Characterization of Putative Iron Responsive Genes as Species-Specific Indicators of Iron Stress in Thalassiosiroid Diatoms

Iron (Fe) availability restricts diatom growth and primary production in large areas of the oceans. It is a challenge to assess the bulk Fe nutritional health of natural diatom populations, since species can differ in their physiological and molecular responses to Fe limitation. We assayed expression of selected genes in diatoms from the Thalassiosira genus to assess their potential utility as species-specific molecular markers to indicate Fe status in natural diatom assemblages. In this study, we compared the expression of the photosynthetic genes encoding ferredoxin (a Fe-requiring protein) and flavodoxin (a Fe-free protein) in culture experiments with Fe replete and Fe stressed Thalassiosira pseudonana (CCMP 1335) isolated from coastal waters and Thalassiosira weissflogii (CCMP 1010) isolated from the open ocean. In T. pseudonana, expression of flavodoxin and ferredoxin genes were not sensitive to Fe status but were found to display diel periodicities. In T. weissflogii, expression of flavodoxin was highly responsive to iron levels and was only detectable when cultures were Fe limited. Flavodoxin genes have been duplicated in most diatoms with available genome data and we show that T. pseudonana has lost its copy related to the Fe-responsive copy in T. weissflogii. We also examined the expression of genes for a putative high affinity, copper (Cu)-dependent Fe uptake system in T. pseudonana. Our results indicate that genes encoding putative Cu transporters, a multi-Cu oxidase, and a Fe reductase are not linked to Fe status. The expression of a second putative Fe reductase increased in Fe limited cultures, but this gene was also highly expressed in Fe replete cultures, indicating it may not be a useful marker in the field. Our findings highlight that Fe metabolism may differ among diatoms even within a genus and show a need to validate responses in different species as part of the development pipeline for genetic markers of Fe status in field populations

Single copy/knock-in models of ALS SOD1 in C. elegans suggest loss and gain of function have different contributions to cholinergic and glutamatergic neurodegeneration

Mutations in Cu/Zn superoxide dismutase 1 (SOD1) lead to Amyotrophic Lateral Sclerosis (ALS), a neurodegenerative disease that disproportionately affects glutamatergic and cholinergic motor neurons. Previous work with SOD1 overexpression models supports a role for SOD1 toxic gain of function in ALS pathogenesis. However, the impact of SOD1 loss of function in ALS cannot be directly examined in overexpression models. In addition, overexpression may obscure the contribution of SOD1 loss of function in the degeneration of different neuronal populations. Here, we report the first single-copy, ALS knock-in models in C. elegans generated by transposon- or CRISPR/Cas9- mediated genome editing of the endogenous sod-1 gene. Introduction of ALS patient amino acid changes A4V, H71Y, L84V, G85R or G93A into the C. elegans sod-1 gene yielded single-copy/knock-in ALS SOD1 models. These differ from previously reported overexpression models in multiple assays. In single-copy/knock-in models, we observed differential impact of sod-1 ALS alleles on glutamatergic and cholinergic neurodegeneration. A4V, H71Y, G85R, and G93A animals showed increased SOD1 protein accumulation and oxidative stress induced degeneration, consistent with a toxic gain of function in cholinergic motor neurons. By contrast, H71Y, L84V, and G85R lead to glutamatergic neuron degeneration due to sod-1 loss of function after oxidative stress. However, dopaminergic and serotonergic neuronal populations were spared in single-copy ALS models, suggesting a neuronal-subtype specificity previously not reported in invertebrate ALS SOD1 models. Combined, these results suggest that knock-in models may reproduce the neurotransmitter-type specificity of ALS and that both SOD1 loss and gain of toxic function differentially contribute to ALS pathogenesis in different neuronal populations.Peer reviewe

l-Fucose Stimulates Utilization of d-Ribose by \u3cem\u3eEscherichia coli\u3c/em\u3e MG1655 ΔfucAO and \u3cem\u3eE. coli\u3c/em\u3e Nissle 1917 ΔfucAO Mutants in the Mouse Intestine and in M9 Minimal Medium

