Associations between memory performance and Bifidobacterium pseudolongum abundance in the canine gut microbiome.

Memory has been identified as the least heritable cognitive trait in canines, suggesting a significant influence of non-genetic factors. We observed a trend that overall memory scores (OMS) improve with age in a cohort of 27 young dogs, but considerable plasticity exists. Employing linear discriminant analysis of gut microbiome data from dogs exhibiting low and high OMS, a single bacterial species, Bifidobacterium pseudolongum, was identified and confirmed to be correlated with elevated OMS. Subsequent analysis using a random forest regression model revealed that sex, litter, and breed identity had minimal predictive importance. Age had some predictive value but failed to achieve statistical significance in this dataset. In sharp contrast, the abundance of 17 bacterial taxa in the microbiome showed a stronger predictive capacity for memory performance. Our findings provide insights into microbiome underpinnings of mammalian cognitive functions and suggest avenues for developing psychobiotics to enhance canine memory and learning.

Hepatic transcript profiling in beef cattle: Effects of rumen-protected niacin supplementation.

The objective of our study was to assess the effect of rumen-protected niacin supplementation on the transcriptome of liver tissue in growing Angus × Simmental steers and heifers through RNA-seq analysis. Consequently, we wanted to assess the known role of niacin in the physiological processes of vasodilation, detoxification, and immune function in beef hepatic tissue. Normal weaned calves (~8 months old) were provided either a control diet or a diet supplemented with rumen-protected niacin (6 g/hd/d) for a 30-day period, followed by a liver biopsy. We observed a significant list of changes at the transcriptome level due to rumen-protected niacin supplementation. Several metabolic pathways revealed potential positive effects to the animal's liver metabolism due to administration of rumen-protected niacin; for example, a decrease in lipolysis, apoptosis, inflammatory responses, atherosclerosis, oxidative stress, fibrosis, and vasodilation-related pathways. Therefore, results from our study showed that the liver transcriptional machinery switched several metabolic pathways to a condition that could potentially benefit the health status of animals supplemented with rumen-protected niacin. In conclusion, based on the results of our study, we can suggest the utilization of rumen-protected niacin supplementation as a nutritional strategy could improve the health status of growing beef cattle in different beef production stages, such as backgrounding operations or new arrivals to a feedlot.

Fascinating Immobilization-Free Electrochemical Immunosensing Strategy Based on the Cooperation of Buoyancy and Magnetism.

Rapid and accurate detection of biomolecules is of vital importance for the diagnosis of disease and for performing timely treatments. The point-of-care analysis of cancer biomarkers in the blood with low cost and easy processing is still challenging. Herein, an advanced and robust strategy, which integrates the buoyant recognition probe with the magnetic reporter probe in one solution, was first proposed for immobilization-free electrochemical immunosensing. The tumor marker of alpha fetoprotein (AFP) can be captured immune-buoyantly, and then a multifunctional magnetic reporter probe in pseudo-homogeneous solution was further captured to fulfill a sandwich-type immunoreaction. The residual magnetic reporter probe can be firmly and efficiently attracted on a magnetic glassy carbon electrode to fulfill the conversion of the target AFP amount into the residual magnetic electrochemical signal indicator. As a result, the electrochemical signal of methylene blue can accurately reflect the original level of target antigen AFP concentration. By integrating buoyancy-driven quasi-homogenous biorecognition with magnetism-mediated amplification and signal output, the proposed immobilization-free electrochemical immunosensing strategy displayed a wide range of linear response (100 fg mL-1 to 10 ng mL-1), low detection limit (14.52 fg mL-1), and good reproducibility, selectivity, and stability. The designed strategy manifests remarkable advantages including assay simplicity, rapidness, and high sensitivity owing to the in-solution instead of on-electrode biorecognition that could accelerate and improve the biorecognition efficiency. To the best of our knowledge, this is the first cooperation of buoyancy-driven biorecognition with magnetism-mediated signal output in bioanalysis, which would be attractive for rapid clinic biomedical application. Thus, this work provides a fresh perspective for convenient and favorable immobilization-free electrochemical biosensing of universal biomolecules.

