The fossilized birth–death process for coherent calibration of divergence-time estimates

Time-calibrated species phylogenies are critical for addressing a wide range of questions in evolutionary biology, such as those that elucidate historical biogeography or uncover patterns of coevolution and diversification. Because molecular sequence data are not informative on absolute time, external data—most commonly, fossil age estimates—are required to calibrate estimates of species divergence dates. For Bayesian divergence time methods, the common practice for calibration using fossil information involves placing arbitrarily chosen parametric distributions on internal nodes, often disregarding most of the information in the fossil record. We introduce the “fossilized birth–death” (FBD) process—a model for calibrating divergence time estimates in a Bayesian framework, explicitly acknowledging that extant species and fossils are part of the same macroevolutionary process. Under this model, absolute node age estimates are calibrated by a single diversification model and arbitrary calibration densities are not necessary. Moreover, the FBD model allows for inclusion of all available fossils. We performed analyses of simulated data and show that node age estimation under the FBD model results in robust and accurate estimates of species divergence times with realistic measures of statistical uncertainty, overcoming major limitations of standard divergence time estimation methods. We used this model to estimate the speciation times for a dataset composed of all living bears, indicating that the genus Ursus diversified in the Late Miocene to Middle Pliocene.A phylogenetic analysis of species has two goals: to infer the evolutionary relationships and the amount of divergence among species. Preferably, divergence is estimated in units proportional to time, thus revealing the times at which speciation events occurred. Once orthologous DNA sequences from the species have been aligned, both goals can be accomplished by assuming that nucleotide substitutions occur at the same rate in all lineages [the “molecular clock” assumption (1)] and that the time of at least one speciation event on the tree is known, i.e., one speciation event acts to “calibrate” the substitution rate.The goal of reconstructing rooted, time-calibrated phylogenies is complicated by substitution rates changing over the tree and by the difficulty of determining the date of any speciation event. Substitution rate variation among lineages is pervasive and has been accommodated in several ways. The most widely used method to account for rate heterogeneity is to assign an independent parameter to each branch of the tree. Branch lengths, then, are the product of substitution rate and time, and usually measured in units of expected number of substitutions per site. This solution allows estimation of the tree topology—which is informative about interspecies relationships—but does not attempt to estimate the rate and time separately. Thus, under this “unconstrained” parameterization, molecular sequence data allow inference of phylogenetic relationships and genetic distances among species, but the timing of speciation events is confounded in the branch-length parameter (2–4). Under a “relaxed-clock” model, substitution rates change over the tree in a constrained manner, thus separating the rate and time parameters associated with each branch and allowing inference of lineage divergence times. A considerable amount of effort has been directed at modeling lineage-specific substitution rate variation, with many different relaxed-clock models described in the literature (5–19). When such models are coupled with a model on the distribution of speciation events over time [e.g., the Yule model (20) or birth–death process (21)], molecular sequence data can inform the relative rates and node ages in a phylogenetic analysis.Estimates of branch lengths in units of absolute time (e.g., millions of years) are required for studies investigating comparative or biogeographical questions (e.g., refs. 22, 23). However, because commonly used diversification priors are imprecise on node ages, external information is required to infer the absolute timing of speciation events. Typically, a rooted time tree is calibrated by constraining the ages of a set of internal nodes. Age constraints may be derived from several sources, but the most common and reliable source of calibration information is the fossil record (24, 25). Despite the prevalence of these data in divergence time analyses, the problem of properly calibrating a phylogenetic tree has received less consideration than the problem of accommodating rate variation. Moreover, various factors may lead to substantial errors in parameter estimates (26–31). When estimating node ages, a calibration node must be identified for each fossil. For a given fossil, the calibration node is the node in the extant species tree that represents the most recent common ancestor (MRCA) of the fossil and a set of extant species. Based on the fossil, the calibration node’s age is estimated on an absolute timescale. Thus, fossil data typically can provide valid minimum-age constraints only on these nodes (24, 27), and erroneous conclusions may result if the calibration node is not specified properly (26).Bayesian inference methods are well adapted to accommodating uncertainty in calibration times by assuming that the age of the calibrated node is a random variable drawn from some parametric probability distribution (10, 14, 29, 31–35). Although this Bayesian approach properly propagates uncertainty in the calibration times through the analysis (reflected in the credible intervals on uncalibrated node ages), two problems remain unresolved.First, these approaches, as they commonly are applied, induce a probability distribution on the age of each calibrated node that comes from both the node-specific calibration prior and the tree-wide prior on node ages, leading to an incoherence in the model of branching times on the tree (35, 36). Typically, a birth–death process of cladogenesis is considered as the generating model for the tree and speciation times (20, 21, 37–40), serving as the tree-wide prior distribution on branch times in a Bayesian analysis. The speciation events acting as calibrations then are considered to be drawn from an additional, unrelated probability distribution intended to model uncertainty in the calibration time. This procedure results in overlaying two prior distributions for a calibration node: one from the tree prior and one from the calibration density (35, 41). Importantly, this incoherence is avoided by partitioning the nodes and applying a birth–death process to uncalibrated nodes conditioned on the calibrated nodes (32), although many divergence time methods do not use this approach. Nevertheless, a single model that acts as a prior on the speciation times for both calibrated and uncalibrated nodes is a better representation of the lineage diversification process and preferable as a prior on branching times when using fossil data.Second, the probability distributions used to model uncertainty in calibration times are poorly motivated. The standard practice in Bayesian divergence time methods is to model uncertainty in calibrated node ages by using simple probability distributions, such as the uniform, log-normal, gamma, or exponential distributions (29). When offset by a minimum age, these “calibration densities” (35) simply seek to characterize the age of the node with respect to its descendant fossil. However, the selection and parameterization of calibration priors rarely are informed by any biological process or knowledge of the fossil record (except see refs. 42–44). A probability model that acts as a fossil calibration prior should have parameters relevant to the preservation history of the group, such as the rate at which fossils occur in the rock record, a task that likely is difficult for most groups without an abundant fossil record (43, 45). Consequently, most biologists are faced with the challenge of choosing and parameterizing calibration densities without an explicit way to describe their prior knowledge about the calibration time. Thus, calibration priors often are specified based on arbitrary criteria or ad hoc validation methods (46), and ultimately, this may lead to arbitrary or ad hoc estimates of divergence times.We provide an alternative method for calibrating phylogenies with fossils. Because fossils and molecular sequences from extant species are different observations of the same diversification process, we use an explicit speciation–extinction–fossilization model to describe the distribution of speciation times and recovered fossils. This model—the fossilized birth–death (FBD) process—acts as a prior for divergence time dating. The parameters of the model—the speciation rate, extinction rate, fossil recovery rate, and proportion of sampled extant species—interact to inform the amount of uncertainty for every speciation event on the tree. These four parameters are the only quantities requiring prior assumptions, compared with assuming separate calibration densities for each fossil. Analyses of simulated data under the FBD model result in reliable estimates of absolute divergence times with realistic measures of statistical uncertainty. Moreover, node age estimates are robust to several biased sampling strategies of fossils and extant species—strategies that may be common practice or artifacts of fossil preservation but heavily violate assumptions of the model.     

