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Opis przypadku Case report
Neosporosis was first recognised in dogs in Norway. Following its identification in dogs in the United States, the genus Neospora and occasionally the type species Neospora caninum (N. caninum) were proposed for this protozoan (6). N. caninum has recently been rec-ognised as a protozoan parasite capable of infecting dogs and other animals. A wide host range is reported, with natural infection observed in dogs, cattle, sheep, goats, horses and deer (8). Dogs are the definitive hosts for N. caninum (15). Currently, there is strong evi-dence that its life cycle can be maintained in dogs and cattle (4, 17). Dogs may be infected by bovine foetal membranes (4). It is also suggested that the ingestion of raw meat may cause infection (17). Tachyzoites and tissue cysts are the only known stages of the life cycle of N. caninum. The presence of tachyzoites has been demonstrated in a variety of host cells, whereas tissue cysts have been found only in the central nervous system (8).
Neospora caninum infection in dogs is characterised mainly by neurological disorders. In adult dogs, a mul-tifocal involvement of the central nervous system, as well as polymyositis, myocarditis and dermatitis can be observed (8). Canine neosporosis has been reported worldwide in serological surveys and case series, but until now there has been little information on canine neosporosis in Turkey. The purpose of the present study is to report the presence of N. caninum infection in Aydin, Turkey, and to describe skin lesions associated with the infection in an adult hunter dog, as well as a partially effective treatment of the animal.
Case descriptions
A 3-year-old female Pointer dog was presented to the Department of Internal Medicine, Faculty of Veterinary Medicine, Adnan Menderes University, with a history of progressive skin lesions. The skin lesions appeared 7 months prior to admission. They emerged first on the feet and
foot-Adjunctive chloroquine as a possible anti-inflammatory
therapy for canine cutaneous neosporosis
ÜMIT KARADEMIR, SERDAR PAŞA*, KEREM URAL*, İBRAHIM AKIN**
Department of Pharmacology and Toxicology, *Department of Internal Medicine, **Department of Surgery, Faculty of Veterinary Medicine, Adnan Menderes University, Aydin-Turkey
Received 08.04.2015 Accepted 04.12.2015
Karademir Ü., Paşa S., Ural K., Akin İ.
Adjunctive chloroquine as a possible anti-inflammatory therapy for canine cutaneous neosporosis Summary
Neospora caninum has recently been recognised as a protozoan parasite capable of infecting dogs and other animals. Naturally occurring intermediate hosts include dogs, cattle, goats, sheep, horses and deer. The purpose of the present study was to report the presence of N. caninum infection in Aydin, Turkey, and to describe clinical, haematological and serological aspects of cutaneous neosporosis.
In March 2011, the Department of Internal Medicine, Faculty of Veterinary Medicine, University of Adnan, Menderes, received a 3-year-old female Pointer dog with a history of severe skin lesions, located mostly on the feet and partly in the abdominal area. The skin lesions appeared 7 months prior to admission. They emerged first on the feet and footpads, spreading to the thorax and ventral abdomen, ulcerating 15 days prior to admission. The dermal lesions took the form of a diffuse pyogranulomatous dermatitis with a dense infiltrate of leucocytes. Histological sections revealed several N. caninum tachyzoites scattered throughout the tissue. Haematological examination involving complete blood counts, as well as serum biochemical and blood gas analysis, showed no abnormalities. The cutaneous lesions partially resolved following therapy with clindamycin hydrochloride, trimethoprim-sulphonamide combination and chloroquine.
The most common clinical presentation reported in N. caninum infection in dogs is limb paralysis, but the disease may be generalised, involving all organs. There are few studies regarding Neospora-associated dermatitis in dogs. The affected dog in the present case was 3 years old, which is consistent with previous reports of cutaneous neosporosis, involving only adult dogs. It would be premature to conclude whether similar cases reported in adult dogs represented primary infection or a relapse of congenital infection. These observations demonstrate the presence of N. caninum in the Aydin province, Turkey, and confirm that neosporosis should be on the list of differential diagnosis for pyogranulomatous dermatitis in dogs.
