The Impact of Essential Amino Acids on the Gut Microbiota of Broiler Chickens.

The research involving the beneficial aspects of amino acids being added to poultry feed pertaining to performance, growth, feed intake, and feed conversion ratio is extensive. Yet currently the effects of amino acids on the gut microbiota aren't fully understood nor have there been many studies executed in poultry to explain the relationship between amino acids and the gut microbiota. The overall outcome of health has been linked to bird gut health due to the functionality of gastrointestinal tract (GIT) for digestion/absorption of nutrients as well as immune response. These essential functions of the GI are greatly driven by the resident microbiota which produce metabolites such as butyrate, propionate, and acetate, providing the microbiota a suitable and thrive driven environment. Feed, age, the use of feed additives and pathogenic infections are the main factors that have an effect on the microbial community within the GIT. Changes in these factors may have potential effects on the gut microbiota in the chicken intestine which in turn may have an influence on health essentially affecting growth, feed intake, and feed conversion ratio. This review will highlight limited research studies that investigated the possible role of amino acids in the gut microbiota composition of poultry.

Advances in the Xenopus immunome: Diversification, expansion, and contraction.

Xenopus is a genus of African clawed frogs including two species, X. tropicalis and X. laevis that are extensively used in experimental biology, immunology, and biomedical studies. The availability of fully sequenced and annotated Xenopus genomes is strengthening genome-wide analyses of gene families and transgenesis to model human diseases. However, inaccuracies in genome annotation for genes involved in the immune system (i.e., immunome) hamper immunogenetic studies. Furthermore, advanced genome technologies (e.g., single-cell and RNA-Seq) rely on well-annotated genomes. The annotation problems of Xenopus immunome include a lack of established orthology across taxa, merged gene models, poor representation in gene pages on Xenbase, misannotated genes and missing gene IDs. The Xenopus Research Resource for Immunobiology in collaboration with Xenbase and a group of investigators are working to resolve these issues in the latest versions of genome browsers. In this review, we summarize the current problems of previously misannotated gene families that we have recently resolved. We also highlight the expansion, contraction, and diversification of previously misannotated gene families.

Molecular diversity and functional implication of amphibian interferon complex: Remarking immune adaptation in vertebrate evolution.

Cross-species comparison of vertebrate genomes has unraveled previously unknown complexities of interferon (IFN) systems in amphibian species. Recent genomic curation revealed that amphibian species have evolved expanded repertoires of four types of intron-containing IFN genes akin to those seen in jawed fish, intronless type I IFNs and intron-containing type III IFNs akin to those seen in amniotes, as well as uniquely intronless type III IFNs. This appears to be the case with at least ten analyzed amphibian species; with distinct species encoding diverse repertoires of these respective IFN gene subsets. Amphibians represent a key stage in vertebrate evolution, and in this context offer a unique perspective into the divergent and converged pathways leading to the emergence of distinct IFN families and groups. Recent studies have begun to unravel the roles of amphibian IFNs during these animals' immune responses in general and during their antiviral responses, in particular. However, the pleiotropic potentials of these highly expanded amphibian IFN repertoires warrant further studies. Based on recent reports and our omics analyses using Xenopus models, we posit that amphibian IFN complex may have evolved novel functions, as indicated by their extensive molecular diversity. Here, we provide an overview and an update of the present understanding of the amphibian IFN complex in the context of the evolution of vertebrate immune systems. A greater understanding of the amphibian IFN complex will grant new perspectives on the evolution of vertebrate immunity and may yield new measures by which to counteract the global amphibian declines.

Immunometabolic Dysregulation at the Intersection of Obesity and COVID-19.

