Effects of udder edema on parlor behavior in first- and second-lactation Holstein dairy cows.

Udder edema (UE) is a metabolic disorder that most commonly occurs around the transition period. However, gaps exist in our knowledge about its effects on parlor behavior. The aim of this study was to examine the prevalence and effects of UE on behavior in the milking parlor during udder preparation and active milking from 2 to 9 days in milk in first- (FL) and second-lactation (SL) Holstein dairy cattle. Each cow (n = 375) was observed once and monitored from the point of first contact during udder preparation through the first 5 min of active milking. Behavior measurements include step, kick, and kicking off the milking unit. Cows were determined to have UE when the rear udder medial suspensory ligament lost definition and was softened due to the presence of interstitial fluid. In this study, 237 out of 247 (95.6%) FL cows and 104 of 128 (81.3%) SL cows presented with UE. First-lactation cows with UE had a higher step rate (3.97 steps/session) when in contact during udder preparation and attachment (2.80 steps/session), and kick rate during milking (2.68 kicks/milking session) compared with SL cows with UE (2.37 steps/session, 1.25 steps/session, 1.24 kicks/milking session). Those FL cows with UE had a lower step rate during milking (6.04 steps/milking session) compared with FL cows without UE (7.20 steps/milking session). The FL cows with UE had a higher average count of kicking off the milking unit (µ = 0.220 kick-offs) than SL cows with UE (µ = 0.029 kick-offs) and FL cows without UE (µ = 0.091 kick-offs). The results indicate that UE is prevalent among dairy cattle, and the disorder has effects on behaviors presented in the milking parlor. Reducing UE has the potential to decrease step and kick behaviors to improve welfare of transition cows and reduce risk to dairy caretakers in the milking parlor.

Tissue predictability of elastography is low in collagenase induced deep digital flexor tendinopathy.

Elastography is an emerging imaging modality for characterizing tendon injury in horses, but its ability to differentiate tissue deformability relative to treatment group and biochemical properties using a prospective, experimental study design remain unknown. Objectives of the current study were to (a) to investigate differences in glycosaminoglycan, DNA, and soluble collagen levels in mesenchymal stem cell (MSC) treated limbs compared to untreated control limbs utilizing a collagenase model of tendinopathy; (b) compare elastographic features between treatment groups; and (c) determine tissue-level predictive capabilities of elastography in relation to biochemical outcomes. Bone marrow was collected for MSC culture and expansion. Tendinopathy of both forelimb deep digital flexor tendons (DDFTs) was induced with collagenase under ultrasonographic guidance. One randomly assigned limb was treated with intra-lesional MSC injection with the opposite limb serving as an untreated control. Horses were placed into a controlled exercise program with elastographic evaluations performed baseline (0) and 14, 60, 90, and 214 days post-treatment. Postmortem biochemical analysis was performed. MSC-treated limbs demonstrated significantly less (42%) glycosaminoglycan (P = .006). Significant differences in elastographic region of interest (ROI) percent hardness, ROI color histogram, and subjective lesion stiffness were appreciated between treatment groups at various study time points. Elastographic outcome parameters were weak predictors of biochemical tissue analysis, with all R2 values ≤ 0.50. Within this range of differences in glycosaminoglycan content between treatment groups, elastography outcomes did not predict biochemical differences. Tissue-specific differences between DDFTs treated with MSCs compared to controls were apparent biochemically, but not predicted by elastography.

Lives saved with vaccination for 10 pathogens across 112 countries in a pre-COVID-19 world.

