CanISO: a database of genomic and transcriptomic variations in domestic dog (Canis lupus familiaris).

BACKGROUND: The domestic dog, Canis lupus familiaris, is a companion animal for humans as well as an animal model in cancer research due to similar spontaneous occurrence of cancers as humans. Despite the social and biological importance of dogs, the catalogue of genomic variations and transcripts for dogs is relatively incomplete. RESULTS: We developed CanISO, a new database to hold a large collection of transcriptome profiles and genomic variations for domestic dogs. CanISO provides 87,692 novel transcript isoforms and 60,992 known isoforms from whole transcriptome sequencing of canine tumors (N = 157) and their matched normal tissues (N = 64). CanISO also provides genomic variation information for 210,444 unique germline single nucleotide polymorphisms (SNPs) from the whole exome sequencing of 183 dogs, with a query system that searches gene- and transcript-level information as well as covered SNPs. Transcriptome profiles can be compared with corresponding human transcript isoforms at a tissue level, or between sample groups to identify tumor-specific gene expression and alternative splicing patterns. CONCLUSIONS: CanISO is expected to increase understanding of the dog genome and transcriptome, as well as its functional associations with humans, such as shared/distinct mechanisms of cancer. CanISO is publicly available at https://www.kobic.re.kr/caniso/ .

Exploring the genetic diversity of genotypes G8 and G10 of the Echinococcus canadensis cluster in Europe based on complete mitochondrial genomes (13 550-13 552 bp).

Echinococcus granulosus sensu lato is a group of tapeworm species known to cause cystic echinococcosis. Within this group, the Echinococcus canadensis cluster includes genotypes G8 and G10 that have a predominantly sylvatic life cycle ­ transmission occurs between wild cervids and wolves. Relatively few studies have explored the genetic variation of the elusive G8 and G10, and their extent of genetic variation is yet to be investigated at the complete mitochondrial (mt) genome level. The aim was to explore the genetic variation of these 2 genotypes in Europe using complete mtDNA sequences and provide a high-quality reference dataset for future studies. Sequences of complete mt genomes were produced for 29 samples of genotype G8 and G10 from wolves, moose, reindeer and roe deer, originating from Finland, Sweden, Russia, Poland, Latvia and Estonia. Genetic variation was explored based on phylogenetic network analysis, revealing marked differences between G8 and G10 (over 400 mutations), and more detailed patterns of variability within the 2 genotypes than previously observed. Understanding the mt genetic composition of a species provides a baseline for future studies aiming to understand whether this mt distinctiveness is mirrored in the nuclear genome and whether it has any impact on any phenotypic traits or parasite transmission.

Grey wolf genomic history reveals a dual ancestry of dogs.

The grey wolf (Canis lupus) was the first species to give rise to a domestic population, and they remained widespread throughout the last Ice Age when many other large mammal species went extinct. Little is known, however, about the history and possible extinction of past wolf populations or when and where the wolf progenitors of the present-day dog lineage (Canis familiaris) lived1-8. Here we analysed 72 ancient wolf genomes spanning the last 100,000 years from Europe, Siberia and North America. We found that wolf populations were highly connected throughout the Late Pleistocene, with levels of differentiation an order of magnitude lower than they are today. This population connectivity allowed us to detect natural selection across the time series, including rapid fixation of mutations in the gene IFT88 40,000-30,000 years ago. We show that dogs are overall more closely related to ancient wolves from eastern Eurasia than to those from western Eurasia, suggesting a domestication process in the east. However, we also found that dogs in the Near East and Africa derive up to half of their ancestry from a distinct population related to modern southwest Eurasian wolves, reflecting either an independent domestication process or admixture from local wolves. None of the analysed ancient wolf genomes is a direct match for either of these dog ancestries, meaning that the exact progenitor populations remain to be located.

Mitochondrial DNA alterations in the domestic dog (Canis lupus familiaris) and their association with development of diseases: A review.