Escherichia coli MG1655 uses several sugars for growth in the mouse intestine. To determine the roles of l-fucose and d-ribose, an E. coli MG1655 ΔfucAO mutant and an E. coli MG1655 ΔrbsK mutant were fed separately to mice along with wild-type E. coli MG1655. The E. coli MG1655 ΔfucAO mutant colonized the intestine at a level 2 orders of magnitude lower than that of the wild type, but the E. coli MG1655 ΔrbsK mutant and the wild type colonized at nearly identical levels. Surprisingly, an E. coli MG1655 ΔfucAO ΔrbsK mutant was eliminated from the intestine by either wild-type E. coli MG1655 or E. coli MG1655 ΔfucAO, suggesting that the ΔfucAO mutant switches to ribose in vivo. Indeed, in vitro growth experiments showed that l-fucose stimulated utilization of d-ribose by the E. coli MG1655 ΔfucAO mutant but not by an E. coli MG1655 ΔfucK mutant. Since the ΔfucK mutant cannot convert l-fuculose to l-fuculose-1-phosphate, whereas the ΔfucAO mutant accumulates l-fuculose-1-phosphate, the data suggest that l-fuculose-1-phosphate stimulates growth on ribose both in the intestine and in vitro. An E. coli Nissle 1917 ΔfucAO mutant, derived from a human probiotic commensal strain, acted in a manner identical to that of E. coli MG1655 ΔfucAO in vivo and in vitro. Furthermore, l-fucose at a concentration too low to support growth stimulated the utilization of ribose by the wild-type E. coli strains in vitro. Collectively, the data suggest that l-fuculose-1-phosphate plays a role in the regulation of ribose usage as a carbon source by E. coli MG1655 and E. coli Nissle 1917 in the mouse intestine

Mouse Intestine Selects Nonmotile \u3cem\u3eflhDC\u3c/em\u3e Mutants of \u3cem\u3eEscherichia coli\u3c/em\u3e MG1655 with Increased Colonizing Ability and Better Utilization of Carbon Sources

d-Gluconate which is primarily catabolized via the Entner-Doudoroff (ED) pathway, has been implicated as being important for colonization of the streptomycin-treated mouse large intestine by Escherichia coli MG1655, a human commensal strain. In the present study, we report that an MG1655 Δedd mutant defective in the ED pathway grows poorly not only on gluconate as a sole carbon source but on a number of other sugars previously implicated as being important for colonization, including l-fucose, d-gluconate, d-glucuronate, N-acetyl-d-glucosamine, d-mannose, and d-ribose. Furthermore, we show that the mouse intestine selects mutants of MG1655 Δedd and wild-type MG1655 that have improved mouse intestine-colonizing ability and grow 15 to 30% faster on the aforementioned sugars. The mutants of MG1655 Δedd and wild-type MG1655 selected by the intestine are shown to be nonmotile and to have deletions in the flhDC operon, which encodes the master regulator of flagellar biosynthesis. Finally, we show that ΔflhDC mutants of wild-type MG1655 and MG1655 Δedd constructed in the laboratory act identically to those selected by the intestine; i.e., they grow better than their respective parents on sugars as sole carbon sources and are better colonizers of the mouse intestine

Respiration of \u3cem\u3eEscherichia coli\u3c/em\u3e in the Mouse Intestine

Mammals are aerobes that harbor an intestinal ecosystem dominated by large numbers of anaerobic microorganisms. However, the role of oxygen in the intestinal ecosystem is largely unexplored. We used systematic mutational analysis to determine the role of respiratory metabolism in the streptomycin-treated mouse model of intestinal colonization. Here we provide evidence that aerobic respiration is required for commensal and pathogenic Escherichia coli to colonize mice. Our results showed that mutants lacking ATP synthase, which is required for all respiratory energy-conserving metabolism, were eliminated by competition with respiratory-competent wild-type strains. Mutants lacking the high-affinity cytochrome bd oxidase, which is used when oxygen tensions are low, also failed to colonize. However, the low-affinity cytochrome bo3 oxidase, which is used when oxygen tension is high, was found not to be necessary for colonization. Mutants lacking either nitrate reductase or fumarate reductase also had major colonization defects. The results showed that the entire E. coli population was dependent on both microaerobic and anaerobic respiration, consistent with the hypothesis that the E. coli niche is alternately microaerobic and anaerobic, rather than static. The results indicate that success of the facultative anaerobes in the intestine depends on their respiratory flexibility. Despite competition for relatively scarce carbon sources, the energy efficiency provided by respiration may contribute to the widespread distribution (i.e., success) of E. coli strains as commensal inhabitants of the mammalian intestine

Comparison of Carbon Nutrition for Pathogenic and Commensal ,\u3cem\u3eEscherichia coli\u3c/em\u3e Strains in the Mouse Intestine