Background: Vaccination is one of the most effective public health interventions. We investigate the impact of vaccination activities for Haemophilus influenzae type b, hepatitis B, human papillomavirus, Japanese encephalitis, measles, Neisseria meningitidis serogroup A, rotavirus, rubella, Streptococcus pneumoniae, and yellow fever over the years 2000-2030 across 112 countries. Methods: Twenty-one mathematical models estimated disease burden using standardised demographic and immunisation data. Impact was attributed to the year of vaccination through vaccine-activity-stratified impact ratios. Results: We estimate 97 (95%CrI[80, 120]) million deaths would be averted due to vaccination activities over 2000-2030, with 50 (95%CrI[41, 62]) million deaths averted by activities between 2000 and 2019. For children under-5 born between 2000 and 2030, we estimate 52 (95%CrI[41, 69]) million more deaths would occur over their lifetimes without vaccination against these diseases. Conclusions: This study represents the largest assessment of vaccine impact before COVID-19-related disruptions and provides motivation for sustaining and improving global vaccination coverage in the future. Funding: VIMC is jointly funded by Gavi, the Vaccine Alliance, and the Bill and Melinda Gates Foundation (BMGF) (BMGF grant number: OPP1157270 / INV-009125). Funding from Gavi is channelled via VIMC to the Consortium's modelling groups (VIMC-funded institutions represented in this paper: Imperial College London, London School of Hygiene and Tropical Medicine, Oxford University Clinical Research Unit, Public Health England, Johns Hopkins University, The Pennsylvania State University, Center for Disease Analysis Foundation, Kaiser Permanente Washington, University of Cambridge, University of Notre Dame, Harvard University, Conservatoire National des Arts et Métiers, Emory University, National University of Singapore). Funding from BMGF was used for salaries of the Consortium secretariat (authors represented here: TBH, MJ, XL, SE-L, JT, KW, NMF, KAMG); and channelled via VIMC for travel and subsistence costs of all Consortium members (all authors). We also acknowledge funding from the UK Medical Research Council and Department for International Development, which supported aspects of VIMC's work (MRC grant number: MR/R015600/1).JHH acknowledges funding from National Science Foundation Graduate Research Fellowship; Richard and Peggy Notebaert Premier Fellowship from the University of Notre Dame. BAL acknowledges funding from NIH/NIGMS (grant number R01 GM124280) and NIH/NIAID (grant number R01 AI112970). The Lives Saved Tool (LiST) receives funding support from the Bill and Melinda Gates Foundation.This paper was compiled by all coauthors, including two coauthors from Gavi. Other funders had no role in study design, data collection, data analysis, data interpretation, or writing of the report. All authors had full access to all the data in the study and had final responsibility for the decision to submit for publication.

Estimating the health impact of vaccination against ten pathogens in 98 low-income and middle-income countries from 2000 to 2030: a modelling study.

BACKGROUND: The past two decades have seen expansion of childhood vaccination programmes in low-income and middle-income countries (LMICs). We quantify the health impact of these programmes by estimating the deaths and disability-adjusted life-years (DALYs) averted by vaccination against ten pathogens in 98 LMICs between 2000 and 2030. METHODS: 16 independent research groups provided model-based disease burden estimates under a range of vaccination coverage scenarios for ten pathogens: hepatitis B virus, Haemophilus influenzae type B, human papillomavirus, Japanese encephalitis, measles, Neisseria meningitidis serogroup A, Streptococcus pneumoniae, rotavirus, rubella, and yellow fever. Using standardised demographic data and vaccine coverage, the impact of vaccination programmes was determined by comparing model estimates from a no-vaccination counterfactual scenario with those from a reported and projected vaccination scenario. We present deaths and DALYs averted between 2000 and 2030 by calendar year and by annual birth cohort. FINDINGS: We estimate that vaccination of the ten selected pathogens will have averted 69 million (95% credible interval 52-88) deaths between 2000 and 2030, of which 37 million (30-48) were averted between 2000 and 2019. From 2000 to 2019, this represents a 45% (36-58) reduction in deaths compared with the counterfactual scenario of no vaccination. Most of this impact is concentrated in a reduction in mortality among children younger than 5 years (57% reduction [52-66]), most notably from measles. Over the lifetime of birth cohorts born between 2000 and 2030, we predict that 120 million (93-150) deaths will be averted by vaccination, of which 58 million (39-76) are due to measles vaccination and 38 million (25-52) are due to hepatitis B vaccination. We estimate that increases in vaccine coverage and introductions of additional vaccines will result in a 72% (59-81) reduction in lifetime mortality in the 2019 birth cohort. INTERPRETATION: Increases in vaccine coverage and the introduction of new vaccines into LMICs have had a major impact in reducing mortality. These public health gains are predicted to increase in coming decades if progress in increasing coverage is sustained. FUNDING: Gavi, the Vaccine Alliance and the Bill & Melinda Gates Foundation.