Obesity prevails worldwide to an increasing effect. For example, up to 42% of American adults are considered obese. Obese individuals are prone to a variety of complications of metabolic disorders including diabetes mellitus, hypertension, cardiovascular disease, and chronic kidney disease. Recent meta-analyses of clinical studies in patient cohorts in the ongoing coronavirus-disease 2019 (COVID-19) pandemic indicate that the presence of obesity and relevant disorders is linked to a more severe prognosis of COVID-19. Given the significance of obesity in COVID-19 progression, we provide a review of host metabolic and immune responses in the immunometabolic dysregulation exaggerated by obesity and the viral infection that develops into a severe course of COVID-19. Moreover, sequela studies of individuals 6 months after having COVID-19 show a higher risk of metabolic comorbidities including obesity, diabetes, and kidney disease. These collectively implicate an inter-systemic dimension to understanding the association between obesity and COVID-19 and suggest an interdisciplinary intervention for relief of obesity-COVID-19 complications beyond the phase of acute infection.

Virus-Targeted Transcriptomic Analyses Implicate Ranaviral Interaction with Host Interferon Response in Frog Virus 3-Infected Frog Tissues.

Ranaviruses (Iridoviridae), including Frog Virus 3 (FV3), are large dsDNA viruses that cause devastating infections globally in amphibians, fish, and reptiles, and contribute to catastrophic amphibian declines. FV3's large genome (~105 kb) contains at least 98 putative open reading frames (ORFs) as annotated in its reference genome. Previous studies have classified these coding genes into temporal classes as immediate early, delayed early, and late viral transcripts based on their sequential expression during FV3 infection. To establish a high-throughput characterization of ranaviral gene expression at the genome scale, we performed a whole transcriptomic analysis (RNA-Seq) using total RNA samples containing both viral and cellular transcripts from FV3-infected Xenopus laevis adult tissues using two FV3 strains, a wild type (FV3-WT) and an ORF64R-deleted recombinant (FV3-∆64R). In samples from the infected intestine, liver, spleen, lung, and especially kidney, an FV3-targeted transcriptomic analysis mapped reads spanning the full-genome coverage at ~10× depth on both positive and negative strands. By contrast, reads were only mapped to partial genomic regions in samples from the infected thymus, skin, and muscle. Extensive analyses validated the expression of almost all of the 98 annotated ORFs and profiled their differential expression in a tissue-, virus-, and temporal class-dependent manner. Further studies identified several putative ORFs that encode hypothetical proteins containing viral mimicking conserved domains found in host interferon (IFN) regulatory factors (IRFs) and IFN receptors. This study provides the first comprehensive genome-wide viral transcriptome profiling during infection and across multiple amphibian host tissues that will serve as an instrumental reference. Our findings imply that Ranaviruses like FV3 have acquired previously unknown molecular mimics, interfering with host IFN signaling during evolution.

Targeted Transcriptomics of Frog Virus 3 in Infected Frog Tissues Reveal Non-Coding Regulatory Elements and microRNAs in the Ranaviral Genome and Their Potential Interaction with Host Immune Response.

Background: Frog Virus 3 (FV3) is a large dsDNA virus belonging to Ranaviruses of family Iridoviridae. Ranaviruses infect cold-blood vertebrates including amphibians, fish and reptiles, and contribute to catastrophic amphibian declines. FV3 has a genome at ~105 kb that contains nearly 100 coding genes and 50 intergenic regions as annotated in its reference genome. Previous studies have mainly focused on coding genes and rarely addressed potential non-coding regulatory role of intergenic regions. Results: Using a whole transcriptomic analysis of total RNA samples containing both the viral and cellular transcripts from FV3-infected frog tissues, we detected virus-specific reads mapping in non-coding intergenic regions, in addition to reads from coding genes. Further analyses identified multiple cis-regulatory elements (CREs) in intergenic regions neighboring highly transcribed coding genes. These CREs include not only a virus TATA-Box present in FV3 core promoters as in eukaryotic genes, but also viral mimics of CREs interacting with several transcription factors including CEBPs, CREBs, IRFs, NF-κB, and STATs, which are critical for regulation of cellular immunity and cytokine responses. Our study suggests that intergenic regions immediately upstream of highly expressed FV3 genes have evolved to bind IRFs, NF-κB, and STATs more efficiently. Moreover, we found an enrichment of putative microRNA (miRNA) sequences in more than five intergenic regions of the FV3 genome. Our sequence analysis indicates that a fraction of these viral miRNAs is targeting the 3'-UTR regions of Xenopus genes involved in interferon (IFN)-dependent responses, including particularly those encoding IFN receptor subunits and IFN-regulatory factors (IRFs). Conclusions: Using the FV3 model, this study provides a first genome-wide analysis of non-coding regulatory mechanisms adopted by ranaviruses to epigenetically regulate both viral and host gene expressions, which have co-evolved to interact especially with the host IFN response.

