Outcome Orientation: A Misconception of Probability That Harms Medical Research and Practice.

Human understanding of randomness and variation is shaped by a number of cognitive biases. In this article, we relate a lesser-known cognitive bias, the "outcome orientation," to medical questions and describe the harm it can do to medical research and practice. An outcome orientation means predicting outcomes one at a time, neglecting the fact that each event may be a member of a group of comparable events. People who reason according to an outcome orientation assign a subjective degree of belief to an outcome, but do so in a way that is incompatible with Bayesian reasoning or any other standard laws of probability. Instead of accepting that uncertainty is inevitable and generalizing from the frequency of similar events, the outcome orientation prefers one-off causal narratives. In medicine, the outcome orientation therefore erodes support for randomized controlled trials in favor of reductionist approaches. The rhetoric of personalized medicine resonates with, and can promote, the outcome orientation by emphasizing how the measurable attributes of individual patients, rather than chance or unknowable factors, causally produce each particular patient's outcome.

Microscale sulfur cycling in the phototrophic pink berry consortia of the Sippewissett Salt Marsh.

Microbial metabolism is the engine that drives global biogeochemical cycles, yet many key transformations are carried out by microbial consortia over short spatiotemporal scales that elude detection by traditional analytical approaches. We investigate syntrophic sulfur cycling in the 'pink berry' consortia of the Sippewissett Salt Marsh through an integrative study at the microbial scale. The pink berries are macroscopic, photosynthetic microbial aggregates composed primarily of two closely associated species: sulfide-oxidizing purple sulfur bacteria (PB-PSB1) and sulfate-reducing bacteria (PB-SRB1). Using metagenomic sequencing and (34) S-enriched sulfate stable isotope probing coupled with nanoSIMS, we demonstrate interspecies transfer of reduced sulfur metabolites from PB-SRB1 to PB-PSB1. The pink berries catalyse net sulfide oxidation and maintain internal sulfide concentrations of 0-500 µm. Sulfide within the berries, captured on silver wires and analysed using secondary ion mass spectrometer, increased in abundance towards the berry interior, while δ(34) S-sulfide decreased from 6‰ to -31‰ from the exterior to interior of the berry. These values correspond to sulfate-sulfide isotopic fractionations (15-53‰) consistent with either sulfate reduction or a mixture of reductive and oxidative metabolisms. Together this combined metagenomic and high-resolution isotopic analysis demonstrates active sulfur cycling at the microscale within well-structured macroscopic consortia consisting of sulfide-oxidizing anoxygenic phototrophs and sulfate-reducing bacteria.

Diversity and abundance of phyllosphere bacteria are linked to insect herbivory.

Simultaneous or sequential attack by herbivores and microbes is common in plants. Many seed plants exhibit a defence trade-off against chewing herbivorous insects and leaf-colonizing ('phyllosphere') bacteria, which arises from cross-talk between the phytohormones jasmonic acid (JA, induced by many herbivores) and salicylic acid (SA, induced by many bacteria). This cross-talk may promote reciprocal susceptibility in plants between phyllosphere bacteria and insect herbivores. In a population of native bittercress (Cardamine cordifolia, Brassicaceae), we tested whether simulating prior damage with JA or SA treatment induced resistance or susceptibility (respectively) to chewing herbivores. In parallel, we conducted culture-dependent surveys of phyllosphere bacteria to test the hypothesis that damage by chewing herbivores correlates positively with bacterial abundance in leaves. Finally, we tested whether bacterial infection induced susceptibility to herbivory by a major chewing herbivore of bittercress, Scaptomyza nigrita (Drosophilidae). Overall, our results suggest that reciprocal susceptibility to herbivory and microbial attack occurs in bittercress. We found that JA treatment reduced and SA treatment increased S. nigrita herbivory in bittercress in the field. Bacterial abundance was higher in herbivore-damaged vs. undamaged leaves (especially Pseudomonas syringae). However, Pedobacter spp. and Pseudomonas fluorescens infections were negatively associated with herbivory. Experimental Pseudomonas spp. infections increased S. nigrita herbivory in bittercress. Thus, plant defence signalling trade-offs can have important ecological consequences in nature that may be reflected in a positive correlation between herbivory and phyllosphere bacterial abundance and diversity. Importantly, the strength and direction of this association varies within and among prevalent bacterial groups.