Med. Weter. 2016, 72 (2), 125-127 126
pads, spreading to the thorax and ventral abdomen, ulcerat-ing 15 days prior to admission. On physical examination, except for the skin lesions, the dog was found to be healthy. Body temperature was slightly elevated (39.4°C), heart and respiratory rates were within normal limits. Multiple pyogranulomatous skin nodules were observed all over the body, and generalised lymphadenomegaly was noted. Haematological examination involving complete blood counts, as well as serum biochemical and blood gas analy-sis, showed no abnormalities.
The dog was negative for the canine heartworm anti-gen, and no antibodies were detected against Anaplasma
phagocytophilum, Ehrlichia canis or Borrelia burgdorferi
(Snap 4Dx). IFAT was negative for leishmaniasis. Blood smear evaluation for possible haematological pathogens was unremarkable.
The differential diagnosis considered to account for pododermatitis, other than by cutaneous neosporosis in dogs, included pyoderma, dermatophytosis, demodicosis, as well as Pelodera strongyloides and hookworm infections. Antibiogram studies of the lesional sites were unremarkable, except for Staphylococcus aureus. Wood’s lamp examina-tion and histopathology were negative for dermatophytosis. Deep skin scrapings revealed no mites. Polymerase chain reaction and the immune fluorescence antibody technique (IFAT) were both negative for leishmaniasis. A result of 1 : 50 on the IFAT was positive evidence of Neospora
caninum infection in the present case, whereas the IFAT
was negative for Toxoplasma gondii. In addition, the his-topathology of a punch biopsy sample revealed pyogranu-lomatous necrotising dermatitis caused by N. caninum. Immunohistochemistry provided definitive diagnosis in this clinical case, where tachyzoites were stained with specific anti-N. caninum antiserum, but not with antibodies against
T. gondii. Interestingly, numerous N. caninum tachyzoites
were found in fluid squeezed from dermal lesions on the feet, as described previously (7, 17).
Initial therapy consisted of clindamycin hydrochloride (15 mg/kg q 12 h subcutaneously for 3 weeks) (Klindan amp., 600 mg, Bilim, Turkey), trimethoprim sulphonamide (20 mg/kg q 12 h, subcutaneously for 4 weeks) (Bakteral
flk. 50 ml., Topkim, Turkey) and chloroquine phosphate (2.5 mg/kg q 12 h, perorally for 4 weeks) (Kutlu tblt. 250 mg., Abdi İbrahim, Turkey). The cutaneous lesions partially resolved following 3 weeks of the combination therapy. After 7 weeks of therapy, most of the cutaneous lesions disappeared, whereas pododermatitis was still evident with a promising scar and healthy tissue formation. After 12 weeks of therapy, the lesions were 60-70% resolved. The owner of the patient was lost for follow-up after that time.
Discussion
Although the most common clinical presentation of N. caninum infection in dogs is the paralysis of limbs, the disease may be generalised, involving almost all organs (8). Neospora-associated dermatitis is not very common, and up to date some case series have been reported in the literature, elsewhere reviewed (17). There are very few studies regarding the presence of neosporosis in dogs in Turkey, including only one clinical case report (3).
The clinical case enrolled in this study was a 3-year-old dog, which is consistent with the other reports of cutaneous neosporosis, involving exclusively adult dogs. It is unknown whether such cases reported in adult animals represented a relapse of congenital infection or whether they were primary infections. Neurological signs, which are generally observed in dogs under 6 months of age, were not evident in the present case. Dogs that consume raw meat may prob-ably become infected with N. caninum (13, 14). The higher N. caninum seroprevalence among hunting dogs compared with pet dogs in a study in the United Kingdom supports the additional diagnostic importance of raw meat for transmission (11). According to the owner, the present case, a hunter dog, was frequently fed with raw meat. Other modes of transmission, such as transplacental transmission, were unknown in the
Fig. 1. Pyogranulomatous dermatitis a) in thoracal area b) on rear legs with oozing c) on claws and footpads d) on footpads in lateral view
Fig. 2. Pyogranulomatous necrotising dermatitis caused by Neospora caninum (HE × 400). Dermal pyogranulomatous inflammation with clusters of Neospora caninum tachyzoites
Med. Weter. 2016, 72 (2), 125-127 127 present case, as there was no information regarding
offspring.