Estimation and prediction for a mechanistic model of measles transmission using particle filtering and maximum likelihood estimation.

Disease incidence reported directly within health systems frequently reflects a partial observation relative to the true incidence in the population. State-space models present a general framework for inferring both the dynamics of infectious disease processes and the unobserved burden of disease in the population. Here, we present a state-space model of measles transmission and vaccine-based interventions at the country-level and a particle filter-based estimation procedure. Our dynamic transmission model builds on previous work by incorporating population age-structure to allow explicit representation of age-targeted vaccine interventions. We illustrate the performance of estimators of model parameters and predictions of unobserved states on simulated data from two dynamic models: one on the annual time-scale of observations and one on the biweekly time-scale of the epidemiological dynamics. We show that our model results in approximately unbiased estimates of unobserved burden and the underreporting rate. We further illustrate the performance of the fitted model for prediction of future disease burden in the next one to 15 years.

Changes in Shear Modulus of the Lumbar Multifidus Muscle During Different Body Positions.

Multifidus function is important for active stabilization of the spine, but it can be compromised in patients with chronic low back pain and other spine pathologies. Force production and strength of back muscles are often evaluated using isometric or isokinetic tests, which lack the ability to quantify multifidi contribution independent of the erector spinae and adjacent hip musculature. The objective of this study is to evaluate localized force production capability in multifidus muscle using ultrasound shear wave elastography (SWE) in healthy individuals. Three different body positions were considered: lying prone, sitting up, and sitting up with the right arm lifted. These positions were chosen to progressively increase multifidus contraction and to minimize body motion during measurements. Shear modulus was measured at the superficial and deeper layers of the multifidus. Repeatability and possible sources of error of the shear modulus measurements were analyzed. Multifidus shear modulus (median (interquartile range)) increased from prone, i.e., 16.15 (6.69) kPa, to sitting up, i.e., 27.28 (15.72) kPa, to sitting up with the right arm lifted position, i.e., 45.02 (25.27) kPa. Multifidi shear modulus in the deeper layer of the multifidi was lower than the superficial layer, suggesting lower muscle contraction. Intraclass correlation coefficients (ICCs) for evaluation of shear modulus by muscle layer were found to be excellent (ICC = 0.76-0.80). Results suggest that the proposed protocol could quantify local changes in spinal muscle function in healthy adults; further research in patients with spine pathology is warranted.

Impact of Helminth Infections and Nutritional Constraints on the Small Intestine Microbiota.

Helminth infections and nutrition can independently alter the composition and abundance of the gastrointestinal microbiota, however, their combined effect is poorly understood. Here, we used the T. retortaeformis-rabbit system to examine how the helminth infection and host restriction from coprophagy/ready-to-absorb nutrients affected the duodenal microbiota, and how these changes related to the acquired immune response at the site of infection. A factorial experiment was performed where the bacterial community, its functionality and the immune response were examined in four treatments (Infect, Infect+Collar, Control+Collar and Control). Helminths reduced the diversity and abundance of the microbiota while the combination of parasites and coprophagic restriction led to a more diversified and abundant microbiota than infected cases, without significantly affecting the intensity of infection. Animals restricted from coprophagy and free from parasites exhibited the richest and most abundant bacterial community. By forcing the individuals to absorb nutrients from less digested food, the coprophagic restriction appears to have facilitated the diversity and proliferation of bacteria in the duodenum. Changes in the microbiota were more clearly associated with changes in the immune response for the infected than the nutrient restricted animals. The functional and metabolic characteristics of the duodenal microbiota were not significantly different between treatments. Overall, infection and diet affect the gut microbiota but their interactions and outcome can be complex. These findings can have important implications for the development of control measures to helminth infections where poor nutrition/malnutrition can also be a concern.

