Quantitative and descriptive histological aspects of jaguar (Panthera onca Linnaeus, 1758) ear skin as a step towards formation of biobanks

Skin of mammals vulnerable to extinction, such as the jaguar, is used as a source of material in conservation strategies. The composition of skin is not uniform among species, and the ability to distinguish similarities in skin morphology in animal groups is fundamental in the application of skin tissue for use in biobanks. The aim of our study was to evaluate the structure, composition and capacity for culture of ear skin from the yellow and black jaguars. Both qualitative and quantitative methods were used, focusing on skin thickness, cell quantification and distribution, collagen density, proliferative activity and viability. Histomorphometrical study of the skin showed a total thickness of 273.2 and 274.6 µm for the yellow and black jaguars, respectively. Melanocytes and fibroblasts were, respectively, 9.7 and 23.0 for the yellow jaguar and 11.3 and 26.8 for the black jaguar. A collagen density of 67.0% and 49.0% was observed for yellow and black jaguars, respectively. Both animals presented a proliferative activity varying between 1.20 and 1.30. All tissues could promote cellular detachment, reaching subconfluence in 10–15 days. This kind of information from histomorphometrical features and cell cultures can be essential for a more targeted application of ear skin cryopreservation in this species, as such information will enable understanding the action of substances on tissues during the conservation process.     

Different isolates from Leishmania braziliensis complex induce distinct histopathological features in a murine model of infection

The aim of this study was to evaluate the histopathological features in tissues of mice infected by human isolates (I, II, and III) or the reference M2903 strain of Leishmania braziliensis complex. BALB/c and C57Bl/6 mice were infected in the hind footpad with 10⁶ stationary-phase promastigotes of L. braziliensis complex. The evolution of lesions was observed for 10 weeks and the animals were then euthanized and liver, spleen and popliteal lymph nodes were collected. Tissues were stained with hematoxylin and eosin and analyzed by immunohistochemistry assay. Increased thickness of infected footpads was observed in all animals, lesions were nodular and non-ulcerated. Mice infected with isolate I presented inflammatory infiltrates consisting predominantly of mononuclear cells in all tissues examined, and also a great number of megakaryocytes, compared with other isolates. Infection with isolate II led to an infected footpad enlargement not seen in other isolates. In addition, mononuclear infiltrates in the liver and hemosiderin in spleen were noted. Conversely, mice infected with either isolate III or M2903 strain only showed an increased number of megakaryocytes in spleen. All tissues examined had detectable amastigote forms of Leishmania by immunohistochemistry in all groups. Taking together, our results showed an unforeseen behavior of different isolates of L. braziliensis complex that led to diverse pathological findings.     

Ixodid ticks parasitizing Iberian red deer (Cervus elaphus hispanicus) and European wild boar (Sus scrofa) from Spain: geographical and temporal distribution

Commercial hunting of Spanish wild ungulates has made them an important economic resource. Wild ungulates may have an important role in the maintenance of ixodid tick populations, and also as reservoirs of pathogens. We studied the ixodid ticks that parasitize Iberian red deer and European wild boar from Spain. Ixodid ticks (n=6,336) were collected from 431 Iberian red deer and 142 wild boar in different regions of Spain. We found 10 different ixodid tick species parasitizing Iberian red deer, mainly Hyalomma marginatum marginatum (63.7%), Rhipicephalus (Boophilus) annulatus (7.9%) and R. bursa (7.5%). R. (Boophilus) annulatus was only collected in the province of Cádiz (southern Spain). We found 8 ixodid tick species on the wild boar, mainly Hy. m. marginatum (68.7%), R. bursa (14.6%) and Dermacentor marginatus (9.3%). We found one adult Hy. marginatum rufipes and one adult Hy. anatolicum excavatum parasitizing wild boar from south-central Spain. Mean prevalence of ixodid ticks was 41.3+/-0.08% (n=475) and 31+/-0.09% (n=284) and intensity of parasitization was 13.9+/-0.2 (n=283) and 13.6+/-0.3 (n=130) ticks/animal for Iberian red deer and wild boar, respectively. Only 5 of the 13 ixodid tick species found were shared by Iberian red deer and wild boar. This finding could indicate a host preference when Iberian red deer and wild boar share common habitats. In both Iberian red deer and wild boar from south-central Spain the monthly relative frequencies of Hy. m. marginatum and R. bursa presented an inverse pattern. The highest Hy. m. marginatum relative frequencies coincided with the lowest R. bursa relative frequencies along the year. R. bursa and I. ricinus were present in areas from northern to southern Spain while Hyalomma sp. and D. marginatus were exclusively collected in the two southern thirds of Spain. Haemaphysalis sp. and D. reticulatus were collected in northern Spain. Hy. m. marginatum and R. bursa were present during the whole year in red deer and wild boar from south-central Spain, showing more than one life cycle per year. These results are important for understanding the role of wild ungulates in the maintenance of tick infestations and to improve tick control programmes.     

