High-fat diet disrupts the gut microbiome, leading to inflammation, damage to tight junctions, and apoptosis and necrosis in Nyctereutes procyonoides intestines.

Given the burgeoning Nyctereutes procyonoides breeding industry and its growing scale, it is imperative to investigate the impact of high-fat diets on the health of these animals. This study involved 30 male Nyctereutes procyonoides of comparable weights (3 kg ±0.5), randomly assigned to either a control group or a high-fat diet group (n = 15 each). The latter group was fed a mixture of lard and basal diet in a 2:5 ratio, establishing a high-fat diet model in Nyctereutes procyonoides. This diet induced diarrhea and histopathological changes in the Nyctereutes procyonoides. Analysis of the small intestine contents using 16S rRNA sequencing revealed a high-fat diet-induced disruption in the gut microbiota. Specifically, Escherichia-Shigella emerged as the biomarker in the high-fat diet group (P = 0.049), while Vagococcus was prevalent in the control group (P = 0.049), indicating a significant increase in harmful bacteria in the high-fat diet group. Furthermore, this disrupted gut flora correlated with inflammation and oxidative stress, as evidenced by marked increases in TNF-α (P < 0.01), IL-1ß (P < 0.05), and IL-6 (P < 0.05) levels, measured via q-PCR, Western blot, and oxidative stress assays. In addition, q-PCR analysis revealed significant upregulation of apoptosis and necrosis markers, including Bax, Caspase3, Caspase9, Caspase12, RIPK3, and RIPK1 (P < 0.01 to P < 0.001), and a concurrent downregulation of the anti-apoptotic gene Bcl-2 (P < 0.01) in the high-fat diet group, consistent with protein expression trends. These findings suggest that a high-fat diet alters the gut microbiome toward a more harmful bacterial composition, escalating inflammatory responses and intestinal tissue permeability, culminating in intestinal cell apoptosis and necrosis.IMPORTANCEThis study examines the impact of high-fat diets on Nyctereutes procyonoides. Our research established a Nyctereutes procyonoides model on a high-fat diet, revealing significant health impacts, such as diarrhea, histological anomalies, and alterations in the gut microbiota. These findings emphasize the importance of preventing health issues and promoting sustainable industry growth. They highlight the significant impact of diet on gut microbiota and overall animal health.

Multi-Omics Analysis Reveals the Pathogenesis of Growth-Disordered Raccoon Dog.

Microorganisms of the genus Eperythrozoon are a zoonotic chronic infectious disease with wide distribution. We found that raccoons infected with Eperythrozoon showed obvious stunting, which seriously affected the economic benefits of raccoon dogs. To investigate the pathogenesis of the raccoon dog, we used transcriptome and proteome sequencing to analyze the changes in mRNA, miRNA, and protein expression in raccoon dogs infected with Eperythrozoon and normal raccoons. The results showed that the expression levels of genes related to immunity, metabolism, and enzyme activity were significantly changed. Among these, ERLIN1, IGF1R, CREB3L1, TNS1, TENC1, and mTOR play key roles. Additionally, the miR-1268, miR-125b, miR-10-5p, and miR-10 as central miRNAs regulate the expression of these genes. Integrated transcriptomic and proteomic analyses revealed consistent trends in mRNA and protein changes in MYH9, FKBP1A, PRKCA, and CYP11B2. These results suggest that Eperythrozoon may contribute to the slow development of raccoons by affecting the expression of mRNAs and miRNAs, reducing their immunity and causing metabolic abnormalities.

Using DNA metabarcoding and a novel canid-specific blocking oligonucleotide to investigate the composition of animal diets of raccoon dogs (Nyctereutes procyonoides) inhabiting the waterside area in Korea.

The raccoon dog (Nyctereutes procyonoides) is known to be an opportunistic generalist who feeds on a wide variety of foods. Historically, their diet has been investigated by morphological observation of undigested remains in feces, requiring specialized knowledge such as osteology, zoology, and phytology. Here, we used DNA metabarcoding of vertebrate 12S rRNA gene and invertebrate 16S rRNA gene to investigate their fecal contents. Additionally, we developed a blocking oligonucleotide that specifically inhibits the amplification of the canid 12S rRNA gene. We confirmed that the blocking oligonucleotide selectively inhibit the amplification of raccoon dog's DNA without significantly changing the composition of the preys' DNA. We found that the main foods of raccoon dogs in our study area, the waterside of paddy fields in Korea, were fishes such as Cyprinidae and insects such as mole crickets, which makes sense given the Korean fauna and their well-known opportunistic feeding behaviors. As a method to conveniently and objectively investigate feeding habits of raccoon dogs, this study provided baseline information on DNA metabarcoding. By using DNA metabarcoding, it is expected that the diet habits and ecology of raccoon dogs will be better understood by future research.