Nonoperative management of splenic injuries: significance of age.

BACKGROUND: In the nonoperative management (NOM) of blunt splenic injuries (BSI), the clinical relevance of age as a risk factor has not been well studied. METHODS: Using the 2011 National Trauma Data Bank data set, age was analyzed both as a continuous variable and a categorical variable (group 1 [13-54 y], group 2 [55-74 y], and group 3 [≥75 y]). BSI severity was stratified by abbreviated injury scale (AIS): group 1 (AIS ≤2), group 2 (AIS 3), and group 3 (AIS ≥4). A semiparametric proportional odds model was used to model NOM outcomes and effects due to age and BSI severity. RESULTS: Of 15,113 subjects, 15.3% failed NOM. The odds of failure increased by a factor of 1.014 for each year of age, or factor of 1.5 for groups 2 and 3 each. BSI severity groups 2 and 3 had increases in the odds of failure by factors of 3.9 and 13, respectively, compared with those of group 1. Most failures occurred by 48 h irrespective of age. The effect of age was most pronounced in age groups 2 and 3 with the most severe BSI, where a NOM failure rate of >50% was seen. Both age and failure of NOM were independent predictors of mortality. CONCLUSIONS: Age is associated with failure of NOM but its effect seems more clinically relevant only in high-grade BSI. Factors that could influence NOM success in elderly patients with high-grade injuries deserve further study.

Delayed administration of alpha-difluoromethylornithine prevents hippocampus-dependent cognitive impairment after single and combined injury in mice.

Radiation exposure due to radiological terrorism and military circumstances are a continuing threat for the civilian population. In an uncontrolled radiation event, it is likely that there will be other types of injury involved, including trauma. While radiation combined injury is recognized as an area of great significance, overall there is a paucity of information regarding the mechanisms underlying the interactions between irradiation and other forms of injury, or what countermeasures might be effective in ameliorating such changes. The objective of this study was to determine if difluoromethylornithine (DFMO) could reduce the adverse effects of single or combined injury if administered beginning 24 h after exposure. Eight-week-old C57BL/J6 young-adult male mice received whole-body cesium-137 ((137)Cs) irradiation with 4 Gy. Immediately after irradiation, unilateral traumatic brain injury was induced using a controlled cortical impact system. Forty-four days postirradiation, animals were tested for hippocampus-dependent cognitive performance in the Morris water maze. After cognitive testing, animals were euthanized and their brains snap frozen for immunohistochemical assessment of neuroinflammation (activated microglia) and neurogenesis in the hippocampal dentate gyrus. Our data show that single and combined injuries induced variable degrees of hippocampus-dependent cognitive dysfunction, and when given 24 h post trauma, DFMO treatment ameliorated those effects. Cellular changes including neurogenesis and numbers of activated microglia were generally not associated with the cognitive changes. Further analyses also revealed that DFMO increased hippocampal protein levels of the antioxidants thioredoxin 1 and peroxiredoxin 3 compared to vehicle treated animals. While the mechanisms responsible for the improvement in cognition after DFMO treatment are not yet clear, these results constitute a basis for further development of DFMO as a countermeasure for ameliorating the of risks for cognitive dysfunction in individuals subjected to trauma and radiation combined injury.

Life extension factor klotho enhances cognition.

Aging is the primary risk factor for cognitive decline, an emerging health threat to aging societies worldwide. Whether anti-aging factors such as klotho can counteract cognitive decline is unknown. We show that a lifespan-extending variant of the human KLOTHO gene, KL-VS, is associated with enhanced cognition in heterozygous carriers. Because this allele increased klotho levels in serum, we analyzed transgenic mice with systemic overexpression of klotho. They performed better than controls in multiple tests of learning and memory. Elevating klotho in mice also enhanced long-term potentiation, a form of synaptic plasticity, and enriched synaptic GluN2B, an N-methyl-D-aspartate receptor (NMDAR) subunit with key functions in learning and memory. Blockade of GluN2B abolished klotho-mediated effects. Surprisingly, klotho effects were evident also in young mice and did not correlate with age in humans, suggesting independence from the aging process. Augmenting klotho or its effects may enhance cognition and counteract cognitive deficits at different life stages.