Preservation of cadavers for surgical technique training

OBJECTIVE: To evaluate a technique for preservation of organoleptic tissue characteristics (color, odor, texture, and flexibility) in cadavers used for surgical instruction. STUDY DESIGN: Experimental study. ANIMALS: Forty-three canine cadavers. METHODS: Cadavers were preserved with a modified Larssen solution of the Hospital Cochim, Paris and cryopreservation. Tissue handling qualities were evaluated in surgical laboratory sessions. RESULTS: All cadavers kept texture and tissues consistency, especially skin and muscle, similar to those of live animals. Some skin desquamation and pallor of the mucous membranes occurred with repetitive freeze-thaw cycles. CONCLUSIONS: This preservation technique provides acceptable cadaver quality and tissue handling for use in surgical instruction. CLINICAL RELEVANCE: Preparation of patient cadavers by intravascular injection of modified Larssen solution yielded suitable instructional models for surgical training.     

Molecular characterization of Haemophilus parasuis ferric hydroxamate uptake (fhu) genes and constitutive expression of the FhuA receptor

Bacteria have evolved a set of highly specialized proteins to capture iron in iron-depleted environments. The acquisition and uptake of iron present in the extracellular milieu of eukaryotic organisms is indispensable for the growth and survival of microbial pathogens in the course of infection. Haemophilus parasuis is the causative agent of Gl?sser disease, which is responsible for considerable financial losses in pig-rearing worldwide. To gain insight into the mechanisms involved in siderophore-mediated iron uptake in H. parasuis, genes in the H. parasuis ferric hydroxamate uptake (Fhu) region were amplified in the work being reported here. As has been described in A. pleuropneumoniae, an Fhu genomic region was also present in H. parasuis, being composed of four potential consecutive open reading frames (ORF) designated as fhuC, fhuD, fhuB, and fhuA, respectively. By immunoblotting, using a cross-reactive polyclonal antibody raised against Actinobacillus pleuropneumoniae FhuA protein, it was demonstrated that this protein was constitutively expressed in H. parasuis and its level of expression was not modified under conditions of restricted iron availability. This is the first report describing the presence of the fhu genes in H. parasuis. Our results indicate that FhuA protein expression is not affected under iron-restricted conditions, however, it is one of the targets of the humoral immune response.     

Application of 10% imidacloprid/50% permethrin to prevent Ehrlichia canis exposure in dogs under natural conditions

Canine monocytic ehrlichiosis (CME) caused by Ehrlichia canis is the most known canine tick-borne disease (TBD) spread throughout the world. Preventing tick bites is a priority to reduce the risk of TBDs and it was the aim of the present study to evaluate the efficacy of a combination of imidacloprid 10% and permethrin 50% (ImPer) (Advantix; Bayer AG, Germany) in a spot-on formulation to control CME under field conditions. On January-March 2005, 845 dogs from two kennels in southern Italy (kennels of Bari (KB)- and Ginosa (KG)), with a history of tick infestation were initially tested by serology and PCR assay for E. canis infection. Data on Leishmania infantum infection were also available from a previous study carried out on the same dog population. One hundred twenty-six dogs (14.9%) presented anti-E. canis antibodies with a relative prevalence of 15.6% (n=65 dogs in KB) and 14.2% (n=61 dogs in KG). Five hundred thirty-five animals found negative both for E. canis and L. infantum infections were enrolled in three groups (Group A--treated with ImPer once a month; Group B--treated every 2 weeks; and Group C--untreated control animals) and monitored for E. canis infection by serology and PCR in November 2005 (first follow-up) and in March 2006 (second follow-up). The E. canis infection was serologically revealed, at the first and/or second follow-up, in 26 animals from Group C in KB and KG (mean incidence density rate (IDR), 13.24%) while in none of the animals from Group A (KB and KG) and only in one animal from Group B (IDR 1.13%) in KG. The final protection efficacy of ImPer ranged from 95.57% to 100% in Groups B and A. At PCR only 15 dogs from KG were positive for Rickettsiales only at the first follow-up and at the sequence analysis two (both in Group C) revealed 100% homology with E. canis sequences while 13 with Anaplasma platys. Four out of 13 A. platys PCR-positive dogs were also seropositive for E. canis at one or both follow-ups. ImPer, by virtue of its repellent and acaricidal activity against ticks, has been shown to be efficacious to prevent E. canis infection in treated dogs living under natural conditions in endemic areas.     

Influence of murine Toxocara canis infection on plasma and bronchoalveolar lavage fluid eosinophil numbers and its correlation with cytokine levels

Toxocara canis is a nematode of the Ascaridae family that normally parasites the small intestine of canid species. Humans are accidentally infected upon ingestion of embryonated eggs, and can manifest several clinical alterations such as fever, hepatomegaly, splenomegaly, respiratory symptoms, muscle pain and anorexia. In the present work, we investigated the kinetics of tissue distribution of L2 larva in lungs, liver, kidney, brain, skeletal muscle and myocardium. Also, we analyzed the blood and bronchoalveolar lavage fluid (BAL) for levels of IL-6, IFN-gamma, eotaxin and Regulated on Activation Normal T Cell Expressed and Secreted (RANTES) in experimental murine T. canis infection. We observed liver, lung and kidney lesions correlated to larva migration as early as the first day of infection. After the seventh post-infection day, larva could also be detected in brain, skeletal muscle and heart, as an indicator of biphasic migration pattern. Increased inflammatory activity was detected in BAL and plasma of infected animals, as was an intense eosinophil migration associated with an increase in the levels of all the cytokines studied. In conclusion, our results establish a tight correlation between tissue lesions caused by larva migration and increased plasma levels of pro-inflammatory and eosinophil chemotactic cytokines. Thus, murine T. canis infection may prove to be useful in understanding the role of cytokines in infection.