia, and mature into germ cells. We then show that the PGC clusters arise at the interface between hedgehog1 and patched domains in the developing mesenteries and use gene knockdown, knockout and inhibitor experiments to demonstrate that Hh signaling is required for both PGC specification and general endomesodermal patterning. These results provide evidence that the Nematostella germline is specified by inductive signals rather than maternal factors, and support the existence of zygotically-induced PGCs in the eumetazoan common ancestor.

There is a pressing need to increase the rigor of research in the life and biomedical sciences. To address this issue, we propose that communities of 'rigor champions' be established to campaign for reforms of the research culture that has led to shortcomings in rigor. These communities of rigor champions would also assist in the development and adoption of a comprehensive educational platform that would teach the principles of rigorous science to researchers at all career stages.

Summary: C57BL/6N (N) and C57BL/6J (J) mice possess key genetic differences, including a deletion in the Nicotinamide nucleotide transhydrogenase (Nnt) gene that results in a non-functional protein in J mice. NNT regulates mitochondrial oxidative stress. Although elevated oxidative stress can compromise hematopoietic stem and progenitor cell (HSPC) function, it is unknown whether N- and J-HSPCs are functionally equivalent. Here, we report that J-HSPCs display compromised short-term hematopoietic repopulating activity relative to N-HSPCs that is defined by a delay in lymphoid reconstitution and impaired function of specific multi-potent progenitor populations post transplant. J-HSPCs also displayed elevated reactive oxygen species (ROS) relative to N-HSPCs post transplant and upregulate ROS levels more in response to hematopoietic stress. Nnt knockdown in N-HSPCs recapitulated J-HSPCs’ short-term repopulating defect, indicating that NNT loss contributes to this defect. In summary, C57BL/6N and C57BL/6J HSPCs are not functionally equivalent, which should be considered when determining the substrain most appropriate for investigations of HSPC biology. : By directly assessing the repopulating capability of mouse HSPCs in vivo, Morales-Hernández and colleagues describe critical functional differences between two widely used C57BL/6 substrains, including the ability of HSPCs to resolve oxidative stress during HSCT. This finding highlights the critical importance of genetic background when studying HSPCs, especially their metabolism, mitochondrial function, or response to hematopoietic stress. Keywords: hematopoietic stem cell transplantation, oxidative stress, nicotinamide nucleotide transhydrogenase, Nnt, C57BL/6

Abstract Background The ability to efficiently visualize and manipulate chromosomes is fundamental to understanding the genome architecture of organisms. Conventional chromosome preparation protocols developed for mammalian cells and those relying on species-specific conditions are not suitable for many invertebrates. Hence, a simple and inexpensive chromosome preparation protocol, adaptable to multiple invertebrate species, is needed. Results We optimized a chromosome preparation protocol and applied it to several planarian species (phylum Platyhelminthes), the freshwater apple snail Pomacea canaliculata (phylum Mollusca), and the starlet sea anemone Nematostella vectensis (phylum Cnidaria). We demonstrated that both mitotically active adult tissues and embryos can be used as sources of metaphase chromosomes, expanding the potential use of this technique to invertebrates lacking cell lines and/or with limited access to the complete life cycle. Simple hypotonic treatment with deionized water was sufficient for karyotyping; growing cells in culture was not necessary. The obtained karyotypes allowed the identification of differences in ploidy and chromosome architecture among otherwise morphologically indistinguishable organisms, as in the case of a mixed population of planarians collected in the wild. Furthermore, we showed that in all tested organisms representing three different phyla this protocol could be effectively coupled with downstream applications, such as chromosome fluorescent in situ hybridization. Conclusions Our simple and inexpensive chromosome preparation protocol can be readily adapted to new invertebrate research organisms to accelerate the discovery of novel genomic patterns across the branches of the tree of life.

