Genetic and metabolic links between the murine microbiome and memory.

BackgroundRecent evidence has linked the gut microbiome to host behavior via the gut-brain axis [1-3]; however, the underlying mechanisms remain unexplored. Here, we determined the links between host genetics, the gut microbiome and memory using the genetically defined Collaborative Cross (CC) mouse cohort, complemented with microbiome and metabolomic analyses in conventional and germ-free (GF) mice.ResultsA genome-wide association analysis (GWAS) identified 715 of 76,080 single-nucleotide polymorphisms (SNPs) that were significantly associated with short-term memory using the passive avoidance model. The identified SNPs were enriched in genes known to be involved in learning and memory functions. By 16S rRNA gene sequencing of the gut microbial community in the same CC cohort, we identified specific microorganisms that were significantly correlated with longer latencies in our retention test, including a positive correlation with Lactobacillus. Inoculation of GF mice with individual species of Lactobacillus (L. reuteri F275, L. plantarum BDGP2 or L. brevis BDGP6) resulted in significantly improved memory compared to uninoculated or E. coli DH10B inoculated controls. Untargeted metabolomics analysis revealed significantly higher levels of several metabolites, including lactate, in the stools of Lactobacillus-colonized mice, when compared to GF control mice. Moreover, we demonstrate that dietary lactate treatment alone boosted memory in conventional mice. Mechanistically, we show that both inoculation with Lactobacillus or lactate treatment significantly increased the levels of the neurotransmitter, gamma-aminobutyric acid (GABA), in the hippocampus of the mice.ConclusionTogether, this study provides new evidence for a link between Lactobacillus and memory and our results open possible new avenues for treating memory impairment disorders using specific gut microbial inoculants and/or metabolites. Video Abstract

Variability in the use of pulse oximeters with children in Kenyan hospitals: A mixed-methods analysis.

BACKGROUND: Pulse oximetry, a relatively inexpensive technology, has the potential to improve health outcomes by reducing incorrect diagnoses and supporting appropriate treatment decisions. There is evidence that in low- and middle-income countries, even when available, widespread uptake of pulse oximeters has not occurred, and little research has examined why. We sought to determine when and with which children pulse oximeters are used in Kenyan hospitals, how pulse oximeter use impacts treatment provision, and the barriers to pulse oximeter use. METHODS AND FINDINGS: We analyzed admissions data recorded through Kenya's Clinical Information Network (CIN) between September 2013 and February 2016. We carried out multiple imputation and generated multivariable regression models in R. We also conducted interviews with 30 healthcare workers and staff from 14 Kenyan hospitals to examine pulse oximetry adoption. We adapted the Integrative Model of Behavioural Prediction to link the results from the multivariable regression analyses to the qualitative findings. We included 27,906 child admissions from 7 hospitals in the quantitative analyses. The median age of the children was 1 year, and 55% were male. Three-quarters had a fever, over half had a cough; other symptoms/signs were difficulty breathing (34%), difficulty feeding (34%), and indrawing (32%). The most common diagnoses were pneumonia, diarrhea, and malaria: 45%, 35%, and 28% of children, respectively, had these diagnoses. Half of the children obtained a pulse oximeter reading, and of these, 10% had an oxygen saturation level below 90%. Children were more likely to receive a pulse oximeter reading if they were not alert (odds ratio [OR]: 1.30, 95% confidence interval (CI): 1.09, 1.55, p = 0.003), had chest indrawing (OR: 1.28, 95% CI: 1.17, 1.40, p < 0.001), or a very high respiratory rate (OR: 1.27, 95% CI: 1.13, 1.43, p < 0.001), as were children admitted to certain hospitals, at later time periods, and when a Paediatric Admission Record (PAR) was used (OR PAR used compared with PAR not present: 2.41, 95% CI: 1.98, 2.94, p < 0.001). Children were more likely to be prescribed oxygen if a pulse oximeter reading was obtained (OR: 1.42, 95% CI:1.25, 1.62, p < 0.001) and if this reading was below 90% (OR: 3.29, 95% CI: 2.82, 3.84, p < 0.001). The interviews indicated that the main barriers to pulse oximeter use are inadequate supply, broken pulse oximeters, and insufficient training on how, when, and why to use pulse oximeters and interpret their results. According to the interviews, variation in pulse oximeter use between hospitals is because of differences in pulse oximeter availability and the leadership of senior doctors in advocating for pulse oximeter use, whereas variation within hospitals over time is due to repair delays. Pulse oximeter use increased over time, likely because of the CIN's feedback to hospitals. When pulse oximeters are used, they are sometimes used incorrectly and some healthcare workers lack confidence in readings that contradict clinical signs. The main limitations of the study are that children with high levels of missing data were not excluded, interview participants might not have been representative, and the interviews did not enable a detailed exploration of differences between counties or across senior management groups. CONCLUSIONS: There remain major challenges to implementing pulse oximetry-a cheap, decades old technology-into routine care in Kenya. Implementation requires efficient and transparent procurement and repair systems to ensure adequate availability. Periodic training, structured clinical records that include prompts, the promotion of pulse oximetry by senior doctors, and monitoring and feedback might also support pulse oximeter use. Our findings can inform strategies to support the use of pulse oximeters to guide prompt and effective treatment, in line with the Sustainable Development Goals. Without effective implementation, the potential benefits of pulse oximeters and possible hospital cost-savings by targeting oxygen therapy might not be realized