Currently, the issue of the aetiology of mitochondrial diseases resulting from mitochondrial DNA (mtDNA) defects is underestimated. Genetic research is mostly focused on alterations in the nuclear genome (nDNA), and its impact on disease development as well as further health consequences without considering mtDNA abnormalities. However, in the case of energy-dependent diseases, it is important to understand the bioenergetic pathophysiology and its relation with mtDNA changes. In the current animal research, there is limited data about mtDNA defects and their association with the development of bioenergetic diseases in the domestic dog (Canis lupus familiaris) in contrast to human medicine, where mitochondrial genetics research has recently increased. Molecular findings about mtDNA indicate that improper functioning of mitochondria resulting from genetic defects of mtDNA has a severe impact on cells and tissues, especially those that are heavily dependent on oxidative metabolism such as the brain, skeletal and cardiac muscles and, consequently, the whole organism. The aim of this paper is to highlight the role of defects of mitochondria and mtDNA on the development and course of different diseases in the domestic dog. The field of canine mitochondrial genetics and genomics is definitely inexhaustible and it is worth drawing attention to the importance and consequences of the mitochondrial genome alterations. This review collects scientific data on mitochondrial DNA with special regard to the structure, features of canine mtDNA, and abnormalities in the mitochondrial genome and their association with the course and development of diseases, including mitochondrial myopathies, encephalopathies, and tumours.

Variants That Differentiate Wolf and Dog Populations Are Enriched in Regulatory Elements.

Research on the genetics of domestication most often focuses on the protein-coding exons. However, exons cover only a minor part (1-2%) of the canine genome, whereas functional mutations may be located also in regions beyond the exome, in regulatory regions. Therefore, a large proportion of phenotypical differences between dogs and wolves may remain genetically unexplained. In this study, we identified variants that have high allelic frequency differences (i.e., highly differentiated variants) between wolves and dogs across the canine genome and investigated the potential functionality. We found that the enrichment of highly differentiated variants was substantially higher in promoters than in exons and that such variants were enriched also in enhancers. Several enriched pathways were identified including oxytocin signaling, carbohydrate digestion and absorption, cancer risk, and facial and body features, many of which reflect phenotypes of potential importance during domestication, including phenotypes of the domestication syndrome. The results highlight the importance of regulatory mutations during dog domestication and motivate the functional annotation of the noncoding part of the canine genome.

The evolutionary history of dogs in the Americas.

Dogs were present in the Americas before the arrival of European colonists, but the origin and fate of these precontact dogs are largely unknown. We sequenced 71 mitochondrial and 7 nuclear genomes from ancient North American and Siberian dogs from time frames spanning ~9000 years. Our analysis indicates that American dogs were not derived from North American wolves. Instead, American dogs form a monophyletic lineage that likely originated in Siberia and dispersed into the Americas alongside people. After the arrival of Europeans, native American dogs almost completely disappeared, leaving a minimal genetic legacy in modern dog populations. The closest detectable extant lineage to precontact American dogs is the canine transmissible venereal tumor, a contagious cancer clone derived from an individual dog that lived up to 8000 years ago.

DNA Sequence Variants in the Five Prime Untranslated Region of the Cyclooxygenase-2 Gene Are Commonly Found in Healthy Dogs and Gray Wolves.

The aim of this study was to investigate the frequency of regional DNA variants upstream to the translation initiation site of the canine Cyclooxygenase-2 (Cox-2) gene in healthy dogs. Cox-2 plays a role in various disease conditions such as acute and chronic inflammation, osteoarthritis and malignancy. A role for Cox-2 DNA variants in genetic predisposition to canine renal dysplasia has been proposed and dog breeders have been encouraged to select against these DNA variants. We sequenced 272-422 bases in 152 dogs unaffected by renal dysplasia and found 19 different haplotypes including 11 genetic variants which had not been described previously. We genotyped 7 gray wolves to ascertain the wildtype variant and found that the wolves we analyzed had predominantly the second most common DNA variant found in dogs. Our results demonstrate an elevated level of regional polymorphism that appears to be a feature of healthy domesticated dogs.

HIV/AIDS epidemic in the State of Amazonas: characteristics and trends from 2001 to 2012

A scoping review was conducted to describe the epidemiological characteristics of the human immunodeficiency virus/acquired immunodeficiency syndrome (HIV/AIDS) epidemic in the State of Amazonas, Brazil, from 2001 to 2012, and temporary patterns were estimated from surveillance data. The results suggest that in its third decade, the Amazon HIV/AIDS epidemic is far from being stabilized and displays rising AIDS incidence and mortality rates and late diagnoses. The data suggest that AIDS cases are hitting mostly young adults and have recently shifted toward men, both homosexual and heterosexual. AIDS cases among the indigenous people have remained stable and low. However, the epidemic has disseminated to the interior of the state, which adds difficulties to its control, given the geographical isolation, logistical barriers, and culturally and ethnically diverse population. Antiretroviral (ARV) therapy has been decentralized, but peripheral ARV services are still insufficient and too distant from people who need them. Recently, the expansion of point-of-care (POC) rapid HIV testing has been contributing to overcoming logistical barriers. Other new POC devices, such as the PIMA CD4 analyzer, will bring the laboratory to the patient. AIDS uniquely coexists with other tropical infections, sharing their epidemiological profiles. The increased demand for HIV/AIDS care services can only be satisfied through increased decentralization to peripheral health units, which can also naturally integrate care with other tropical infections and can promote a shift from vertical to integrated programming. Future challenges involve building surveillance data on HIV case notification and covering the spectrum of engagement in care, including adherence to treatment and follow-up loss.