Radiation exposure prior to traumatic brain injury induces responses that differ as a function of animal age.

PURPOSE: Uncontrolled radiation exposure due to radiological terrorism, industrial accidents or military circumstances is a continuing threat for the civilian population. Age plays a major role in the susceptibility to radiation; younger children are at higher risk of developing cognitive deterioration when compared to adults. Our objective was to determine if an exposure to radiation affected the vulnerability of the juvenile hippocampus to a subsequent moderate traumatic injury. MATERIALS AND METHODS: Three-week-old (juvenile) and eight-week-old young adult C57BL/J6 male mice received whole body cesium-137 ((137)Cs) irradiation with 4 gray (Gy). One month later, unilateral traumatic brain injury was induced using a controlled cortical impact system. Two months post-irradiation, animals were tested for hippocampus-dependent cognitive performance in the Morris water-maze. After cognitive testing, animals were euthanized and their brains frozen for immunohistochemical assessment of activated microglia and neurogenesis in the hippocampal dentate gyrus. RESULTS: All animals were able to learn the water maze task; however, treatment effects were seen when spatial memory retention was assessed. Animals that received irradiation as juveniles followed by a moderate traumatic brain injury one month later did not show spatial memory retention, i.e., were cognitively impaired. In contrast, all groups of animals that were treated as adults showed spatial memory retention in the probe trials. CONCLUSION: Although the mechanisms involved are not clear, our results suggest that irradiation enhanced a young animal's vulnerability to develop cognitive injury following a subsequent traumatic injury.

Population genomic analysis reveals a rich speciation and demographic history of orang-utans (Pongo pygmaeus and Pongo abelii).

To gain insights into evolutionary forces that have shaped the history of Bornean and Sumatran populations of orang-utans, we compare patterns of variation across more than 11 million single nucleotide polymorphisms found by previous mitochondrial and autosomal genome sequencing of 10 wild-caught orang-utans. Our analysis of the mitochondrial data yields a far more ancient split time between the two populations (~3.4 million years ago) than estimates based on autosomal data (0.4 million years ago), suggesting a complex speciation process with moderate levels of primarily male migration. We find that the distribution of selection coefficients consistent with the observed frequency spectrum of autosomal non-synonymous polymorphisms in orang-utans is similar to the distribution in humans. Our analysis indicates that 35% of genes have evolved under detectable negative selection. Overall, our findings suggest that purifying natural selection, genetic drift, and a complex demographic history are the dominant drivers of genome evolution for the two orang-utan populations.

Effects of radiation combined injury on hippocampal function are modulated in mice deficient in chemokine receptor 2 (CCR2).

Chemokines and their receptors play a crucial role in normal brain function as well as in pathological conditions such as injury and disease-associated neuroinflammation. Chemokine receptor-2 (CCR2), which mediates the recruitment of infiltrating and resident microglia to sites of central nervous system (CNS) inflammation, is upregulated by ionizing irradiation and traumatic brain injury. Our objective was to determine if a deficiency in CCR2 and subsequent effects on brain microglia affect neurogenesis and cognitive function after radiation combined injury (RCI). CCR2 knock-out ⁻/⁻ and wild-type (WT) mice received 4 Gy of whole body ¹³7Cs irradiation. Immediately after irradiation, unilateral traumatic brain injury was induced using a controlled cortical impact system. Forty-four days postirradiation, animals were tested for hippocampus-dependent cognitive performance in the Morris water-maze. After cognitive testing, animals were euthanized and their brains snap frozen for immunohistochemical assessment of neuroinflammation (activated microglia) and neurogenesis in the hippocampal dentate gyrus. All animals were able to locate the visible and hidden platform locations in the water maze; however, treatment effects were seen when spatial memory retention was assessed in the probe trials (no platform). In WT animals that received combined injury, a significant impairment in spatial memory retention was observed in the probe trial after the first day of hidden platform training (first probe trial). This impairment was associated with increased neurogenesis in the ipsilateral hemisphere of the dentate gyrus. In contrast, CCR2⁻/⁻ mice, independent of insult showed significant memory retention in the first probe trial and there were no differences in the numbers of newly born neurons in the animals receiving irradiation, trauma or combined injury. Although the mechanisms involved are not clear, our data suggests that CCR2 deficiency can exert a protective effect preventing the impairment of cognitive function after combined injury.