A mutant gene for albino body color is widespread in natural populations of tanuki (Japanese raccoon dog).

Albino mutants (white coat and red eyes) of tanuki (Nyctereutes procyonoides viverrinus) have been repeatedly found in the Central Alps area of Japan. We recently reported that an albino tanuki from Iida, a city in this area, lacks the third exon of the TYR gene encoding tyrosinase, which is essential for melanin synthesis. The absence of this exon was due to the chromosomal deletion of a complex structure. In the present study, we analyzed TYR of another albino tanuki that was found in Matsusaka, a city located outside the mountainous area. In this animal, the third exon was also lost, and the loss was due to a deletion in which the structure was identical to that of the Iida mutant. Our results indicate, in consideration of the complex structure of the deletion, that the two albino animals inherited a single deletion that arose in their common ancestor. Iida and Matsusaka are approximately 170 km apart. This is, to our knowledge, the first report of an albino mutant gene that is widely distributed in mammalian natural populations. As the origin of this mutation is not known, the distance covered by the mutant gene remains unclear. If we assume that the mutation occurred halfway between Iida and Matsusaka, we can predict the migration distance to be approximately 85 km; however, if the mutation occurred at any other place, a longer distance would be predicted. Natural selection against albino tanuki may be relaxed because of a recent increase in food resources and refuge in urban areas.

Phylogenetic relationships between different raccoon dog (Nyctereutes procyonoides) populations based on four nuclear and Y genes.

BACKGROUND: The raccoon dog (Nyctereutes procyonoides), endemic to East Asia, is classified as six subspecies according to their geographical distribution including a population introduced to Europe. Studies on phylogenetic relationship or population genetics in both native and introduced areas have been carried out recently. Lately, opinions that Japanese raccoon dogs should be classified as a different species were asserted based on several studies using karyotypes, morphometric characters, mtDNA, and microsatellites analysis. However, no data pertaining to the nuclear DNA (nDNA) or Y chromosome are available. OBJECTIVE: To estimate the relationship among the species using different genes is necessary in understanding of the history of this species. METHOD: Therefore, we investigated nDNA and Y chromosomes in our study to define relationships: (1) between continental raccoon dog populations, (2) between original and introduced groups, and (3) between continental and Japanese groups. RESULTS: The analysis of four nuclear (CHRNA1, VTN, TRSP, WT1) and ZFY genes indicated that there had been no genetic differentiation among the continental populations. However, significant differences were observed between continental and Japanese raccoon dogs in VTN and ZFY genes implying genetic differentiation has been going between them. CONCLUSION: To better understand the phylogenetic relationship among raccoon dog populations, further study will be necessary.

Population structure of the raccoon dog (Nyctereutes procyonoides) using microsatellite loci analysis in South Korea: Implications for disease management.

The prevention and control of infectious diseases transmitted by wildlife are gaining importance. To establish effective management strategies, it is essential to understand the population structure of animals. Raccoon dogs (Nyctereutes procyonoides) in South Korea play a key role in the maintenance of food web stability and possess genetic compositions that are unique compared to those in other areas. However, wild raccoon dogs play another role as the main host of various infectious diseases. To establish long-term strategies for disease management, we investigated the genetic structure and possible geographic barriers that influence the raccoon dog population in South Korea by analyzing 16 microsatellite loci. The present study showed that mountains were the major factors responsible for genetic structuring, along with distance. We proposed potential management units (MUs) for raccoon dogs based on the genetic structuring and gene-flow barrier data obtained in this study. Four MUs were suggested for the Korean raccoon dog population (Northern, Central, Southwestern, and Southeastern). The Korean raccoon dog population structure determined in this study and the proposed MUs will be helpful to establish pragmatic strategies for managing Korean raccoon dog population and for preventing the transmission of infectious diseases.

Genetic Diversity and Population Structure of East Asian Raccoon Dog (Nyctereutes procyonoides): Genetic Features in Central and Marginal Populations.

