Sequences of canine glutamate decarboxylase (GAD) 1 and GAD2 genes, and variation of their genetic polymorphisms among five dog breeds

Glutamate decarboxylase (GAD) is the primary enzyme in the brain that catalyzes the synthesis of gamma-aminobutyric acid (GABA), the main inhibitory neurotransmitter. There are two isoforms named according to their molecular weights, GAD67 and GAD65, which are encoded by GAD1 and GAD2, respectively. To investigate the association between GAD genes and temperament in domestic dogs, Canis familiaris, we sequenced the full lengths of the coding regions of these genes and identified three single nucleotide polymorphisms (SNPs) in GAD1 and one in GAD2. When comparing genotype and allele frequencies of SNPs among five breeds with different behavioral traits, statistically significant interbreed differences were observed for three SNPs in GAD1. These results suggest that GAD1 SNPs may be useful for behavioral genetic studies in dogs.     

Estimated frequency of the canine hyperuricosuria mutation in different dog breeds

Background: Hyperuricosuria is a condition that predisposes dogs to urate urolithiasis. A mutation that causes canine hyperuricosuria was previously identified in 3 unrelated dog breeds. The occurrence of the mutation in additional breeds was not determined. Hypothesis/Objectives: Identify additional breeds that have the hyperuricosuria mutation and estimate the mutant allele frequency in those breeds. Animals: Three thousand five hundred and thirty dogs from 127 different breeds were screened for the hyperuricosuria mutation. Methods: DNA samples were genotyped by pyrosequencing and allele‐specific polymerase chain reaction methods. Results: Mutant allele frequencies that range from 0.001 to 0.15 were identified in the American Staffordshire Terrier, Australian Shepherd, German Shepherd Dog, Giant Schnauzer, Parson (Jack) Russell Terrier, Labrador Retriever, Large Munsterlander, Pomeranian, South African Boerboel, and Weimaraner breeds. Conclusions and Clinical Importance: The hyperuricosuria mutation has been identified in several unrelated dog breeds. The mutant allele frequencies vary among breeds and can be used to determine an appropriate breeding plan for each breed. A DNA test is available and may be used by breeders to decrease the mutant allele frequency in breeds that carry the mutation. In addition, veterinarians may use the test as a diagnostic tool to identify the cause of urate urolithiasis.     

Genetic structure and diversity among sheep breeds in the United States: identification of the major gene pools

Understanding existing levels of genetic diversity of sheep breeds facilitates in situ and ex situ conservation activities. A comprehensive evaluation of US sheep breeds has not been previously performed; therefore, we evaluated the genetic diversity among and within 28 US sheep breeds. Both major and minor breeds were included in the analysis and consisted of 666 animals from 222 producers located in 38 states. The level of within-breed genetic diversity was variable and not dependent upon status of a breed as a major or minor breed. Bayesian cluster analysis indicated the breeds were grouped more by physiological differences (meat vs. wool production) rather than geographic origin. Results suggest several actionable items to improve in situ and ex situ conservation. The results clearly identify breeds in need of increased in situ and ex situ management (e.g., Hog Island and Karakul) and allow several suggestions for in situ management of flocks. Conversely, several of the breeds appear genetically similar and therefore require less emphasis on collecting germplasm samples for the gene bank. Commercially important breeds (e.g., Rambouillet and Suffolk) were found to have substantial variation, which should enable breeders to proceed, unencumbered by genetic diversity concerns, with selection strategies that maximize profit.     

Canine tyrosine hydroxylase (TH) gene and dopamine beta -hydroxylase (DBH) gene: their sequences, genetic polymorphisms, and diversities among five different dog breeds

Dopamine and noradrenaline are catecholamine neurotransmitters that are produced by biosynthetic enzymes such as tyrosine hydroxylase (TH) and dopamine beta -hydroxylase (DBH). As a first step to elucidate the genetic background of canine behavioral traits, we selected these genes as targets and sequenced these canine genes, and found that both were highly homologous with those of human beings. Then brain cDNAs derived from ten unrelated Beagles were used to search for polymorphisms in these genes. Four single nucleotide polymorphisms (SNPs) (C97T, G168A, G180A and C264T), one of which (C97T) will cause amino acid substitution in the TH gene, and two SNPs (C789A and A1819G), both of which will cause amino acid substitutions in the DBH gene were identified. The allelic frequencies among five dog breeds (47 Golden Retrievers, 41 Labrador Retrievers, 40 Malteses, 26 Miniature Schnauzers, and 39 Shibas) were examined and found to have significant variation between them with regards to all these SNPs, except for C97T in the TH gene and A1819G in the DBH gene. The polymorphisms of C97T and A1819G were found only in the Shiba. The present results suggest that the polymorphisms of the genes encoding catecholamine biosynthetic enzymes may become important markers for examining the genetic background of behavioral characteristics in dogs.     