OspC is potent plasminogen receptor on surface of Borrelia burgdorferi.

Host-derived proteases are crucial for the successful infection of vertebrates by several pathogens, including the Lyme disease spirochete bacterium, Borrelia burgdorferi. B. burgdorferi must traverse tissue barriers in the tick vector during transmission to the host and during dissemination within the host, and it must disrupt immune challenges to successfully complete its infectious cycle. It has been proposed that B. burgdorferi can accomplish these tasks without an endogenous extra-cytoplasmic protease by commandeering plasminogen, the highly abundant precursor of the vertebrate protease plasmin. However, the molecular mechanism by which B. burgdorferi immobilizes plasminogen to its surface remains obscure. The data presented here demonstrate that the outer surface protein C (OspC) of B. burgdorferi is a potent plasminogen receptor on the outer membrane of the bacterium. OspC-expressing spirochetes readily bind plasminogen, whereas only background levels of plasminogen are detectable on OspC-deficient strains. Furthermore, plasminogen binding by OspC-expressing spirochetes can be significantly reduced using anti-OspC antibodies. Co-immunofluorescence staining assays demonstrate that wild-type bacteria immobilize plasminogen only if they are actively expressing OspC regardless of the expression of other surface proteins. The co-localization of plasminogen and OspC on OspC-expressing spirochetes further implicates OspC as a biologically relevant plasminogen receptor on the surface of live B. burgdorferi.

Evolution of haploid and diploid populations reveals common, strong, and variable pleiotropic effects in non-home environments.

Adaptation is driven by the selection for beneficial mutations that provide a fitness advantage in the specific environment in which a population is evolving. However, environments are rarely constant or predictable. When an organism well adapted to one environment finds itself in another, pleiotropic effects of mutations that made it well adapted to its former environment will affect its success. To better understand such pleiotropic effects, we evolved both haploid and diploid barcoded budding yeast populations in multiple environments, isolated adaptive clones, and then determined the fitness effects of adaptive mutations in 'non-home' environments in which they were not selected. We find that pleiotropy is common, with most adaptive evolved lineages showing fitness effects in non-home environments. Consistent with other studies, we find that these pleiotropic effects are unpredictable: they are beneficial in some environments and deleterious in others. However, we do find that lineages with adaptive mutations in the same genes tend to show similar pleiotropic effects. We also find that ploidy influences the observed adaptive mutational spectra in a condition-specific fashion. In some conditions, haploids and diploids are selected with adaptive mutations in identical genes, while in others they accumulate mutations in almost completely disjoint sets of genes.

Phenotypic and molecular evolution across 10,000 generations in laboratory budding yeast populations.

Laboratory experimental evolution provides a window into the details of the evolutionary process. To investigate the consequences of long-term adaptation, we evolved 205 Saccharomyces cerevisiae populations (124 haploid and 81 diploid) for ~10,000,000 generations in three environments. We measured the dynamics of fitness changes over time, finding repeatable patterns of declining adaptability. Sequencing revealed that this phenotypic adaptation is coupled with a steady accumulation of mutations, widespread genetic parallelism, and historical contingency. In contrast to long-term evolution in E. coli, we do not observe long-term coexistence or populations with highly elevated mutation rates. We find that evolution in diploid populations involves both fixation of heterozygous mutations and frequent loss-of-heterozygosity events. Together, these results help distinguish aspects of evolutionary dynamics that are likely to be general features of adaptation across many systems from those that are specific to individual organisms and environmental conditions.

Insect herbivory reshapes a native leaf microbiome.

