The effects of dog breed development on genetic diversity and the relative influences of performance and conformation breeding.

Genetic diversity was compared among eight dog breeds selected primarily for conformation (Standard Poodle, Italian Greyhound and show English Setter), conformation and performance (Brittany), predominantly performance (German Shorthaired and Wirehaired Pointers) or solely performance (field English Setter and Red Setter). Modern village dogs, which better reflect ancestral genetic diversity, were used as the standard. Four to seven maternal and one to two Y haplotypes were found per breed, with one usually dominant. Diversity of maternal haplotypes was greatest in village dogs, intermediate in performance breeds and lowest in conformation breeds. Maternal haplotype sharing occurred across all breeds, while Y haplotypes were more breed specific. Almost all paternal haplotypes were identified among village dogs, with the exception of the dominant Y haplotype in Brittanys, which has not been identified heretofore. The highest heterozygosity based on 24 autosomal microsatellites was found in village dogs and the lowest in conformation (show) breeds. Principal coordinate analysis indicated that conformation-type breeds were distinct from breeds heavily used for performance, the latter clustering more closely with village dogs. The Brittany, a well-established dual show and field breed, was also genetically intermediate between the conformation and performance breeds. The number of DLA-DRB1 alleles varied from 3 to 10 per breed with extensive sharing. SNPs across the wider DLA region were more frequently homozygous in all pure breeds than in village dogs. Compared with their village dog relatives, all modern breed dogs exhibit reduced genetic diversity. Genetic diversity was even more reduced among breeds under selection for show/conformation.

Possible relationships between canine hematopoietic neoplasia, other malignancies and immune mediated diseases.

In the last ten years few retroviruses have been isolated. The reasons for this lack of new discoveries are obvious, most retroviral investigations have been directed to HIV, HTLVI and HTLVII research with little research attention given to other species that may have retroviral infections, such as the dog. The domesticated dog is particularly important to investigate because they live in close proximity to human beings and have done so for at least the last 14 thousand years (1). This report gives relationships between canine hematopoietic, other neoplasia and immune mediated diseases in two families of inbred dogs that possibly were also infected with a yet to be fully described retrovirus. These findings should lead to a renewed interest in studying retroviral infections in dogs. The heterospecies relationship of retroviral infections closely related to human AIDS virus is of obvious scientific importance.

Risk factors associated with the incidence of seroconversion to caprine arthritis-encephalitis virus in goats on California dairies.

Incidence of seroconversion to caprine arthritis-encephalitis virus (CAEV) was determined for 1,194 goats on 11 dairies, using 2 repeated annual herd tests for CAEV. Current life table methods were used to compare age-specific incidence of seroconversion for pasteurized milk-raised and unpasteurized milk-raised goats. Logistic regression models were used to determine the risk factors associated with CAEV seroconversion, and to estimate odds ratios for seroconversion for various factor levels. Goats raised by unpasteurized milk-feeding methods were 2.5 to 6.7 times more likely to seroconvert than were goats raised by pasteurized milk-feeding methods, depending on the method of comparison. Similarly, 61.6 to 85.0% of seroconversions in yearling goats possibly were attributable to unpasteurized milk feeding. Among yearling goats, CAEV seroconversion was associated with feeding method, breed, and source of goat (herd of origin) when the effect of dairy was considered. In addition to the 6.7 times greater risk of seroconversion for unpasteurized milk-raised goats, yearling goats of the Saanen and Toggenburg breeds were 2.2 and 3.3 times, respectively, more likely to seroconvert than were Alpine yearling goats. Yearling goats purchased from another source were less likely to seroconvert than were yearlings raised on the dairy where they were studied. Among goats > 1 year old, age was associated with risk of seroconversion. Goats that were 3 years old or were > or = 4 years old were 1.7 and 3.2 times, respectively, more likely to seroconvert than were 2-year-old goats, when adjusted for effect of dairy. The effects of dairy were significant (P < or = 0.001) in yearling and older goats.

Preclinical study of sequential tumor necrosis factor and interleukin 2 in the treatment of spontaneous canine neoplasms.