Sequences and polymorphisms of the canine monoamine transporter genes SLC6A2, SLC6A3, and SLC6A4 among five dog breeds

Monoamine transporters have roles to regulate the monoamine concentrations in synaptic clefts in the central nervous system and are the targets of antidepressants and psychostimulants. They include transporters for norepinephrine, dopamine and serotonin, which are encoded by the SLC6A2, SLC6A3, and SLC6A4 genes, respectivily. We sequenced the full lengths of the coding regions of these genes for dogs and identified four single nucleotide polymorphisms (SNPs) in SLC6A2 and four in SLC6A3. One SLC6A3 SNP was non-synonymous and caused an amino acid substitution from threonine to serine. The genotype frequencies of these polymorphisms differed significantly among five breeds with different behavioral traits, suggesting that novel SLC6A2 and SLC6A3 SNPs would provide additional useful information for behavioral genetic studies in dogs.     

Population parameters to compare dog breeds: differences between five Dutch purebred populations

Differences in five purebred dog populations born in 1994 in the Netherlands were evaluated using different parameters. Numerically, the Golden Retriever was the largest breed (840 litters of 234 sires) and the Kooiker Dog (101 litters of 41 sires) the smallest. The litter per sire ratio was largest in the Bernese Mountain Dog (4.59) and lowest in the Kooiker Dog (2.46). The mean relatedness and the actual mean level of inbreeding in the studied generations were 0.102 and 0.056 respectively for the Bernese Mountain Dog, 0.041 and 0.046 for the Bouvier des Flandres, 0.087 and 0.061 for the Boxer, 0.020 and 0.018 for the Golden Retriever, and 0.146 and 0.070 for the Kooiker Dog. Quantification and visualization of population parameters for purebred dogs will facilitate the comparison of breeds and the comparison of breeds in different periods or countries. It appears unlikely that the increase in inbreeding is a major determinant of the possible increase in the frequency of genetic diseases.     

利用PCR-RFLP检测五品种警犬遗传缺陷——血管性假性血友病

犬血管性假性血友病(vWD)是常染色体不完全显性遗传性出血病.血管性假血友病因子(vWF)的数量和质量正常与否决定着是否患有vWD,而vWF基因的表达又控制着vWF的数量和质量.本研究应用DNA测序技术和PCR-RFLP方法检测德国牧羊犬、杜伯文犬、罗威纳犬、史宾格犬和马里努阿犬等5个品种共132头犬的vWF基因5个候选区域.结果显示,德国牧羊犬、罗威纳犬、史宾格犬和马里努阿犬未发现vWF突变基因,杜伯文犬中有2头患病和3头携带者,携带频率为5.16%.     

Analysis of a deletion in the nephronophthisis 4 gene in different dog breeds

Cone‐rod dystrophy is a progressive inherited retinal degenerative disorder that occurs in humans and dogs. The deletion in the nephronophthisis 4 (NPHP4) gene was established as a causative mutation in standard wire‐haired Dachshunds. We analyzed all varieties of Dachshunds from the Czech Republic and five other dog breeds and found that the deletion in the NPHP4 (in heterozygous state) is present not only in standard‐, but also in miniature wire‐haired Dachshunds, but not in other varieties of Dachshunds or in other breeds.     