Summary: Sequences with the capacity to adopt alternative DNA structures have been implicated in cancer etiology; however, the mechanisms are unclear. For example, H-DNA-forming sequences within oncogenes have been shown to stimulate genetic instability in mammals. Here, we report that H-DNA-forming sequences are enriched at translocation breakpoints in human cancer genomes, further implicating them in cancer etiology. H-DNA-induced mutations were suppressed in human cells deficient in the nucleotide excision repair nucleases, ERCC1-XPF and XPG, but were stimulated in cells deficient in FEN1, a replication-related endonuclease. Further, we found that these nucleases cleaved H-DNA conformations, and the interactions of modeled H-DNA with ERCC1-XPF, XPG, and FEN1 proteins were explored at the sub-molecular level. The results suggest mechanisms of genetic instability triggered by H-DNA through distinct structure-specific, cleavage-based replication-independent and replication-dependent pathways, providing critical evidence for a role of the DNA structure itself in the etiology of cancer and other human diseases. : DNA sequences that can adopt alternative structures, such as H-DNA, have been implicated in cancer etiology. Zhao et al. found that such sequences are enriched at translocation breakpoints in human cancer genomes and that repair- and replication-related nucleases cleave H-DNA in both error-free “replication-related” and mutagenic “replication-independent” mechanisms. Keywords: DNA structure, H-DNA, chromosome translocation, mutation hotspot, DNA repair, nuclease, structural simulation

Regulatory T cells (Tregs) play a pivotal role in maintaining immunological tolerance, but they can also play a detrimental role by preventing antitumor responses. Here, we characterized T helper (Th)-like Treg subsets to further delineate their biological function and tissue distribution, focusing on their possible contribution to disease states. RNA sequencing and functional assays revealed that Th2-like Tregs displayed higher viability and autocrine interleukin-2 (IL-2)-mediated activation than other subsets. Th2-like Tregs were preferentially found in tissues rather than circulation and exhibited the highest migratory capacity toward chemokines enriched at tumor sites. These cellular responses led us to hypothesize that this subset could play a role in maintaining a tumorigenic environment. Concurrently, Th2-like Tregs were enriched specifically in malignant tissues from patients with melanoma and colorectal cancer compared to healthy tissue. Overall, our results suggest that Th2-like Tregs may contribute to a tumorigenic environment due to their increased cell survival, higher migratory capacity, and selective T-effector suppressive ability.

Measurement of the apparent conductivity of salt marsh sediments using electromagnetic induction (EMI) is a rapid alternative to traditional methods of salinity determination that can be used to map soil salinity across a marsh surface. Soil salinity measures can provide information about marsh processes, since salinity is important in determining the structure and function of tidally influenced marsh communities. While EMI has been shown to accurately reflect salinity to a specified depth, more information is needed on the potential for spatial and temporal variability in apparent conductivity measures that may impact the interpretation of salinity data. In this study we mapped soil salinity at two salt marshes in the Narragansett Bay, RI estuary monthly over the course of several years to examine spatial and temporal trends in marsh salinity. Mean monthly calculated salinity was 25.8 ± 5.5 ppt at Narrow River marsh (NAR), located near the mouth of the Bay, and 17.7 ± 5.3 ppt at Passeonkquis marsh (PAS) located in the upper Bay. Salinity varied seasonally with both marshes, showing the lowest values (16.3 and 8.3 ppt, respectively) in April and highest values (35.4 and 26.2 ppt, respectively) in August. Contour plots of calculated salinities showed that while the mean whole-marsh calculated salinity at both sites changed over time, within-marsh patterns of higher versus lower salinity were maintained at NAR but changed over time at PAS. Calculated salinity was significantly negatively correlated with elevation at NAR during a sub-set of 12 sample events, but not at PAS. Best-supported linear regression models for both sites included one-month and 6-month cumulative rainfall, and tide state as potential factors driving observed changes in calculated salinity. Mapping apparent conductivity of salt marsh sediments may be useful both identifying within-marsh micro-habitats, and documenting marsh-wide changes in salinity over time.

Exposing chicks to one hour of light during the night disrupts the release of a hormone that is needed by cells in the developing brain to survive.