Attention‐network specific alterations of structural connectivity in the undamaged white matter in acute neglect

Visual neglect results from dysfunction within the spatial attention network. The structural connectivity in undamaged brain tissue in neglect has barely been investigated until now. In the present study, we explored the microstructural white matter characteristics of the contralesional hemisphere in relation to neglect severity and recovery in acute stroke patients. We compared age‐matched healthy subjects and three groups of acute stroke patients (9 ± 0.5 days after stroke): (i) patients with nonrecovered neglect (n = 12); (ii) patients with rapid recovery from initial neglect (within the first week post‐stroke, n = 7), (iii) stroke patients without neglect (n = 17). We analyzed the differences between groups in grey and white matter density and fractional anisotropy (FA) and used fiber tracking to identify the affected fibers. Patients with nonrecovered neglect differed from those with rapid recovery by FA‐reduction in the left inferior parietal lobe. Fibers passing through this region connect the left‐hemispheric analogues of the ventral attention system. Compared with healthy subjects, neglect patients with persisting neglect had FA‐reduction in the left superior parietal lobe, optic radiation, and left corpus callosum/cingulum. Fibers passing through these regions connect centers of the left dorsal attention system. FA‐reduction in the identified regions correlated with neglect severity. The study shows for the first time white matter changes within the spatial attention system remote from the lesion and correlating with the extent and persistence of neglect. The data support the concept of neglect as disintegration within the whole attention system and illustrate the dynamics of structural‐functional correlates in acute stroke. Hum Brain Mapp 35:4678–4692, 2014. © 2014 Wiley Periodicals, Inc .     