Structure and sequence variation of the canine perforin gene

Lymphocyte-mediated cytotoxicity is essential to control viral infections, limit lymphocyte expansion and activation, and survey for malignant cells. Humans with defects in lymphocyte cytotoxicity have reduced perforin function resulting in uncontrolled lymphocyte expansion, leading to excessive histiocyte activation and a hemophagocytic disorder. Dog breeds such as Bernese mountain dogs (BMD) have a high incidence of reactive and malignant diseases affecting histiocytes. This study addressed the hypothesis that changes in the perforin gene contribute to the development of hemophagocytic histiocytic sarcoma (HHS) in BMD. Canine perforin DNA was amplified and sequenced through multiple PCR assays from healthy and diseased dogs, and the gene structure determined by rapid amplification of cDNA ends. The coding component of the gene consists of 1679 bp, with two exons of 536 bp and 1143 bp separated by an intron of 865 bp. Gene configuration and location differ from that in other species although the coding sequence is highly conserved. Three silent single nucleotide polymorphisms (SNP) were identified. Analysis of their distribution indicated a consistent genotype among 6 middle-aged to older BMD without histiocytic diseases. Among samples from 10 dogs with HHS and 10 without histiocytic diseases SNP occurred with variable frequency. It was concluded that changes in the amino acid sequence of perforin were not associated with HHS but that a constellation of SNP may characterize BMD without histiocytic disease. Investigation of more dogs is required to confirm a specific genotype. Future studies should focus on the potential contribution of reduced perforin expression and/or function to HHS in dogs.     

Polymorphisms in the canine glucocorticoid receptor alpha gene (NR3C1α)

Corticosteroids are one of the most extensively used class of therapeutic agents in dogs. In human patients, response to corticosteroid therapy has been correlated with the presence of certain polymorphisms of the glucocorticoid receptor gene (NR3C1). Depending on the polymorphism present, patients may show either increased sensitivity to glucocorticoid‐induced adverse effects or resistance to their therapeutic effects. Because response to corticosteroid therapy in dogs can also be variable and unpredictable, we hypothesized that genetic variability exists in the canine NR3C1 gene. The aim of this study was to sequence the coding regions of the canine NR3C1 gene in a representative sample of dogs. Samples from 97 dogs from four previously identified genetic groupings of domestic breeds (Asian/Ancient, Herding, Hunting, and Mastiff) were sequenced and evaluated. Four exons contained polymorphisms and four exons showed no variation from the reference sequence. A total of six single nucleotide polymorphisms (SNPs) were identified including four synonymous SNPs and two nonsynonymous SNPs (c.811A>T and c.2111T>C). No dogs were homozygous for either variant allele, while 23 dogs were heterozygous for the c.811A>T allele and 2 were heterozygous for c.2111T>C allele. The amino acid changes caused by c.811A>T (serine to cysteine) and c.2111T>C (isoleucine to threonine) were both predicted by in silico analysis to be ‘probably damaging’ to structure and function of the resulting protein. We conclude that NR3C1 polymorphisms occur in dogs and may cause individual variation in response to corticosteroid therapy.     

Characterization of canine filaggrin: gene structure and protein expression in dog skin

Background – Filaggrin (FLG) is a key protein for skin barrier formation and hydration of the stratum corneum. In humans, a strong association between FLG gene mutations and atopic dermatitis has been reported. Although similar pathogenesis and clinical manifestation have been argued in canine atopic dermatitis, our understanding of canine FLG is limited. Hypothesis/Objectives – The aim of this study was to determine the structure of the canine FLG gene and to raise anti‐dog FLG antibodies, which will be useful to detect FLG protein in dog skin. Methods – The structure of the canine FLG gene was determined by analysing the publicly available canine genome DNA sequence. Polyclonal anti‐dog FLG antibodies were raised based on the canine FLG sequence analysis and used for defining the FLG expression pattern in dog skin by western blotting and immunohistochemistry. Results – Genomic DNA sequence analysis revealed that canine FLG contained four units of repeated sequences corresponding to FLG monomer protein. Western blots probed with anti‐dog FLG monomer detected two bands at 59 and 54 kDa, which were estimated sizes. The results of immunohistochemistry showed that canine FLG was expressed in the stratum granulosum of the epidermis as a granular staining pattern in the cytoplasmic region. Conclusions and clinical importance – This study revealed the unique gene structure of canine FLG that results in production of FLG monomers larger than those of humans or mice. The anti‐dog FLG antibodies raised in this study identified FLG in dog skin. These antibodies will enable us to screen FLG‐deficient dogs with canine atopic dermatitis or ichthyosis.     