Recombinant human tumor necrosis factor and recombinant human interleukin 2 were administered in a sequential schedule to 30 dogs with a variety of spontaneous neoplasms. Dose escalation of both drugs was performed, and a maximally tolerated dose of recombinant human tumor necrosis factor of 125 mg/m2 i.v. for 3 days, followed by 1.5 x 10(6) units/m2 of recombinant human interleukin 2 s.c. for 9 days, was derived. Dose-limiting toxicities were primarily gastrointestinal; however, weakness and malaise were seen during therapy at doses higher than the maximally tolerated dose. No clinically significant hematological toxicities were seen at any dose level. Objective tumor responses were seen in dogs with oral mucosal melanoma and cutaneous mastocytoma. Because of the histological, behavioral, and epidemiological similarities between human and canine tumor types, the canine cancer patient provides a unique model for the preclinical evaluation of recombinant cytokine therapy.

Studies with canine sera that contain antibodies which recognize human immunodeficiency virus structural proteins.

In a serological survey, using the immunoblotting technique, we found that substantial numbers of dog sera from both normal and diseased dogs, including dogs with neoplasia, reacted with one or more human immunodeficiency virus (HIV) recombinant proteins. A total of 144 dog sera were tested, and 72 (50%) of them reacted with one or more HIV recombinant structural proteins. Ten dog sera were also tested for reactivity with simian immunodeficiency virus (SIV), feline immunodeficiency virus (FIV), and caprine arthritis encephalitis virus (CAEV). Six dog sera reacted with at least the major core protein of HIV, while one of the dog sera tested reacted with SIV core protein, and there were no reactions with the viral proteins of either FIV or CAEV. Cell extracts from canine peripheral blood lymphocytes cocultivated with human cells and an extract of human cells infected with HIV were immunoblotted against dog sera which previously tested positive or negative on HIV recombinant protein commercially available Western blot strips. Two lymphocyte lysates and the HIV-infected Hut cell lysate reacted with the Western blot strip-positive dog serum; however, no reactions were seen with the Western blot strip-negative dog serum.

Clinical and lymphohematologic responses after bone marrow transplantation in sibling and unrelated donor-recipient pairs of cats.

Conditions necessary for establishment of a graft, posttransplant supportive care and complications, and lymphohematopoietic reconstitution after bone marrow transplantation were evaluated in 7 cats. Donor-recipient pairs were selected on the basis of low mutual reactivity in one-way mixed lymphocyte reactions. Before transplantation, cats were given marrow ablative (7 Gray) total-body gamma irradiation. Cyclosporine A was administered to cat 7, which was given marrow from an unrelated donor. Rapid hematologic recovery was attained in 5 of 5 (cats 1 to 5) sibling bone marrow recipients and 1 (cat 7; cyclosporine A-treated) of 2 recipients from unrelated donors. Lymphocyte recovery was prolonged, requiring up to 100 days to attain reference concentrations. Lymphocyte blastogenic responses were below reference range in 2 of 3 cats (cats 1 and 3) examined approximately 1 to 3 months after transplantation. Serum IgG concentrations determined 1 to 6 months after transplantation were within reference range in cats 1 to 5 which were given sibling bone marrow. Fatal infections did not develop in cats that had established grafts. Antimicrobial-responsive fevers did develop, but were generally detected only when granulocyte counts were low (less than 1 x 10(9) cells/L). Clinical signs of disease in the immediate posttransplant period consisted of hepatic lipidosis (fatal) in cat 4, hepatitis (mild graft-vs-host disease) in cat 3, and immune-mediated hemolytic anemia and thrombocytopenia in cat 7. Cats with hepatitis and immune-mediated disease responded to immunosuppressive therapy.

Immunohistological demonstration of virus and tumor associated antigens in tissues experimental and spontaneous bovine leukemia virus (BLV) infection.

