Multi-omics strategy reveals potential role of antimicrobial resistance and virulence factor genes responsible for Simmental diarrheic calves caused by Escherichia coli.

Escherichia coli (E. coli) is reported to be an important pathogen associated with calf diarrhea. Antibiotic resistance genes (ARGs) and virulence factor genes (VFGs) pose a considerable threat to both animal and human health. However, little is known about the characterization of ARGs and VFGs presented in the gut microbiota of diarrheic calves caused by E. coli. In this study, we used multi-omics strategy to analyze the ARG and VFG profiles of Simmental calves with diarrhea caused by E. coli K99. We found that gut bacterial composition and their microbiome metabolic functions varied greatly in diarrheic calves compared to healthy calves. In total, 175 ARGs were identified, and diarrheal calves showed a significantly higher diversity and abundance of ARGs than healthy calves. Simmental calves with diarrhea showed higher association of VFGs with pili function, curli assembly, and ferrienterobactin transport of E. coli. Co-occurrence patterns based on Pearson correlation analysis revealed that E. coli had a highly significant (P < 0.0001) correlation coefficient (>0.8) with 16 ARGs and 7 VFGs. Metabolomics analysis showed that differentially expressed metabolites in Simmental calves with diarrhea displayed a high correlation with the aforementioned ARGs and VFGs. Phylotype analysis of E. coli genomes showed that the predominant phylogroup B1 in diarrheic Simmental calves was associated with 10 ARGs and 3 VFGs. These findings provide an overview of the diversity and abundance of the gut microbiota in diarrheic calves caused by E. coli and pave the way for further studies on the mechanisms of antibiotic resistance and virulence in the calves affected with diarrhea.IMPORTANCESimmental is a well-recognized beef cattle breed worldwide. They also suffer significant economic losses due to diarrhea. In this study, fecal metagenomic analysis was applied to characterize the antibiotic resistance gene (ARG) and virulence factor gene (VFG) profiles of diarrheic Simmental calves. We identified key ARGs and VFGs correlated with Escherichia coli isolated from Simmental calves. Additionally, metabolomics analysis showed that differentially expressed metabolites in Simmental calves with diarrhea displayed a high correlation with the aforementioned ARGs and VFGs. Our findings provide an insight into the diversity and abundance of the gut microbiota in diarrheic calves caused by Escherichia coli and pave the way for further studies on the mechanisms of antibiotic resistance and virulence in the diarrheal calves from cattle hosts.

The Identification of Functional Genes Affecting Fat-Related Meat Traits in Meat-Type Pigeons Using Double-Digest Restriction-Associated DNA Sequencing and Molecular Docking Analysis.

The Chinese indigenous Shiqi (SQ) pigeon and the imported White King (WK) pigeon are two meat-type pigeon breeds of economical and nutritional importance in China. They displayed significant differences in such meat quality traits as intramuscular fat (IMF) content and fatty acid (FA) compositions in the breast muscles. In this study, we aimed to screen candidate genes that could affect fat-related meat quality traits in meat-type pigeons. We investigated the polymorphic variations at the genomic level using double-digest restriction-associated DNA (ddRAD) sequencing in 12 squabs of SQ and WK pigeons that exhibited significant inter-breed differences in IMF content as well as FA and amino acid compositions in the breast muscles, and screened candidate genes influencing fat-related traits in squabs through gene ontology analysis and pathway analysis. By focusing on 6019 SNPs, which were located in genes with correct annotations and had the potential to induce changes in the encoded proteins, we identified 19 genes (ACAA1, ACAA2, ACACB, ACADS, ACAT1, ACOX3, ACSBG1, ACSBG2, ACSL1, ACSL4, ELOVL6, FADS1, FADS2, HACD4, HADH, HADHA, HADHB, MECR, OXSM) as candidate genes that could affect fat-related traits in squabs. They were significantly enriched in the pathways of FA metabolism, degradation, and biosynthesis (p < 0.05). Results from molecular docking analysis further revealed that three non-synonymous amino acid alterations, ACAA1(S357N), ACAA2(T234I), and ACACB(H1418N), could alter the non-bonding interactions between the enzymatic proteins and their substrates. Since ACAA1, ACAA2, and ACACB encode rate-limiting enzymes in FA synthesis and degradation, alterations in the enzyme-substrate binding affinity may subsequently affect the catalytic efficiency of enzymes. We suggested that SNPs in these three genes were worthy of further investigation into their roles in explaining the disparities in fat-related traits in squabs.

MAEL gene contributes to bovine testicular development through the m5C-mediated splicing.