This 8-page document discusses common biological, chemical, and physical hazards that may need preventive measures during the manufacture of swine feeds. Written by Taylor Langford, Morgan McKinney, Chad Carr, and Jason M. Scheffler, and published by the UF/IFAS Department of Animal Sciences, September 2019. https://edis.ifas.ufl.edu/an357

Variety choice is a critical management decision for peanut production. There are several good peanut varieties to choose from today. This document is SS-AGR-13, one of a series of the Agronomy Department, UF/IFAS Extension. Originally published as Marianna NFREC Research Report 06-1. Publication date January 2006.

Variety choice is a critical management decision for peanut production. There are several good peanut varieties to choose from today. This document is SS-AGR-13, one of a series of the Agronomy Department, UF/IFAS Extension. Originally published as Marianna NFREC Research Report 06-1. Publication date January 2006.

Variety choice is a critical management decision for peanut production. There are several good peanut varieties to choose from today. This document is SS-AGR-13, one of a series of the Agronomy Department, UF/IFAS Extension. Originally published as Marianna NFREC Research Report 06-1. Publication date January 2006.

Mutation signatures in cancer genomes reflect endogenous and exogenous mutational processes, offering insights into tumour etiology, features for prognostic and biologic stratification and vulnerabilities to be exploited therapeutically. We present a novel machine learning formalism for improved signature inference, based on multi-modal correlated topic models (MMCTM) which can at once infer signatures from both single nucleotide and structural variation counts derived from cancer genome sequencing data. We exemplify the utility of our approach on two hormone driven, DNA repair deficient cancers: breast and ovary (n = 755 samples total). We show how introducing correlated structure both within and between modes of mutation can increase accuracy of signature discovery, particularly in the context of sparse data. Our study emphasizes the importance of integrating multiple mutation modes for signature discovery and patient stratification, and provides a statistical modeling framework to incorporate additional features of interest for future studies.

Animal disease traceability has become an increasingly important topic within animal agriculture. Florida's breeders are encouraged to apply premise ID before marketing pigs to youth who will exhibit in terminal shows to assist with traceability. If pigs do not already have a premise ID tag, personnel from that terminal show should apply a plastic premise ID tag associated with that particular fair.

Animal disease traceability has become an increasingly important topic within animal agriculture. Florida's breeders are encouraged to apply premise ID before marketing pigs to youth who will exhibit in terminal shows to assist with traceability. If pigs do not already have a premise ID tag, personnel from that terminal show should apply a plastic premise ID tag associated with that particular fair.

Animal disease traceability has become an increasingly important topic within animal agriculture. Florida's breeders are encouraged to apply premise ID before marketing pigs to youth who will exhibit in terminal shows to assist with traceability. If pigs do not already have a premise ID tag, personnel from that terminal show should apply a plastic premise ID tag associated with that particular fair.

Summary: Dynamic trafficking of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid glutamate receptors (AMPARs) to synapses is critical for activity-dependent synaptic plasticity underlying learning and memory, but the identity of key molecular effectors remains elusive. Here, we demonstrate that membrane depolarization and N-methyl-D-aspartate receptor (NMDAR) activation triggers secretion of the chemotropic guidance cue netrin-1 from dendrites. Using selective genetic deletion, we show that netrin-1 expression by excitatory neurons is required for NMDAR-dependent long-term potentiation (LTP) in the adult hippocampus. Furthermore, we demonstrate that application of exogenous netrin-1 is sufficient to trigger the potentiation of excitatory glutamatergic transmission at hippocampal Schaffer collateral synapses via Ca2+-dependent recruitment of GluA1-containing AMPARs, promoting the maturation of immature or nascent synapses. These findings identify a central role for activity-dependent release of netrin-1 as a critical effector of synaptic plasticity in the adult hippocampus. : Glasgow et al. demonstrate depolarization- and NMDAR-dependent secretion of netrin-1 from the dendrites of hippocampal neurons. They report that genetic deletion of netrin-1 from excitatory neurons impairs hippocampal synaptic plasticity and that netrin-1 application is sufficient to trigger the potentiation of excitatory synaptic transmission via the insertion of GluA1 AMPARs. Keywords: CA1 pyramidal neurons, long-term potentiation, axon guidance, AMPA receptor trafficking, GluR1, deleted-in-colorectal cancer, DCC