Insect herbivory is pervasive in plant communities, but its impact on microbial plant colonizers is not well-studied in natural systems. By calibrating sequencing-based bacterial detection to absolute bacterial load, we find that the within-host abundance of most leaf microbiome (phyllosphere) taxa colonizing a native forb is amplified within leaves affected by insect herbivory. Herbivore-associated bacterial amplification reflects community-wide compositional shifts towards lower ecological diversity, but the extent and direction of such compositional shifts can be interpreted only by quantifying absolute abundance. Experimentally eliciting anti-herbivore defences reshaped within-host fitness ranks among Pseudomonas spp. field isolates and amplified a subset of putatively phytopathogenic P. syringae in a manner causally consistent with observed field-scale patterns. Herbivore damage was inversely correlated with plant reproductive success and was highly clustered across plants, which predicts tight co-clustering with putative phytopathogens across hosts. Insect herbivory may thus drive the epidemiology of plant-infecting bacteria as well as the structure of a native plant microbiome by generating variation in within-host bacterial fitness at multiple phylogenetic and spatial scales. This study emphasizes that 'non-focal' biotic interactions between hosts and other organisms in their ecological settings can be crucial drivers of the population and community dynamics of host-associated microbiomes.

Complete Genome Sequences for Pseudomonas sp. Strains 29A and 43A.

Pseudomonas sp. strains 29A and 43A were originally isolated from the phyllosphere of individual plants of Cardamine cordifolia (Brassicaceae). Here, we report complete genome sequences for these two closely related strains, assembled using a hybrid approach combining Illumina paired-end reads and longer reads sequenced on an Oxford Nanopore MinION flow cell.

Habitat preference of an herbivore shapes the habitat distribution of its host plant.

Plant distributions can be limited by habitat-biased herbivory, but the proximate causes of such biases are rarely known. Distinguishing plant-centric from herbivore-centric mechanisms driving differential herbivory between habitats is difficult without experimental manipulation of both plants and herbivores. Here we tested alternative hypotheses driving habitat-biased herbivory in bittercress (Cardamine cordifolia), which is more abundant under shade of shrubs and trees (shade) than in nearby meadows (sun) where herbivory is intense from the specialist fly Scaptomyza nigrita. This system has served as a textbook example of habitat-biased herbivory driving a plant's distribution across an ecotone, but the proximate mechanisms underlying differential herbivory are still unclear. First, we found that higher S. nigrita herbivory in sun habitats contrasts sharply with their preference to attack plants from shade habitats in laboratory choice experiments. Second, S. nigrita strongly preferred leaves in simulated sun over simulated shade habitats, regardless of plant source habitat. Thus, herbivore preference for brighter, warmer habitats overrides their preference for more palatable shade plants. This promotes the sun-biased herbivore pressure that drives the distribution of bittercress into shade habitats.

Early onset of inflammation during ontogeny of bipolar disorder: the NLRP2 inflammasome gene distinctly differentiates between patients and healthy controls in the transition between iPS cell and neural stem cell stages.

Neuro-inflammation and neuronal communication are considered as mis-regulated processes in the aetiology and pathology of bipolar disorder (BD). Which and when specific signal pathways become abnormal during the ontogeny of bipolar disorder patients is unknown. To address this question, we applied induced pluripotent stem cell (iPSC) technology followed by cortical neural differentiation on adipocyte-derived cells from BD type I patients (with psychotic episodes in psychiatric history) and healthy volunteers (controls). RNA sequencing in iPSC and cortical neural stem cell (NSC) lines were used to examine alterations between the transcriptomes from BD I and control samples during transition from the pluripotent stage towards the neural developmental stage. At the iPSC stage, the most highly significant differentially expressed gene (DEG) was the NLRP2 inflammasome (P=2.66 × 10-10). Also among 42 DEGs at the NSC stage, NLRP2 showed the strongest statistical significance (P=3.07 × 10-19). In addition, we have also identified several cytoskeleton-associated genes as DEGs from the NSC stage, such as TMP2, TAGLN and ACTA2; the former two genes are recognised for the first time to be associated with BD. Our results also suggest that iPSC-derived BD-cortical NSCs carry several abnormalities in dopamine and GABA receptor canonical pathways, underlining that our in vitro BD model reflects pathology in the central nervous system. This would indicate that mis-regulated gene expression of inflammatory, neurotransmitter and cytoskeletal signalling occurs during early fetal brain development of BD I patients.

The Green Berry Consortia of the Sippewissett Salt Marsh: Millimeter-Sized Aggregates of Diazotrophic Unicellular Cyanobacteria.