Knowledge of RNA molecules regulating testicular development and spermatogenesis in bulls is essential for elite bull selection and an ideal breeding program. Herein, we performed direct RNA sequencing (DRS) to explore the functional characterization of RNA molecules produced in the testicles of 9 healthy Simmental bulls at three testicular development stages (prepuberty, puberty, and postpuberty). We identified 5,043 differentially expressed genes associated with testicular weight. These genes exhibited more alternative splicing at sexual maturity, particularly alternative 3' (A3) and 5' (A5) splice sites usage and exon skipping (SE). The expression of hub genes in testicular developmental stages was also affected by both m6A and m5C RNA modifications. We found m5C-mediated splicing events significantly (p < 0.05) increased MAEL gene expression at the isoform level, likely promoting spermatogenesis. Our findings highlight the complexity of RNA processing and expression as well as the regulation of transcripts involved in testicular development and spermatogenesis.

Heat Stress Induces Shifts in the Rumen Bacteria and Metabolome of Buffalo.

Exposure to the stress (HS) negatively affects physiology, performance, reproduction and welfare of buffalo. However, the mechanisms by which HS negatively affects rumen bacteria and its associated metabolism in buffalo are not well known yet. This study aimed to gain insight into the adaption of bacteria and the complexity of the metabolome in the rumen of six buffalo during HS using 16S rDNA and gas chromatography metabolomics analyses. HS increased respiratory rate (p < 0.05) and skin temperature (p < 0.01), and it decreased the content of acetic acid (p < 0.05) and butyric acid (p < 0.05) in the rumen. Omics sequencing revealed that the relative abundances of Lachnospirales, Lachnospiraceae, Lachnospiraceae_NK3A20_group and Clostridia_UCG-014 were significantly (p < 0.01) higher under HS than non-heat stress conditions. Several bacteria at different levels, such as Lactobacillales, Streptococcus, Leuconostocaceae and Leissella, were significantly (p < 0.05) more abundant in the rumen of the non-heat stress than HS condition. Thirty-two significantly different metabolites closely related to HS were identified (p < 0.05). Metabolic pathway analysis revealed four key pathways: D-Alanine metabolism; Lysine degradation, Tropane; piperidine and pyridine alkaloid biosynthesis; and Galactose metabolism. In summary, HS may negatively affected rumen fermentation efficiency and changed the composition of rumen community and metabolic function.

Whole-Genome Resequencing of Worldwide Wild and Domestic Sheep Elucidates Genetic Diversity, Introgression, and Agronomically Important Loci.

Domestic sheep and their wild relatives harbor substantial genetic variants that can form the backbone of molecular breeding, but their genome landscapes remain understudied. Here, we present a comprehensive genome resource for wild ovine species, landraces and improved breeds of domestic sheep, comprising high-coverage (∼16.10×) whole genomes of 810 samples from 7 wild species and 158 diverse domestic populations. We detected, in total, ∼121.2 million single nucleotide polymorphisms, ∼61 million of which are novel. Some display significant (P < 0.001) differences in frequency between wild and domestic species, or are private to continent-wide or individual sheep populations. Retained or introgressed wild gene variants in domestic populations have contributed to local adaptation, such as the variation in the HBB associated with plateau adaptation. We identified novel and previously reported targets of selection on morphological and agronomic traits such as stature, horn, tail configuration, and wool fineness. We explored the genetic basis of wool fineness and unveiled a novel mutation (chr25: T7,068,586C) in the 3'-UTR of IRF2BP2 as plausible causal variant for fleece fiber diameter. We reconstructed prehistorical migrations from the Near Eastern domestication center to South-and-Southeast Asia and found two main waves of migrations across the Eurasian Steppe and the Iranian Plateau in the Early and Late Bronze Ages. Our findings refine our understanding of genome variation as shaped by continental migrations, introgression, adaptation, and selection of sheep.

Evolutionary Analysis of OAT Gene Family in River and Swamp Buffalo: Potential Role of SLCO3A1 Gene in Milk Performance.

The organic anion transporter (OAT) family is the subfamily of the solute carrier (SLC) superfamily, which plays a vital role in regulating essential nutrients in milk. However, little is known about the members' identification, evolutionary basis, and function characteristics of OAT genes associated with milk performance in buffalo. Comparative genomic analyses were performed to identify the potential role of buffalo OAT genes in milk performance in this study. The results showed that a total of 10 and 7 OAT genes were identified in river buffalo and swamp buffalo, respectively. These sequences clustered into three groups based on their phylogenetic relationship and had similar motif patterns and gene structures in the same groups. Moreover, the river-specific expansions and homologous loss of OAT genes occurred in the two buffalo subspecies during the evolutionary process. Notably, the duplicated SLCO3A1 gene specific to river buffalo showed higher expression level in mammary gland tissue than that of swamp buffalo. These findings highlight some promising candidate genes that could be potentially utilized to accelerate the genetic progress in buffalo breeding programs. However, the identified candidate genes require further validation in a larger cohort for use in the genomic selection of buffalo for milk production.

