Occurrence and molecular characterization of Cryptosporidium spp. isolated from domestic animals in a rural area surrounding Atlantic dry forest fragments in Teodoro Sampaio municipality, State of São Paulo, Brazil

The aim of this study was to assess the occurrence of Cryptosporidium in domestic animals in rural properties surrounding rain forest fragments within the municipality of Teodoro Sampaio, southeastern Brazil. Conventional sucrose flotation method followed by molecular characterization of the parasites by sequencing PCR products amplified from SSU rRNA gene were used. Stool samples were collected from domestic animals raised as pets and livestock in all rural properties surrounding three forest fragments. Samples from cattle (197), equine (63), pigs (25), sheep (11), and dogs (28) were collected from 98 rural properties. The frequency of occurrence of Cryptosporidium within each animal species was 3.0% (6/197) among cattle and 10.7% (3/28) among dogs. Cryptosporidium was not detected in stool samples from equine, sheep, and pigs. All sequences obtained from the six samples of calves showed molecular identity with Cryptosporidium andersoni while all sequences from dog samples were similar to C. canis. The frequency of occurrence of Cryptosporidium in these domestic animal species was low. The absence of C. parvum in the present study suggests that the zoonotic cycle of cryptosporidiosis may not be relevant in the region studied. The presence of Cryptosporidium species seldom described in humans may be, otherwise, important for the wild fauna as these animals are a source of infection and dissemination of this protozoan to other animal species. The impact and magnitude of infection by C. andersoni in wild ruminants and C. canis in wild canids have to be assessed in future studies to better understand the actual importance of these species in this region.

Modulation of extracellular matrix by nutritional hepatotrophic factors in thioacetamide-induced liver cirrhosis in the rat

Nutritional substances associated to some hormones enhance liver regeneration when injected intraperitoneally, being denominated hepatotrophic factors (HF). Here we verified if a solution of HF (glucose, vitamins, salts, amino acids, glucagon, insulin, and triiodothyronine) can revert liver cirrhosis and how some extracellular matrices are affected. Cirrhosis was induced for 14 weeks in 45 female Wistar rats (200 mg) by intraperitoneal injections of thioacetamide (200 mg/kg). Twenty-five rats received intraperitoneal HF twice a day for 10 days (40 mL·kg-1·day-1) and 20 rats received physiological saline. Fifteen rats were used as control. The HF applied to cirrhotic rats significantly: a) reduced the relative mRNA expression of the genes: Col-α1 (-53%), TIMP-1 (-31.7%), TGF-β1 (-57.7%), and MMP-2 (-41.6%), whereas Plau mRNA remained unchanged; b) reduced GGT (-43.1%), ALT (-17.6%), and AST (-12.2%) serum levels; c) increased liver weight (11.3%), and reduced liver collagen (-37.1%), regenerative nodules size (-22.1%), and fibrous septum thickness. Progranulin protein (immunohistochemistry) and mRNA (in situ hybridization) were found in fibrous septa and areas of bile duct proliferation in cirrhotic livers. Concluding, HF improved the histology and serum biochemistry of liver cirrhosis, with an important reduction of interstitial collagen and increased extracelullar matrix degradation by reducing profibrotic gene expression.

Neurobehavioral, reflexological and physical development of Wistar rat offspring exposed to ayahuasca during pregnancy and lactation

Ayahuasca is a hallucinogenic beverage prepared by the decoction of plants native to the Amazon Basin region. The beverage has been used throughout the world by members of some syncretic religious movements. Despite the recent legalization of ayahuasca in Brazil for religious purposes, there is little pre-clinical and clinical information attesting to its safety, particularly in relation to the use during pregnancy. The aim of the current work was to determine the effects of perinatal exposure to ayahuasca (from the 6th day of pregnancy to the 10th day of lactation) on physical, reflexology and neurobehavioral parameters of the Wistar rat offspring. The offspring showed no statistically significant changes in the physical and reflexology parameters evaluated. However, in adult rats, perinatally exposed to ayahuasca, an increase in frequency of entries in open arms in elevated plus-maze test, a decrease in total time of interaction in social interaction test, a decrease in time of latency for the animal to start swimming and a decrease of the minimum convulsant dose induced by pentylenetetrazol were observed. In conclusion, our results showed that the use of ayahuasca by mothers during pregnancy and lactation reduced the general anxiety and social motivation of the rat offspring. Besides, it promoted a higher sensitivity for initiation and spread of seizure activity.

