Jeanine Peters-Kennedy

Consistent Detection of Felis Domesticus Papillomavirus 2 DNA Sequences Within Feline Viral Plaques

Viral plaques are well recognized skin lesions of cats. They are thought to be caused by papillomavirus infection; however, the causative papillomavirus is uncertain. In the current study, polymerase chain reaction using 2 consensus primer sets and 1 primer set specific for Felis domesticus papillomavirus 2 (FdPV-2) was used to amplify DNA from a series of 14 feline viral plaques. The FdPV-2 sequences were detected in all 14 viral plaques by the specific primers but in only 1 of 14 feline cutaneous trichoblastomas. Papillomavirus DNA was amplified from 8 plaques using the consensus primers. Sequences from FdPV-2 were amplified using the consensus primers from 4 plaques. In addition, 3 plaques contained papillomavirus DNA sequences from Felis domesticus papillomavirus sequence MY1, and a previously unreported papillomavirus DNA sequence was amplified from 1 plaque. As FdPV-2 was consistently present within the plaques, this suggests that this papillomavirus is the likely etiologic agent. Feline viral plaques can undergo neoplastic transformation to Bowenoid in situ carcinomas (BISCs). As FdPV-2 DNA is frequently present within BISCs, this suggests that FdPV-2 induces viral plaque formation and then remains detectible after neoplastic transformation.

Apparent clinical resolution of pinnal actinic keratoses and squamous cell carcinoma in a cat using topical imiquimod 5% cream.

Imiquimod is a topical immune response modifier and stimulator used in humans to treat a number of cutaneous neoplasms. This case report describes a cat with actinic keratoses and squamous cell carcinoma of the pinnae. The pinnal lesions were treated with topical 5% imiquimod three times per week. Treatment was discontinued after 82 days of therapy. Twelve weeks of topical imiquimod application resulted in clinical resolution of the pinnal lesions. Although no post-treatment biopsies were performed, there was no relapse of the pinnal lesions in 5 months of clinical follow-up. Expected side effects were limited to erythema, crusting, alopecia, and mild discomfort at the sites of application during the first 3 weeks of application. These results suggest that topical imiquimod, although unproven, might be a therapeutic option or adjunct to therapy for cats with actinic keratoses and squamous cell carcinoma, especially those cats for whom surgery and radiation therapy are not an option.

Investigation of an outbreak of besnoitiosis in donkeys in northeastern Pennsylvania.

OBJECTIVE To describe the clinical, endoscopic, and serologic features of an outbreak of besnoitiosis in 2 donkey operations in northeastern Pennsylvania and to report the outcome of attempted treatment of 1 naturally infected individual. DESIGN Observational study. ANIMALS 29 donkeys (Equus asinus) in northeastern Pennsylvania. PROCEDURES Donkeys were examined for lesions suggestive of besnoitiosis in an outbreak investigation. Information was collected regarding the history and signalment of animals on each premises. Rhinolaryngoscopy was performed to identify nasopharyngeal and laryngeal lesions. Serum samples were collected for immunofluorescent antibody testing and immunoblotting for Besnoitia spp. Skin biopsy samples were obtained from 8 animals with lesions suggestive of besnoitiosis for histologic examination. Quantitative real-time PCR assay for Besnoitia spp was performed on tissue samples from 5 animals. RESULTS Besnoitiosis was confirmed in 6 of the 8 suspected cases. The most common lesion site was the nares, followed by the skin and sclera. Donkeys with clinical signs of disease had higher serum antibody titers and tested positive for a greater number of immunoblot bands than did donkeys without clinical signs of disease. All animals evaluated by PCR assay tested positive. Putative risk factors for disease included age and sex. Ponazuril was not effective at treating besnoitiosis in a naturally infected donkey. CONCLUSIONS AND CLINICAL RELEVANCE Knowledge of clinical and serologic features of besnoitiosis in donkeys will assist clinicians in the diagnosis and prevention of this disease in donkey populations. Besnoitiosis may be an emerging disease of donkeys in the United States.

Diagnosis of Parelaphostrongylus spp. infection as a cause of meningomyelitis in calves

Migration of Parelaphostrongylus spp. has been documented to cause central nervous system damage in a number of aberrant host species but appears to be uncommon in cattle. The current report describes the clinical and laboratory findings, antemortem and definitive diagnosis, and response to treatment of Parelaphostrongylus spp. infection in five 3–7- month-old Limousin calves from 2 farms. All calves had signs of acute (n = 2) and chronic (n = 3) progressive spinal cord dysfunction. Cerebrospinal fluid analysis revealed a marked eosinophilic (acute cases) or lymphocytic (chronic cases) pleocytosis and elevated protein in all calves. A necropsy and histopathologic evaluation was performed on 2 euthanized calves, and histopathology revealed lymphoplasmacytic and eosinophilic meningomyelitis with multiple intradural and intramedullary expansile hyperplastic lymphoid nodules containing germinal centers and nematode fragments. DNA sequencing was performed on nested polymerase chain reaction products amplified with parasite-specific primers obtained from formalin-fixed and frozen spinal cord; PCR products from these 2 calves were 100% identical to Parelaphostrongylus species on DNA sequencing, confirming the diagnosis. Surviving calves rapidly improved following treatment with anthelmintics and corticosteroids. This case series identified Parelaphostrongylus spp. (likely P. tenuis) as a cause of spinal cord disease in calves and highlights the need for vigilance against aberrant parasite migration in calves grazing wet, snail-infested pastures. Cerebrospinal fluid eosinophilia is useful for supporting an antemortem diagnosis of Parelaphostrongylus in calves with acute neurologic disease; however, a lymphocytosis is observed in chronic or treated cases.

