Detection of equine X chromosome mosaicism in a mare using an equine X whole chromosome painting probe (WCPP)--a case report

An infertile mare with hypoplastic ovaries was subjected to cytogenetic analysis. Fluorescence in situ hybridisation (FISH) using the equine X whole chromosome painting probe (WCPP) was carried out on a chromosome preparation obtained from blood lymphocyte culture. The number of analysed spreads was high (235) and in the X chromosome aneuploidy in mosaic form was diagnosed. The karyotype formula was 63,X / 64,XX / 65,XXX. The ratio of the three lines was 15%, 82% and 3%, respectively. The application of the FISH technique with WCPP is discussed.     

Gait abnormalities and ridden horse behaviour in a convenience sample of the United Kingdom ridden sports horse and leisure horse population

The objectives of this study were to compare horses’ gaits in hand and when ridden; to assess static and dynamic saddle fit for each horse and rider; to apply the Ridden Horse Pain Ethogram (RHpE) and relate the findings to gait abnormalities consistent with musculoskeletal pain, rider position and balance and saddle fit; and to document noseband use and its relationship with mouth opening during ridden exercise. Data were acquired prospectively from a convenience sample of horses believed by their owners to be working comfortably. All assessments were subjective. Gait in hand and when ridden were evaluated independently, by two assessors, and compared using McNemar’s test. Static tack fit and noseband type were recorded. Movement of the saddle during ridden exercise, rider position, balance and size relative to the saddle was documented. RHpE scores were based on assessment of video recordings. Multivariable Poisson regression analysis was used to determine factors which influenced the RHpE scores. Of 148 horses, 28.4% were lame in hand, whereas 62.2% were lame ridden (P<0.001). Sixty per cent of horses showed gait abnormalities in canter. The median RHpE score was 8/24 (interquartile range 5, 9; range 0, 15). There was a positive association between lameness and the RHpE score (P<0.001). Riding School horses had higher RHpE scores compared with General Purpose horses (P = 0.001). Saddles with tight tree points (P = 0.001) and riders seated at the back of the saddle rather than the middle (P = 0.001) were associated with higher RHpE scores. Horses wearing crank cavesson compared with cavesson nosebands had higher RHpE scores (P = 0.006). There was no difference in mouth opening, as defined by the RHpE, in horses with a noseband with the potential to restrict mouth opening, compared with a correctly fitted cavesson noseband, or no noseband. It was concluded that lameness or gait abnormalities in canter may be missed unless horses are assessed ridden.     

Further cases of equine sex chromosome abnormalities

Sex chromosome abnormalities have been detected in a further five mares with clinical histories of small ovaries and absent or irregular oestrous cycles. Three mares had 63,XO karyotypes (X monosomy) and two were sex chromosome mosaics with karyotypes of 63,XO/64,XY and 63,XO/64,XX/64,XY respectively. A sex chromosome abnormality (X monosomy) has also been found in a filly where it was suspected because of her short stature.     

Teratoma Associated With Testicular Tissue in a Female-Like Horse With 64,XY (SRY-Positive) Disorder of Sex Development

A four-year-old female-like Quarter Horse was admitted for clinical evaluation because of masculinized-aggressive behavior and lack of estrous signs. On external inspection, a hypertrophied clitoris and prominent muscles were observed. On gynecological examination, apparently normal mammary glands, vulva, vagina, and cervix were noted. On the other hand, the uterus had no tone and was smaller than normal. The left gonad was very soft on palpation and the right gonad was mostly firm, irregular, and wider than the left gonad. On ultrasound examination, there were no signs of regular ovarian structure or follicular activity. Because of the different shapes and consistencies of the gonads and a suspicion of increased testosterone production, a bilateral gonadectomy was recommended. Blood was collected for testosterone levels quantification and for cytogenetic and molecular investigations. After removal, gonads were analyzed macroscopically and sections were sent for histopathological examination. A final diagnosis of benign adult teratoma associated with seminiferous tubules and Leydig cells was made. On cytogenetic and molecular analyses, a normal diploid number of 64 chromosomes and the presence of the XY sex chromosomes were seen in all cells, as well as the SRY gene. Testosterone levels were higher than normal before surgery and were reduced after gonads removal. In conclusion, the masculinized behavior was probably caused by increased testosterone levels produced by testicular tissue, in a female-like horse with 64,XY SRY-positive disorder of sex development, which was associated with a teratoma.     