The raccoon dog (Nyctereutes procyonoides) is endemic to East Asia but has been introduced in Europe. Its high adaptability enabled its rapid colonization of European countries, where population growth has been raising concerns regarding ecosystem disturbance and the spread of zoonotic diseases. The genetic diversity and structure of endemic, source, and introduced populations from seven locations across South Korea, China, Russian Far East, Finland (spread to Finland after introduction to European part of Russia from Russian Far East), Vietnam, and Japan (Honshu and Hokkaido) were examined based on 16 microsatellite loci. Two major and significantly different (FST = 0.236) genetic clusters were found: continental (South Korean, Chinese, Russian, Finnish, and Vietnamese) and island (Japanese) populations. The continental raccoon dog population comprises three subpopulations (Chinese_Russian_Finnish, South Korean, and Vietnamese) and the Japanese population consists of Honshu and Hokkaido subpopulations. The genetic diversity and geographic structure of raccoon dogs in East Asia has been influenced by natural barriers to gene flow and reveals a typical central-marginal trend in genetic diversity (continental vs. island, and central vs. marginal or source vs. introduced within continental populations). The detected differences between continental and island populations agree with those reported in previous studies that considered these populations as different species.

Population Structure of the Raccoon Dog on the Grounds of the Imperial Palace, Tokyo, Revealed by Microsatellite Analysis of Fecal DNA.

The raccoon dog (Nyctereutes procyonoides, Canidae, Carnivora) is highly adaptable to urban environments. Populations of carnivorans inhabiting urban areas sometimes differ ecologically and genetically from those in rural areas. However, there is little information on urban raccoon dogs. This study focused on raccoon dog populations in Tokyo, one of the most highly urbanized cities in the world. We examined the genotypes of 10 microsatellites for 101 fecal samples from raccoon dogs inhabiting the grounds of the Imperial Palace, a green space in central Tokyo. We successfully genotyped 58 samples originating from 31 individuals. We also analyzed muscle tissue samples from raccoon dogs from the grounds of the Imperial Palace, the Akasaka Imperial Grounds (a green space close to the Imperial Palace), and the surrounding urban area, and then investigated the genetic structure and diversity of these populations, and the genetic differentiation among them. The population on the grounds of the Imperial Palace was genetically differentiated from that in the Akasaka Imperial Grounds, suggesting that the roads and buildings act as barriers to gene flow. In addition, the population on the grounds of the Imperial Palace showed greater genetic difference from that in the surrounding area than that in the Akasaka Imperial Grounds. We speculate that the moats around the Imperial Palace restrict individual ranges within the palace grounds and limit migration and gene flow to other areas.

Identification and characterization of the toll-like receptor 8 gene in the Chinese raccoon dog (Nyctereutes procyonoides).

TLR8 is an important sensor of single-stranded RNA (ssRNA) from the viral genome and plays an essential role in innate antiviral responses via the recognition of conserved viral molecular patterns. In this report, TLR8 in the Chinese raccoon dog was characterized and analyzed for the first time. The full-length sequence of raccoon dog TLR8 (RdTLR8) cDNA was cloned by rapid amplification of cDNA ends (RACE) and is 3191bp with a 3117-bp open reading frame (ORF) encoding 1038 amino acids. The putative protein exhibits typical features of the TLR families, with 19 leucine-rich repeats (LRRs) in the extracellular domain and a cytoplasmic TIR domain. Comparative analyses of the RdTLR8 amino acid sequence indicated a 73.6-99.4% sequence identity with dog, horse, pig, sheep, cattle, human and mouse TLR8. Phylogenetic analysis grouped 71 mammalian TLR proteins into five sub-families, wherein RdTLR8 was clustered into a monophyletic TLR8 clade in the TLR9 family, which was completely coincident with the evolutionary relationship among mammals. Quantitative real-time PCR analysis revealed extensive expression of RdTLR8 in tissues from healthy Chinese raccoon dogs with the highest expression in the peripheral blood mononuclear cells (PBMCs) and the lowest expression in the skeletal muscle. HEK293 cells cotransfected with a RdTLR8 expression plasmid and an NF-κB-luciferase reporter plasmid significantly responded to the agonist 3M-002, indicating a functional TLR8 homolog. In addition, raccoon dog PBMCs exposed to the canine distemper virus (CDV) wild strain CDV-PS and the TLR8 agonist 3M-002 showed significant upregulation of RdTLR8 mRNA and proinflammatory cytokines TNF-α and IFN-α, suggesting that RdTLR8 might play an important role in the immune response to viral infections in the Chinese raccoon dog.