Comparison of the levels of Equus caballus papillomavirus type 2 (EcPV-2) DNA in equine squamous cell carcinomas and non-cancerous tissues using quantitative PCR.

Equus caballus papillomavirus type 2 (EcPV-2) infection has been associated with equine genital squamous cell carcinomas (SCCs). However, quantitative PCR (qPCR) has not been performed to determine viral copy numbers within these lesions. Additionally, the frequency with which EcPV-2 can be detected in other common sites of equine SCC development remains uncertain. The aim of this study was to develop a qPCR assay to estimate the viral load in a variety of equine tissue samples. These included 40 SCC lesions, 19 penile non-SCC or precursor disease lesions, and 222 tissues without observable lesions from SCC-prone sites on clinically normal horses. EcPV-2 DNA was present significantly more frequently, and in higher copy numbers, in equine penile SCC lesions than in either healthy penile mucosa or non-SCC penile lesions. This supports the hypothesis that EcPV-2 is involved in development of penile SCCs and suggests that penile EcPV-2 infection is rare in the absence of SCCs. Samples of normal vulval mucosa rarely contained EcPV-2 DNA and none of the nictitating membrane samples contained EcPV-2 DNA, indicating that asymptomatic EcPV-2 infection is uncommon at these sites. EcPV-2 DNA was detected in a proportion of both SCCs and normal samples from the oral cavity or pharynx, although there were no significant differences in the rate of infection or viral copy number between the SCCs and the normal mucosal samples. As such, the role of EcPV-2 in development of SCCs in this location remains to be established.

Serological diagnosis of Besnoitia bennetti infection in donkeys (Equus asinus)

Besnoitiosis is an emerging infectious disease of donkeys (Equus asinus) in the United States for which there are currently no serologic methods of diagnosis. A study was performed to evaluate physical examination findings and 3 serologic assays for the detection of Besnoitia bennetti infection in donkeys. A prospective study of 416 donkeys from 6 privately owned herds across 5 U.S. states (New York, Pennsylvania, Vermont, Oregon, and Washington) was performed. Donkeys were examined for clinical lesions suggestive of besnoitiosis and evaluated for antibodies against B. bennetti using a fluorescent antibody test (FAT) and 2 immunoblot assays specific for bradyzoite and tachyzoite antigens, respectively. Donkeys were confirmed to be infected with B. bennetti by histology (cases; n = 32) and were compared to those with no clinical signs of besnoitiosis (controls; n = 384). Identifying clinical lesions in 2 or more locations correctly identified infected donkeys 83% of the time. Donkeys with besnoitiosis had significantly higher FAT titers (P < 0.001) and numbers of bradyzoite (P < 0.001) and tachyzoite (P < 0.001) immunoblot bands than control donkeys. The sensitivity and specificity of the serologic assays for detecting besnoitiosis was 88% and 96% for FAT, 81% and 91% for bradyzoite immunoblot, and 91% and 92% for tachyzoite immunoblot, respectively. Fluorescent antibody and immunoblot assays are effective at identifying donkeys with besnoitiosis and provide a more efficient and less invasive diagnostic alternative to histology.

Resident lymphocytes in the dermis of the normal dorsolateral thoracic skin of alpacas.

BACKGROUND Small numbers of resident T lymphocytes are present in the dermis of normal skin of humans, cattle and sheep. HYPOTHESIS/OBJECTIVES We wanted to determine the prevalence, numbers and immunophenotype of lymphocytes in the dermis of healthy skin from alpacas. ANIMALS Skin biopsy specimens were collected from the dorsolateral thorax of 31 alpacas with normal skin. METHODS Skin biopsy specimens were evaluated for the prevalence and numbers of CD3+ and CD79a+ lymphocytes. RESULTS Resident CD3+ and CD79a+ lymphocytes were found around the superficial and deep dermal blood vessels. The CD3+ lymphocytes were more numerous than CD79a+ lymphocytes. Both CD3+ and CD79a+ lymphocytes were more numerous around superficial dermal blood vessels. CONCLUSIONS AND CLINICAL IMPORTANCE Resident CD3+ and CD79a+ lymphocytes are present around superficial and deep dermal blood vessels in normal skin from alpacas; hence, the presence of lymphocytes in these locations without obvious features of inflammation must be interpreted cautiously when evaluating skin biopsy specimens from alpacas with skin disease.