Treatment of clinically diagnosed equine sarcoid with a mistletoe extract (Viscum album austriacus)

Background: Equine sarcoids (ES) are common, difficult to treat, and have high recurrence rates. Viscum album extracts (VAE) are used in human cancer treatment. Hypothesis: That therapy with VAE (Iscador P) is effective in the treatment of ES. Animals: Fifty‐three horses (444 ES); 42 were treated with VAE or placebo as monotherapy; 11 were treated with VAE or placebo after selective excision of ES. Methods: Prospective, randomised, blinded, clinical trial. Horses were randomly assigned to treatment (VAE; n = 32) or control group (Placebo; n = 21). One milliliter of VAE (Iscador P) in increasing concentrations from 0.1 to 20 mg/mL or physiological NaCl solution was given SC 3 times a week over 105 days. Number, localization, and type of the ES were documented over 12 months. A subset of 163 clinically diagnosed equine sarcoid (CDES) lesions (95 VAE, 68 Placebo) was evaluated in detail, considering clinical findings and tumor volume. Results: No undesired adverse effects were observed except for mild edema at the injection site in 5 of 32 horses (16%). Complete or partial regression was observed in 13 horses of the VAE group (41%) and in 3 of the control horses (14%; P < .05). After VAE treatment, 48 of 95 CDES (67%) showed an improvement compared with 17 of 68 CDES in the control group (40%; P < .01). Twenty‐seven CDES had disappeared completely in the VAE group (38%) compared with 9 CDES in the control group (13% NS). Conclusions and Clinical Importance: VAE (Iscador P) represents a safe and effective treatment for CDES.     

Preliminary analysis of genomic abnormalities in canine meningiomas

Cytogenetic detection of unbalanced genomic aberrations in tumours is a strategy for the identification of tumour suppressor genes and oncogenes. When considered in concert with clinical data, the approach also represents a means of identifying markers of prognosis. In a preliminary investigation of the molecular basis of canine meningioma tumorigenesis, we profiled three tumours by comparative genomic hybridization. Distinct patterns of sub-chromosomal deletions were identified suggesting alternative mechanisms of tumour initiation. The deleted chromosomal segments encompass two regions (10q23.1 and 17q22-q23) that are syntenic to the chromosomes (22 and 1p) most often deleted in human meningiomas. A number of genes associated with DNA repair, cell cycle progression and apoptosis are located on both the deleted canine chromosomal segments and the syntenic regions deleted in human meningiomas. This study represents the first report of chromosomal copy number abnormalities in non-cultured canine brain tumour tissue.     

Pathological studies of cheek teeth apical infections in the horse: 2. Quantitative measurements in normal equine dentine

Measurements of primary, regular and irregular secondary dentine and pulp dimensions were made on transverse, sub-occlusal and mid-tooth sections, of 40 maxillary and 42 mandibular control equine cheek teeth (CT) of different ages. Maxillary and mandibular CT primary dentine in different age groups had a mean thickness of 922–1065 μm and 1099–1179 μm, respectively, on the lateral aspects, and 1574–2035 μm and 1155–1330 μm, respectively, on the medial aspects of pulp horns. Surprisingly, some increase in thickness was found in some mandibular CT primary enamel in the first few years following eruption.Regular secondary dentine thickness increased with age, for example at mid-tooth level in mandibular CT from 124 μm at 3 years dental age to 290 μm at >7 years dental age on the lateral aspect of pulp horns, and from 166 μm to 509 μm on the medial aspects of pulp horns, indicating a deposition rate of 0.5–10 μm/day. This type of dentine was thicker sub-occlusally than in the mid-tooth region. Maxillary dentinal dimensions showed a similar age-related increase in thickness. Maxillary CT dentine was significantly thicker (72% in primary, 43% in regular secondary dentine) on the medial compared to the lateral aspects of pulp horns, but mandibular CT dentine was just 15% and 14% thicker in primary and regular secondary dentine thickness, respectively, on the their medial as compared to their lateral aspects. Dentinal and pulp dimensions varied between individual pulp horns, Triadan tooth position, and dental age, with complex interactions between these variables for some parameters.     

Detection of Brucella abortus in mammalian tissue, using biotinylated, whole genomic DNA as a molecular probe

A method has been developed for the detection of Brucella abortus in complex tissue homogenates. The technique uses tissue homogenization in the presence of sucrose and Triton X-100 and subsequent filtration through a 5-microns pore size filter to remove mammalian nuclei and cellular debris. The DNA from the bacteria is then extracted, dot blotted onto nitrocellulose, and hybridized with a biotinylated probe of B abortus strain 19 DNA. In the present study, BALB/C mice were inoculated intraperitoneally with either 10(9) or 10(11) B abortus strain 2308S organisms. After 6 days, the mice were euthanatized by cervical dislocation and the livers were removed, weighed, and the appearance of each was noted. The tissues were homogenized, and a viable cell count was performed to determine the number of bacteria in each organ. The DNA was extracted, blotted onto nitrocellulose, and hybridized with the Brucella probe. The biotin label was detected by use of a commercially available streptavidin/alkaline phosphatase system. In control experiments, the technique detected 10(5) organisms in a mixture of bacteria and 1 g of rat liver. The technique also detected 10(7) B abortus organisms/g of tissue from experimentally inoculated mice. The probe was specific for Brucella and had no affinity for contaminating bovine or bacterial DNA.     