Modelo de suplementação nutricional com fatores hepatotróficos aumenta proliferação celular em fígado de ratos sadios

Foram avaliados dois protocolos de administração, em ratos sadios, de uma solução de fatores hepatotróficos (FH), composta por aminoácidos, vitaminas, sais minerais, glicose, insulina, glucagon e triiodotironina (T3). A solução foi administrada durante 10 dias, 40mg/kg/dia, i.p., em duas, grupo 2xFH (n=15), ou três doses, grupo 3xFH (n=15), diárias. Foram observados os efeitos na proliferação celular dos hepatócitos, na angiogênese e na matriz extracelular hepática, assim como as possíveis reações adversas. Os animais dos grupos 2xFH e 3xFH apresentaram aumento da massa hepática de 30,1% e 22,5%, respectivamente, em relação ao grupo-controle (CT; n=15). O índice de proliferação hepatocelular foi maior nos grupos 2xFH (1,4%) e 3xFH (1,2%) em relação ao grupo CT (0,53%), e a densitometria relativa do fator de crescimento do endotélio vascular pelo imunoblot não revelou diferença estatística entre os três grupos. Nos grupos 2xFH e 3xFH, houve redução do colágeno intersticial em relação ao grupo CT. A solução de FH estimulou o crescimento hepático e reduziu o volume de colágeno perissinusoidal. A administração em três doses diárias resultou em mortalidade de 26,7%, possivelmente pelo excessivo estresse da manipulação e pela menor adaptação fisiológica dos ratos, o que não ocorreu nos grupos 2xFH e CT. Para esse tipo de abordagem em ratos, o procedimento experimental mais apropriado, seguro, com melhor chance de adaptação dos animais e com resultados significativos é a aplicação dos FH em duas doses diárias.

Clonal relationship among atypical enteropathogenic Escherichia coli strains isolated from different animal species and humans

Forty-nine typical and atypical enteropathogenic Escherichia coli (EPEC) strains belonging to different serotypes and isolated from humans, pets (cats and dogs), farm animals (bovines, sheep, and rabbits), and wild animals (monkeys) were investigated for virulence markers and clonal similarity by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). The virulence markers analyzed revealed that atypical EPEC strains isolated from animals have the potential to cause diarrhea in humans. A close clonal relationship between human and animal isolates was found by MLST and PFGE. These results indicate that these animals act as atypical EPEC reservoirs and may represent sources of infection for humans. Since humans also act as a reservoir of atypical EPEC strains, the cycle of mutual infection of atypical EPEC between animals and humans, mainly pets and their owners, cannot be ruled out since the transmission dynamics between the reservoirs are not yet clearly understood.

Extended-spectrum beta-lactamases among Enterobacteriaceae isolated in a public hospital in Brazil

Extended-spectrum beta-lactamases (ESBL) in enterobacteria are recognized worldwide as a great hospital problem. In this study, 127 ESBL-producing Enterobacteriaceae isolated in one year from inpatients and Outpatients at a public teaching hospital at Sao Paulo, Brazil, were Submitted to analysis by PCR with specific primers for bla(SHV), bla(TEM) and bla(CTX-M) genes. From the 127 isolates, 96 (75.6%) Klebsiella pneumoniae, 12 (9.3%) Escherichia coli, 8 (6.2%) Morganella morganii, 3 (2.3%) Proteus mirabilis, 2 (1.6%) Klebsiella oxytoca, 2 (1.6%) Providencia rettgeri, 2 (1.6%) Providencia stuartti, 1 (0.8%) Enterobacter aerogenes and 1 (0.8%) Enterobacter cloacae were identified as ESBL producers. Bla(SHV), bla(TEM), and bla(CTX-M) were detected in 63%, 17.3% and 33.9% strains, respectively. Pulsed field get eletrophoresis genotyping of K. pneumoniae revealed four main molecular patterns and 29 unrelated profiles. PCR results showed a high variety of ESBL groups among strains, in nine different species. The results Suggest the spread of resistance genes among genetically different strains of ESBL-producing K. pneumoniae in some hospital wards, and also that some strongly related strains were identified in different hospital wards, Suggesting clonal spread in the institutional environment