Microbial interactions driving key biogeochemical fluxes often occur within multispecies consortia that form spatially heterogeneous microenvironments. Here, we describe the "green berry" consortia of the Sippewissett salt marsh (Falmouth, MA, United States): millimeter-sized aggregates dominated by an uncultured, diazotrophic unicellular cyanobacterium of the order Chroococcales (termed GB-CYN1). We show that GB-CYN1 is closely related to Crocosphaera watsonii (UCYN-B) and "Candidatus Atelocyanobacterium thalassa" (UCYN-A), two groups of unicellular diazotrophic cyanobacteria that play an important role in marine primary production. Other green berry consortium members include pennate diatoms and putative heterotrophic bacteria from the Alphaproteobacteria and Bacteroidetes. Tight coupling was observed between photosynthetic oxygen production and heterotrophic respiration. When illuminated, the green berries became supersaturated with oxygen. From the metagenome, we observed that GB-CYN1 encodes photosystem II genes and thus has the metabolic potential for oxygen production unlike UCYN-A. In darkness, respiratory activity rapidly depleted oxygen creating anoxia within the aggregates. Metagenomic data revealed a suite of nitrogen fixation genes encoded by GB-CYN1, and nitrogenase activity was confirmed at the whole-aggregate level by acetylene reduction assays. Metagenome reads homologous to marker genes for denitrification were observed and suggest that heterotrophic denitrifiers might co-occur in the green berries, although the physiology and activity of facultative anaerobes in these aggregates remains uncharacterized. Nitrogen fixation in the surface ocean was long thought to be driven by filamentous cyanobacterial aggregates, though recent work has demonstrated the importance of unicellular diazotrophic cyanobacteria (UCYN) from the order Chroococcales. The green berries serve as a useful contrast to studies of open ocean UCYN and may provide a tractable model system to investigate microbial dynamics within phytoplankton aggregates, a phenomenon of global importance to the flux of particulate organic carbon and nitrogen in surface waters.

Pseudomonas syringae enhances herbivory by suppressing the reactive oxygen burst in Arabidopsis.

Plant-herbivore interactions have evolved in the presence of plant-colonizing microbes. These microbes can have important third-party effects on herbivore ecology, as exemplified by drosophilid flies that evolved from ancestors feeding on plant-associated microbes. Leaf-mining flies in the genus Scaptomyza, which is nested within the paraphyletic genus Drosophila, show strong associations with bacteria in the genus Pseudomonas, including Pseudomonas syringae. Adult females are capable of vectoring these bacteria between plants and larvae show a preference for feeding on P. syringae-infected leaves. Here we show that Scaptomyza flava larvae can also vector P. syringae to and from feeding sites, and that they not only feed more, but also develop faster on plants previously infected with P. syringae. Our genetic and physiological data show that P. syringae enhances S. flava feeding on infected plants at least in part by suppressing anti-herbivore defenses mediated by reactive oxygen species.

Aversion and attraction to harmful plant secondary compounds jointly shape the foraging ecology of a specialist herbivore.

Most herbivorous insect species are restricted to a narrow taxonomic range of host plant species. Herbivore species that feed on mustard plants and their relatives in the Brassicales have evolved highly efficient detoxification mechanisms that actually prevent toxic mustard oils from forming in the bodies of the animals. However, these mechanisms likely were not present during the initial stages of specialization on mustard plants ~100 million years ago. The herbivorous fly Scaptomyza nigrita (Drosophilidae) is a specialist on a single mustard species, bittercress (Cardamine cordifolia; Brassicaceae) and is in a fly lineage that evolved to feed on mustards only in the past 10-20 million years. In contrast to many mustard specialists, S. nigrita does not prevent formation of toxic breakdown products (mustard oils) arising from glucosinolates (GLS), the primary defensive compounds in mustard plants. Therefore, it is an appealing model for dissecting the early stages of host specialization. Because mustard oils actually form in the bodies of S. nigrita, we hypothesized that in lieu of a specialized detoxification mechanism, S. nigrita may mitigate exposure to high GLS levels within plant tissues using behavioral avoidance. Here, we report that jasmonic acid (JA) treatment increased GLS biosynthesis in bittercress, repelled adult female flies, and reduced larval growth. S. nigrita larval damage also induced foliar GLS, especially in apical leaves, which correspondingly displayed the least S. nigrita damage in controlled feeding trials and field surveys. Paradoxically, flies preferred to feed and oviposit on GLS-producing Arabidopsis thaliana despite larvae performing worse in these plants versus non-GLS-producing mutants. GLS may be feeding cues for S. nigrita despite their deterrent and defensive properties, which underscores the diverse relationship a mustard specialist has with its host when lacking a specialized means of mustard oil detoxification.