Studies on growth cartilage in the horse and their application to aetiopathogenesis of dyschondroplasia (osteochondrosis)

The importance of osteochondrosis (dyschondroplasia) to the horse industry has been well documentedsince it was first recorded 50 years ago. The condition is known to be multifactorial in origin, arising from focal failure of endochondral ossification at predilection sites in articular/epiphyseal growth cartilage, but specific information on its aetiopathogenesis is sparse. This paper reviews the current knowledge of growth cartilate metabolism and the process of normal endochondral ossification in the horse. It highlights the localization of various protein products of chondrocytes and the differences in the zones of articular cartilage. In the early focal lesions (referred to as dyschondroplasia) there are alterations in the chondrocytes, extracellular matrix and some of the local protein products. The most obvious feature is an alteration in matrix metabolism which may be responsible for triggering a range of other factors leading to the development of a retained core of cartilage and a primary lesion of dyschondroplasia.Based on available evidence, a preliminary hypothesis for pathogenesis is presented. This suggests thatthere are a number of factors capable of initiating the condition. One of these involves high circulating insulin levels from high energy feeding which may affect chondrocyte maturation leading to altered matrix metabolism and faulty mineralization resulting in the formation of cartilage cores which characterize the condition. Further research to test this hypothesis is needed before there can be a rational basis for prophylaxis.     

Diagnosis of equine monocytic ehrlichiosis (Potomac horse fever) by indirect immunofluorescence

The recent establishment of a system for the continuous in vitro propagation of Ehrlichia risticii, the causative agent of equine monocytic ehrlichiosis (EME; synonym, Potomac horse fever), has facilitated the development of an indirect fluorescent antibody test for the diagnosis of this disease under laboratory and field conditions. The field diagnostic application of the test has aided in the recognition of the disease in 16 states of the United States and in 1 province of Canada. A limited epidemiologic study conducted between January and September 1985, in an area where the disease is known to be enzootic, revealed that conversion from seronegative to seropositive status is not always accompanied by clinical manifestations of the disease. Confirmatory findings in experimentally inoculated horses suggest the existence of clinically undetectable infections.     

Biological control of horse cyathostomin (Nematoda: Cyathostominae) using the nematophagous fungus Duddingtonia flagrans in tropical southeastern Brazil

The viability of a fungal formulation using the nematode-trapping fungus Duddingtonia flagrans was assessed for the biological control of horse cyathostomin. Two groups (fungus-treated and control without fungus treatment), consisting of eight crossbred mares (3–18 years of age) were fed on Cynodon sp. pasture naturally infected with equine cyathostome larvae. Each animal of the treated group received oral doses of sodium alginate mycelial pellets (1 g/(10 kg live weight week)), during 6 months. Significant reduction (p < 0.01) in the number of eggs per gram of feces and coprocultures was found for animals of the fungus-treated group compared with the control group. There was difference (p < 0.01) of 78.5% reduction in herbage samples collected up to (0–20 cm) between the fungus-treated group and the control group, during the experimental period (May–October). Difference of 82.5% (p < 0.01) was found between the fungus-treated group and the control group in the sampling distance (20–40 cm) from fecal pats. During the last 3 months of the experimental period (August, September and October), fungus-treated mares had significant weight gain (p < 0.01) compared with the control group, an increment of 38 kg. The treatment with sodium alginate pellets containing the nematode-trapping fungus D. flagrans reduced cyathostomin in tropical southeastern Brazil and could be an effective tool for biological control of this parasitic nematode in horses.     

Surgical treatment of tarsometatarsal joint luxation in a miniature horse foal

A 2-week-old Miniature Horse foal was referred for evaluation and treatment of a luxated right tarsometatarsal joint. Treatment consisted of closed reduction and internal fixation using two partially threaded Steinmann pins placed in normograde fashion through the tuber calcis into the proximal third metatarsus. Traumatic luxation has been reported to occur in the tarsocrural, proximal intertarsal and tarsometatarsal joints within the equine tarsus. Treatment for luxation of the distal intertarsal joint has not been documented. The treatment method most commonly suggested for tarsal luxation is closed reduction and cast immobilisation. Internal fixation using lag screws and plating has also been described. A combination of internal fixation and external coaptation is thought to achieve maximal stability and allow faster convalescence in cases of tarsal luxation. This case report describes for the first time a technique using two Steinmann pins to achieve successful internal fixation of a traumatic tarsometatarsal joint luxation in a 2-week-old Miniature Horse foal.     