Expression of bovine leukemia virus (BLV) antigens in vivo has not been shown. After BLV infection, however, production of antibodies directed towards BLV proteins (e.g. gp51) can be easily demonstrated. Thus, production of BLV proteins has to take place somewhere in infected cattle. Tissues and organs of experimentally infected cattle were fixed in acetone and embedded in paraffin. Monoclonal antibodies directed to gp51 were used to demonstrate BLV expression immunohistologically by the peroxidase-antiperoxidase (PAP) method. The same samples were also used to demonstrate a tumor associated antigen (TAA) employing a monoclonal antibody. Our results indicate that very few cells, found in the intestinal mucosa, produce gp51 in vivo. The expression of TAA, however, increases significantly shortly after infection with BLV and remains high throughout life.

The TP1 isolate of feline sarcoma virus encodes a fgr-related oncogene lacking gamma actin sequences.

We have isolated a new feline sarcoma virus, TP1-FeSV. The virus encodes a myristilated 83 kD gag-onc fusion protein displaying tyrosine kinase activity. We have established nonproducer cell lines lacking the TP1-FeSV associated helper virus (FeLV) and TP1-FeSV transfected NIH cell lines. Southern Blot analysis of genomic DNA and Northern Blot analysis of RNA isolated from these cell lines revealed that the oncogene of the TP1-FeSV isolate is related to the fgr oncogene of the GR-FeSV, but shows no hybridization to the gamma actin homologous sequences of the GR-FeSV. We have isolated TP1-FeSV specific clones from a genomic library. Restriction enzyme and sequence analysis showed that the TP1-FeSV genome consists of the first 1651 nucleotides of the gag gene, followed directly by fgr sequences. The TP1-FeSV fgr sequence starts 43 nucleotides after the beginning of the GR-FeSV fgr sequence. In contrast to the GR-FeSV fgr which has lost 13 amino acids of the c-fgr carboxy terminus, the TP1-FeSV fgr contains the complete carboxy terminus of the cellular fgr gene. The TP1-FeSV fgr sequence is followed by a unique 328 nucleotide long sequence of unknown origin. The 3' recombination site occurs within the pol gene, 460 nucleotides from the start of the env leader sequence. Comparison of the subcellular localization of the transforming proteins of TP1-FeSV and GR-FeSV show no striking difference; both molecules are in part associated with subcellular membrane/cytoskeletal fractions and form complexes with the cellular pp90 and pp50.

Cisplatin for treatment of transitional cell and squamous cell carcinomas in dogs.

Thirteen dogs with tumors were treated with monthly infusions of cisplatin. Complete responses were not observed. Of 8 dogs with urinary tract transitional cell carcinomas, 1 (12.5%) had a partial response of 31 weeks' duration, and 4 (50%) had stable disease for 12, 30, 32, and 34 weeks. Three (60%) of 5 dogs with head and neck squamous cell carcinomas had partial responses for 2, 10, and 15 weeks. All 13 dogs were evaluated for signs of toxicosis. Transient episodes of vomiting were recorded for 7 dogs (54%), and 2 dogs (15%) had mild thrombocytopenia. Although renal function gradually decreased in 2 dogs (15%), none of the dogs had an episode of acute renal failure attributable to cisplatin. These findings suggest that cisplatin may be a safe and potentially effective agent for treatment of transitional cell and squamous cell carcinomas in dogs.

Feline immunodeficiency syndrome--a comparison between feline T-lymphotropic lentivirus and feline leukemia virus.

A feline T-lymphotrophic lentvirus (FTLV) has recently been isolated from a domestic cat free of feline leukemia virus (FeLV). This virus is distinct from FeLV (an oncornavirus), although they share a common denominator, namely, the ability to cause immunosuppression and induce lymphadenopathy and anemia. Their differences can be revealed by examining the following: the metal requirement for reverse transcriptase activity, the antigenic comparison by Western blot analysis, the different susceptibilities of a variety of feline cells, and the morphology based on electron microscopy. In the serological survey of 1,612 cats surveyed in the USA, 232 (14.4%) were seropositive for antibodies to FTLV, which was lower than for the 42 Canadian cats surveyed of which 8 (19%) were seropositive. Of the 61 cats positive for FeLV, 15 (25%) were also positive for FTLV, giving the impression that coinfection between these two retroviruses plays an important role in the cliniocpathological signs of what was previously thought to be solely an FeLV syndrome.

Use of cisplatin for treatment of appendicular osteosarcoma in dogs.