Characterization and allelic variation of the transporters associated with antigen processing (TAP) genes in the domestic dog (Canis lupus familiaris).

The function of the transporters associated with antigen processing (TAP) complex is to shuttle antigenic peptides from the cytosol to the endoplasmic reticulum to load MHC class I molecules for CD8(+) T-cell immunosurveillance. Here we report the promoter and coding regions of the canine TAP1 and TAP2 genes, which encode the homologous subunits forming the TAP heterodimer. By sampling genetically divergent breeds, polymorphisms in both genes were identified, although there were few amino acid differences between alleles. Splice variants were also found. When aligned to TAP genes of other species, functional regions appeared conserved, and upon phylogenetic analysis, canine sequences segregated appropriately with their orthologs. Transfer of the canine TAP2 gene into a murine TAP2-defective cell line rescued surface MHC class I expression, confirming exporter function. This data should prove useful in investigating the association of specific TAP defects or alleles with immunity to intracellular pathogens and cancer in dogs.

The genomics of selection in dogs and the parallel evolution between dogs and humans.

The genetic bases of demographic changes and artificial selection underlying domestication are of great interest in evolutionary biology. Here we perform whole-genome sequencing of multiple grey wolves, Chinese indigenous dogs and dogs of diverse breeds. Demographic analysis show that the split between wolves and Chinese indigenous dogs occurred 32,000 years ago and that the subsequent bottlenecks were mild. Therefore, dogs may have been under human selection over a much longer time than previously concluded, based on molecular data, perhaps by initially scavenging with humans. Population genetic analysis identifies a list of genes under positive selection during domestication, which overlaps extensively with the corresponding list of positively selected genes in humans. Parallel evolution is most apparent in genes for digestion and metabolism, neurological process and cancer. Our study, for the first time, draws together humans and dogs in their recent genomic evolution.

Mitochondrial capture by a transmissible cancer.

Canine transmissible venereal tumor (CTVT) is an infectious cell line circulating in many feral dog populations. It originated once, about 10,000 years ago. Phylogenetic analyses of mitochondrial sequences from dogs, wolves, and a geographically diverse collection of CTVT samples indicate that the cancer has periodically acquired mitochondria from its host. We suggest that this may be because the cancer's own mitochondria have a tendency to degenerate, due to high mutation rates and relaxed selection, resulting in host mitochondria being more fit.

Analysis of Canis mitochondrial DNA demonstrates high concordance between the control region and ATPase genes.

BACKGROUND: Phylogenetic studies of wild Canis species have relied heavily on the mitochondrial DNA control region (mtDNA CR) to infer species relationships and evolutionary lineages. Previous analyses of the CR provided evidence for a North American evolved eastern wolf (C. lycaon), that is more closely related to red wolves (C. rufus) and coyotes (C. latrans) than grey wolves (C. lupus). Eastern wolf origins, however, continue to be questioned. Therefore, we analyzed mtDNA from 89 wolves and coyotes across North America and Eurasia at 347 base pairs (bp) of the CR and 1067 bp that included the ATPase6 and ATPase8 genes. Phylogenies and divergence estimates were used to clarify the evolutionary history of eastern wolves, and regional comparisons of nonsynonomous to synonomous substitutions (dN/dS) at the ATPase6 and ATPase8 genes were used to elucidate the potential role of selection in shaping mtDNA geographic distribution. RESULTS: We found high concordance across analyses between the mtDNA regions studied. Both had a high percentage of variable sites (CR = 14.6%; ATP = 9.7%) and both phylogenies clustered eastern wolf haplotypes monophyletically within a North American evolved lineage apart from coyotes. Divergence estimates suggest the putative red wolf sequence is more closely related to coyotes (DxyCR = 0.01982 +/- 0.00494 SD; DxyATP = 0.00332 +/- 0.00097 SD) than the eastern wolf sequences (DxyCR = 0.03047 +/- 0.00664 SD; DxyATP = 0.00931 +/- 0.00205 SD). Neutrality tests on both genes were indicative of the population expansion of coyotes across eastern North America, and dN/dS ratios suggest a possible role for purifying selection in the evolution of North American lineages. dN/dS ratios were higher in European evolved lineages from northern climates compared to North American evolved lineages from temperate regions, but these differences were not statistically significant. CONCLUSIONS: These results demonstrate high concordance between coding and non-coding regions of mtDNA, and provide further evidence that the eastern wolf possessed distinct mtDNA lineages prior to recent coyote introgression. Purifying selection may have influenced North American evolved Canis lineages, but detection of adaptive selection in response to climate is limited by the power of current statistical tests. Increased sampling and development of alternative analytical tools will be necessary to disentangle demographic history from processes of natural selection.