Effects of Lactobacillus reuteri and Streptomyces coelicolor on Growth Performance of Broiler Chickens.

There are well documented complications associated with the continuous use of antibiotics in the poultry industry. Over the past few decades, probiotics have emerged as viable alternatives to antibiotics; however, most of these candidate probiotic microorganisms have not been fully evaluated for their effectiveness as potential probiotics for poultry. Recent evaluation of a metagenome of broiler chickens in our laboratory revealed a prevalence of Lactobacillus reuteri (L. reuteri) and Actinobacteria class of bacteria in their gastrointestinal tract. In this study Lactobacillus reuteri and Streptomyces coelicolor (S. coelicolor) were selected as probiotic bacteria, encapsulated, and added into broiler feed at a concentration of 100 mg/kg of feed. In an 8-week study, 240 one day-old chicks were randomly assigned to four dietary treatments. Three dietary treatments contained two probiotic bacteria in three different proportions (L. reuteri and S. coelicolor individually at 100 ppm, and mixture of L. reuteri and S. coelicolor at 50 ppm each). The fourth treatment had no probiotic bacteria and it functioned as the control diet. L. reuteri and S. coelicolor were added to the feed by using wheat middlings as a carrier at a concentration of 100 ppm (100 mg/kg). Chickens fed diets containing L. reuteri and S. coelicolor mixture showed 2% improvement in body weight gain, 7% decrease in feed consumption, and 6-7% decrease in feed conversion ratios. This research suggests that L. reuteri and S. coelicolor have the potential to constitute probiotics in chickens combined or separately, depending on the desired selection of performance index.

Recent Advances in the Application of CRISPR/Cas9 Gene Editing System in Poultry Species.

CRISPR/Cas9 system genome editing is revolutionizing genetics research in a wide spectrum of animal models in the genetic era. Among these animals, is the poultry species. CRISPR technology is the newest and most advanced gene-editing tool that allows researchers to modify and alter gene functions for transcriptional regulation, gene targeting, epigenetic modification, gene therapy, and drug delivery in the animal genome. The applicability of the CRISPR/Cas9 system in gene editing and modification of genomes in the avian species is still emerging. Up to date, substantial progress in using CRISPR/Cas9 technology has been made in only two poultry species (chicken and quail), with chicken taking the lead. There have been major recent advances in the modification of the avian genome through their germ cell lineages. In the poultry industry, breeders and producers can utilize CRISPR-mediated approaches to enhance the many required genetic variations towards the poultry population that are absent in a given poultry flock. Thus, CRISPR allows the benefit of accessing genetic characteristics that cannot otherwise be used for poultry production. Therefore CRISPR/Cas9 becomes a very powerful and robust tool for editing genes that allow for the introduction or regulation of genetic information in poultry genomes. However, the CRISPR/Cas9 technology has several limitations that need to be addressed to enhance its use in the poultry industry. This review evaluates and provides a summary of recent advances in applying CRISPR/Cas9 gene editing technology in poultry research and explores its potential use in advancing poultry breeding and production with a major focus on chicken and quail. This could aid future advancements in the use of CRISPR technology to improve poultry production.

