Hemorrhagic disease in dogs infected with an unclassified intraendothelial piroplasm in southern Brazil

A hemorrhagic disease affecting dogs in Brazil, referred to popularly as "nambiuvú" (bloody ears) and believed to be transmitted by ticks, has been observed in animals infected with an organism described originally in 1910 as a piroplasm, and known locally as Rangelia vitalli. In this series of 10 cases, the disease was characterized by anaemia, jaundice, fever, spleno- and lymphadenomegaly, hemorrhage in the gastrointestinal tract, and persistent bleeding from the nose, oral cavity and tips, margins and outer surface of the pinnae. The ixodid ticks Rhipicephalus sanguineus and Amblyomma aureolatum infested affected dogs from suburban and rural areas, respectively. Laboratory findings included regenerative anaemia, spherocytosis, icteric plasma and bilirubinuria. Those intracellular organisms were found in bone marrow smears but not in blood smears. Microscopically, zoites were seen within the cytoplasm of blood capillary endothelial cells. Parasitized and non-parasitized endothelial cells were positive immunohistochemically for von Willebrand factor (vWF). Langhans-type multinucleate giant cells were observed in the lymph nodes and choroid plexus. There was prominent erythrophagocytosis by macrophages in the lymph node sinuses and infiltration of the medullary cords by numerous plasma cells. Ultrastructurally, this organism had an apical complex that included a polar ring and rhoptries but no conoid. This parasite was contained within a parasitophorous vacuole that had a trilaminar membrane with villar protrusions and was situated in the cytoplasm of capillary endothelial cells. This organism tested positive by immunohistochemistry for Babesia microti. This pathogen was also positive by in situ hybridization for B. microti. Tentative clinical diagnosis in these cases was based on the history, clinical picture, haemogram and favorable response to therapy, and confirmed through microscopic examination of smears from the bone marrow or histological sections of multiple tissues, especially lymph nodes where zoites were most frequently found. The disease was reproduced by intravenous inoculation of blood from a naturally infected dog into an experimental dog. The authors demonstrate in this study that this organism is a protozoa of the phylum Apicomplexa, order Piroplasmorida. This piroplasm seems to be different from Babesia since it has an intraendothelial stage. Molecular phylogenetic analysis is necessary to better characterize this parasite and clarify its taxonomic status.     

Use of autogenous fascia lata graft for perineal herniorrhaphy in dogs

OBJECTIVE: To evaluate the effectiveness of a herniorrhaphy technique, using an autogenous fascia lata graft (FLG) for perineal hernia (PH) repair in dogs. STUDY DESIGN: Prospective clinical study. ANIMALS: Twelve dogs with PH. METHOD: PHs were repaired with FLG harvested from the dog's ipsilateral thigh and sutured directly into the perineal defect. Correction of associated conditions, and castration were performed. Surgical time, pain, inflammation, pattern of defecation, lameness, hospitalization time, postoperative complications, and owner satisfaction were recorded. Histopathologic examination was performed in 1 dog euthanatized 10 months after repair. RESULTS: Hernia did not recur (mean follow-up, 5.8 months). Lameness was the most frequent minor complication, and was resolved within a few days. Transient rectal prolapse occurred in 2 dogs with bilateral PH. The mean (+/-SD) hospitalization was 1.8+/-0.9 days, and the surgical time was 76.5+/-9.8 minutes. Histopathologic examination in 1 dog revealed perfect integration of FLG into adjacent tissues without substantial tissue reaction. CONCLUSIONS: FLG reconstruction of PH is a simple, effective method of treatment. CLINICAL RELEVANCE: FLG can be used without major complications for primary repair of PH, as an augmentation procedure when the internal obturator muscle is thin or friable, or when herniation has recurred after another repair technique.     

Attaching and effacing Escherichia coli isolated from dogs in Brazil: characteristics and serotypic relationship to human enteropathogenic E. coli (EPEC)