Blood transcriptome responses to PFOA and GenX treatment in the marsupial biomedical model Monodelphis domestica.

Introduction: Perfluoroalkyl and poly-fluoroalkyl substances (PFASs) are widely used in industrial and consumer products. Due to their environmental persistence and bioaccumulation, PFASs can be found in the blood of humans and wild animals all over the world. Various fluorinated alternatives such as GenX have been developed to replace the long-chain PFASs, but there is limited information about their potential toxicity. Methods:The current study developed blood culture protocols to assess the response to toxic compounds in the marsupial, Monodelphis domestica. After whole-blood culture conditions were tested and optimized, changes in gene expression in response to PFOA and GenX treatment were assessed. Results: More than 10,000 genes were expressed in the blood transcriptomes with and without treatment. Both PFOA and GenX treatment led to significant changes in the whole blood culture transcriptomes. A total of 578 and 148 differentially expressed genes (DEGs) were detected in the PFOA and GenX treatment groups, 32 of which overlapped. Pathway enrichment analysis revealed that DEGs involved in developmental processes were upregulated after PFOA exposure, while those enriched for metabolic and immune system processes were downregulated. GenX exposure upregulated genes associated with fatty acid transport pathways and inflammatory processes, which is consistent with previous studies using rodent models. Discussion: To our knowledge, this study is the first to investigate the effect of PFASs in a marsupial model. The findings provide supportive evidence for significant transcriptomic alterations, suggesting that this mammalian model may provide a mechanism for exploring the potential toxicity of PFOA and GenX.

Origin and Evolution of Marsupial-specific Imprinting Clusters Through Lineage-specific Gene Duplications and Acquisition of Promoter Differential Methylation.

Genomic imprinting is a parent-of-origin-specific expression phenomenon that plays fundamental roles in many biological processes. In animals, imprinting is only observed in therian mammals, with ∼200 imprinted genes known in humans and mice. The imprinting pattern in marsupials has been minimally investigated by examining orthologs to known eutherian imprinted genes. To identify marsupial-specific imprinting in an unbiased way, we performed RNA-seq studies on samples of fetal brain and placenta from the reciprocal cross progeny of two laboratory opossum stocks. We inferred allele-specific expression for >3,000 expressed genes and discovered/validated 13 imprinted genes, including three previously known imprinted genes, Igf2r, Peg10, and H19. We estimate that marsupials imprint ∼60 autosomal genes, which is a much smaller set compared with eutherians. Among the nine novel imprinted genes, three noncoding RNAs have no known homologs in eutherian mammals, while the remaining genes have important functions in pluripotency, transcription regulation, nucleolar homeostasis, and neural differentiation. Methylation analyses at promoter CpG islands revealed differentially methylated regions in five of these marsupial-specific imprinted genes, suggesting that differential methylation is a common mechanism in the epigenetic regulation of marsupial imprinting. Clustering and co-regulation were observed at marsupial imprinting loci Pou5f3-Npdc1 and Nkrfl-Ipncr2, but eutherian-type multi-gene imprinting clusters were not detected. Also differing from eutherian mammals, the brain and placenta imprinting profiles are remarkably similar in opossums, presumably due to the shared origin of these organs from the trophectoderm. Our results contribute to a fuller understanding of the origin, evolution, and mechanisms of genomic imprinting in therian mammals.

Effect of mineral oil as a lubricant to collect feces from cats for microbiome studies.