Transcriptome Analysis and Expression of Selected Cationic Amino Acid Transporters in the Liver of Broiler Chicken Fed Diets with Varying Concentrations of Lysine.

Amino acids are known to play a key role in gene expression regulation. Amino acid signaling is mediated via two pathways: the mammalian target of rapamycin complex 1 (mTORC1) and the amino acid responsive (AAR) pathways. Cationic amino acid transporters (CATs) are crucial in these pathways due to their sensing, signaling and transport functions. The availability of certain amino acids plays a key role in the intake of other amino acids, hence affecting growth in young birds. However, the specific mechanism for regulating lysine transport for growth is not clear. In this study, we analyze the transcriptome profiles and mRNA expression of selected cationic amino acid transporters in the livers of broilers fed low and high lysine diets. Birds consumed high-lysine (1.42% lysine) or low-lysine (0.85% lysine) diets while the control group consumed 1.14% lysine diet. These concentrations of lysine represent 125% (high lysine), 75% (low lysine) and 100% (control), respectively, of the National Research Council's (NRC) recommendation for broiler chickens. After comparing the two groups, 210 differentially expressed genes (DEGs) were identified (fold change >1 and false discovery rate (FDR) <0.05). When comparing the high lysine and the low lysine treatments, there were 67 upregulated genes and 143 downregulated genes among these DEGs. Analysis of Kyoto Encyclopedia of Genes and Genomes (KEGG) and the Gene Ontology (GO) enrichment analysis show that cellular growth, lipid metabolism and lysine metabolism pathways were among the significantly enriched pathways. This study contributes to a better understanding of the potential molecular mechanisms underlying the correlation between lysine intake, body weight gain (BWG) and feed intake (FI) in broiler chickens. Moreover, the DEGs obtained in this study may be used as potential candidate genes for further investigation of broiler growth customized responses to individualized nutrients such as amino acids.

Expression of lysine-mediated neuropeptide hormones controlling satiety and appetite in broiler chickens.

Lysine is the second most limiting amino acid after methionine and is considered the most limiting amino acid for growth in poultry. Lysine requirement for broiler chickens has changed over the years. Leptin and adiponectin represent 2 adipokines that mediate metabolism by eliciting satiety effects whereas ghrelin peptide hormone influences appetite. We hypothesize that this affects growth performance of chicks. This study evaluates the effect of varying dietary lysine homeostasis on performance of broiler chickens through satiety- and appetite-mediating hormones. In 3 replications, 270 one-day-old chicks were reared for 8 wk feeding on diets comprising 0.85, 1.14, and 1.42% lysine during the starter period and 0.75, 1.00, and 1.25% lysine during the grower period. These concentrations of lysine represent 75% (low lysine), 100% (control), and 125% (high lysine) of National Research Council recommendation for broiler chickens. Feed and water were provided for ad libitum consumption. At 8 wk of age, liver, pancreas, brain, and hypothalamus tissues were collected from 18 birds randomly selected from each treatment, snap frozen in liquid nitrogen, and stored at -80°C until use. Total RNA was extracted, and cDNA was synthesized for quantitative real-time PCR assays. Low lysine concentration caused slow growth and high mortality. There was significant upregulation of ghrelin in the hypothalamus and pancreas, and leptin and adiponectin in the hypothalamus and liver, and downregulation of ghrelin in the intestines. At low lysine concentrations, adiponectin was not expressed in both pancreas and intestines. High lysine concentration exhibited increased growth, upregulation of ghrelin in the liver, and downregulation of ghrelin in the intestines, and both adiponectin and leptin in the liver. The expression of ghrelin was negatively correlated with the expression of adiponectin and leptin (P < 0.05) in the liver, hypothalamus, and pancreas. Expression of leptin was positively correlated with adiponectin in the hypothalamus and liver (P < 0.05), exhibiting satiety effects when the concentrations of lysine were low.