The carbon sources that support the growth of pathogenic Escherichia coli O157:H7 in the mammalian intestine have not previously been investigated. In vivo, the pathogenic E. coli EDL933 grows primarily as single cells dispersed within the mucus layer that overlies the mouse cecal epithelium. We therefore compared the pathogenic strain and the commensal E. coli strain MG1655 modes of metabolism in vitro, using a mixture of the sugars known to be present in cecal mucus, and found that the two strains used the 13 sugars in a similar order and cometabolized as many as 9 sugars at a time. We conducted systematic mutation analyses of E. coli EDL933 and E. coli MG1655 by using lesions in the pathways used for catabolism of 13 mucus-derived sugars and five other compounds for which the corresponding bacterial gene system was induced in the transcriptome of cells grown on cecal mucus. Each of 18 catabolic mutants in both bacterial genetic backgrounds was fed to streptomycin-treated mice, together with the respective wild-type parent strain, and their colonization was monitored by fecal plate counts. None of the mutations corresponding to the five compounds not found in mucosal polysaccharides resulted in colonization defects. Based on the mutations that caused colonization defects, we determined that both E. coli EDL933 and E. coli MG1655 used arabinose, fucose, and N-acetylglucosamine in the intestine. In addition, E. coli EDL933 used galactose, hexuronates, mannose, and ribose, whereas E. coli MG1655 used gluconate and N-acetylneuraminic acid. The colonization defects of six catabolic lesions were found to be additive with E. coli EDL933 but not with E. coli MG1655. The data indicate that pathogenic E. coli EDL933 uses sugars that are not used by commensal E. coli MG1655 to colonize the mouse intestine. The results suggest a strategy whereby invading pathogens gain advantage by simultaneously consuming several sugars that may be available because they are not consumed by the commensal intestinal microbiota

Many Labs 5:Testing pre-data collection peer review as an intervention to increase replicability

Replication studies in psychological science sometimes fail to reproduce prior findings. If these studies use methods that are unfaithful to the original study or ineffective in eliciting the phenomenon of interest, then a failure to replicate may be a failure of the protocol rather than a challenge to the original finding. Formal pre-data-collection peer review by experts may address shortcomings and increase replicability rates. We selected 10 replication studies from the Reproducibility Project: Psychology (RP:P; Open Science Collaboration, 2015) for which the original authors had expressed concerns about the replication designs before data collection; only one of these studies had yielded a statistically significant effect (p < .05). Commenters suggested that lack of adherence to expert review and low-powered tests were the reasons that most of these RP:P studies failed to replicate the original effects. We revised the replication protocols and received formal peer review prior to conducting new replication studies. We administered the RP:P and revised protocols in multiple laboratories (median number of laboratories per original study = 6.5, range = 3?9; median total sample = 1,279.5, range = 276?3,512) for high-powered tests of each original finding with both protocols. Overall, following the preregistered analysis plan, we found that the revised protocols produced effect sizes similar to those of the RP:P protocols (?r = .002 or .014, depending on analytic approach). The median effect size for the revised protocols (r = .05) was similar to that of the RP:P protocols (r = .04) and the original RP:P replications (r = .11), and smaller than that of the original studies (r = .37). Analysis of the cumulative evidence across the original studies and the corresponding three replication attempts provided very precise estimates of the 10 tested effects and indicated that their effect sizes (median r = .07, range = .00?.15) were 78% smaller, on average, than the original effect sizes (median r = .37, range = .19?.50)

The psychological science accelerator’s COVID-19 rapid-response dataset

In response to the COVID-19 pandemic, the Psychological Science Accelerator coordinated three large-scale psychological studies to examine the effects of loss-gain framing, cognitive reappraisals, and autonomy framing manipulations on behavioral intentions and affective measures. The data collected (April to October 2020) included specific measures for each experimental study, a general questionnaire examining health prevention behaviors and COVID-19 experience, geographical and cultural context characterization, and demographic information for each participant. Each participant started the study with the same general questions and then was randomized to complete either one longer experiment or two shorter experiments. Data were provided by 73,223 participants with varying completion rates. Participants completed the survey from 111 geopolitical regions in 44 unique languages/dialects. The anonymized dataset described here is provided in both raw and processed formats to facilitate re-use and further analyses. The dataset offers secondary analytic opportunities to explore coping, framing, and self-determination across a diverse, global sample obtained at the onset of the COVID-19 pandemic, which can be merged with other time-sampled or geographic data

A global experiment on motivating social distancing during the COVID-19 pandemic

Finding communication strategies that effectively motivate social distancing continues to be a global public health priority during the COVID-19 pandemic. This cross-country, preregistered experiment (n = 25,718 from 89 countries) tested hypotheses concerning generalizable positive and negative outcomes of social distancing messages that promoted personal agency and reflective choices (i.e., an autonomy-supportive message) or were restrictive and shaming (i.e., a controlling message) compared with no message at all. Results partially supported experimental hypotheses in that the controlling message increased controlled motivation (a poorly internalized form of motivation relying on shame, guilt, and fear of social consequences) relative to no message. On the other hand, the autonomy-supportive message lowered feelings of defiance compared with the controlling message, but the controlling message did not differ from receiving no message at all. Unexpectedly, messages did not influence autonomous motivation (a highly internalized form of motivation relying on one’s core values) or behavioral intentions. Results supported hypothesized associations between people’s existing autonomous and controlled motivations and self-reported behavioral intentions to engage in social distancing. Controlled motivation was associated with more defiance and less long-term behavioral intention to engage in social distancing, whereas autonomous motivation was associated with less defiance and more short- and long-term intentions to social distance. Overall, this work highlights the potential harm of using shaming and pressuring language in public health communication, with implications for the current and future global health challenges