Perianesthetic development of diaphragmatic hernia in a horse with equine pituitary pars intermedia dysfunction (PPID)

A 21-year-old Thoroughbred gelding with a history of equine pituitary pars intermedia dysfunction (PPID) presented with priapism of 2 days’ duration. The horse received a caudal morphine epidural and then underwent corpus cavernosum lavage and phallectomy under general anesthesia. The patient’s recovery featured multiple unsuccessful attempts to stand and his respiratory distress persisted for several hours until he acutely developed severe colic and was euthanized. Necropsy findings revealed a pituitary adenoma of the pars intermedia, bilateral adrenal cortical hyperplasia, and diaphragmatic herniation. This report suggests that horses with PPID may present a greater risk for diaphragmatic hernia under general anesthesia or during procedures placing stress on the diaphragm, including anesthetic recovery.     

Improved detection of equine antibodies against Sarcocystis neurona using polyvalent ELISAs based on the parasite SnSAG surface antigens

Equine protozoal myeloencephalitis (EPM) is a common neurologic disease of horses that is caused by the apicomplexan pathogen Sarcocystis neurona. To help improve serologic diagnosis of S. neurona infection, we have modified existing enzyme-linked immunosorbent assays (ELISAs) based on the immunogenic parasite surface antigens SnSAG2, SnSAG3, and SnSAG4 to make the assays polyvalent, thereby circumventing difficulties associated with parasite antigenic variants and diversity in equine immune responses. Two approaches were utilized to achieve polyvalence: (1) mixtures of the individual recombinant SnSAGs (rSnSAGs) were included in single ELISAs; (2) a collection of unique SnSAG chimeras that fused protein domains from different SnSAG surface antigens into a single recombinant protein were generated for use in the ELISAs. These new assays were assessed using a defined sample set of equine sera and cerebrospinal fluids (CSFs) that had been characterized by Western blot and/or were from confirmed EPM horses. While all of the polyvalent ELISAs performed relatively well, the highest sensitivity and specificity (100%/100%) were achieved with assays containing the rSnSAG4/2 chimera (Domain 1 of SnSAG4 fused to SnSAG2) or using a mixture of rSnSAG3 and rSnSAG4. The rSnSAG4 antigen alone and the rSnSAG4/3 chimera (Domain 1 of SnSAG4 fused to Domain 2 of SnSAG3) exhibited the next best accuracy at 95.2% sensitivity and 100% specificity. Binding ratios and percent positivity (PP) ratios, determined by comparing the mean values for positive versus negative samples, showed that the most advantageous signal to noise ratios were provided by rSnSAG4 and the rSnSAG4/3 chimera. Collectively, our results imply that a polyvalent ELISA based on SnSAG4 and SnSAG3, whether as a cocktail of two proteins or as a single chimeric protein, can give optimal results in serologic testing of serum or CSF for the presence of antibodies against S. neurona. The use of polyvalent SnSAG ELISAs will enhance the reliability of serologic testing for S. neurona infection, which should lead to improved diagnosis of EPM.     

Detection of equine infectious anemia virus in a horse with an equivocal agar gel immunodiffusion test reaction

A horse whose serum reacted equivocally in the agar gel immunodiffusion (AGID) test for equine infectious anemia was studied over a 3-year period. The horse remained afebrile and virus was detected in only 1 of 6 horse inoculation tests. The intensity of AGID test reactions increased temporarily following this evidence for virus. Although the AGID test reaction was equivocal and 5 of the 6 transmission attempts failed, the 1 successful transmission proved the horse was infected.     

Aggravating factors in the development of ocular abnormalities in farmed Atlantic halibut (Hippoglossus hippoglossus)

Cystic lesions in the eyes of farmed halibut with a number of ocular sequelae have been reported previously with evidence that the condition can be associated with increased activity of choroidal carbonic anhydrase and elevated oxygen tension in the aqueous humour of affected fish. These changes may be compared to the 'bends' in human divers which are characterized by elevated circulating levels of nitrogen together with aggravating factors such as physical exercise. It is postulated that bubble formation and subsequent development of cysts in halibut require not only high aqueous humour oxygen tension but also the imposition of factors such as handling or intraspecific aggression which lead to bubble formation. A significantly higher incidence of lesions was noted in fish handled, weighed and measured every two weeks compared with a control group of unhandled fish, suggesting that physical activity associated with handled and possibly also intraspecific aggression is a factor in generation of ocular lesions in these fish.