Normal canine excretory urogram: effects of dose, time, and individual dog variation

Ten healthy, young, adult mongrel dogs were given sodium iothalamate at dose levels of 200, 400, and 800 mg of iodine/0.45 kg of body weight on separate occasions by rapid IV injection; urinary bladders of the dogs were empty before injections were begun. Seven of the ten dogs were given an additional dose of sodium iothalamate (400 mg of iodine/0.45 kg) with the bladder partially distended with sterilized saline solution. Ventrodorsal abdominal radiographs were obtained immediately and at 5, 10, 20, 40, 60, and 120 minutes after injection of contrast medium. The kidneys, renal pelves, pelvic diverticula, and ureters were evaluated for radiographic density (radiopacity). The lengths and widths of the kidneys, pelves, and diverticula and the width of the ureters were determined, and those measurements were standardized by dividing the values by the corresponding length of the second lumbar vertebral body. From these evaluations, it was determined that postinjection radiographs should be obtained immediately and at 5, 20, and 40 minutes. The optimal dose of contrast medium was 400 mg of iodine/0.45 kg of body weight. It was also determined that the dose of contrast medium, as well as the time of postinjection radiography, significantly influenced many of the measurements (both linear and density) in the excretory urogram of normal dogs. Values for the measurements of the urinary structures based on the results of the present study are also presented.     

Achondroplastic dog breeds have no mutations in the transmembrane domain of the FGFR-3 gene.

One of the most common skeletal affections in humans is achondroplasia, a short-limbed dwarfism that is, in most cases, caused by mutations in the transmembrane domain of the fibroblast growth factor receptor-3 (FGFR-3) gene. Due to the lack of sufficient radiological, genetic, and molecular studies, most types of skeletal anomalies in dogs are classified as achondroplasia. To initiate the molecular characterization of some osteochondrodysplastic dog breeds, we obtained the DNA sequence of the transmembrane domain of the FGFR-3 gene from the dachshund, basset hound, bulldog, and German shepherd dogs. All 4 breeds showed no mutation in the evaluated region. This indicates that the mutation responsible for the osteochondrodysplastic phenotype in the tested dog breeds lies either elsewhere in the FGFR-3 gene or in other ones involved in the formation and development of endochondral bone.     

Allelic variation of the D4 dopamine receptor polymorphic region in two dog breeds, Golden retriever and Shiba

The D4 dopamine receptor (D4DR) polymorphic region, which is possibly related to the personality trait known as novelty seeking in humans, was examined in 34 dogs from two breeds (Golden retriever and the Japanese indigenous breed, Shiba) by the polymerase chain reaction (PCR), and the DNA sequences of each allele were determined. The polymorphic region of the dog D4DR gene was composed of 39- and 12- base pair (bp) units, and four alleles (A-D) were identified based on the number and/or order of these units. Intra- and inter-breed allele variations were observed. The frequency of the short A allele was dominant (78.9%) in the Golden retriever, while the long D allele was most common (46.7%) in the Shiba. These findings suggested that the allele frequency varied significantly between different breeds, and that analysis of the polymorphism in D4DR might be of use for understanding the behavioral traits of dogs.     

鸡形目黑素皮质素受体1基因多态性研究

为了对已知的鸡形目禽类黑素皮质素受体1(MC1R)基因进行生物信息学和分子进化分析,根据已发表的12种鸡形目物种MC1R基因序列(包括棕色来航鸡、红色原鸡、土佐本地鸡、日系乌骨鸡、白色来航鸡、日本鹌鹑、火鸡、蓝胸鹑、珠鸡、朝鲜鹌鹑、河北柴鸡和略阳乌鸡),利用多种生物学软件进行遗传多态性研究。结果显示:12种鸡形目物种MC1R基因编码区碱基序列同源性和氨基酸序列同源性均较高(分别为98.84%,98.91%),蛋白质二级结构相似度极高,以α构螺旋为主(57.32%⁶7.83%),无规则卷曲次之(25.80%67.83%),无规则卷曲次之(25.80%²9.62%),β-折叠最少(6.05%29.62%),β-折叠最少(6.05%¹5.61%)。红色原鸡、土佐本地鸡和棕色来航鸡的二级结构组成相同,其中红色原鸡和土佐本地鸡的碱基CDS序列完全相同。分子进化分析发现,12个鸡形目物种可以明显地分为A、B、C 3个类群,火鸡、珠鸡、鹌鹑、蓝胸鹑、棕色来航鸡、红色原鸡和土佐本地鸡聚为A群;河北柴鸡、朝鲜鹌鹑和日系乌骨鸡聚为B群;白色来航鸡和略阳乌鸡聚为C群。火鸡较其他鸡形目起源较早,棕色来航鸡,白色来航鸡和略阳乌鸡起源较迟。但聚类结果中白色来航鸡和棕色来航鸡却属于不同类群,这可能是由于MC1R基因参与家禽羽色的调控,而白色来航鸡和棕色来航鸡羽毛色素存在明显差异,从而造成了二者在鸡形目MC1R基因分子进化树中的遗传距离较远。     