BACKGROUND: Fecal specimens are critical for disease screening, diagnosis, and gut microbiome research. For domestic cats, lubricants are often necessary to obtain a sufficient quantity of sample. However, the effect of lubrication on feline microbiome analysis has not been assessed. OBJECTIVES: To evaluate if lubrication using mineral oil during cat feces sample collection affects the DNA extraction, metagenomic sequencing yield, and the microbial composition and diversity in subsequent gut microbiome analyses. ANIMALS: Eight 6-year-old male, neutered, domestic short-haired cats housed in a research facility. METHODS: Cohort study. The gut microbiomes were investigated for fecal sample collection with and without lubrication using whole-genome shotgun metagenomic sequencing. RESULTS: Fecal specimens were collected using a fecal loop under sedation without lubrication and with mineral oil lubrication. There were no significant differences between the 2 groups in the microbial DNA yield in ng/mg fecal sample (75.75 [25.8-125.7] vs 60.72 [33.49-87.95], P = .95), metagenomic sequencing yield in Gbp (10.31 [6.29-14.32] vs 13.53 [12.04-15.02], P = .2), proportion of host contamination (0.1 [0.02-0.18] vs 0.15 [0-0.3], P = .84), relative taxonomy abundance (P > .8), or the number of microbial genes covered (408 132 [341 556-474 708] vs 425 697 [358 505-492 889], P = .31). CONCLUSIONS AND CLINICAL IMPORTANCE: Fecal sampling with mineral oil lubrication did not change the microbial DNA extraction yield, metagenomic sequencing yield, level of host contamination, the microbial composition and diversity in subsequent gut microbiome analyses. Here we reported a proven cat-friendly protocol for fecal sample collection in clinical and research setting for gut microbiome analyses.

Whole-Genome Shotgun Metagenomic Sequencing Reveals Distinct Gut Microbiome Signatures of Obese Cats.

Overweight and obesity are growing health problems in domestic cats, increasing the risks of insulin resistance, lipid dyscrasias, neoplasia, cardiovascular disease, and decreasing longevity. The signature of obesity in the feline gut microbiota has not been studied at the whole-genome metagenomic level. We performed whole-genome shotgun metagenomic sequencing in the fecal samples of eight overweight/obese and eight normal cats housed in the same research environment. We obtained 271 Gbp of sequences and generated a 961-Mbp de novo reference contig assembly, with 1.14 million annotated microbial genes. In the obese cat microbiome, we discovered a significant reduction in microbial diversity (P < 0.01) and Firmicutes abundance (P = 0.005), as well as decreased Firmicutes/Bacteroidetes ratios (P = 0.02), which is the inverse of obese human/mouse microbiota. Linear discriminant analysis and quantitative PCR (qPCR) validation revealed significant increases of Bifidobacterium sp., Olsenella provencensis, Dialister sp.CAG:486, and Campylobacter upsaliensis as the hallmark of obese microbiota among 400 enriched species, whereas 1,525 bacterial species have decreased abundance in the obese microbiome. Phascolarctobacterium succinatutens and an uncharacterized Erysipelotrichaceae bacterium are highly abundant (>0.05%) in the normal gut with over 400-fold depletion in the obese microbiome. Fatty acid synthesis-related pathways are significantly overrepresented in the obese compared with the normal cat microbiome. In conclusion, we discovered dramatically decreased microbial diversity in obese cat gut microbiota, suggesting potential dysbiosis. A panel of seven significantly altered, highly abundant species can serve as a microbiome indicator of obesity. Our findings in the obese cat microbiome composition, abundance, and functional capacities provide new insights into feline obesity. IMPORTANCE Obesity affects around 45% of domestic cats, and licensed drugs for treating feline obesity are lacking. Physical exercise and calorie restrictions are commonly used for weight loss but with limited efficacy. Through comprehensive analyses of normal and obese cat gut bacteria flora, we identified dramatic shifts in the obese gut microbiome, including four bacterial species significantly enriched and two species depleted in the obese cats. The key bacterial community and functional capacity alterations discovered from this study will inform new weight management strategies for obese cats, such as evaluations of specific diet formulas that alter the microbiome composition, and the development of prebiotics and probiotics that promote the increase of beneficial species and the depletion of obesity-associated species. Interestingly, these bacteria identified in our study were also reported to affect the weight loss success in human patients, suggesting translational potential in human obesity.