In most well-studied rod-shaped bacteria, peptidoglycan is primarily crosslinked by penicillin-binding proteins (PBPs). However, in mycobacteria, crosslinks formed by L,D-transpeptidases (LDTs) are highly abundant. To elucidate the role of these unusual crosslinks, we characterized Mycobacterium smegmatis cells lacking all LDTs. We find that crosslinks generate by LDTs are required for rod shape maintenance specifically at sites of aging cell wall, a byproduct of polar elongation. Asymmetric polar growth leads to a non-uniform distribution of these two types of crosslinks in a single cell. Consequently, in the absence of LDT-mediated crosslinks, PBP-catalyzed crosslinks become more important. Because of this, Mycobacterium tuberculosis (Mtb) is more rapidly killed using a combination of drugs capable of PBP- and LDT- inhibition. Thus, knowledge about the spatial and genetic relationship between drug targets can be exploited to more effectively treat this pathogen.

Southern New England salt marsh vegetation and habitats are changing rapidly in response to sea-level rise. At the same time, fiddler crab (Uca spp.) distributions have expanded and purple marsh crab (Sesarma reticulatum) grazing on creekbank vegetation has increased. Sea-level rise and reduced predation pressure drive these changing crab populations but most studies focus on one species; there is a need for community-level assessments of impacts from multiple crab species. There is also a need to identify additional factors that can affect crab populations. We sampled crabs and environmental parameters in four Rhode Island salt marshes in 2014 and compiled existing data to quantify trends in crab abundance and multiple factors that potentially affect crabs. Crab communities were dominated by fiddler and green crabs (Carcinus maenas); S. reticulatum was much less abundant. Burrow sizes suggest that Uca is responsible for most burrows. On the marsh platform, burrows and Carcinus abundance were negatively correlated with elevation, soil moisture, and soil percent organic matter and positively correlated with soil bulk density. Uca abundance was negatively correlated with Spartina patens cover and height and positively correlated with Spartina alterniflora cover and soil shear strength. Creekbank burrow density increased dramatically between 1998 and 2016. During the same time, fishing effort and the abundance of birds that prey on crabs decreased, and water levels increased. Unlike in other southern New England marshes where recreational overfishing is hypothesized to drive increasing marsh crab abundance, we propose that changes in crab abundance were likely unrelated to recreational finfish over-harvest; instead, they better track sea-level rise and changing abundances of alternate predators, such as birds. We predict that marsh crab abundance will continue to expand with ongoing sea-level rise, at least until inundation thresholds for crab survival are exceeded.

Neural crest cells migrate throughout the embryo, but how cells move in a directed and collective manner has remained unclear. Here, we perform the first single-cell transcriptome analysis of cranial neural crest cell migration at three progressive stages in chick and identify and establish hierarchical relationships between cell position and time-specific transcriptional signatures. We determine a novel transcriptional signature of the most invasive neural crest Trailblazer cells that is consistent during migration and enriched for approximately 900 genes. Knockdown of several Trailblazer genes shows significant but modest changes to total distance migrated. However, in vivo expression analysis by RNAscope and immunohistochemistry reveals some salt and pepper patterns that include strong individual Trailblazer gene expression in cells within other subregions of the migratory stream. These data provide new insights into the molecular diversity and dynamics within a neural crest cell migratory stream that underlie complex directed and collective cell behaviors.

Planarian neoblasts are pluripotent, adult somatic stem cells and lineage-primed progenitors that are required for the production and maintenance of all differentiated cell types, including the germline. Neoblasts, originally defined as undifferentiated cells residing in the adult parenchyma, are frequently compared to embryonic stem cells yet their developmental origin remains obscure. We investigated the provenance of neoblasts during Schmidtea mediterranea embryogenesis, and report that neoblasts arise from an anarchic, cycling piwi-1+ population wholly responsible for production of all temporary and definitive organs during embryogenesis. Early embryonic piwi-1+ cells are molecularly and functionally distinct from neoblasts: they express unique cohorts of early embryo enriched transcripts and behave differently than neoblasts in cell transplantation assays. Neoblast lineages arise as organogenesis begins and are required for construction of all major organ systems during embryogenesis. These subpopulations are continuously generated during adulthood, where they act as agents of tissue homeostasis and regeneration.