Escherichia coli isolates recovered from 182 fecal specimens from dogs up to five months old from the cities of S?o Paulo and Campinas, SP, Brazil, were examined by polymerase chain reaction (PCR) for several virulence factors and properties. The eae gene was found in 23 isolates of E. coli from 22 dogs, 19 of 146 (13%) from dogs with diarrhea and 3 of 36 (8.3%) from dogs with no diarrhea. Two different eae+ isolates were recovered from one dog with diarrhea. Isolates from two dogs with diarrhea harbored the bfpA gene, and none of the isolates possessed genes for enterotoxins, the EAF plasmid or Shiga toxins. PCR showed that, among the 23 isolates, eight were positive for beta intimin, six for gamma, two for, one for alpha, one for kappa, and five showed no amplification with any of the nine pairs of specific intimin primers used. PCR also showed that the LEE (locus of enterocyte effacement) was inserted in selC in four isolates, likely in pheU in seven isolates, and in undetermined sites in twelve isolates. Fifteen isolates adhered to HEp-2 cells and were fluorescence actin staining (FAS) positive. The predominant adherence pattern was the localized adherence-like (LAL) pattern. The eae-positive isolates belonged to a wide diversity of serotypes, including O111:H25, O119:H2 and O142:H6, which are serotypes that are common among human EPEC. These results confirmed the presence of EPEC in dogs (DEPEC) with and without diarrhea. The virulence factors found in these strains were similar to those in human EPEC, leading to the possibility that EPEC may move back and forth among human and canine populations.     

Cell proliferation in feline normal, hyperplastic and neoplastic mammary tissue--an immunohistochemical study

The expression of a proliferation cell marker in neoplastic and non-neoplastic mammary tissue in 31 cats was assessed by immunohistochemistry in formalin-fixed paraffin-embedded samples using a monoclonal antibody against nuclear antigen Ki-67 (MIB-1). The results revealed that cell proliferation was more intense in hyperplastic than in normal mammary tissue. Carcinomas exhibited a higher MIB-1 index than benign tumours. There was also a positive correlation between proliferative activity and the histological grade of carcinomas. Fibroadenomatous change, which is considered to be hyperplastic and associated with favourable biological behaviour, exhibited high proliferative activity involving both epithelial and mesenchymal components. MIB-1 detected in formalin-fixed material with pre-treatment with antigen retrieval solution appeared to be a reliable marker of proliferation in feline mammary tumours, but further studies are needed to investigate the value of this proliferation marker in predicting clinical outcome and/or as a prognostic factor in feline mammary lesions.     

Adenosine deaminase activity in serum, erythrocytes and lymphocytes of rats infected with Leptospira icterohaemorrhagiae

Leptospirosis is a systemic disease of humans and domestic animals, mainly dogs, cattle and swine. The course of human leptospirosis varies from mild to severe fatal forms and the most severe form of human leptospirosis is principally caused by Leptospira interrogans serovar icterohaemorrhagiae (L. icterohaemorrhagiae). The enzyme adenosine deaminase (ADA) plays an important role in the production and differentiation of blood cells. The aim of this study was to evaluate the activity of ADA in serum, erythrocytes and lymphocytes of rats infected with L. icterohaemorrhagiae, as compared with non-infected rats. Twenty-four adult rats, divided into two uniform groups (A and B) were used for the enzymatic assays. The animals in Group B were inoculated intraperitoneally with 2×10(8) leptospires/rat, and the rodents in Group A (control) were not-inoculated. Blood collection was performed on days 5 and 15 post-infection (PI) and the blood used to assess the ADA activity. The infection by L.icterohaemorrhagiae altered erythrocyte count, hemoglobin concentration and hematocrit, causing a decrease in all these parameters on day 15 PI. Lymphocytes decreased significantly on day 15 PI, and ADA activity in serum was inhibited in infected rats on days 5 and 15 PI and its activity in erythrocytes were increased on day 5 PI. On day 5 PI, we found an increase in ADA activity in erythrocytes of infected rats. No correlation was observed between hematocrit and erythrocyte ADA activity on days 5 and 15 PI. The ADA activity was inhibited in rats infected on day 15 PI. A positive correlation (r(2)=60) was also observed between the number of lymphocytes and ADA activity in lymphocytes on day 15 PI (P<0.05). In conclusion, our results showed that the ADA activity is altered in serum, lymphocytes and erythrocytes in experimental infection by L.icterohaemorrhagiae in rats, concomitantly with hematological parameters.     

Toxic effects of domoic acid in the seabream Sparus aurata

Neurotoxicity induced in fish by domoic acid (DA) was assessed with respect to occurrence of neurotoxic signs, lethality, and histopathology by light microscopy. Sparus aurata were exposed to a single dose of DA by intraperitoneal (i.p.) injection of 0, 0.45, 0.9, and 9.0 mg DA kg(-1) bw. Mortality (66.67 ± 16.67%) was only observed in dose of 9.0 mg kg(-1) bw. Signs of neurological toxicity were detected for the doses of 0.9 and 9.0 mg DA kg(-1) bw. Furthermore, the mean concentrations (±SD) of DA detected by HPLC-UV in extracts of brain after exposure to 9.0 mg DA kg(-1) bw were 0.61 ± 0.01, 0.96 ± 0.00, and 0.36 ± 0.01 mg DA kg(-1) tissue at 1, 2, and 4 hours. The lack of major permanent brain damage in S. aurata, and reversibility of neurotoxic signs, suggest that lower susceptibility to DA or neuronal recovery occurs in affected individuals.     