A Novel Spherical Crystallization Method Using Pickering Emulsions.

Spherical crystallization is a promising process intensification technique, where surfactant is an important ingredient in formulation but needs to be used carefully due to toxicological reasons. This work proposes to adopt colloidal particles stabilized Pickering emulsions for spherical crystallization, in order to eliminate or reduce the surfactant use. A representative system is selected for study, where silica nanoparticles are prepared to stabilize emulsions and evaporative crystallization of ibuprofen is carried out. Depletion attraction is exploited to improve the Pickering emulsion stability for better confining on crystallization with two depletants PEG and PVA tested. Crystal products from the emulsions prepared with silica nanoparticles and the non-ionic surfactant Tween 20 are compared. The results show that depletion attraction is helpful for producing stable Pickering emulsions with high dispersed phase fraction and mono-dispersed ibuprofen spherical agglomerates. Silica nanoparticles contribute to reduced induction time by boosting heterogeneous nucleation and mitigate secondary agglomeration possibly by steric effects.

Genetic and genomic architecture in eight strains of the laboratory opossum Monodelphis domestica.

The gray short-tailed opossum (Monodelphis domestica) is an established laboratory-bred marsupial model for biomedical research. It is a critical species for comparative genomics research, providing the pivotal phylogenetic outgroup for studies of derived vs ancestral states of genomic/epigenomic characteristics for eutherian mammal lineages. To characterize the current genetic profile of this laboratory marsupial, we examined 79 individuals from eight established laboratory strains. Double digest restriction site-associated DNA sequencing and whole-genome resequencing experiments were performed to investigate the genetic architecture in these strains. A total of 66,640 high-quality single nucleotide polymorphisms (SNPs) were identified. We analyzed SNP density, average heterozygosity, nucleotide diversity, and population differentiation parameter Fst within and between the eight strains. Principal component and population structure analysis clearly resolve the strains at the level of their ancestral founder populations, and the genetic architecture of these strains correctly reflects their breeding history. We confirmed the successful establishment of the first inbred laboratory opossum strain LSD (inbreeding coefficient F > 0.99) and a nearly inbred strain FD2M1 (0.98 < F < 0.99), each derived from a different ancestral background. These strains are suitable for various experimental protocols requiring controlled genetic backgrounds and for intercrosses and backcrosses that can generate offspring with informative SNPs for studying a variety of genetic and epigenetic processes. Together with recent advances in reproductive manipulation and CRISPR/Cas9 techniques for Monodelphis domestica, the existence of distinctive inbred strains will enable genome editing on different genetic backgrounds, greatly expanding the utility of this marsupial model for biomedical research.

Placental mtDNA copy number and methylation in association with macrosomia in healthy pregnancy.

INTRODUCTION: Fetal growth and development depend on metabolic energy from placental mitochondria. However, the impact of placental mitochondria on the occurrence of macrosomia remains unclear. We aimed to explore the association between macrosomia without gestational diabetes mellitus (non-GDM) and changes in placental mitochondrial DNA (mtDNA) copy number and methylation. METHODS: Fifty-four newborns with macrosomia and 54 normal birthweight controls were enrolled in this study. Placental mtDNA copy number and mRNA expression of nuclear genes related to mitochondrial replication or ATP synthesis-related genes were measured by real-time quantitative polymerase chain reaction (qPCR). Methylation levels of the non-coding regulatory region D-loop and ATP synthesis-related genes were detected by targeted bisulfite sequencing. RESULTS: Newborns with macrosomia had lower placental mtDNA copy number and higher methylation rates of the CpG15 site in the D-loop region (D-CpG15) and CpG6 site in the cytochrome C oxidase III (COX3) gene (COX3-CpG6) than normal birth weight newborns. After adjusting for potential covariates (gestational age, prepregnancy BMI, and infant sex), decreased placental mtDNA copy number (adjusted odds ratio [aOR] = 2.09, 95% confidence interval [CI] 1.03-4.25), elevated methylation rate of D-CpG15 (aOR = 2.06, 95% CI 1.03-4.09) and COX3-CpG6 (aOR = 2.13, 95% CI 1.08-4.20) remained significantly associated with a higher risk of macrosomia. DISCUSSION: Reduced mtDNA copy number and increased methylation levels of specific loci at mtDNA would increase the risk of macrosomia. However, the detailed molecular mechanism needs further identification.

