Effects of a mildly cooked human-grade dog diet on gene expression, skin and coat health measures, and fecal microbiota of healthy adult dogs.

Purported benefits of human-grade pet foods include reduced inflammation, enhanced coat quality, and improved gut health, but research is scarce. Therefore, we compared gene expression, skin and coat health measures, and the fecal microbiome of dogs consuming a mildly cooked human-grade or extruded kibble diet. Twenty beagles (BW = 10.25 ± 0.82 kg; age = 3.85 ± 1.84 yr) were used in a completely randomized design. Test diets included: 1) chicken and brown rice recipe [feed-grade; extruded; blue buffalo (BB)]; and 2) chicken and white rice [human-grade; mildly cooked; Just Food for Dogs (JFFD)]. The study consisted of a 4-week baseline when all dogs ate BB, and a 12-week treatment phase when dogs were randomized to either diet (n = 10/group). After the baseline and treatment phases, fresh fecal samples were scored and collected for pH, dry matter (DM), and microbiome analysis; blood samples were collected for gene expression analysis; hair samples were microscopically imaged; and skin was analyzed for delayed-type hypersensitivity (DTH), sebum concentration, hydration status, and transepidermal water loss (TEWL). Data were analyzed as a change from baseline (CFB) using the Mixed Models procedure of SAS (version 9.4). At baseline, fecal pH was higher (P < 0.05) and hair surface score, superoxide dismutase (SOD) expression, and tumor necrosis factor-α (TNF-α) expression was lower (P < 0.05) in dogs allotted to JFFD. The decrease in CFB fecal pH and DM was greater (P < 0.05) in dogs fed JFFD, but fecal scores were not different. The increase in CFB hair surface score was higher (P < 0.05) in dogs fed JFFD. The decrease in CFB TEWL (back region) was greater (P < 0.05) in dogs fed JFFD, but TEWL (inguinal and ear regions), hydration status, and sebum concentrations in all regions were not different. Hair cortex scores and DTH responses were not affected by diet. The increase in CFB gene expression of SOD, COX-2, and TNF-α was greater (P < 0.05) in dogs fed JFFD. PCoA plots based on Bray-Curtis distances of bacterial genera and species showed small shifts over time in dogs fed BB, but dramatic shifts in those fed JFFD. JFFD increased (adj. P < 0.05) relative abundances of 4 bacterial genera, 11 bacterial species, 68 KEGG pathways, and 167 MetaCyc pathways, and decreased (adj. P < 0.05) 16 genera, 25 species, 98 KEGG pathways, and 87 MetaCyc pathways. In conclusion, the JFFD diet dramatically shifted the fecal microbiome but had minor effects on skin and coat measures and gene expression.

This study tested the effects of a mildly cooked human-grade diet and a feed-grade extruded kibble diet on the fecal microbiome, skin and coat health measures, and expression of genes related to inflammation and oxidative stress in healthy adult dogs. During a 4-week baseline, 20 beagles consumed the kibble diet. After baseline, 10 dogs continued to consume that diet, while 10 dogs consumed the mildly cooked diet for 12 weeks. After baseline and treatment phases, fresh fecal, blood, and hair samples were collected and skin was analyzed. The mildly cooked diet led to lower fecal pH and dry matter percentage, but fecal scores were not affected. The mildly cooked diet dramatically altered the fecal microbiome, shifting the relative abundances of over 30 bacterial species and 165 bacterial metabolic pathways. Measures of skin sebum content and hydration status were not different between groups, but skin water loss was lower in dogs consuming the mildly cooked diet. Baseline and post-treatment gene expression and hair surface scores were noted, but hair cortex and delayed-type hypersensitivity testing were not altered by diet. Our results demonstrate that mildly cooked diets dramatically change the fecal microbiome, but may not impact skin and coat in healthy adult dogs over a short time period.

Sequencing of Argonaute-bound microRNA/mRNA hybrids reveals regulation of the unfolded protein response by microRNA-320a.

MicroRNAs (miRNA) are short non-coding RNAs widely implicated in gene regulation. Most metazoan miRNAs utilize the RNase III enzymes Drosha and Dicer for biogenesis. One notable exception is the RNA polymerase II transcription start sites (TSS) miRNAs whose biogenesis does not require Drosha. The functional importance of the TSS-miRNA biogenesis is uncertain. To better understand the function of TSS-miRNAs, we applied a modified Crosslinking, Ligation, and Sequencing of Hybrids on Argonaute (AGO-qCLASH) to identify the targets for TSS-miRNAs in HCT116 colorectal cancer cells with or without DROSHA knockout. We observed that miR-320a hybrids dominate in TSS-miRNA hybrids identified by AGO-qCLASH. Targets for miR-320a are enriched for the eIF2 signaling pathway, a downstream component of the unfolded protein response. Consistently, in miR-320a mimic- and antagomir- transfected cells, differentially expressed gene products are associated with eIF2 signaling. Within the AGO-qCLASH data, we identified the endoplasmic reticulum (ER) chaperone calnexin as a direct miR-320a down-regulated target, thus connecting miR-320a to the unfolded protein response. During ER stress, but not amino acid deprivation, miR-320a up-regulates ATF4, a critical transcription factor for resolving ER stress. In summary, our study investigates the targetome of the TSS-miRNAs in colorectal cancer cells and establishes miR-320a as a regulator of unfolded protein response.

A noncanonical microRNA derived from the snaR-A noncoding RNA targets a metastasis inhibitor.