The von Hippel-Lindau tumor suppressor gene protects cells from UV-mediated apoptosis.

The familial cancer syndrome, von Hippel-Lindau (VHL) disease, characterized by a predisposition to renal cell carcinoma and certain other tumor types, is caused by mutational inactivation of the VHL tumor suppressor gene. Loss of VHL gene function is detected also in the vast majority of sporadic renal cell carcinomas. Previous reports have determined a protective role for VHL in response to serum withdrawal and glucose deprivation. In this study, the effect of UV irradiation on VHL-negative and VHL-positive renal carcinoma cells was examined. VHL-negative 786-O renal carcinoma cells underwent apoptosis following UV irradiation. In contrast, reintroduction of wild-type VHL expression protected 786-O cells from UV-mediated cell death. p53 and Bax levels were equivalent in VHL-negative and VHL-positive 786-O cells. Strikingly, cyclin-dependent kinase inhibitors p21 and p27 underwent proteasome-dependent degradation in VHL-negative 786-O cells following UV treatment. However, p21 and p27 protein levels were stable in VHL-positive cells. Also, levels of the anti-apoptotic proteins, Bcl-2 and Bcl-xL were elevated in VHL-positive cells, consistent with the protection from apoptotic stimuli. UV treatment led to increased S phase in VHL-negative, but not VHL-positive cells. Thus, following UV irradiation, diminution of p21 and p27 levels resulted in a hyperproliferative state in VHL-negative cells, leading to apoptosis. These results suggest that loss of VHL function promotes apoptosis and may provide selective pressure toward cells that are able to escape apoptosis, leading to tumorigenesis.

Simultaneous gout and pyarthrosis.

Although infrequently reported, the simultaneous occurrence of gout and infection in the same joint has been noted recently in three patients at the University of Virginia Hospital. Our report suggests that gout and pyarthrosis may coexist more often than is generally recognized and emphasizes the necessity of carefully searching for both conditions in an inflamed joint. The importance of adequately treating gout occurring with infection is also illustrated. Although speculative, factors such as synovial fluid pH, endotoxin, and enzymatic digestion of articular cartilage may predispose an infected joint to acute gout.

Diagnosis of superior vena caval obstruction by transesophageal echocardiography.

Superior vena caval (SVC) syndrome may be caused by extravascular compression or intravascular obstruction. Knowing the mechanism of SVC syndrome enables the physician to choose appropriate treatment. Transesophageal echocardiography (TEE) is a safe bedside procedure that is excellent for evaluating the SVC and its surrounding structures. We report the valuable role of TEE in demonstrating the mechanism of SVC syndrome.

Pre-operative intra-aortic balloon counterpulsation in stable patients with left main coronary disease.

OBJECTIVE: To determine whether prophylactic, pre-operative, intra-aortic balloon counterpulsation (IABC) improves clinical outcome in stable patients with severe left main coronary artery disease. METHODS: A post-hoc analysis of 457 prospectively tracked, non-randomized patients undergoing coronary artery bypass graft surgery (CABG) for left main stenoses 50% and multivessel coronary disease, but without any hemodynamic compromise or ongoing angina, was conducted. Patients with heart failure, shock, ongoing ischemia or previous CABG were excluded. In 287 patients, pre-operative IABC was not used (Group 1), while IABC was initiated in 170 patients for "prophylaxis" (Group 2). RESULTS: Groups 1 and 2 were similar in age (67 +/- 10 years versus 67 +/- 11 years, respectively), sex (72% male versus 71% male, respectively), and body mass index (28 +/- 5.5 versus 27 +/- 5.1, respectively). However, more Group 1 patients had peripheral vascular disease (PVD) (25% versus 11%), but more Group 2 patients had diabetes (37% versus 29%), and a lower left ventricular ejection fraction. The unadjusted 30-day mortality was significantly higher in Group 1 [16 (5.6%) versus 2 (1.2%); p = 0.02]. Cardiopulmonary bypass time and post-operative length of stay did not differ between the two groups. After adjusting for PVD in the multivariate analysis, the p-value for the no IABP versus IABP comparison in the presence of PVD was 0.10, even though 0/18 patients with PVD and IABC died. CONCLUSION: While unadjusted mortality appears lower with prophylactic IABC, confounding variables such as PVD mandate a larger, randomized clinical trial in order to establish the role of IABC in stable patients with left main disease.