Genomic and in vitro pharmacodynamic analysis of rifampicin resistance in multidrug-resistant canine Staphylococcus pseudintermedius isolates.

BACKGROUND: Antimicrobial resistance is a growing concern in canine Staphylococcus pseudintermedius dermatitis. Treatment with rifampicin (RFP) is considered only in meticillin-resistant and multidrug-resistant S. pseudintermedius (MDR-MRSP). HYPOTHESIS/OBJECTIVES: To determine an optimal RFP dosing for MDR-MRSP treatment without induction of RFP resistance and identify causal mutations for antimicrobial resistance. METHODS AND MATERIALS: Time-kill assays were performed in a control isolate and three MDR-MRSP isolates at six clinically relevant concentrations [32 to 1,024 × MIC (the minimum inhibitory concentration)]. Whole-genome resequencing and bioinformatic analysis were performed in the resistant strains developed in this assay. RESULTS: The genomic analysis identified nine antimicrobial resistance genes (ARGs) in MDR-MRSP isolates, which are responsible for resistance to seven classes of antibiotics. RFP activity against all four isolates was consistent with a time-dependent and bacteriostatic response. RFP resistance was observed in six of the 28 time-kill assays, including concentrations 64 × MIC in MDR-MRSP1 isolates at 24 h, 32 × MIC in MDR-MRSP2 at 48 h, 32 × MIC in MDR-MRSP3 at 48 h and 256 × MIC in MDR-MRSP3 at 24 h. Genome-wide mutation analyses in these RFP-resistant strains discovered the causal mutations in the coding region of the rpoB gene. CONCLUSIONS AND CLINICAL RELEVANCE: A study has shown that 6 mg/kg per os results in plasma concentrations of 600-1,000 × MIC of S. pseudintermedius. Based on our data, this dose should achieve the minimum MIC (×512) to prevent RFP resistance development; therefore, we recommend a minimum daily dose of 6 mg/kg for MDR-MRSP pyoderma treatment when limited antibiotic options are available.

Phylogenomic Analysis of Wolbachia Strains Reveals Patterns of Genome Evolution and Recombination.

Wolbachia are widespread intracellular bacteria that mediate many important biological processes in arthropod species. In this study, we identified 210 conserved single-copy genes in 33 genome-sequenced Wolbachia strains in the A-F supergroups. Phylogenomic analyses with these core genes indicate that all 33 Wolbachia strains maintain the supergroup relationship, which was classified previously based on the multilocus sequence typing (MLST) genes. Using an interclade recombination screening method, 14 inter-supergroup recombination events were discovered in six genes (2.9%) among 210 single-copy orthologs. This finding suggests a relatively low frequency of intergroup recombination. Interestingly, they have occurred not only between A and B supergroups (nine events) but also between A and E supergroups (five events). Maintenance of such transfers suggests possible roles in Wolbachia infection-related functions. Comparisons of strain divergence using the five genes of the MLST system show a high correlation (Pearson correlation coefficient r = 0.98) between MLST and whole-genome divergences, indicating that MLST is a reliable method for identifying related strains when whole-genome data are not available. The phylogenomic analysis and the identified core gene set in our study will serve as a valuable foundation for strain identification and the investigation of recombination and genome evolution in Wolbachia.

