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DNA methylation plays important functions in gene expression regulation that is involved in individual development and various diseases. DNA methylation has been well studied in human and model organisms, but only limited data exist in companion animals like dog. Using methylation-sensitive restriction enzyme-based next generation sequencing (Canine DREAM), we obtained canine DNA methylation maps of 16 somatic tissues from two dogs. In total, we evaluated 130,861 CpG sites. The majority of CpG sites were either highly methylated (> 70%, 52.5-64.6% of all CpG sites analyzed) or unmethylated (< 30%, 22.5-28.0% of all CpG sites analyzed) which are methylation patterns similar to other species. The overall methylation status of CpG sites across the 32 methylomes were remarkably similar. However, the tissue types were clearly defined by principle component analysis and hierarchical clustering analysis with DNA methylome. We found 6416 CpG sites located closely at promoter region of genes and inverse correlation between DNA methylation and gene expression of these genes. Our study provides basic dataset for DNA methylation profiles in dogs.

Although the domestic dog's origin is still unclear, this lineage is believed to have been domesticated from an extinct population of gray wolves, which is expected to be more closely related to dogs than to other populations of gray wolves. Here, we sequence the whole genomes of nine Japanese wolves (7.5–100x: Edo to Meiji periods) and 11 modern Japanese dogs and analyze them together with those from other populations of dogs and wolves. A phylogenomic tree shows that, among the gray wolves, Japanese wolves are closest to the dog, suggesting that the ancestor of dogs is closely related to the ancestor of the Japanese wolf. Based on phylogenetic and geographic relationships, the dog lineage has most likely originated in East Asia, where it diverged from a common ancestor with the Japanese wolf. Since East Eurasian dogs possess Japanese wolf ancestry, we estimate an introgression event from the ancestor of the Japanese wolf to the ancestor of the East Eurasian dog that occurred before the dog's arrival in the Japanese archipelago. The evolutionary origin of the domestic dog is uncertain. Here, the authors sequence the whole genomes of 9 extinct Japanese wolves and 11 modern Japanese dogs, applying a phylogenetic analysis to show that dogs may have originated in East Asia from a common ancestor with the Japanese wolf. [ABSTRACT FROM AUTHOR]

Background: DNA methylation analysis might identify prognostic CpG sites in CHOP‐treated dogs with multicentric high‐grade B‐cell lymphoma (MHGL) with heterogenous prognosis. Objective: To identify prognostic CpG sites of MHGL through genome‐wide DNA methylation analysis with pyrosequencing validation. Animals: Test group: 24 dogs. Validation group: 100 dogs. All client‐owned dogs were diagnosed with MHGL and treated with CHOP chemotherapy. Methods: Cohort study. DNA was extracted from lymph node samples obtained via FNA. Genome‐wide DNA methylation analysis using Digital Restriction Enzyme Analysis of Methylation (DREAM) was performed on the test group to identify differentially methylated CpG sites (DMCs). Bisulfite pyrosequencing was used to measure methylation status of candidate DMCs in the validation group. Median survival times (MST) were analyzed using Kaplan‐Meier (log‐rank) product limit method. Results: DREAM analyzed 101 576 CpG sites. Hierarchical clustering of 16 262 CpG sites in test group identified group with better prognosis (MST = 55‐477 days vs 10‐301 days, P =.007). Volcano plot identified 1371 differentially methylated CpG sites (DMCs). DMC near the genes of FAM213A (DMC‐F) and PHLPP1 (DMC‐P) were selected as candidates. Bisulfite‐pyrosequencing performed on validation group showed group with methylation level of DMC‐F < 40% had favorable prognosis (MST = 11‐1072 days vs 8‐1792 days, P =.01), whereas group with the methylation level combination of DMC‐F < 40% plus DMC‐P < 10% had excellent prognosis (MST = 18‐1072 days vs 8‐1792 days, P =.009). Conclusion and Clinical Importance: Methylation status of prognostic CpG sites delineate canine MGHL cases with longer MST, providing owners with information on expectations of potential improved treatment outcomes. [ABSTRACT FROM AUTHOR]

Simple Summary: The present study aims to establish a method of whole mitochondrial DNA (mtDNA) sequencing for domestic dogs. Prior to proceeding with the experiment, the collection of relevant DNA samples was essential. Therefore, we decided to use oral mucosa DNA, which can be collected non-invasively. A polymerase chain reaction (PCR) was performed using the DNA collected from six dogs raised in Japan and four primer pairs as specific to the mtDNA, after which the amplified products were sequenced using next-generation sequencing. As a result, the whole mtDNA obtained from all dogs was correctly sequenced. Thus, we determined that the method we used in our study may be useful for future research on dog-related medical care and welfare. In human beings, whole mitochondrial DNA (mtDNA) sequencing has been widely used in many research fields, including medicine, forensics, and genetics. With respect to the domestic dog (Canis lupus familiaris), which is commonly recognized as being an additional member of the traditional human family structure, research studies on mtDNA should be developed to expand and improve our collective knowledge of dog medicine and welfare as it seems that there is still room for further development in these areas. Moreover, a simple and robust method for sequencing whole mtDNA that can be applied to various dog breeds has not yet been described in the literature. In the present study, we aim to establish such a method for the whole mtDNA sequencing of the domestic dog. In the experiments we conducted, oral mucosa DNA samples obtained from six Japanese domestic dogs were used as a template. We designed four primer pairs that could amplify approximately 5 kbp from each region of the mtDNA and validated several PCR conditions. Subsequently, the PCR amplicons were pooled and subjected to library preparation. The sequencing of the libraries was performed using next-generation sequencing (NGS), followed by bioinformatics analysis. Our results demonstrate that the proposed method can be used to perform highly accurate resequencing. We believe that this method may be useful for future research conducted to better understand dog medicine and welfare. [ABSTRACT FROM AUTHOR]

The emergence and global spread of extended-spectrum or AmpC β-lactamase (ESBL/AmpC)-producing Enterobacteriaceae in companion animals have led to the hypothesis that companion animals might be reservoirs for cross-species transmission because of their close contact with humans. However, current knowledge in this field is limited; therefore, the role of companion animals in cross-species transmission remains to be elucidated. Herein, we studied ESBL/AmpC-producing Enterobacteriaceae, Escherichia coli in particular, isolated from extraintestinal sites and feces of companion dogs. Whole-genome sequencing analysis revealed that (i) extraintestinal E. coli isolates were most closely related to those isolated from feces from the same dog, (ii) chromosomal sequences in the ST131/C1-M27 clade isolated from companion dogs were highly similar to those in the ST131/C1-M27 clade of human origin, (iii) certain plasmids, such as IncFII/pMLST F1:A2:B20/blaCTX-M-27, IncI1/pMLST16/blaCTX-M-15, or IncI1/blaCMY-2 from dog-derived E. coli isolates, shared high homology with those from several human-derived Enterobacteriaceae, (iv) chromosomal blaCTX-M-14 was identified in the ST38 isolate from a companion dog, and (v) eight out of 14 tested ESBL/AmpC-producing E. coli isolates (i.e., ST131, ST68, ST405, and ST998) belonged to the human extraintestinal pathogenic E. coli (ExPEC) group. All of the bla-coding plasmids that were sequenced genome-wide were capable of horizontal transfer. These results suggest that companion dogs can spread ESBL/AmpC-producing ExPEC via their feces. Furthermore, at least some ESBL/AmpC-producing ExPECs and bla-coding plasmids can be transmitted between humans and companion dogs. Thus, companion dogs can act as an important reservoir for ESBL/AmpC-producing E. coli in the community. [ABSTRACT FROM AUTHOR]