Seroprevalence of canine herpesvirus-1 in Turkish dog population

Canine herpesvirus-1 (CHV-1) is the agent of reproductive and respiratory disorders in adult dogs, and the infection generally results in haemorrhagic disease conditions and neonatal death. In this study, virus neutralisation test that used complement (VNT) as well as in-house ELISA were utilised to investigate the CHV-1 seroprevalence in the Turkish dog population. Among the 560 serum samples, 39.3% of the samples tested by ELISA were CHV-1 positive while 29.4% of the samples tested by VNT were CHV-1 positive. Compared to the individual dogs (39.0%), there was a higher CHV-1 seroprevalence (62.1%) found in the colony dogs (62.1%) (p = 0.0002). However, there was an insignificant difference between male and female dogs. Although the highest antibody prevalence (56.7%) was found in Golden Retrievers, there were no significant variations detected among the dog breeds used in this study. Neutralizing antibody titres were very low (?1:16) in a high portion of the tested animals, confirming the rapid decrease of CHV-1 antibodies after the course of infection. The results of this study show that CHV-1 seroprevalence is moderately high in the Turkish dog population.     

Simultaneous identification of two MspI polymorphisms of the porcine myogenin gene in Hungarian breeds

Simultaneous analysis of two myogenin (MYOG) loci has been carried out using polymerase chain reaction‐restriction fragment length polymorphism (PCR‐RFLP) tests in DNA isolated from blood samples collected from four different pig breeds (Hungarian Large White, Duroc, Mangalica and Hungarian Landrace). The following MYOG^A frequency values have been obtained: Hungarian Large White 0.2416; Duroc 0.0900; Mangalica 0.6617; Hungarian Landrace 0.2361. The MYOG² allele was found in Duroc and Mangalica breeds only. This test allows the analysis of two loci of the MYOG gene in parallel, which might be used in marker assisted selection programmes in the near future. Ongoing studies on the possible effects of the MYOG types on carcass and growth traits are outlined.     

浅议警犬繁育工作中的种群

种群是用以实现某些特定目标而开展品种保持、培育和繁殖的基础群。种群在自然选择中是以“优胜劣汰”为规律,在人工选择中是以“高产、优质、高效、低耗”为准则。有人仍片面地认为,只要能繁殖的犬就可作为种犬,没有认识到种群在繁育工作中的地位,从而使一些低品质犬充斥到种群中,给繁育工作造成了较大的损失,这应引起业内人士的高度重视。     

Allele frequency distribution of the canine dopamine receptor D4 gene exon III and I in 23 breeds

Various canine breeds are remarkably different from each other not only in their sizes and shapes but also in behavioral traits, suggesting that some of them are under genetic control. Although dopaminergic neurotransmission system is considered to affect animal behavior, little is known about related genes in canine. Relations between specific alleles in polymorphic regions of the dopamine receptor D4 gene (DRD4) and personality or psychiatric disorders have been reported in humans, and we first found polymorphism in exon III region of the gene in 4 canine breeds. In this study we surveyed allele frequency distribution in 23 breeds including a total of 1,535 unrelated individuals. In exon III, 8 alleles including a novel allele were identified. A group of breeds in which the alleles 447b, 498 and 549 were frequent tended toward high scores in aggression-related behavioral traits than that with frequent alleles 435 and 447a. Moreover, a polymorphism based on 24 bp insertion/deletion was found in exon I region for the first time in dogs. This information may be of use for candidate gene studies of behavioral variation in dogs.