Frequent MYC coamplification and DNA hypomethylation of multiple genes on 8q in 8p11-p12-amplified breast carcinomas.

Genetic and epigenetic (DNA methylation, histone modifications, microRNA expression) crosstalk promotes inactivation of tumor suppressor genes or activation of oncogenes by gene loss/hypermethylation or duplications/hypomethylation, respectively. The 8p11-p12 chromosomal region is a hotspot for genomic aberrations (chromosomal rearrangements, amplifications and deletions) in several cancer forms, including breast carcinoma where amplification has been associated with increased proliferation rates and reduced patient survival. Here, an integrative genomics screen (DNA copy number, transcriptional and DNA methylation profiling) performed in 229 primary invasive breast carcinomas identified substantial coamplification of the 8p11-p12 genomic region and the MYC oncogene (8q24.21), as well as aberrant methylation and transcriptional patterns for several genes spanning the 8q12.1-q24.22 genomic region (ENPP2, FABP5, IMPAD1, NDRG1, PLEKHF2, RRM2B, SQLE, TAF2, TATDN1, TRPS1, VPS13B). Taken together, our findings suggest that MYC activity and aberrant DNA methylation may also have a pivotal role in the aggressive tumor phenotype frequently observed in breast carcinomas harboring 8p11-p12 regional amplification.

Microbial interactions and the ecology and evolution of Hawaiian Drosophilidae.

Adaptive radiations are characterized by an increased rate of speciation and expanded range of habitats and ecological niches exploited by those species. The Hawaiian Drosophilidae is a classic adaptive radiation; a single ancestral species colonized Hawaii approximately 25 million years ago and gave rise to two monophyletic lineages, the Hawaiian Drosophila and the genus Scaptomyza. The Hawaiian Drosophila are largely saprophagous and rely on approximately 40 endemic plant families and their associated microbes to complete development. Scaptomyza are even more diverse in host breadth. While many species of Scaptomyza utilize decomposing plant substrates, some species have evolved to become herbivores, parasites on spider egg masses, and exploit microbes on living plant tissue. Understanding the origin of the ecological diversity encompassed by these nearly 700 described species has been a challenge. The central role of microbes in drosophilid ecology suggests bacterial and fungal associates may have played a role in the diversification of the Hawaiian Drosophilidae. Here we synthesize recent ecological and microbial community data from the Hawaiian Drosophilidae to examine the forces that may have led to this adaptive radiation. We propose that the evolutionary success of the Hawaiian Drosophilidae is due to a combination of factors, including adaptation to novel ecological niches facilitated by microbes.

Evolution of jasmonate and salicylate signal crosstalk.

The evolution of land plants approximately 470 million years ago created a new adaptive zone for natural enemies (attackers) of plants. In response to attack, plants evolved highly effective, inducible defense systems. Two plant hormones modulating inducible defenses are salicylic acid (SA) and jasmonic acid (JA). Current thinking is that SA induces resistance against biotrophic pathogens and some phloem feeding insects and JA induces resistance against necrotrophic pathogens, some phloem feeding insects and chewing herbivores. Signaling crosstalk between SA and JA commonly manifests as a reciprocal antagonism and may be adaptive, but this remains speculative. We examine evidence for and against adaptive explanations for antagonistic crosstalk, trace its phylogenetic origins and provide a hypothesis-testing framework for future research on the adaptive significance of SA-JA crosstalk.