Unlike many viruses that suppress cellular protein synthesis, host mRNA translation and polyribosome formation are stimulated by human cytomegalovirus (HCMV). How HCMV impacts the translationally regulated cellular mRNA repertoire and its contribution to virus biology remains unknown. Using polysome profiling, we show that HCMV presides over the cellular translational landscape, selectively accessing the host genome to extend its own coding capacity and regulate virus replication. Expression of the HCMV UL38 mTORC1-activator partially recapitulates these translational alterations in uninfected cells. The signature of cellular mRNAs translationally stimulated by HCMV resembles pathophysiological states (such as cancer) where translation initiation factor levels or activity increase. In contrast, cellular mRNAs repressed by HCMV include those involved in differentiation and the immune response. Surprisingly, interfering with the virus-induced activation of cellular mRNA translation can either limit or enhance HCMV growth. The unanticipated extent to which HCMV specifically manipulates host mRNA translation may aid in understanding its association with complex inflammatory disorders and cancer.

Vaccination is an effective prevention of influenza infection. However, certain individuals develop a lower antibody response after vaccination, which may lead to susceptibility to subsequent infection. An important challenge in human health is to find baseline gene signatures to help identify individuals who are at higher risk for infection despite influenza vaccination. We developed a multi-level machine learning strategy to build a predictive model of vaccine response using pre−vaccination antibody titers and network interactions between pre−vaccination gene expression levels. The first-level baseline−antibody model explains a significant amount of variation in post-vaccination response, especially for subjects with large pre−existing antibody titers. In the second level, we clustered individuals based on pre−vaccination antibody titers to focus gene−based modeling on individuals with lower baseline HAI where additional response variation may be predicted by baseline gene expression levels. In the third level, we used a gene−association interaction network (GAIN) feature selection algorithm to find the best pairs of genes that interact to influence antibody response within each baseline titer cluster. We used ratios of the top interacting genes as predictors to stabilize machine learning model generalizability. We trained and tested the multi-level approach on data with young and older individuals immunized against influenza vaccine in multiple cohorts. Our results indicate that the GAIN feature selection approach improves model generalizability and identifies genes enriched for immunologically relevant pathways, including B Cell Receptor signaling and antigen processing. Using a multi-level approach, starting with a baseline HAI model and stratifying on baseline HAI, allows for more targeted gene−based modeling. We provide an interactive tool that may be extended to other vaccine studies.

We performed a quantitative analysis of the HLA restriction, antigen and epitope specificity of human pathogen specific responses in healthy individuals infected with M. tuberculosis (Mtb), in a South African cohort as a test case. The results estimate the breadth of T cell responses for the first time in the context of an infection and human population setting. We determined the epitope repertoire of eleven representative Mtb antigens and a large panel of previously defined Mtb epitopes. We estimated that our analytic methods detected 50-75% of the total response in a cohort of 63 individuals. As expected, responses were highly heterogeneous, with responses to a total of 125 epitopes detected. The 66 top epitopes provided 80% coverage of the responses identified in our study. Using a panel of 48 HLA class II-transfected antigen-presenting cells, we determined HLA class II restrictions for 278 epitope/donor recognition events (36% of the total). The majority of epitopes were restricted by multiple HLA alleles, and 380 different epitope/HLA combinations comprised less than 30% of the estimated Mtb-specific response. Our results underline the complexity of human T cell responses at a population level. Efforts to capture and characterize this broad and highly HLA promiscuous Mtb-specific T cell epitope repertoire will require significant peptide multiplexing efforts. We show that a comprehensive "megapool" of Mtb peptides captured a large fraction of the Mtb-specific T cells and can be used to characterize this response.

