An integrated whole genome analysis of Mycobacterium tuberculosis reveals insights into relationship between its genome, transcriptome and methylome.

Human tuberculosis disease (TB), caused by Mycobacterium tuberculosis (Mtb), is a complex disease, with a spectrum of outcomes. Genomic, transcriptomic and methylation studies have revealed differences between Mtb lineages, likely to impact on transmission, virulence and drug resistance. However, so far no studies have integrated sequence-based genomic, transcriptomic and methylation characterisation across a common set of samples, which is critical to understand how DNA sequence and methylation affect RNA expression and, ultimately, Mtb pathogenesis. Here we perform such an integrated analysis across 22 M. tuberculosis clinical isolates, representing ancient (lineage 1) and modern (lineages 2 and 4) strains. The results confirm the presence of lineage-specific differential gene expression, linked to specific SNP-based expression quantitative trait loci: with 10 eQTLs involving SNPs in promoter regions or transcriptional start sites; and 12 involving potential functional impairment of transcriptional regulators. Methylation status was also found to have a role in transcription, with evidence of differential expression in 50 genes across lineage 4 samples. Lack of methylation was associated with three novel variants in mamA, likely to cause loss of function of this enzyme. Overall, our work shows the relationship of DNA sequence and methylation to RNA expression, and differences between ancient and modern lineages. Further studies are needed to verify the functional consequences of the identified mechanisms of gene expression regulation

The bovine alveolar macrophage DNA methylome is resilient to infection with Mycobacterium bovis

DNA methylation is pivotal in orchestrating gene expression patterns in various mammalian biological processes. Perturbation of the bovine alveolar macrophage (bAM) transcriptome, due to Mycobacterium bovis (M. bovis) infection, has been well documented; however, the impact of this intracellular pathogen on the bAM epigenome has not been determined. Here, whole genome bisulfite sequencing (WGBS) was used to assess the effect of M. bovis infection on the bAM DNA methylome. The methylomes of bAM infected with M. bovis were compared to those of non-infected bAM 24 hours post-infection (hpi). No differences in DNA methylation (CpG or non-CpG) were observed. Analysis of DNA methylation at proximal promoter regions uncovered >250 genes harbouring intermediately methylated (IM) promoters (average methylation of 33-66%). Gene ontology analysis, focusing on genes with low, intermediate or highly methylated promoters, revealed that genes with IM promoters were enriched for immune-related GO categories; this enrichment was not observed for genes in the high or low methylation groups. Targeted analysis of genes in the IM category confirmed the WGBS observation. This study is the first in cattle examining genome-wide DNA methylation at single nucleotide resolution in an important bovine cellular host-pathogen interaction model, providing evidence for IM promoter methylation in bAM

Sentra PM (a Medical Food) and Trazodone in the Management of Sleep Disorders

Sleep disorders are a common and poorly treated disease state. This double blind, four arm placebo-controlled, randomized trial compared (1) low dose trazodone, (2) Sentra PM, a neurotransmitter based medical food, (3) the joint administration of trazodone and the medical food Sentra PM and (4) placebo. There were 111 subjects studied in 12 independent sites. Subjects underwent baseline screening, informed consent and an initial sleep questionnaire. After 14 days subjects underwent a second evaluation by questionnaire. At baseline and Day 14 the subjects underwent 24 hour ECG recordings that were analyzed in the frequency domain of heart rate variability. The specific high frequency parasympathetic autonomic nervous system activity was analyzed. The primary endpoints were sleep latency and parasympathetic autonomic nervous system improvement in sleeping hours. The results showed improvement in sleep latency for the Sentra PM and combination of Sentra PM and trazodone (−41 and −56 minutes P < 0.001). There was an improvement in quality of sleep for the amino acid formulation Sentra PM and the combination (3.86 and 6.48 Likert units on a 10 point scale P < 0.001). There was an activation of circadian activity percent at night in the medical food and combination groups while there was no change in parasympathetic activity in either the placebo or trazodone group. These data indicate that Sentra PM can improve the quality of sleep, the response to trazodone as a sleep medication and parasympathetic autonomic nervous system activity

“Pesticides Protect the Fruit, but Not the People”: Using Community-Based Ethnography to Understand Farmworker Pesticide-Exposure Risks

Objectives. We used community-based ethnography and public health risk assessment to assess beliefs about pesticide exposure risks among farmworkers in the Lower Yakima Valley of Washington State