Draft Genome Assemblies of Two Staphylococcus pseudintermedius Strains Isolated from Canine Skin Biopsy Specimens.

Staphylococcus pseudintermedius is a Gram-positive bacterial species highly relevant to animal and human health. In this study, we report the draft genome sequences of two clinical isolates of S. pseudintermedius from canine skin biopsy specimens at the Dermatology Service of the Auburn University Small Animal Teaching Hospital.

Genome Assembly of the A-Group Wolbachia in Nasonia oneida Using Linked-Reads Technology.

Wolbachia are obligate intracellular bacteria which commonly infect various nematode and arthropod species. Genome sequences have been generated from arthropod samples following enrichment for the intracellular bacteria, and genomes have also been assembled from arthropod whole-genome sequencing projects. However, these methods remain challenging for infections that occur at low titers in hosts. Here we report the first Wolbachia genome assembled from host sequences using 10× Genomics linked-reads technology. The high read depth attainable by this method allows for recovery of intracellular bacteria that are at low concentrations. Based on the depth differences (714× for the insect and 59× for the bacterium), we assembled the genome of a Wolbachia in the parasitoid jewel wasp species Nasonia oneida. The final draft assembly consists of 1,293, 06 bp in 47 scaffolds with 1,114 coding genes and 97.01% genome completeness assessed by checkM. Comparisons of the five Multi Locus Sequence Typing genes revealed that the sequenced Wolbachia genome is the A1 strain (henceforth wOneA1) previously reported in N. oneida. Pyrosequencing confirms that the wasp strain lacks A2 and B types previously detected in this insect, which were likely lost during laboratory culturing. Assembling bacterial genomes from host genome projects can provide an effective method for sequencing bacterial genomes, even when the infections occur at low density in sampled tissues.

Speciation transformation of arsenic in abalone viscera hydrolysate fraction: In vitro digestion and in vivo metabolism.

Speciation transformation of arsenic in the abalone viscera hydrolysate fraction (AVHF) was evaluated using in vitro and in vivo methods to determine its safety given that AVHF is rich in arsenic. The dimethylarsinic acid (DMA) proportion and some free amino acid contents increased, whereas arsenobetaine (AB) proportion decreased when AVHF was digested by pepsin. However, molecular weight distribution was unchanged, and no obvious changes were found in the intestinal medium. In the single-dose experiment, the AB concentration on the mouse plasma rapidly increased, which reached up to 12.53 ng/mL in 2 h after the administration of AVHF (10 g/kg body weight) and reduced to half of the maximum at 8 h after administration. Furthermore, alanine (Ala) content in the urine of mice increased at 8 h after AVHF administration, suggesting that Ala might be chelated with arsenic and could not be absorbed well. Long-term experiments showed that AB was not accumulated in mice tissue/organ. However, some AB could be converted into DMA, which was mainly accumulated in mice hair. The in vivo experiments also suggested that the AVHF is safe as health food.

SIK2 regulates fasting-induced PPARα activity and ketogenesis through p300.

Fatty acid oxidation and subsequent ketogenesis is one of the major mechanisms to maintain hepatic lipid homeostasis under fasting conditions. Fasting hormone glucagon has been shown to stimulate ketone body production through activation of PPARα; however, the signal pathway linking glucagon to PPARα is largely undiscovered. Here we report that a SIK2-p300-PPARα cascade mediates glucagon's effect on ketogenesis. p300 interacts with PPARα through a conserved LXXLL motif and enhances its transcriptional activity. SIK2 disrupts p300-PPARα interaction by direct phosphorylation of p300 at Ser89, which in turn decreases PPARα-mediated ketogenic gene expression. Moreover, SIK2 phosphorylation defective p300 (p300 S89A) shows increased interaction with PPARα and abolishes suppression of SIK2 on PPARα-mediated ketogenic gene expression in liver. Taken together, our results unveil the signal pathway that mediates fasting induced ketogenesis to maintain hepatic lipid homeostasis.