High resolution ultrasound in posterior interosseous nerve syndrome

Introduction: Posterior interosseous nerve (PIN) syndrome is a rare compression neuropathy of the PIN in the region of the supinator muscle, most common by the arcade of Frohse. We aimed to specify ultrasonographic findings in patients with PIN syndrome in comparison to healthy volunteers. Methods: Ultrasound images and clinical data of 13 patients with PIN syndrome confirmed by neurological examination and electrophysiological testing were evaluated retrospectively. Anteroposterior nerve diameters measured at the arcade of Frohse were compared with those of 20 healthy volunteers. The echotexture and the presence of a caliber change of the PIN were additionally assessed. Results: Enlargement of the PIN was seen in all patients with PIN syndrome, but not in volunteers (statistically significant difference in mean diameter P < 0.05). Furthermore, edema and caliber change of the PIN were present in all patients. Conclusions: High‐resolution ultrasound allows for differentiation between patients with PIN syndrome and healthy volunteers. Muscle Nerve 49 : 35–39, 2014     

Gestational sleep deprivation is associated with higher offspring body mass index and blood pressure

Study ObjectivesThe objective of this study was to evaluate the association between gestational sleep deprivation and childhood adiposity and cardiometabolic profile.MethodsData were used from two population-based birth cohorts (Rhea study and Amsterdam Born Children and their Development study). A total of 3,608 pregnant women and their children were followed up until the age of 11 years. Gestational sleep deprivation was defined as 6 or fewer hours of sleep per day, reported by questionnaire. The primary outcomes included repeated measures of body mass index (BMI), waist circumference, body fat, serum lipids, systolic and diastolic blood pressure (DBP) levels in childhood. We performed a pooled analysis with adjusted linear mixed effect and Cox proportional hazards models. We tested for mediation by birthweight, gestational age, and gestational diabetes.ResultsGestational sleep deprivation was associated with higher BMI (beta; 95% CI: 0.7; 0.4, 1.0 kg/m²) and waist circumference (beta; 95% CI: 0.9; 0.1, 1.6 cm) in childhood, and increased risk for overweight or obesity (HR; 95% CI: 1.4; 1.1, 2.0). Gestational sleep deprivation was also associated with higher offspring DBP (beta; 95% CI: 1.6; 0.5, 2.7 mmHg). The observed associations were modified by sex (all p-values for interaction < 0.05); and were more pronounced in girls. Gestational diabetes and shorter gestational age partly mediated the seen associations.ConclusionsThis is the first study showing that gestational sleep deprivation may increase offspring’s adiposity and blood pressure, while exploring possible mechanisms. Attention to glucose metabolism and preterm birth might be extra warranted in mothers with gestational sleep deprivation.     

Genetic variability of serotonin pathway associated with schizophrenia onset,progression, and treatment

Schizophrenia (SZ) onset and treatment outcome have important genetic components, however individual genes do not have strong effects on SZ phenotype. Therefore, it is important to use the pathway‐based approach and study metabolic and signaling pathways, such as dopaminergic and serotonergic. Serotonin pathway has an important role in brain signaling, nevertheless, its role in SZ is not as thoroughly examined as that of dopamine pathway. In this study, we reviewed serotonin pathway genes and genetic variations associated with SZ, including variations at DNA, RNA, and epigenetic level. We obtained 30 serotonin pathway genes from Kyoto encyclopedia of genes and genomes and used these genes for the literature review. We extracted 20 protein coding serotonin pathway genes with genetic variations associated with SZ onset, development, and treatment from 31 research papers. Genes associated with SZ are present on all levels of serotonin pathway: serotonin synthesis, transport, receptor binding, intracellular signaling, and reuptake; however, regulatory genes are poorly researched. We summarized common challenges of genetic association studies and presented some solutions. The analysis of reported serotonin pathway‐SZ associations revealed lack of information about certain serotonin pathway genes potentially associated with SZ. Furthermore, it is becoming clear that interactions among serotonin pathway genes and their regulators may bring further knowledge about their involvement in SZ.     

Analysis of IgG reactivity against human papillomavirus type-16 E7 in patients with cervical intraepithelial neoplasm indicates an association with clearance of viral infection: results of a prospective study