The dog did not suffer from a systemic disease. Detailed clinical and laboratory tests for ehrlichiosis, anaplasmosis, dirofilariosis, borreliosis, leishmaniasis, hepatozoonosis and other relevant systemic/metabolic diseases were also performed with negative results. However, it was impossible to rule out all internal diseases accompanying Neospora infection. No organ samples, other than a skin punch biopsy sample, were taken for histopathology.
Early treatment applications within clindamycin (9) and potentiated sulphonamides may influence clinical remission, as this protocol resulted in clinical improve-ment in 10 out of 16 dogs previously treated (2).
Chloroquine is a 4-aminoquinoline antimalarial drug, first introduced into the market in the late 1940s and used on a massive scale for malaria treatment and prevention in humans (1, 15). Its efficacy, low price and safety have made it the gold standard therapy regime against malaria for many years (1, 15). Among relevant antimalarials, chloroquine is one of the choice with the longest half-lives (approximately 60 days), providing a chemoprophylactic effect during the drug elimination phase (15, 18). Chloroquine also acts on the parasites for an extended time period, after which its concentra-tion falls below the therapeutic level (18). In addiconcentra-tion, chloroquine has been used as a treatment protocol for anti-inflammatory conditions (20).
Published data indicate that serum levels of pro-inflammatory cytokines, including interleukin (IL) -12, interferon (IFN) -γ, and tumour necrosis factor (TNF) -α, are increased in N. caninum infection (10). Cell-mediated immunity involving proinflammatory cytokines, such as interferon gamma (IFN-γ), are likely to be important components of protective immunity to N. caninum (10). Literature data clearly indicate that some proinflammatory cytokines, such as IL-12, IFN-γ and TNF-α, are involved in N. caninum infection and play an important role in killing the parasite (5, 12, 16).
In human patients with systemic lupus erythema-tosus, chloroquine therapy lowers some of the proin-flammatory cytokines as part of its anti-inproin-flammatory effects (20). Moreover, chloroquine has been shown to be effective against a variety of bacterial and fungal pathogens by both direct and indirect mechanisms (19). Therefore this drug has the potential for reducing microbiological infections (19). Because of its proven efficacy against malaria and the above-mentioned potential effects against several agents, chloroquine was chosen against Neospora infection in the present case. Another reason for selecting this drug was the lack of pyrimethamine. Pyrimethamine is a standard treatment protocol for neosporosis, but it is com-mercially unavailable in Turkey. For these reasons, chloroquine was the drug of choice, accompanying the therapy with clindamycin hydrochloride and trimethoprim-sulphonamide combination. Although
the levels of proinflammatory cytokines were not analysed in this case, chloroquine may have helped to reduce the levels of these cytokines as part of its anti-inflammatory effects, and thus may have hastened clinical recovery.
Although several case reports of cutaneous neo-sporosis in dogs have been documented from other countries, the clinical case presented herein is, to the authors’ knowledge, the first case of N. caninum infec-tion in the Aydin province, Turkey. Detailed clinical trials involving the combination of clindamycin hydro-chloride, trimethoprim sulphonamide and chloroquine need to be conducted in a larger dog population with cutaneous neosporosis.
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Corresponding author: Umit Karademir, Department of Pharmacology and Toxicology, Faculty of Veterinary Medicine, Adnan Menderes University, 09016, Isikli, Aydin-Turkey; e-mail: umitkarademir@yahoo.com
