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Background Dissecting the role copy number variants (CNVs) play in disease pathogenesis is directly reliant on accurate methods for quantification. The Shar-Pei dog breed is predisposed to a complex autoinflammatory disease with numerous clinical manifestations. One such sign, recurrent fever, was previously shown to be significantly associated with a novel, but unstable CNV (CNV_16.1). Droplet digital PCR (ddPCR) offers a new mechanism for CNV detection via absolute quantification with the promise of added precision and reliability. The aim of this study was to evaluate ddPCR in relation to quantitative PCR (qPCR) and to assess the suitability of the favoured method as a genetic test for Shar-Pei Autoinflammatory Disease (SPAID). Results One hundred and ninety-six individuals were assayed using both PCR methods at two CNV positions (CNV_14.3 and CNV_16.1). The digital method revealed a striking result. The CNVs did not follow a continuum of alleles as previously reported, rather the alleles were stable and pedigree analysis showed they adhered to Mendelian segregation. Subsequent analysis of ddPCR case/control data confirmed that both CNVs remained significantly associated with the subphenotype of fever, but also to the encompassing SPAID complex (p

Immunoglobulin A deficiency (IgAD) is the most common primary immune deficiency disorder in both humans and dogs, characterized by recurrent mucosal tract infections and a predisposition for allergic and other immune mediated diseases. In several dog breeds, low IgA levels have been observed at a high frequency and with a clinical resemblance to human IgAD. In this study, we used genome-wide association studies (GWAS) to identify genomic regions associated with low IgA levels in dogs as a comparative model for human IgAD. We used a novel percentile groups-approach to establish breed-specific cut-offs and to perform analyses in a close to continuous manner. GWAS performed in four breeds prone to low IgA levels (German shepherd, Golden retriever, Labrador retriever and Shar-Pei) identified 35 genomic loci suggestively associated (p

Autoinflammatory disease (AID) manifests from the dysregulation of the innate immune system and is characterised by systemic and persistent inflammation. Clinical heterogeneity leads to patients presenting with one or a spectrum of phenotypic signs, leading to difficult diagnoses in the absence of a clear genetic cause. We used separate genome-wide SNP analyses to investigate five signs of AID (recurrent fever, arthritis, breed specific secondary dermatitis, otitis and systemic reactive amyloidosis) in a canine comparative model, the pure bred Chinese Shar-Pei. Analysis of 255 DNA samples revealed a shared locus on chromosome 13 spanning two peaks of association. A three-marker haplotype based on the most significant SNP (p

A multiple immunodeficiency, involving antibody- and cell-mediated responses in 10 Chinese Shar-Pei (CSP) dogs is described. Abnormal levels of serum IgM and IgA in most cases, and IgG in fewer cases characterized the immunoglobulin deficiencies. Decreased in vitro proliferative responses of pokeweed mitogen (PWM)-stimulated peripheral blood mononuclear cells (PBMC) were found in nine cases. Clinical presentation involved several organ systems and was associated with recurrent infections and malignancy. Sera from affected dogs suppressed PWM-stimulated cell proliferation of affected and normal dogs, but not cultures stimulated with PWM followed by recombinant IL-2 (rIL-2). In vitro supplementation of PBMC cultures with immunomodulatory guanosine analogs (GA) resulted in increased de novo IgG and/or interleukin-6 (IL-6) synthesis. Cells from five immunodeficient dogs showed in vitro evidence of GA- or rIL-2-dependent enhanced immunological responses. Since rIL-2-mediated activation of the IL-2 receptor and GA-mediated immunomodulation are reported to act through protein kinase C (PKC)-independent pathways, it is concluded that the IL-2 receptor is functional in these dogs and that cell activation through alternative pathways may restore immune responses in affected CSP dogs.