MicroRNAs (miRNAs) are small noncoding RNAs that function as critical posttranscriptional regulators in various biological processes. While most miRNAs are generated from processing of long primary transcripts via sequential Drosha and Dicer cleavage, some miRNAs that bypass Drosha cleavage can be transcribed as part of another small noncoding RNA. Here, we develop the target-oriented miRNA discovery (TOMiD) bioinformatic analysis method to identify Drosha-independent miRNAs from Argonaute crosslinking and sequencing of hybrids (Ago-CLASH) data sets. Using this technique, we discovered a novel miRNA derived from a primate specific noncoding RNA, the small NF90 associated RNA A (snaR-A). The miRNA derived from snaR-A (miR-snaR) arises independently of Drosha processing but requires Exportin-5 and Dicer for biogenesis. We identify that miR-snaR is concurrently up-regulated with the full snaR-A transcript in cancer cells. Functionally, miR-snaR associates with Ago proteins and targets NME1, a key metastasis inhibitor, contributing to snaR-A's role in promoting cancer cell migration. Our findings suggest a functional link between a novel miRNA and its precursor noncoding RNA.

A Staphylococcus pro-apoptotic peptide induces acute exacerbation of pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a chronic and fatal disease of unknown etiology; however, apoptosis of lung alveolar epithelial cells plays a role in disease progression. This intractable disease is associated with increased abundance of Staphylococcus and Streptococcus in the lungs, yet their roles in disease pathogenesis remain elusive. Here, we report that Staphylococcus nepalensis releases corisin, a peptide conserved in diverse staphylococci, to induce apoptosis of lung epithelial cells. The disease in mice exhibits acute exacerbation after intrapulmonary instillation of corisin or after lung infection with corisin-harboring S. nepalensis compared to untreated mice or mice infected with bacteria lacking corisin. Correspondingly, the lung corisin levels are significantly increased in human IPF patients with acute exacerbation compared to patients without disease exacerbation. Our results suggest that bacteria shedding corisin are involved in acute exacerbation of IPF, yielding insights to the molecular basis for the elevation of staphylococci in pulmonary fibrosis.

Genome Sequence of the Soybean Cyst Nematode (Heterodera glycines) Endosymbiont "Candidatus Cardinium hertigii" Strain cHgTN10.

In this study, we present the genome sequence of the "Candidatus Cardinium hertigii" strain cHgTN10, an endosymbiotic bacterium of the plant-parasitic nematode Heterodera glycines This is the first genome assembly reported for an endosymbiont directly sequenced from a tylenchid nematode.

Effect of pesticides on microbial communities in container aquatic habitats.

Container aquatic habitats support a specialized community of macroinvertebrates (e.g. mosquitoes) that feed on microbial communities associated with decaying organic matter. These aquatic habitats are often embedded within and around agricultural lands and are frequently exposed to pesticides. We used a microcosm approach to examine the single and combined effects of two herbicides (atrazine, glyphosate), and three insecticides (malathion, carbaryl, permethrin) on microbial communities of container aquatic habitats. MiSeq sequencing of the V4 region of both bacterial and archaeal 16S rRNA gene was used to characterize the microbial communities of indoor microcosms that were either exposed to each pesticide alone, a mix of herbicides, a mix of insecticides, or a mix of all five insecticides. Individual insecticides but not herbicides reduced the microbial diversity and richness and two insecticides, carbaryl and permethrin, also altered the microbial community structure. A mixture of herbicides had no effect on microbial diversity or structure but a mixture of insecticides or all five pesticides reduced microbial diversity and altered the community structure. These findings suggest that exposure of aquatic ecosystems to individual pesticides or their mixtures can disrupt aquatic microbial communities and there is need to decipher how these changes affect resident macroinvertebrate communities.

Plasma Exosomal miRNAs in Persons with and without Alzheimer Disease: Altered Expression and Prospects for Biomarkers.

To assess the value of exosomal miRNAs as biomarkers for Alzheimer disease (AD), the expression of microRNAs was measured in a plasma fraction enriched in exosomes by differential centrifugation, using Illumina deep sequencing. Samples from 35 persons with a clinical diagnosis of AD dementia were compared to 35 age and sex matched controls. Although these samples contained less than 0.1 microgram of total RNA, deep sequencing gave reliable and informative results. Twenty miRNAs showed significant differences in the AD group in initial screening (miR-23b-3p, miR-24-3p, miR-29b-3p, miR-125b-5p, miR-138-5p, miR-139-5p, miR-141-3p, miR-150-5p, miR-152-3p, miR-185-5p, miR-338-3p, miR-342-3p, miR-342-5p, miR-548at-5p, miR-659-5p, miR-3065-5p, miR-3613-3p, miR-3916, miR-4772-3p, miR-5001-3p), many of which satisfied additional biological and statistical criteria, and among which a panel of seven miRNAs were highly informative in a machine learning model for predicting AD status of individual samples with 83-89% accuracy. This performance is not due to over-fitting, because a) we used separate samples for training and testing, and b) similar performance was achieved when tested on technical replicate data. Perhaps the most interesting single miRNA was miR-342-3p, which was a) expressed in the AD group at about 60% of control levels, b) highly correlated with several of the other miRNAs that were significantly down-regulated in AD, and c) was also reported to be down-regulated in AD in two previous studies. The findings warrant replication and follow-up with a larger cohort of patients and controls who have been carefully characterized in terms of cognitive and imaging data, other biomarkers (e.g., CSF amyloid and tau levels) and risk factors (e.g., apoE4 status), and who are sampled repeatedly over time. Integrating miRNA expression data with other data is likely to provide informative and robust biomarkers in Alzheimer disease.