IgG reactivity against the immunodominant region aa6-35 of Human Papillomavirus (HPV) type-16 E7 was determined in a peptide-based ELISA in a cohort study of women with initial mild to moderate cervical dyskaryosis. On the basis of HPV DNA patterns, as determined by PCR in cervical smears prior to IgG testing, HPV-16-positive patients were grouped as having either a cleared, a fluctuating, or a persistent HPV-16 infection. In a cross-sectional study at the start of serological follow-up, positive IgG reactivities were found more often in the total group of HPV-16-positive patients (20.0%) than in patients consistently typed as HPV-negative over a period of at least 12 months prior to testing (3.1%, p < 0.04). The highest proportion of positive responders was found in patients with a cleared HPV-16 infection (29.4%). Also, IgG reactivities found in HPV-16 clearance patients were significantly higher than in patients with a persistent infection (p < 0.008). In a subsequent longitudinal study over a period of up to 27 months, consistently positive reactivities were observed in patients with cleared viral infections who showed seroreactivity in the cross-sectional study, while mostly negative reactivities were found in patients with viral persistence. HPV-16 E7-specific IgG subclass responses were determined in a selection of 19 CIN and 11 HPV- 16-positive cervical carcinoma (CeCa) patients with positive E7-specific IgG responses. IgG₂ was predominant in the CIN patients, suggesting the presence of IFNγ (Th1) at the site of HPV infection. In the CeCa patients IgG₁ and IgG₂ were produced equally, possibly indicating a rise in Th2 cytokines. Our data suggest that HPV-16 E7 IgG reactivity in a subset of CIN patients with viral clearance may result from successful Th1 responses. © 1996 Wiley-Liss, Inc.     

A Synbiotic Formulation Comprising Bacillus subtilis DSM 32315 and L-Alanyl-L-Glutamine Improves Intestinal Butyrate Levels and Lipid Metabolism in Healthy Humans

The gut microbiota is a crucial modulator of health effects elicited by food components, with SCFA (short chain fatty acids), especially butyrate, acting as important mediators thereof. We therefore developed a nutritional synbiotic composition targeted at shifting microbiome composition and activity towards butyrate production. An intestinal screening model was applied to identify probiotic Bacillus strains plus various amino acids and peptides with suitable effects on microbial butyrate producers and levels. A pilot study was performed to test if the synbiotic formulation could improve fecal butyrate levels in healthy humans. A combination of Bacillus subtilis DSM (Number of German Collection of Microorganisms and Cell Cultures) 32315 plus L-alanyl-L-glutamine resulted in distinctly increased levels of butyrate and butyrate-producing taxa (Clostridium group XIVa, e.g., Faecalibacterium prausnitzii), both in vitro and in humans. Moreover, circulating lipid parameters (LDL-, and total cholesterol and LDL/HDL cholesterol ratio) were significantly decreased and further metabolic effects such as glucose-modulation were observed. Fasting levels of PYY (Peptide YY) and GLP-1 (Glucagon-like Peptide 1) were significantly reduced. In conclusion, our study indicates that this synbiotic composition may provide an effective and safe tool for stimulation of intestinal butyrate production with effects on e.g., lipid and glucose homeostasis. Further investigations in larger cohorts are warranted to confirm and expand these findings.     

Invasive Breast Carcinoma with Neuroendocrine Differentiation: A Single-Center Analysis of Clinical Features and Prognosis

Introduction Invasive breast cancer with neuroendocrine differentiation is a rare subtype of breast malignancy. Due to frequent changes in the definition of these lesions, the correct diagnosis, estimation of exact prevalence, and clinical behaviour of this entity may be challenging. The aim of this study was to evaluate the prevalence, clinical features, and outcomes in a large cohort of patients with breast cancer with neuroendocrine differentiation. Patients Twenty-seven cases of breast cancer with neuroendocrine differentiation have been included in this analysis. Twenty-one cases were identified by systematic immunohistochemical re-evaluation of 465 breast cancer specimens using the neuroendocrine markers chromogranin A and synaptophysin, resulting in a prevalence of 4.5%. A further six cases were identified by a review of clinical records. Results Median age at the time of diagnosis was 61 years. 70% of patients had T2 – 4 tumors and 37% were node-positive. The most common immunohistochemical subtype was HR-positive/HER2-negative (85%). 93% were positive for synaptophysin and 48% for chromogranin A. Somatostatin receptor type 2A status was positive in 12 of 24 analyzed tumors (50%). Neuroendocrine-specific treatment with somatostatin analogues was administered in two patients. The 5-year survival rate was 70%. Conclusions Breast cancer with neuroendocrine differentiation is mostly HR-positive/HER2-negative and the diagnosis is made at a higher TNM stage than in patients with conventional invasive breast carcinoma. Moreover, breast cancer with neuroendocrine differentiation was found to be associated with impaired prognosis in several retrospective trials. Due to somatostatin receptor 2A expression, somatostatin receptor-based imaging can be used and somatostatin receptor-targeted therapy can be offered in selected cases. Key words: neuroendocrine neoplasia of the breast, invasive breast cancer with neuroendocrine differentiation, neuroendocrine breast cancer, neuroendocrine markers, somatostatin receptor 2A