Intestinal Anti-Inflammatory Activity of Baccharis dracunculifolia in the Trinitrobenzenesulphonic Acid Model of Rat Colitis

Baccharis dracunculifolia DC (Asteraceae) is a Brazilian medicinal plant popularly used for its antiulcer and anti-inflammatory properties. This plant is the main botanical source of Brazilian green propolis, a natural product incorporated into food and beverages to improve health. The present study aimed to investigate the chemical profile and intestinal anti-inflammatory activity of B. dracunculifolia extract on experimental ulcerative colitis induced by trinitrobenzenosulfonic acid (TNBS). Colonic damage was evaluated macroscopically and biochemically through its evaluation of glutathione content and its myeloperoxidase (MPO) and alkaline phosphatase activities. Additional in vitro experiments were performed in order to test the antioxidant activity by inhibition of induced lipid peroxidation in the rat brain membrane. Phytochemical analysis was performed by HPLC using authentic standards. The administration of plant extract (5 and 50 mgkg(-1)) significantly attenuated the colonic damage induced by TNBS as evidenced both macroscopically and biochemically. This beneficial effect can be associated with an improvement in the colonic oxidative status, since plant extract prevented glutathione depletion, inhibited lipid peroxidation and reduced MPO activity. Caffeic acid, p-coumaric acid, aromadendrin-4-O-methyl ether, 3-prenyl-p-coumaric acid, 3,5-diprenyl-p-coumaric acid and baccharin were detected in the plant extract.

Comparative study of the hygienic behavior of Carniolan and Africanized honey bees directed towards grouped versus isolated dead brood cells

In Apis mellifera, hygienic behavior involves recognition and removal of sick, damaged or dead brood from capped cells. We investigated whether bees react in the same way to grouped versus isolated damaged capped brood cells. Three colonies of wild-type Africanized honey bees and three colonies of Carniolan honey bees were used for this investigation. Capped worker brood cells aged 12 to 14 days old were perforated with the pin-killing method. After making holes in the brood cells, the combs were placed back into the hives; 24 h later the number of cleaned cells was recorded in areas with pin-killed and control brood cells. Four repetitions were made in each colony. Isolated cells were more frequently cleaned than grouped cells, though variance analysis showed no significant difference (P = 0.1421). Carniolan bees also were somewhat, though not significantly more hygienic than Africanized honey bees (P = 0.0840). We conclude that honey bees can detect and remove both isolated and grouped dead brood. The tendency towards greater hygienic efficiency directed towards grouped pin-killed brood may be a consequence of a greater concentration of volatiles emanating from the wounds in the dead pupae.

Antileishmanial Activity of the Hydroalcoholic Extract of Miconia langsdorffii, Isolated Compounds, and Semi-Synthetic Derivatives

The in vitro activity of the crude hydroalcoholic extract of the aerial parts of Miconia langsdorffii Cogn. was evaluated against the promastigote forms of L. amazonensis, the causative agent of cutaneous leishmaniasis in humans. The bioassay-guided fractionation of this extract led to identification of the triterpenes ursolic acid and oleanolic acid as the major compounds in the fraction that displayed the highest activity. Several ursolic acid semi-synthetic derivatives were prepared, to find out whether more active compounds could be obtained. Among these ursolic acid-derived substances, the C-28 methyl ester derivative exhibited the best antileishmanial activity.

Signatures of oxygen on Cu(3)Au(100): From isolated impurity to oxide regimes

We analyze the scanning tunneling microscopy (STM) signatures for the O/Cu(3)Au(100) surface from the low-coverage (isolated impurity) to high-coverage (oxide) regimes. First-principles calculations show that oxygen signatures switch from dark to bright spots as the oxygen coverage increases. This behavior is nicely traced back to a change in the oxygen orbital character of the Fermi-level electronic states. Our results allow for the chemical identification by STM of oxygen and copper atoms in the fully ordered O/Cu(3)Au(100)-c(2x2) surface.