Homogenous Population Genetic Structure of the Non-Native Raccoon Dog (Nyctereutes procyonoides) in Europe as a Result of Rapid Population Expansion.

The extent of gene flow during the range expansion of non-native species influences the amount of genetic diversity retained in expanding populations. Here, we analyse the population genetic structure of the raccoon dog (Nyctereutes procyonoides) in north-eastern and central Europe. This invasive species is of management concern because it is highly susceptible to fox rabies and an important secondary host of the virus. We hypothesized that the large number of introduced animals and the species' dispersal capabilities led to high population connectivity and maintenance of genetic diversity throughout the invaded range. We genotyped 332 tissue samples from seven European countries using 16 microsatellite loci. Different algorithms identified three genetic clusters corresponding to Finland, Denmark and a large 'central' population that reached from introduction areas in western Russia to northern Germany. Cluster assignments provided evidence of long-distance dispersal. The results of an Approximate Bayesian Computation analysis supported a scenario of equal effective population sizes among different pre-defined populations in the large central cluster. Our results are in line with strong gene flow and secondary admixture between neighbouring demes leading to reduced genetic structuring, probably a result of its fairly rapid population expansion after introduction. The results presented here are remarkable in the sense that we identified a homogenous genetic cluster inhabiting an area stretching over more than 1500km. They are also relevant for disease management, as in the event of a significant rabies outbreak, there is a great risk of a rapid virus spread among raccoon dog populations.

Identification of polymorphisms and transcriptional activity of the proto-oncogene KIT located on both autosomal and B chromosomes of the Chinese raccoon dog.

B chromosomes are dispensable and co-exist with autosomal and sex chromosomes. The karyotype of the Chinese raccoon dog (Nyctereutes procyonoides procyonoides) comprises 0-4 B chromosomes. The proto-oncogene KIT is found on all B chromosomes of the Chinese raccoon dog. In the present study, partial DNA and mRNA sequences of KIT were amplified and sequenced from four individuals containing B chromosomes. Sequence analyses revealed that polymorphisms including single nucleotide polymorphisms (SNPs) and inserts/deletions were rich in the KIT gene of Chinese raccoon dog at the genomic level. However, no polymorphism was detected at the mRNA level. A comparison of mRNA sequences from Chinese raccoon dogs with the corresponding sequences derived from arctic fox and dog, which do not contain B chromosomes, revealed the mRNA sequences of the 10 SNPs to be identical between these three species. Therefore, these findings suggest that KIT located on the B chromosomes in Chinese raccoon dog lacks transcriptional activity.

Cloning and identification of the ASIP gene in Chinese raccoon dog (Nyctereutes procyonoides procyonoides).

The quantity, quality, and distribution of eumelanin and pheomelanin determine a wide variety of coat colors in animals. Three coat color variants exist in farmed wild-type Chinese raccoon dog (Nyctereutes procyonoides procyonoides), which is an important fur-bearing animal species. The ASIP gene is an important candidate gene for coat color variation in some species. In this study, the complete cDNA sequences of ASIP were amplified from a wild-type Chinese raccoon dog. Sequence analysis revealed the coding region of ASIP in Chinese raccoon dog to be 396-bp in length and two transcripts (accession Nos. KT224450 and KT224451) were identified due to the alternative use of exon 1 (1A and 1C). However, the alternative splicing pattern and the coding sequence of ASIP in three types of coat color variants were the same as those identified in the wild-type individual. Based on the results obtained in this study, we can exclude a role for alternative splicing of exon 1 and the coding sequence of ASIP in coat color variation in Chinese raccoon dog.

Cloning and association analysis of KIT and EDNRB polymorphisms with dominant white coat color in the Chinese raccoon dog (Nyctereutes procyonoides procyonoides).