Charcot–Leyden crystals: do they exist in veterinary species? A case report and literature review

The Charcot–Leyden crystal (CLC) is a major human eosinophil protein that readily crystallizes; these crystals are common in eosinophilic diseases. Although anecdotal existence of these crystals is known in veterinary pathology, definitive reports do not exist, to our knowledge. We identified eosinophilic crystals in a laryngeal myxosarcoma from a 2-y-old, spayed female, Labrador Retriever dog that were tentatively interpreted as CLCs. However, Ziehl–Neelsen acid-fast stain was negative, arguing against CLCs. The crystals stained red with Masson trichrome, precluding collagen. Periodic acid–Schiff and alcian blue were negative. The crystals stained positively with Okajima, and no myoglobin immunoreactivity was detected, supporting their identity as hemoglobin crystals. In the absence of a hematologic abnormality, these crystals were interpreted to be abnormal hemoglobin breakdown products. Protein sequence comparison was pursued to determine whether a protein similar to CLC exists in mammals. Only 3 nonhuman primate species, the Sumatran orangutan (Pongo abelii), rhesus macaque (Macaca mulatta), and cynomolgus monkey (Macaca fascicularis), had a sequence similarity of >80%. Of the crystal-forming residues, 12 of 54 (22%) were different in the Sumatran orangutan and 15 of 54 (28%) were different in the Macaca spp., which may affect the crystallization process. The lack of reports of CLCs in nonhuman species and our results collectively suggest that CLCs are human-specific.

Hepatocutaneous syndrome in Shih Tzus: 31 cases (1996–2014)

OBJECTIVE To characterize findings in Shih Tzus with progressive superficial necrolytic dermatitis and degenerative vacuolar hepatopathy consistent with hepatocutaneous syndrome. DESIGN Retrospective case series. ANIMALS 31 Shih Tzus. PROCEDURES Medical records were reviewed to obtain information on signalment, history, treatment, outcome, and results of clinicopathologic testing, abdominal ultrasonography, and histologic examination of skin and liver specimens. A pedigree analysis was performed. RESULTS There were 16 males and 15 females. Median age at the time of diagnosis was 8 years (range, 5 to 14 years). Common clinical signs included lethargy, inappetence, weight loss, and lameness. Twenty-five dogs had cutaneous lesions consistent with hepatocutaneous syndrome; the remaining 6 initially only had hepatic abnormalities, but 3 of the 6 subsequently developed cutaneous lesions. Common clinicopathologic abnormalities included microcytosis (15/24 [63%] dogs) and high serum alkaline phosphatase activity (24/24 [100%] dogs). Hepatic ultrasonographic findings included a hyperechoic or heteroechoic appearance to the parenchyma with innumerable hypoechoic nodules. Histologic hepatic lesions consisted of degenerative vacuolar (glycogen and lipid) hepatopathy associated with minimally fibrotic to nonfibrotic, noninflammatory, proliferative nodules. Pedigree analysis confirmed a common ancestry in 12 of 18 dogs. Median survival time was 3 months (range, 1 to 36 months). CONCLUSIONS AND CLINICAL RELEVANCE Results suggested that HCS may have a heritable component in Shih Tzus, although the condition may also be identified in Shih Tzus without affected relatives. Clinical, clinicopathologic, ultrasonographic, and histologic abnormalities in affected Shih Tzus were similar to those previously reported for dogs of other breeds with HCS.

Atypical cutaneous cryptococcosis in four cats in the USA.

BACKGROUND Cryptococcosis is an uncommon fungal infection in humans and mammals. Occasionally, cryptococcosis manifests as cutaneous lesions, either as an extension of nasal disease or as stand alone lesions unassociated with the nose. Histologically, these lesions are typically characterized by abundant organisms with mild granulomatous dermatitis. Herein, four feline cases of atypical cutaneous cryptococcal infections are described. METHODS Skin punch biopsies from four client owned cats were submitted for histological evaluation between 2006 and 2015. Histological examination, including histochemical stains, was performed in all cases. Immunohistochemical stains and PCR were performed in three of four cases. Fungal culture was performed in two cases and transmission electron microscopy was performed in one case. RESULTS Grossly, the cutaneous lesions were papular to nodular with occasional ulceration and were located predominantly on the trunk. Histological examination revealed severe granulomatous to pyogranulomatous and eosinophilic dermatitis with rare, capsule-deficient yeasts. Immunohistochemistry, PCR and fungal culture confirmed Cryptococcus spp. to be the aetiological agent in these cases. CONCLUSIONS AND CLINICAL IMPORTANCE In cutaneous lesions, capsule-deficient strains of Cryptococcus spp. may induce a severe inflammatory response with rare intralesional organisms that may not be readily identified on routine haematoxylin and eosin stained slides. Special stains with careful examination and ancillary tests (PCR, immunohistochemistry, fungal culture or antigen testing) should be performed when pyogranulomatous and eosinophilic dermatitis is encountered without an identifiable cause.