Physiologic brain activity causes DNA double-strand breaks in neurons, with exacerbation by amyloid-ß.

We show that a natural behavior, exploration of a novel environment, causes DNA double-strand breaks (DSBs) in neurons of young adult wild-type mice. DSBs occurred in multiple brain regions, were most abundant in the dentate gyrus, which is involved in learning and memory, and were repaired within 24 h. Increasing neuronal activity by sensory or optogenetic stimulation increased neuronal DSBs in relevant but not irrelevant networks. Mice transgenic for human amyloid precursor protein (hAPP), which simulate key aspects of Alzheimer's disease, had increased neuronal DSBs at baseline and more severe and prolonged DSBs after exploration. Interventions that suppress aberrant neuronal activity and improve learning and memory in hAPP mice normalized their levels of DSBs. Blocking extrasynaptic NMDA-type glutamate receptors prevented amyloid-ß (Aß)-induced DSBs in neuronal cultures. Thus, transient increases in neuronal DSBs occur as a result of physiological brain activity, and Aß exacerbates DNA damage, most likely by eliciting synaptic dysfunction.

Evolutionary and population genomics of the cavity causing bacteria Streptococcus mutans.

Streptococcus mutans is widely recognized as one of the key etiological agents of human dental caries. Despite its role in this important disease, our present knowledge of gene content variability across the species and its relationship to adaptation is minimal. Estimates of its demographic history are not available. In this study, we generated genome sequences of 57 S. mutans isolates, as well as representative strains of the most closely related species to S. mutans (S. ratti, S. macaccae, and S. criceti), to identify the overall structure and potential adaptive features of the dispensable and core components of the genome. We also performed population genetic analyses on the core genome of the species aimed at understanding the demographic history, and impact of selection shaping its genetic variation. The maximum gene content divergence among strains was approximately 23%, with the majority of strains diverging by 5-15%. The core genome consisted of 1,490 genes and the pan-genome approximately 3,296. Maximum likelihood analysis of the synonymous site frequency spectrum (SFS) suggested that the S. mutans population started expanding exponentially approximately 10,000 years ago (95% confidence interval [CI]: 3,268-14,344 years ago), coincidental with the onset of human agriculture. Analysis of the replacement SFS indicated that a majority of these substitutions are under strong negative selection, and the remainder evolved neutrally. A set of 14 genes was identified as being under positive selection, most of which were involved in either sugar metabolism or acid tolerance. Analysis of the core genome suggested that among 73 genes present in all isolates of S. mutans but absent in other species of the mutans taxonomic group, the majority can be associated with metabolic processes that could have contributed to the successful adaptation of S. mutans to its new niche, the human mouth, and with the dietary changes that accompanied the origin of agriculture.

SnIPRE: selection inference using a Poisson random effects model.

We present an approach for identifying genes under natural selection using polymorphism and divergence data from synonymous and non-synonymous sites within genes. A generalized linear mixed model is used to model the genome-wide variability among categories of mutations and estimate its functional consequence. We demonstrate how the model's estimated fixed and random effects can be used to identify genes under selection. The parameter estimates from our generalized linear model can be transformed to yield population genetic parameter estimates for quantities including the average selection coefficient for new mutations at a locus, the synonymous and non-synynomous mutation rates, and species divergence times. Furthermore, our approach incorporates stochastic variation due to the evolutionary process and can be fit using standard statistical software. The model is fit in both the empirical Bayes and Bayesian settings using the lme4 package in R, and Markov chain Monte Carlo methods in WinBUGS. Using simulated data we compare our method to existing approaches for detecting genes under selection: the McDonald-Kreitman test, and two versions of the Poisson random field based method MKprf. Overall, we find our method universally outperforms existing methods for detecting genes subject to selection using polymorphism and divergence data.