The Chinese raccoon dog (Nyctereutes procyonoides procyonoides) is one of the most important fur-bearing animal species. The dominant white individual, a coat color variant in farmed Chinese raccoon dog, shows a completely white phenotype over the entire body. The KIT and EDNRB genes have been reported to be associated with the dominant white coat color in some mammalian species. In the present study, the full-length coding sequences of KIT and EDNRB were amplified from a dominant white and a wild-type Chinese raccoon dog. Sequence analysis revealed that the coding region of KIT and EDNRB in Chinese raccoon dog was 2919 and 1332 base pairs in length (accession No. KM083121 and KM083122), respectively, and 2 single-nucleotide polymorphisms (SNPs; c.600C>T and c.967G>A) in KIT and 1 SNP (c.259A>C) in EDNRB was found only in the dominant white individual. An alternative splicing site at the boundary of 4 and 5 of the KIT gene was identified in both individuals. We further investigated the association between the 3 SNPs of KIT and EDNRB and dominant white coat color by genotyping 18 individuals. We found no association between these SNPs and dominant white coat color. Based on these results, we can exclude the coding regions of the KIT and EDNRB genes as determinants of the dominant white coat color in Chinese raccoon dog.

Intraspecific comparison of complete mitogenome sequences from two Asian raccoon dogs (Canidae: Nyctereutes procyonoides).

We determined the complete mitochondrial genome (GenBank accession number: KF709435) of the Korean raccoon dog Nyctereutes procyonoides koreensis and compared it with a previously published mitogenome (GenBank accession number: GU256221) of a Chinese raccoon dog. The total length of N. p. koreensis mitogenome is 16,802 bp, with a base composition of 32.1% A, 26.9% T, 26.8% C and 14.2% G. High similarity of 98.7% was found between the complete mitogenome sequences of Korean and Chinese raccoon dogs. Sequence similarity of the two mitogenomes was 99.3% in the other gene regions except for D-loop. The sequence similarity of 99.1% was found in the 13 protein-coding gene regions, whereas 99.6% was identical in mtDNA regions covering all the 22 tRNA genes. There was no variation between 12S rRNAs, whereas 0.5% difference was found between 16S rRNAs.

Genome organization and DNA methylation patterns of B chromosomes in the red fox and Chinese raccoon dogs.

The molecular structure of B chromosomes (Bs) is relatively well studied. Previous research demonstrates that Bs of various species usually contain two types of repetitive DNA sequences, satellite DNA and ribosomal DNA, but Bs also contain genes encoding histone proteins and many others. However, many questions remain regarding the origin and function of these chromosomes. Here, we focused on the comparative cytogenetic characteristics of the red fox and Chinese raccoon dog B chromosomes with particular attention to the distribution of repetitive DNA sequences and their methylation status. We confirmed that the small Bs of the red fox show a typical fluorescent telomeric distal signal, whereas medium-sized Bs of the Chinese raccoon dog were characterized by clusters of telomeric sequences along their length. We also found different DNA methylation patterns for the B chromosomes of both species. Therefore, we concluded that DNA methylation may maintain the transcriptional inactivation of DNA sequences localized to B chromosomes and may prevent genetic unbalancing and several negative phenotypic effects.

Population genetic study of the raccoon dog (Nyctereutes procyonoides) in South Korea using newly developed 12 microsatellite markers.

The raccoon dog (Nyctereutes procyonoides) is distributed from southeastern Siberia to northern Vietnam, including Korea and Japan, as well as Europe. In Korea, most of its predators and competitors are extinct, which has resulted in rapid growth of the raccoon dog population. This population increase has raised concerns about its role in the ecosystem and the zoonotic transfer of various contagious diseases, and thus an effective method of raccoon dog population control in Korea is required. To investigate the genetic diversity and structure of raccoon dog populations, 12 polymorphic microsatellite loci were identified and characterized. These novel microsatellite markers were employed to obtain basic population genetic parameters for 104 N. procyonoides specimens from five locations in South Korea. The mean allele number of 12 loci across samples was 8.7, and the number of alleles per locus ranged 2-13. Mean expected and observed heterozygosities were 0.723 and 0.619, respectively. Genetic differentiation, estimated by pairwise FST, was significant for all population pairs excepting Seoul/Gyeonggi and Gangwon pair, with a moderate level of genetic differentiation for all the population pairs (mean FST = 0.054), but little differentiation between Seoul/Gyeonggi and Gangwon (FST = 0.024). Bayesian-based clustering analysis predicted that Korean raccoon dog population is composed of four distinct genetic subpopulations. These genetic information and structure of raccoon dog will be very useful to prevent spreading infectious diseases.

Missense polymorphisms in the MC1R gene of the dog, red fox, arctic fox and Chinese raccoon dog.