The interrelationship between endogenous microbiota, the immune system, and tissue regeneration is an area of intense research due to its potential therapeutic applications. We investigated this relationship in Schmidtea mediterranea, a model organism capable of regenerating any and all of its adult tissues. Microbiome characterization revealed a high Bacteroidetes to Proteobacteria ratio in healthy animals. Perturbations eliciting an expansion of Proteobacteria coincided with ectopic lesions and tissue degeneration. The culture of these bacteria yielded a strain of Pseudomonas capable of inducing progressive tissue degeneration. RNAi screening uncovered a TAK1 innate immune signaling module underlying compromised tissue homeostasis and regeneration during infection. TAK1/MKK/p38 signaling mediated opposing regulation of apoptosis during infection versus normal tissue regeneration. Given the complex role of inflammation in either hindering or supporting reparative wound healing and regeneration, this invertebrate model provides a basis for dissecting the duality of evolutionarily conserved inflammatory signaling in complex, multi-organ adult tissue regeneration.

Neoblasts are an abundant, heterogeneous population of adult stem cells (ASCs) that facilitate the maintenance of planarian tissues and organs, providing a powerful system to study ASC self-renewal and differentiation dynamics. It is unknown how the collective output of neoblasts transit through differentiation pathways to produce specific cell types. The planarian epidermis is a simple tissue that undergoes rapid turnover. We found that as epidermal progeny differentiate, they progress through multiple spatiotemporal transition states with distinct gene expression profiles. We also identified a conserved early growth response family transcription factor, egr-5, that is essential for epidermal differentiation. Disruption of epidermal integrity by egr-5 RNAi triggers a global stress response that induces the proliferation of neoblasts and the concomitant expansion of not only epidermal, but also multiple progenitor cell populations. Our results further establish the planarian epidermis as a novel paradigm to uncover the molecular mechanisms regulating ASC specification in vivo.

Cystic kidney diseases (CKDs) affect millions of people worldwide. The defining pathological features are fluid-filled cysts developing from nephric tubules due to defective flow sensing, cell proliferation and differentiation. The underlying molecular mechanisms, however, remain poorly understood, and the derived excretory systems of established invertebrate models (Caenorhabditis elegans and Drosophila melanogaster) are unsuitable to model CKDs. Systematic structure/function comparisons revealed that the combination of ultrafiltration and flow-associated filtrate modification that is central to CKD etiology is remarkably conserved between the planarian excretory system and the vertebrate nephron. Consistently, both RNA-mediated genetic interference (RNAi) of planarian orthologues of human CKD genes and inhibition of tubule flow led to tubular cystogenesis that share many features with vertebrate CKDs, suggesting deep mechanistic conservation. Our results demonstrate a common evolutionary origin of animal excretory systems and establish planarians as a novel and experimentally accessible invertebrate model for the study of human kidney pathologies.

Variety choice is a critical management decision in producing a peanut crop. Since several good peanut varieties are available, it is essential to know each variety’s attributes and how different varieties might fit into a farm plan. This 7-page fact sheet provides data conducted from trials in Florida at UF/IFAS research centers located in Gainesville (Citra), Marianna, and Jay from 2010–2013. Written by Barry Tillman, Mark Gomillion, Justin McKinney, and George Person, and published by the UF Department of Agronomy, May 2014.

Variety choice is a critical management decision in producing a peanut crop. Since several good peanut varieties are available, it is essential to know each variety’s attributes and how different varieties might fit into a farm plan. This 7-page fact sheet provides data conducted from trials in Florida at UF/IFAS research centers located in Gainesville (Citra), Marianna, and Jay from 2010–2013. Written by Barry Tillman, Mark Gomillion, Justin McKinney, and George Person, and published by the UF Department of Agronomy, May 2014.

Variety choice is a critical management decision in producing a peanut crop. Since several good peanut varieties are available, it is essential to know each variety’s attributes and how different varieties might fit into a farm plan. This 7-page fact sheet provides data conducted from trials in Florida at UF/IFAS research centers located in Gainesville (Citra), Marianna, and Jay from 2010–2013. Written by Barry Tillman, Mark Gomillion, Justin McKinney, and George Person, and published by the UF Department of Agronomy, May 2014.