Dynamic and coordinated epigenetic regulation of developmental transitions in the cardiac lineage.

Heart development is exquisitely sensitive to the precise temporal regulation of thousands of genes that govern developmental decisions during differentiation. However, we currently lack a detailed understanding of how chromatin and gene expression patterns are coordinated during developmental transitions in the cardiac lineage. Here, we interrogated the transcriptome and several histone modifications across the genome during defined stages of cardiac differentiation. We find distinct chromatin patterns that are coordinated with stage-specific expression of functionally related genes, including many human disease-associated genes. Moreover, we discover a novel preactivation chromatin pattern at the promoters of genes associated with heart development and cardiac function. We further identify stage-specific distal enhancer elements and find enriched DNA binding motifs within these regions that predict sets of transcription factors that orchestrate cardiac differentiation. Together, these findings form a basis for understanding developmentally regulated chromatin transitions during lineage commitment and the molecular etiology of congenital heart disease.

Derivation conditions impact X-inactivation status in female human induced pluripotent stem cells.

Female human induced pluripotent stem cell (hiPSC) lines exhibit variability in X-inactivation status. The majority of hiPSC lines maintain one transcriptionally active X (Xa) and one inactive X (Xi) chromosome from donor cells. However, at low frequency, hiPSC lines with two Xas are produced, suggesting that epigenetic alterations of the Xi occur sporadically during reprogramming. We show here that X-inactivation status in female hiPSC lines depends on derivation conditions. hiPSC lines generated by the Kyoto method (retroviral or episomal reprogramming), which uses leukemia inhibitory factor (LIF)-expressing SNL feeders, frequently had two Xas. Early passage Xa/Xi hiPSC lines generated on non-SNL feeders were converted into Xa/Xa hiPSC lines after several passages on SNL feeders, and supplementation with recombinant LIF caused reactivation of some of X-linked genes. Thus, feeders are a significant factor affecting X-inactivation status. The efficient production of Xa/Xa hiPSC lines provides unprecedented opportunities to understand human X-reactivation and -inactivation.

A bayesian mixture model for comparative spectral count data in shotgun proteomics.

Recent developments in mass-spectrometry-based shotgun proteomics, especially methods using spectral counting, have enabled large-scale identification and differential profiling of complex proteomes. Most such proteomic studies are interested in identifying proteins, the abundance of which is different under various conditions. Several quantitative methods have recently been proposed and implemented for this purpose. Building on some techniques that are now widely accepted in the microarray literature, we developed and implemented a new method using a Bayesian model to calculate posterior probabilities of differential abundance for thousands of proteins in a given experiment simultaneously. Our Bayesian model is shown to deliver uniformly superior performance when compared with several existing methods.

Successful computational prediction of novel imprinted genes from epigenomic features.

Approximately 100 mouse genes undergo genomic imprinting, whereby one of the two parental alleles is epigenetically silenced. Imprinted genes influence processes including development, X chromosome inactivation, obesity, schizophrenia, and diabetes, motivating the identification of all imprinted loci. Local sequence features have been used to predict candidate imprinted genes, but rigorous testing using reciprocal crosses validated only three, one of which resided in previously identified imprinting clusters. Here we show that specific epigenetic features in mouse cells correlate with imprinting status in mice, and we identify hundreds of additional genes predicted to be imprinted in the mouse. We used a multitiered approach to validate imprinted expression, including use of a custom single nucleotide polymorphism array and traditional molecular methods. Of 65 candidates subjected to molecular assays for allele-specific expression, we found 10 novel imprinted genes that were maternally expressed in the placenta.