Coat colour variation is determined by many genes, one of which is the melanocortin receptor type 1 (MC1R) gene. In this study, we examined the whole coding sequence of this gene in four species belonging to the Canidae family (dog, red fox, arctic fox and Chinese raccoon dog). Although the comparative analysis of the obtained nucleotide sequences revealed a high conservation, which varied between 97.9 and 99.1%, we altogether identified 22 SNPs (10 in dogs, six in farmed red foxes, two in wild red foxes, three in arctic foxes and one in Chinese raccoon dog). Among them, seven appeared to be novel: one silent in the dog, three missense and one silent in the red fox, one in the 3'-flanking region in the arctic fox and one silent in the Chinese raccoon dog. In dogs and red foxes, the SNPs segregated as 10 and four haplotypes, respectively. Taking into consideration the published reports and results of this study, the highest number of missense polymorphisms was until now found in the dog (9) and red fox (7).

Three-dimensional positioning of B chromosomes in fibroblast nuclei of the red fox and the chinese raccoon dog.

Great progress has been achieved over the last years in studies on chromosome arrangement in mammalian cell nuclei. Growing evidence indicates that the genome's spatial organization is of functional relevance. So far, no attention has been paid to the nuclear organization of B chromosomes (Bs). In this study we have examined nuclear positioning of Bs in 2 species from the Canidae family--the red fox and the Chinese raccoon dog. Using 2D and 3D fluorescence in situ hybridization and 2 gene-specific probes (C-KIT and PDGFRA), we analyzed the location of Bs in fibroblast nuclei. We found that small Bs of the red fox occupied mostly the interior of the nucleus, while medium-sized Bs of the Chinese raccoon dog were observed in the peripheral area of the nucleus as well as in intermediate and interior locations. The more uniform distribution of B chromosomes in the Chinese raccoon dog may be the result of differences in their size, since 3 morphological types of Bs are distinguished in this species. Our results indicate that 3D positioning of B chromosomes in fibroblast nuclei of the 2 canid species is in agreement with the chromosome size-dependent theory.

Development and characterization of polymorphic microsatellite markers for Chinese raccoon dog (Nyctereutes procyonoides procyonoides).

Chinese raccoon dog (Nyctereutes procyonoides procyonoides) is one of the most important fur-bearing animal species. Information about the genetic background of farmed Chinese raccoon dogs is limited. In this study, 17 polymorphic microsatellite markers were isolated and identified from an (AC)n-microsatellite-enriched library of Chinese raccoon dogs. The number of alleles per locus ranged from 2 to 8 based on 48 individuals tested. The expected and observed heterozygosity and polymorphism information content per locus ranged from 0.383 to 0.8378, 0.3200 to 0.8696, and 0.3047 to 0.7947, respectively. Cross-species amplification of these loci in 2 other Canidae species indicated that 9 and 11 of these loci could also be amplified successfully in the arctic and silver fox, respectively. These microsatellite loci developed in the present report will provide useful tools for population genetic studies, individual identification, and phylogenetic analysis in the Chinese raccoon dog and other Canidae species.

Altered expression of melanocortin-1 receptor (MC1R) in a yellow-coloured wild raccoon dog (Nyctereutes procyonoides).

BACKGROUND: The melanocortin 1 receptor (MC1R) gene plays a key role in determining coat colour in mammals by controlling the proportion of eumelanin and pheomelanin granules. Wild raccoon dogs have a mixed coat colour, with black to brown and grey hairs. HYPOTHESIS/OBJECTIVES: The study was performed to identify the cause of the variant yellow coat colour in a wild raccoon dog. ANIMALS: A wild raccoon dog that showed coat colour change to yellow and four wild-type raccoon dogs that showed normal coat colour were included. METHODS: To identify the cause of the variant yellow coat colour, we examined the sequence of the MC1R gene and its expression at the mRNA and protein levels. RESULTS: The coding region of the MC1R gene of this raccoon dog comprised 954 bp, the same as for wild-type raccoon dogs and domestic dogs. By comparing the gene with that in the wild-type raccoon dog, a 2 bp deletion was detected in the 5'-untranslated region, positioned 152 bp upstream of the start codon. However, there was no significant difference in the mRNA expression level. The yellow raccoon dog revealed a significantly decreased MC1R protein level compared with the wild-type raccoon dogs, indicating an increase in pheomelanin synthesis. CONCLUSIONS AND CLINICAL IMPORTANCE: These results suggest that the variant coat colour in the yellow raccoon dog was associated with decreased MC1R function.