Origins and evolution of a transmissible cancer.

Canine transmissible venereal tumor (CTVT) is an infectious disease of dogs. Remarkably, the infectious agent is the cancerous cell itself. To investigate its origin and spread, we collected 37 tumor samples from four continents and determined their evolutionary relationships using microsatellite length differences and microarray-based comparative genomic hybridization (aCGH). The different tumors show very little microsatellite variation, and the pattern of variation that does exist is consistent with a purely asexual mode of transmission. Approximately one quarter of the loci scored by aCGH show copy number variation relative to normal dogs, again with little variation among different tumor samples. Sequence analysis of the RPPH1 gene indicates an origin from either dogs or wolves, and microsatellite analysis indicates that the tumor is more than 6000 years old, and perhaps originated when dogs were first domesticated. By contrast, the common ancestor of extant tumors lived within the last few hundred years, long after the first tumor. The genetic and genomic patterns we observe are typical of those expected of asexual pathogens, and the extended time since first origin may explain the many remarkable adaptations that have enabled this mammalian cell lineage to live as a unicellular pathogen.

Endangered wolves cloned from adult somatic cells.

Over the world, canine species, including the gray wolf, have been gradually endangered or extinct. Many efforts have been made to recover and conserve these canids. The aim of this study was to produce the endangered gray wolf with somatic cell nuclear transfer (SCNT) for conservation. Adult ear fibroblasts from a female gray wolf (Canis lupus) were isolated and cultured in vitro as donor cells. Because of limitations in obtaining gray wolf matured oocytes, in vivo matured canine oocytes obtained by flushing the oviducts from the isthmus to the infundibulum were used. After removing the cumulus cells, the oocyte was enucleated, microinjected, fused with a donor cell, and activated. The reconstructed cloned wolf embryos were transferred into the oviducts of the naturally synchronized surrogate mothers. Two pregnancies were detected by ultrasonography at 23 days of gestation in recipient dogs. In each surrogate dog, two fetal sacs were confirmed by early pregnancy diagnosis at 23 days, but only two cloned wolves were delivered. The first cloned wolf was delivered by cesarean section on October 18, 2005, 60 days after embryo transfer. The second cloned wolf was delivered on October 26, 2005, at 61 days postembryo transfer. Microsatellite analysis was performed with genomic DNA from the donor wolf, the two cloned wolves, and the two surrogate female recipients to confirm the genetic identity of the cloned wolves. Analysis of 19 microsatellite loci confirmed that the cloned wolves were genetically identical to the donor wolf. In conclusion, we demonstrated live birth of two cloned gray wolves by nuclear transfer of wolf somatic cells into enucleated canine oocyte, indicating that SCNT is a practical approach for conserving endangered canids.

Transferability of short tandem repeat markers for two wild Canid species inhabiting the Brazilian Cerrado

The maned wolf (Chrysocyon brachyurus) and the crab-eating fox (Cerdocyon thous) are two wild-canid species found in the Brazilian Cerrado. We tested cross-amplification and transferability of 29 short tandem repeat primers originally developed for cattle and domestic dogs and cats on 38 individuals of each of these two species, collected in the Emas National Park, which is the largest national park in the Cerrado region. Six of these primers were successfully transferred (CSSM-038, PEZ-05, PEZ-12, LOCO-13, LOCO-15, and PEZ-20); five of which were found to be polymorphic. Genetic parameter values (number of alleles per locus, observed and expected heterozygosities, and fixation indices) were within the expected range reported for canid populations worldwide.