Transcriptome-Wide m6A Analysis Provides Novel Insights Into Testicular Development and Spermatogenesis in Xia-Nan Cattle.

Testis is the primary organ of the male reproductive tract in mammals that plays a substantial role in spermatogenesis. Improvement of our knowledge regarding the molecular mechanisms in testicular development and spermatogenesis will be reflected in producing spermatozoa of superior fertility. Evidence showed that N6-Methyladenosine (m6A) plays a dynamic role in post-transcription gene expression regulation and is strongly associated with production traits. However, the role of m6A in bovine testis has not been investigated yet. In this study, we conducted MeRIP-Seq analysis to explore the expression profiles of the m6A and its potential mechanism underlying spermatogenesis in nine bovine testes at three developmental stages (prepuberty, puberty and postpuberty). The experimental animals with triplicate in each stage were chosen based on their semen volume and sperm motility except for the prepuberty bulls and used for testes collection. By applying MeRIP-Seq analysis, a total of 8,774 m6A peaks and 6,206 m6A genes among the studied groups were identified. All the detected peaks were found to be mainly enriched in the coding region and 3'- untranslated regions. The cross-analysis of m6A and mRNA expression exhibited 502 genes with concomitant changes in the mRNA expression and m6A modification. Notably, 30 candidate genes were located in the largest network of protein-protein interactions. Interestingly, four key node genes (PLK4, PTEN, EGR1, and PSME4) were associated with the regulation of mammal testis development and spermatogenesis. This study is the first to present a map of RNA m6A modification in bovine testes at distinct ages, and provides new insights into m6A topology and related molecular mechanisms underlying bovine spermatogenesis, and establishes a basis for further studies on spermatogenesis in mammals.

Paternal Origins and Migratory Episodes of Domestic Sheep.

The domestication and subsequent global dispersal of livestock are crucial events in human history, but the migratory episodes during the history of livestock remain poorly documented [1-3]. Here, we first developed a set of 493 novel ovine SNPs of the male-specific region of Y chromosome (MSY) by genome mapping. We then conducted a comprehensive genomic analysis of Y chromosome, mitochondrial DNA, and whole-genome sequence variations in a large number of 595 rams representing 118 domestic populations across the world. We detected four different paternal lineages of domestic sheep and resolved, at the global level, their paternal origins and differentiation. In Northern European breeds, several of which have retained primitive traits (e.g., a small body size and short or thin tails), and fat-tailed sheep, we found an overrepresentation of MSY lineages y-HC and y-HB, respectively. Using an approximate Bayesian computation approach, we reconstruct the demographic expansions associated with the segregation of primitive and fat-tailed phenotypes. These results together with archaeological evidence and historical data suggested the first expansion of early domestic hair sheep and the later expansion of fat-tailed sheep occurred ∼11,800-9,000 years BP and ∼5,300-1,700 years BP, respectively. These findings provide important insights into the history of migration and pastoralism of sheep across the Old World, which was associated with different breeding goals during the Neolithic agricultural revolution.

Detection of polymorphism within leptin gene in Egyptian river buffalo and predict its effects on different molecular levels.

BACKGROUND: Leptin (LEP) regulates the glucose homeostasis directly and centrally by the regulation of the insulin levels or indirectly by alternation of the levels of the other glucose metabolism regulator hormones. The present investigation studied the polymorphism in LEP gene which is related to fertility in 81 female Egyptian river buffalo. RESULTS: The PCR-RFLP pattern of the gene using the restriction enzyme Eco91I showed that all the animals had monomorphic pattern in the studied gene which consists of CC. A 511-bp fragment from LEP gene was amplified and sequenced. The homology between the amplified LEP gene fragment in buffalo and cattle, sheep, goat, human, and mouse on the nucleotides sequence level was 99, 97, 97, 87, and 79%, respectively, and on the translated amino acids sequence level was 100, 98, 98, 85, and 82%, respectively. Several SNPs were detected; among them, the T27C SNP disrupted an intronic splicing silencer. The A114G, A310G, G263A, and G379A SNPs disrupt exonic splicing enhancers, and the last two SNPs create new exonic splicing enhancers. The A114G, C163A, A211G, G288A, A310G, A322G, G330C, C348T, T360C, and G379A SNPs cause S71G, T87 N, N103S, E129K, E136G, Y140C, E143Q, R149W, S153P, and R159Q amino acids mutations. N103S, E129K, E136G, Y140C, E143Q, and S153P were classified as deleterious mutations. Y140, E143, N103, and R149 were the most conserved among the mutated amino acids. S71G only increased the stability of the leptin protein while the remaining mutations decreased it. CONCLUSION: Four SNPs were revealed among the tested animals. Twenty-one SNPs were found between the sequenced amplicon and the buffalo records in the Genbank. Some SNPs were predicted to have several effects on different biological processes like mRNA splicing, protein stability, and the gene functions.