Detection of intracellular IFN-γ and IL-4 cytokines in CD4+ and CD8+ T cells in the peripheral blood of dogs naturally infected with Leishmania infantum.

Canine leishmaniosis (CanL) is a systemic zoonotic disease the clinical manifestations of which can range from self-healing cutaneous lesions to disseminated visceral disease. Effective activation of cellular immunity is the cornerstone of resistance against Leishmania infantum in infected dogs. The aim of this cross-sectional, controlled study was the intracellular detection of interleukin 4 (IL-4) and interferon-γ (IFN-γ) in CD4+ and CD8+ lymphocytes in the peripheral blood of 40 dogs naturally infected with L. infantum by applying flow cytometry. The percentage of CD4+IL-4+ and CD8+IL-4+ lymphocytes (with or without immunostimulation) was low in the clinically healthy and subclinically infected dogs in contrast to clinically affected ones. In the same groups of dogs, the percentage of CD4+IFN-γ+ and CD8+IFN-γ+ T cells in their resting phase and following specific immunostimulation with Leishmania soluble antigen (LSA) was also low. CD4+IL-4+ and CD8+IL-4+ T cell percentage was higher in sick compared to clinically healthy and subclinically infected dogs, after immunostimulation. The corresponding figure of CD8+IL-4+ cells in sick dogs after LSA immunostimulation was also increased thus underlining the important role these cells may play in humoral immunity and perhaps the progression of CanL.

Pharmacokinetics and tolerability of aminosidine after repeated administrations using an optimal dose regimen in healthy dogs and in dogs with leishmaniosis.

Optimisation of dose schedules of aminoglycosides is required in order to increase efficacy and prevent their toxicity. The objective of this study was to determine the pharmacokinetic profile and the safety of aminosidine in dogs with naturally occurring leishmaniosis and in healthy dogs after once daily administration. Six young-adult, male, healthy, Beagle dogs and 12 dogs with clinical signs of canine leishmaniosis without azotemia and proteinuria were included in the study. Diagnosis of the disease was confirmed by serology, parasitology and molecular techniques. Pharmacokinetics and evaluation of renal function after repeated (once daily for 21 consecutive days) subcutaneous administration of aminosidine, at the dose of 15 mg/kg b.w. in both the healthy and the diseased animals were compared. Concentrations of aminosidine were determined by high-performance liquid chromatography and pharmacokinetic analysis was performed by the non-compartmental method. No significant differences were observed between healthy and diseased dogs considering all pharmacokinetic parameters. In general, mean Cmax ranged between 46.41 and 54.32 µg/mL and between 38.69 and 40.73 µg/mL in healthy dogs and in dogs with canine leishmaniosis, respectively. No accumulation of the drug was observed in either group since total elimination of aminosidine and half-life lambda z were not modified throughout the administration period. Aminosidine was well tolerated in all dogs with no clinical and clinicopathological signs of nephrotoxicity. Once daily administration of high dose of aminoglycosides, resulted in effective serum concentrations and absence of nephrotoxicity.

Re-emergence of animal rabies in northern Greece and subsequent human exposure, October 2012 - March 2013.

Greece has been rabies-free since 1987 with no human cases since 1970. During 2012 to 2013, rabies has re-emerged in wild and domestic animals in northern Greece. By end March 2013, rabies was diagnosed in 17 animals including 14 red foxes, two shepherd dogs and one cat; 104 subsequent human exposures required post-exposure prophylaxis according to the World Health Organization criteria. Human exposures occurred within 50 km radius of a confirmed rabies case in a wild or domestic animal, and most frequently stray dogs were involved.

Treatment of canine leishmaniosis with aminosidine at an optimized dosage regimen: a pilot open clinical trial.