The role of sequence, gene orientation, and intergenic distance in chromatin structure and function

Remarkable conservation of patterns of histone modifications and variants has been observed at active genes [1,2]. We show that genome-wide average distributions of chromatin marks relative to transcription start sites (TSSs) in early Drosophila melanogaster embryos are largely consistent with previously observed patterns in other organisms. This work describes qualitatively different chromatin domains and draws with functional inferences about these regional differences in modification patterns. We find `plateaus' of the histone variant, H2Av spanning transcriptionally inactive loci, as observed in mammalian cells [3]. These mixed cell populations from embryos exhibit distributions of chromatin marks similar to bivalent domains, specialized regions which extend across and beyong genes involved in developmental regulation and cell differentiation in mammalian stem cells; 5' ends of genes in these regions lack H2Av, regardless of the transcriptional state of the genes. In spite of the agreement of our initial studies with similar work in other organisms or tissues, analysis of chromatin patterns for genes grouped with respect to promoter orientation and distance from the adjacent upstream gene identifies differences in the positions and enrichment levels of active mark peaks, as well as levels of gene activity. Analysis of published data from human CD4+ cells and Saccharomyces cerevisiae reveals that many of the relationships between promoter context and the distributions of active marks surrounding TSSs are conserved among widely divergent eukaryotes. We propose that short-range, distance-dependent synergistic interactions between neighboring promoters impact both chromatin state and gene activity. The functional consequences of DNA sequence variation for the specificity and stability of nucleosome associations are largely unknown but open to new avenues of investigation based on emerging DNA sequencing technologies. We present preliminary results on interaction between molecular evolution and nucleosome positioning in Drosophila. Base composition surrounding nucleosomes isolated from melanogaster embryos supports some level sequence-encoded higher order structure with a periodicity of ~200bp. Analysis of the interspecific divergence shows that the rates of change in these regions support an equilibrium model maintaining these patterns. We note a substantially accelerated GC-&gt;AT rate on the melanogaster lineage. Dinucleotide periodicities across nucleosomal fragments are quite similar between melanogaster and simulans, and we find good support for conserved positional changes in in simulans relative to nucleosomes isolated from melanogaster

The role of sequence, gene orientation, and intergenic distance in chromatin structure and function

Remarkable conservation of patterns of histone modifications and variants has been observed at active genes [1,2]. We show that genome-wide average distributions of chromatin marks relative to transcription start sites (TSSs) in early Drosophila melanogaster embryos are largely consistent with previously observed patterns in other organisms. This work describes qualitatively different chromatin domains and draws with functional inferences about these regional differences in modification patterns. We find `plateaus' of the histone variant, H2Av spanning transcriptionally inactive loci, as observed in mammalian cells [3]. These mixed cell populations from embryos exhibit distributions of chromatin marks similar to bivalent domains, specialized regions which extend across and beyong genes involved in developmental regulation and cell differentiation in mammalian stem cells; 5' ends of genes in these regions lack H2Av, regardless of the transcriptional state of the genes. In spite of the agreement of our initial studies with similar work in other organisms or tissues, analysis of chromatin patterns for genes grouped with respect to promoter orientation and distance from the adjacent upstream gene identifies differences in the positions and enrichment levels of active mark peaks, as well as levels of gene activity. Analysis of published data from human CD4+ cells and Saccharomyces cerevisiae reveals that many of the relationships between promoter context and the distributions of active marks surrounding TSSs are conserved among widely divergent eukaryotes. We propose that short-range, distance-dependent synergistic interactions between neighboring promoters impact both chromatin state and gene activity. The functional consequences of DNA sequence variation for the specificity and stability of nucleosome associations are largely unknown but open to new avenues of investigation based on emerging DNA sequencing technologies. We present preliminary results on interaction between molecular evolution and nucleosome positioning in Drosophila. Base composition surrounding nucleosomes isolated from melanogaster embryos supports some level sequence-encoded higher order structure with a periodicity of ~200bp. Analysis of the interspecific divergence shows that the rates of change in these regions support an equilibrium model maintaining these patterns. We note a substantially accelerated GC-&gt;AT rate on the melanogaster lineage. Dinucleotide periodicities across nucleosomal fragments are quite similar between melanogaster and simulans, and we find good support for conserved positional changes in in simulans relative to nucleosomes isolated from melanogaster

Nucleosomes shape DNA polymorphism and divergence.