Trypanocidal Action of (?)-Elatol Involves an Oxidative Stress Triggered by Mitochondria Dysfunction

Natural compounds have shown good potential for the discovery of new chemotherapeutics for the treatment of Chagas’ disease. Recently, our group reported the effective trypanocidal activity of (−)-elatol, extracted from the red macroalgae Laurencia dendroidea present in the Brazilian coast against Trypanosoma cruzi. However, the mechanism of action of this compound has remained unclear. There are only hypotheses concerning its action on mitochondrial function. Here, we further investigated the mechanisms of action of (−)-elatol on trypomastigotes of T. cruzi. For this, we evaluated some biochemical alterations in trypomastigotes treated with (−)-elatol. Our results show that (−)-elatol induced depolarization of the mitochondrial membrane, an increase in the formation of mitochondrial superoxide anion and loss of cell membrane and DNA integrity. Additionally, (−)-elatol induced formation of autophagic vacuoles and a decrease in cell volume. All together, these results suggest that the trypanocidal action of (−)-elatol involves multiple events and mitochondria might be the initial target organelle. Our hypothesis is that the mitochondrial dysfunction leads to an increase of ROS production through the electron transport chain, which affects cell membrane and DNA integrity leading to different types of parasite death.     

Antiproliferative Activity of Fucan Nanogel

Sulfated fucans comprise families of polydisperse natural polysaccharides based on sulfated L-fucose. Our aim was to investigate whether fucan nanogel induces cell-specific responses. To that end, a non toxic fucan extracted from Spatoglossum schröederi was chemically modified by grafting hexadecylamine to the polymer hydrophilic backbone. The resulting modified material (SNFuc) formed nanosized particles. The degree of substitution with hydrophobic chains was close to 100%, as estimated by elemental analysis. SNFfuc in aqueous media had a mean diameter of 123 nm and zeta potential of −38.3 ± 0.74 mV, as measured by dynamic light scattering. Nanoparticles conserved their size for up to 70 days. SNFuc cytotoxicity was determined using the MTT assay after culturing different cell lines for 24 h. Tumor-cell (HepG2, 786, H-S5) proliferation was inhibited by 2.0%–43.7% at nanogel concentrations of 0.05–0.5 mg/mL and rabbit aorta endothelial cells (RAEC) non-tumor cell line proliferation displayed inhibition of 8.0%–22.0%. On the other hand, nanogel improved Chinese hamster ovary (CHO) and monocyte macrophage cell (RAW) non-tumor cell line proliferation in the same concentration range. The antiproliferative effect against tumor cells was also confirmed using the BrdU test. Flow cytometric analysis revealed that the fucan nanogel inhibited 786 cell proliferation through caspase and caspase-independent mechanisms. In addition, SNFuc blocks 786 cell passages in the S and G2-M phases of the cell cycle.     

Increased Antioxidant Content in Juice Enriched with Dried Extract of Pomegranate (Punica granatum) Peel

Antioxidants are compounds responsible for free radical scavenging in the body. They protect the organism from oxidative modification of cells and tissues. These modifications have been associated with degenerative diseases, atherosclerosis and carcinogenesis. Punica granatum displays high antioxidant potential due to the presence of phenolic compounds, which are capable of disease prevention. The present study showed the highest antioxidant activity in pomegranate peel than in seeds and pulp. Based on these results, pomegranate peel was used to produce dried extract that was added to commercial tomato juice and orange juice with strawberries. Analysis to determine the content of phenolic compounds and antioxidant activity was performed on pomegranate pulp, seeds and peel and in juices enriched with dried extract of pomegranate peel. The dried extract was responsible for a significant increase in antioxidant activity of the juices, proportional to the concentrations added. However, although both flavors of enriched juices displayed high antioxidant levels, the samples with higher dried extract concentrations received the lowest scores from sensory analysis participants due to the characteristic astringent flavor of pomegranate peels. Therefore, to obtain greater acceptance in the consumer market, we concluded that the maximum addition of dried pomegranate peel extract is 0.5% in tomato juice and orange juice with strawberries.