Leishmaniosis due to Leishmania infantum (Syn: L. chagasi) is one of the most common diseases of dogs in Mediterranean countries and also has zoonotic potential. The aim of this study was to evaluate the efficacy of an optimized dosage regimen of aminosidine for the treatment of canine leishmaniosis (CanL) in terms of clinical remission, restoration of clinicopathological abnormalities, evolution of antibody titer, lymph node and bone marrow parasitic density and of PCR-based parasitological cure. Twelve non-uremic dogs without proteinuria, presenting clinical signs of CanL were included in the study. The diagnosis was confirmed by serology, microscopy and PCR of lymph node and bone marrow samples. Aminosidine was administered subcutaneously at the dose of 15 mg/kg body weight, once daily, for 21 consecutive days. A partial remission of the clinical signs, amelioration of clinicopathological abnormalities such as anemia, lymphopenia, hyperproteinemia, hyperglobulinemia, and reduced albumin/globulin ratio and reduced lymph node and bone marrow parasitic density were witnessed, although parasitological cure was not achieved. Since data are not supportive enough for the use of aminosidine as an alternative treatment, a large-scale controlled clinical trial using this optimized dosage regimen of aminosidine is warranted to compare efficacy against currently used drugs.

A cross-sectional sero-epidemiological study of canine leishmaniasis in Greek mainland.

Canine leishmaniasis is endemic in Greece as in other countries of the Mediterranean basin. In this study, the regional prevalence of canine seropositivity to Leishmania spp. in Greek mainland was simultaneously assessed in 7 different regions. A total of 2620 serum samples were collected from clinically healthy dogs and were tested for anti-L. infantum antibodies by indirect immunofluorescence antibody test (IFAT) and enzyme linked immunosorbent assay (ELISA). A high degree of agreement (κ=0.96) was observed between these two tests. The seroprevalence rate in the whole study population was nearly 20%, being highest among dogs living in Attiki (30.12%) and lowest for those living in Florina (2.05%). There was no difference in terms of the gender of the dogs, their length of hair coat or their utility; on the contrary, seroprevalence rates were significantly higher among dogs in the age groups of 1-3 years (23.39%) and 3-9 years (23.35%) than in younger (2.26%) or older (6.03%) dogs. Epidemiological data on the seroprevalence of canine leishmaniasis provide indirect information on the prevalence of the infection and the disease and are necessary to implement and then to evaluate the effectiveness of control measures.

Molecular detection of Leishmania infantum in wild rodents (Rattus norvegicus) in Greece.

Leishmania infantum and Leishmania tropica are the species responsible for visceral leishmaniasis and cutaneous leishmaniasis respectively. In Greece, both diseases are endemic. The dog is considered the main reservoir of L. infantum, whereas the role of other animals for both L. infantun and L. tropica is unknown. Spleens from wild Rattus norvegicus, live trapped in Greece, were examined for the presence of Leishmania parasites by PCR. Out of 16 samples examined, only one was found positive for L. infantum with scant amount of parasitic DNA present. This is the first documented case of detection of L. infantum in R. norvegicus in Greece. The results of this preliminary study indicate that R. norvegicus is unlikely to be a reservoir for Leishmania parasites in Greece.

Determination of CD4+ and CD8+ T cells in the peripheral blood of dogs with leishmaniosis before and after prolonged allopurinol monotherapy.

Canine leishmaniosis (CL) is a common systemic parasitic disease that is endemic in many Mediterranean countries including Greece. The immune reaction to the parasite is critical to the outcome of the infection and the response to treatment. Some studies have shown a reduction of circulating CD4+ T cells and of the CD4+/CD8+ ratio in dogs with CL and these changes normalised following treatment with meglumine antimoniate or amphotericin B. Allopurinol is used as a monotherapy for the chronic treatment of CL. The aim of the present study was to determine the circulating CD4+ and CD8+ T lymphocyte numbers and the CD4+/CD8+ ratio in 19 dogs diagnosed with CL before and after prolonged allopurinol monotherapy (18 months). A significant decrease in circulating CD4+ T cells was observed in dogs with CL before treatment. Prolonged allopurinol monotherapy improved the number of circulating CD4+ T cells, but did not restore their number to within the normal range.

Providencia stuartii infection in a dog with severe skin ulceration and cellulitis.

The aim of this report is to present a case of severe skin ulceration and cellulitis in a seven year old mongrel dog following cystitis. The Providencia stuartii strain was the only isolate from skin lesions. This bacterium is a Gram-negative rod belonging to the family of Enterobacteriaceae. To the best of our knowledge, even though there are some reports in the literature concerning animal infections with Providencia species, there have been no cases of cellulitis in which this pathogen is involved. The Providencia stuartii strain was highly sensitive to amikacin, and the dog responded quickly to this antibiotic.