Nineteen dogs were treated for osteosarcoma of the appendicular skeleton. Eleven dogs treated by amputation and adjunctive cisplatin chemotherapy had a significantly longer (P less than 0.003) median survival time of 43 weeks (range, 20 to 108 weeks) than did 8 dogs whose median survival time was 14.5 weeks (range, 8 to 46 weeks) after amputation alone. All 11 dogs given cisplatin were evaluated for signs of drug toxicosis. Transient episodes of vomiting were recorded in 9 of 11 dogs. Additional toxic effects included gradual decreases in endogenous creatinine clearance in 3 dogs and thrombocytopenia in 1 dog. On the basis of prolonged survival times and minimal adverse effects, we concluded that cisplatin has promise as an effective and relatively nontoxic agent, when combined with amputation, for treatment of dogs with osteosarcoma of the appendicular skeleton.

Frequency and significance of feline leukemia virus infection in necropsied cats.

Feline leukemia virus (FeLV) infection was diagnosed immunohistologically on paraffin-embedded tissues obtained from 1,095 necropsied cats. Significant association of FeLV infection was demonstrated by chi 2 and Fisher's tests with various conditions and diseases (ie, anemia, tumors of the leukemia/lymphoma complex, feline infectious peritonitis, bacterial infections, emaciation, FeLV-associated enteritis, lymphatic hyperplasia, and hemorrhage). Unexpected findings associated with FeLV infection were icterus, several types of hepatitis, and liver degeneration. A negative association with FeLV infection was found for most parasitic and viral infections, including feline panleukopenia. Neither positive nor negative associations were established for FeLV infection and most forms of nephritis, including severe glomerulonephritis. Feline leukemia virus-infected cats were significantly (Kruskal-Wallis test) older than were FeLV-negative cats with the same nonneoplastic FeLV-associated diseases.

Response to isoantigens and mitogens in the cat: effects of cyclosporin A.

Growing interest in the use of domesticated cats to study immune-mediated disease and in the research and clinical application of organ and tissue transplantation provided the need for in vitro assays estimating histocompatibility and cell-mediated immunity. In the present study, a whole-blood lymphocyte-stimulation microassay and mixed lymphocyte-response assay were adapted and used to measure significant responses from feline lymphocytes to mitogen and lymphocyte-defined histocompatibility antigens. The addition of cyclosporin A to whole-blood lymphocyte-stimulation microassay and mixed lymphocyte-response assay produced similar non-cytotoxic decrease in lymphocyte responses seen in other species.

Production of monoclonal antibodies. The effect of hybridoma concentration on the yield of antibody-producing clones.

The relationship between the number of hybrid clones in each well of the primary culture, the yield of antibody-producing wells and the short-term stability of the polyclonal cultures was tested. The results of the study demonstrate that a higher yield of antibody-secreting clones was obtained by employing lower density of hybrids and that instability of antibody secretion by polyclonal cultures is not the result of overgrowth of non-secreting clones.

Monoclonal antibodies to the v-fes product and to feline leukemia: virus P27 interspecies-specific determinants encoded by feline sarcoma viruses.

Monoclonal antibodies to p27 gag and v-fes specific determinants on the gag-onc poly-protein encoded by Snyder-Theilen feline sarcoma virus (ST-FeSV) were prepared. In order to obtain hybridoma clones specific to the antigenic determinants encoded by the FeSV genome, Lou rats were immunized with ST-FeSV-transformed, virus-nonproducing syngeneic cells, and boosted with either the same cells or affinity-purified feline leukemia virus (FeLV) p27. Three distinct clones reactive to both FeLV p27 and p85gag-fes, and one clone specific for a p85fes determinant were established. The anti-p27 monoclonal antibodies also reacted with the polyproteins p95gag-fes and p83gag-fgr, from Gardner-Arnstein (GA) and Theilen-Pedersen (TP1) FeSV, respectively. The anti-p27 monoclonal antibodies reacted with MuLV p30 and RD114 p28 but not with RSV, MMTV, or BLV. These results indicated that the part of the p27 gag gene that is preserved in ST-, GA, and TP1-FeSV encodes interspecies-specific p27 determinants.