An estimated 80% of genomic DNA in eukaryotes is packaged as nucleosomes, which, together with the remaining interstitial linker regions, generate higher order chromatin structures [1]. Nucleosome sequences isolated from diverse organisms exhibit ∼10 bp periodic variations in AA, TT and GC dinucleotide frequencies. These sequence elements generate intrinsically curved DNA and help establish the histone-DNA interface. We investigated an important unanswered question concerning the interplay between chromatin organization and genome evolution: do the DNA sequence preferences inherent to the highly conserved histone core exert detectable natural selection on genomic divergence and polymorphism? To address this hypothesis, we isolated nucleosomal DNA sequences from Drosophila melanogaster embryos and examined the underlying genomic variation within and between species. We found that divergence along the D. melanogaster lineage is periodic across nucleosome regions with base changes following preferred nucleotides, providing new evidence for systematic evolutionary forces in the generation and maintenance of nucleosome-associated dinucleotide periodicities. Further, Single Nucleotide Polymorphism (SNP) frequency spectra show striking periodicities across nucleosomal regions, paralleling divergence patterns. Preferred alleles occur at higher frequencies in natural populations, consistent with a central role for natural selection. These patterns are stronger for nucleosomes in introns than in intergenic regions, suggesting selection is stronger in transcribed regions where nucleosomes undergo more displacement, remodeling and functional modification. In addition, we observe a large-scale (∼180 bp) periodic enrichment of AA/TT dinucleotides associated with nucleosome occupancy, while GC dinucleotide frequency peaks in linker regions. Divergence and polymorphism data also support a role for natural selection in the generation and maintenance of these super-nucleosomal patterns. Our results demonstrate that nucleosome-associated sequence periodicities are under selective pressure, implying that structural interactions between nucleosomes and DNA sequence shape sequence evolution, particularly in introns

Haplotypes spanning centromeric regions reveal persistence of large blocks of archaic DNA.

Despite critical roles in chromosome segregation and disease, the repetitive structure and vast size of centromeres and their surrounding heterochromatic regions impede studies of genomic variation. Here we report the identification of large-scale haplotypes (cenhaps) in humans that span the centromere-proximal regions of all metacentric chromosomes, including the arrays of highly repeated α-satellites on which centromeres form. Cenhaps reveal deep diversity, including entire introgressed Neanderthal centromeres and equally ancient lineages among Africans. These centromere-spanning haplotypes contain variants, including large differences in α-satellite DNA content, which may influence the fidelity and bias of chromosome transmission. The discovery of cenhaps creates new opportunities to investigate their contribution to phenotypic variation, especially in meiosis and mitosis, as well as to more incisively model the unexpectedly rich evolution of these challenging genomic regions

Nucleosomes Shape DNA Polymorphism and Divergence

<div><p>An estimated 80% of genomic DNA in eukaryotes is packaged as nucleosomes, which, together with the remaining interstitial linker regions, generate higher order chromatin structures <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1004457#pgen.1004457-Lee1" target="_blank">[1]</a>. Nucleosome sequences isolated from diverse organisms exhibit ∼10 bp periodic variations in AA, TT and GC dinucleotide frequencies. These sequence elements generate intrinsically curved DNA and help establish the histone-DNA interface. We investigated an important unanswered question concerning the interplay between chromatin organization and genome evolution: do the DNA sequence preferences inherent to the highly conserved histone core exert detectable natural selection on genomic divergence and polymorphism? To address this hypothesis, we isolated nucleosomal DNA sequences from <i>Drosophila melanogaster</i> embryos and examined the underlying genomic variation within and between species. We found that divergence along the <i>D. melanogaster</i> lineage is periodic across nucleosome regions with base changes following preferred nucleotides, providing new evidence for systematic evolutionary forces in the generation and maintenance of nucleosome-associated dinucleotide periodicities. Further, Single Nucleotide Polymorphism (SNP) frequency spectra show striking periodicities across nucleosomal regions, paralleling divergence patterns. Preferred alleles occur at higher frequencies in natural populations, consistent with a central role for natural selection. These patterns are stronger for nucleosomes in introns than in intergenic regions, suggesting selection is stronger in transcribed regions where nucleosomes undergo more displacement, remodeling and functional modification. In addition, we observe a large-scale (∼180 bp) periodic enrichment of AA/TT dinucleotides associated with nucleosome occupancy, while GC dinucleotide frequency peaks in linker regions. Divergence and polymorphism data also support a role for natural selection in the generation and maintenance of these super-nucleosomal patterns. Our results demonstrate that nucleosome-associated sequence periodicities are under selective pressure, implying that structural interactions between nucleosomes and DNA sequence shape sequence evolution, particularly in introns.</p></div