Evaluation of cytology in the diagnosis of acute canine monocytic ehrlichiosis (Ehrlichia canis): a comparison between five methods.

The purpose of this study was the comparison of the diagnostic sensitivity between buffy coat (BC), peripheral blood (PB), lymph node (LN), bone marrow (BM) and short-term culture (P-D) cytology that has been based on the detection of Ehrlichia canis morulae, in the acute phase of canine monocytic ehrlichiosis (CME). Their cellular localization, total numbers and microscopic differentials were also investigated. The highest sensitivities were achieved after evaluating 1000 oil immersion fields (OIFs) in BC (66%) and an equal number in LN (60.9%) smears, separately or together (74%). The morulae were more often detected into lymphocytes than monocytes. The highest total number of morulae (n=143) were found in P-D smears. Finally, to avoid false positive diagnoses, platelets, lymphocytic azurophilic granules, lymphoglandular bodies and phagocytosed nuclear material should not be confused with the morulae.

Relation of clinical signs to pathological changes in 19 cases of canine distemper encephalomyelitis.

In an attempt to associate the clinical neurological syndromes with the neuropathological features of canine distemper (CD), 19 spontaneous cases with neurological involvement were examined, before and after euthanasia. Seventeen dogs were less than one year of age and all except two (89.4%) were unvaccinated against CD. Various extraneural signs associated with CD encephalomyelitis (CDE) were seen in 15 dogs. Generalized or localized myoclonus was the most common sign observed (13/19). Seventeen of the dogs presented with signs suggestive of one neuroanatomical location of lesions. Of these animals, seven had signs of cerebral, two of cerebellar, four of cervical, one of cervicothoracic, two of thoracolumbar and two of lumbosacral syndrome. The diagnosis of CD was confirmed immunohistochemically (detection of CD viral antigen), serologically (neutralizing serum antibody titre > or = 16) and histopathologically (CDV inclusion bodies, type of central nervous system lesions). An association of the neuroanatomical lesion location and the histopathological findings was noted in 14 out of 17 dogs (82.3%). Myoclonus could be attributed to lower motor neuron damage in eight out of 13 dogs (61.5%).

Trichinellosis in Greece: a review.

Trichinellosis which constitutes a public health problem in many countries seems to be of no importance on both pig industry and public health in Greece, where in spite of a law requiring mandatory use of trichinoscopy during meat inspection, muscle larvae have not been found in slaughtered pigs since 1957 in Thessaloniki and 1967 in Athens. Since its first recovery in 1946 and up to 1952, human trichinellosis has been found or suspected in 22 persons in the area of Athens and Thessaloniki. Moreover, in 1968, T. spiralis larvae were found incidentally in a human with laryngeal tumor and in 1971, living larvae were postmortem recovered in the diaphragm of a 70-year-old man. The average incidence of infection in pigs at that time was 0.02-2.2%. Since then, no other clinical case had been reported up to 1982-1984 when 15 people were found to harbor the parasite and fourteen of them were part of an outbreak which occurred in a small village in Northern Greece. Moreover, 1.07% of the pig serum samples which came from the same area, showed the presence of specific antibodies. Because of the above data, it is generally accepted that in Greece T. spiralis is only rarely spread in man.

Whole blood lead concentration and erythrocyte delta-aminolevulinic acid dehydratase (ALAD) activity in selected canine populations in Greece.

In a total number of 275 dogs of various ages, sex and breed, blood lead concentrations (BLC) and erythrocyte ALAD activity were measured. Sixty-six of the dogs were living in lead mining areas (Group A), 157 in urban areas (Group B) and 52 in rural areas (Group C) of Greece. Mean BLC differed significantly (P < 0.05) between locations and were 326,97 and 68 micrograms/L, respectively. Mean ALAD activity was significantly different (P < 0.05) only between Groups A and B as between groups A and C. A significant (P < 0.05) negative correlation existed between BLC and ALAD activity. A normal range of erythrocyte ALAD activity of 807-992 mumol/PBG/LRBC/h was established for dogs. None of the 33 Group A dogs and 2 of the Group B dogs that had a BLC of 350 micrograms/L presented clinical signs indicating acute or chronic lead intoxication. No erythrocyte basophilic stippling or large number of nucleated red blood cells were seen in the 30 dogs of Group A with BLC > 350 micrograms/L.