Familial Mediterranean Fever (FMF) is a human disorder characterized by recurrent fever of unknown origin (RFUO), renal amyloidosis, and evidence of peritonitis, pleuritis, and/or synovitis. This report suggests that Chinese Shar-pei (CSP) dogs suffer from a similar syndrome. CSP dogs with RFUO (n = 15) showed greater levels of IL-6 in serum than normal controls, hypergammaglobulinemia, and normal or supranormal in vitro lymphocyte blastogenesis in response to mitogen stimulation, when compared to healthy afebrile dogs. In patients 2 years old or older, RFUO was associated with renal failure, renal amyloidosis, and swollen joints. An epidemiological survey of privately owned dogs indicated a RFUO prevalence of 23% in CSP dogs (n = 132) and 1% in dogs of all breeds (n = 98). Increased levels of circulating cytokines, such as IL-6, have been shown to influence such processes as the febrile response, antibody production, and the synthesis of amyloid precursors. We propose that CSP dogs with RFUO, renal amyloidosis, and joint inflammation may serve as an animal model of FMF and that the clinical syndrome is associated with elevated levels of circulating IL-6.

Hereditary periodic fever syndromes are characterized by recurrent episodes of fever and inflammation with no known pathogenic or autoimmune cause. In humans, several genes have been implicated in this group of diseases, but the majority of cases remain unexplained. A similar periodic fever syndrome is relatively frequent in the Chinese Shar-Pei breed of dogs. In the western world, Shar-Pei have been strongly selected for a distinctive thick and heavily folded skin. In this study, a mutation affecting both these traits was identified. Using genome-wide SNP analysis of Shar-Pei and other breeds, the strongest signal of a breed-specific selective sweep was located on chromosome 13. The same region also harbored the strongest genome-wide association (GWA) signal for susceptibility to the periodic fever syndrome (praw = 2.3×10−6, pgenome = 0.01). Dense targeted resequencing revealed two partially overlapping duplications, 14.3 Kb and 16.1 Kb in size, unique to Shar-Pei and upstream of the Hyaluronic Acid Synthase 2 (HAS2) gene. HAS2 encodes the rate-limiting enzyme synthesizing hyaluronan (HA), a major component of the skin. HA is up-regulated and accumulates in the thickened skin of Shar-Pei. A high copy number of the 16.1 Kb duplication was associated with an increased expression of HAS2 as well as the periodic fever syndrome (p, This work was supported by the Swedish Research Council; FORMAS; the Swedish Research Council for Environment, Agricultural Sciences, and Spatial Planning; the Swedish Foundation for Strategic Research; in part by the Intramural Research Program of the National Institute of Arthritis and Musculoskeletal and Skin Diseases and the National Human Genome Research Institute of the National Institutes of Health; the Chinese Shar-Pei Charitable Trust and the European Commission (FP7-LUPA, GA-201370). KL-T is the recipient of a EURYI award from the European Science Foundation.

Renal amyloidosis (RA) and recurrent fever of unknown origin (RFUO) are characteristics of familial Mediterranean fever (FMF), a human disorder inherited as an autosomal-recessive trait. Although no animal model has been established for FMF, a similar syndrome of RFUO and RA has been reported in Chinese Shar-pei (CSP) dogs. This report addresses two questions: (1) Is RA inherited in CSP dogs? (2) If it is, is it possible to hypothesize the type of inheritance involved? Two studies were conducted to answer these questions: a historical cross-sectional comparison, which included CSP and non-CSP dogs with RA; and a prospective study that included CSP dogs with RA, RFUO, or both. The cross-sectional comparison resulted in an odds ratio of 10 for RA in CSP dogs under 7 years of age. The prospective study of 28 dogs with RA or RFUO identified 20 that had RFUO and RA, three with RA alone, and five with RFUO alone. RFUO preceded RA in all cases with both conditions. The RFUO/RA combination was observed in both sexes. Four dogs with RFUO with or without RA were born to parents that either were alive at age 7 or had died because of conditions other than kidney failure/RA. When one parent was known to express one of these conditions, the prevalence of RA was between 25% and 50% among littermates.(ABSTRACT TRUNCATED AT 250 WORDS)

This study illustrates the use of a student-managed response-cost system using free tokens to modify the on-task behavior of 2 mainstreamed students-an emotionally disturbed fourth grader and a learning disabled sixth grader-in regular classrooms. After the program was implemented, the subjects' on-task behavior as well as their academic performance improved. Applications of the findings to regular classrooms are discussed.