Differential regulation of PGC-1 alpha expression in rat liver and skeletal muscle in response to voluntary running

Background: The beneficial actions of exercise training on lipid, glucose and energy metabolism and insulin sensitivity appear to be in part mediated by PGC-1 alpha. Previous studies have shown that spontaneously exercised rats show at rest enhanced responsiveness to exogenous insulin, lower plasma insulin levels and increased skeletal muscle insulin sensitivity. This study was initiated to examine the functional interaction between exercise-induced modulation of skeletal muscle and liver PGC-1 alpha protein expression, whole body insulin sensitivity, and circulating FFA levels as a measure of whole body fatty acid (lipid) metabolism. Methods: Two groups of male Wistar rats (2 Mo of age, 188.82 +/- 2.77 g BW) were used in this study. One group consisted of control rats placed in standard laboratory cages. Exercising rats were housed individually in cages equipped with running wheels and allowed to run at their own pace for 5 weeks. At the end of exercise training, insulin sensitivity was evaluated by comparing steady-state plasma glucose (SSPG) concentrations at constant plasma insulin levels attained during the continuous infusion of glucose and insulin to each experimental group. Subsequently, soleus and plantaris muscle and liver samples were collected and quantified for PGC-1 alpha protein expression by Western blotting. Collected blood samples were analyzed for glucose, insulin and FFA concentrations. Results: Rats housed in the exercise wheel cages demonstrated almost linear increases in running activity with advancing time reaching to maximum value around 4 weeks. On an average, the rats ran a mean (Mean +/- SE) of 4.102 +/- 0.747 km/day and consumed significantly more food as compared to sedentary controls (P < 0.001) in order to meet their increased caloric requirement. Mean plasma insulin (P < 0.001) and FFA (P < 0.006) concentrations were lower in the exercise-trained rats as compared to sedentary controls. Mean steady state plasma insulin (SSPI) and glucose (SSPG) concentrations were not significantly different in sedentary control rats as compared to exercise-trained animals. Plantaris PGC-1 alpha protein expression increased significantly from a 1.11 +/- 0.12 in the sedentary rats to 1.74 +/- 0.09 in exercising rats (P < 0.001). However, exercise had no effect on PGC-1 alpha protein content in either soleus muscle or liver tissue. These results indicate that exercise training selectively up regulates the PGC-1 alpha protein expression in high-oxidative fast skeletal muscle type such as plantaris muscle. Conclusion: These data suggest that PGC-1 alpha most likely plays a restricted role in exercise-mediated improvements in insulin resistance (sensitivity) and lowering of circulating FFA levels.

The Effects of a 785-nm AlGaInP Laser on the Regeneration of Rat Anterior Tibialis Muscle After Surgically-Induced Injury

Objective: This study aims to investigate the effects of low-level laser therapy (LLLT) on muscle regeneration. For this purpose, the anterior tibialis muscle of 48 male Wistar rats received AlGaInP laser treatment (785 nm) after surgically-induced injury. Background Data: Few studies have been conducted on the effects of LLLT on muscle regeneration at different irradiation doses. Materials and Methods: The animals were randomized into four groups: uninjured rats (UN); uninjured and laser-irradiated rats (ULI); injured rats (IN); and injured and laser-irradiated rats (ILI). The direct contact laser treatment was started 24 h after surgery. An AlGaInP diode laser emitting 75 mW of continuous power at 785 nm was used for irradiation. The laser probe was placed at three treatment points to deliver 0.9 J per point, for a total dose of 2.7 J per treatment session. The animals were euthanized after treatment sessions 1, 2, and 4. Mounted sections were stained with hematoxylin and eosin and used for quantitative morphological analysis, in which the number of leukocytes and fibroblasts were counted over an area of 4480 mu m(2). The data were statistically analyzed by analysis of variance (ANOVA) and the Bonferroni t-test. Results: Quantitative data showed that the number of both polymorphonuclear and mononuclear leukocytes in the inflammatory infiltrate at the injury site was smaller in the ILI(1), ILI(2), and ILI(4) subgroups compared with their respective control subgroups (IN(1), IN(2), and IN(4)) for sessions 1, 2, and 4, respectively (p < 0.05). On the other hand, the number of fibroblasts increased after the fourth treatment session (p < 0.05). With regard to the regeneration of muscle fibers following injury, only after the fourth treatment session was it possible to find muscle precursor cells such as myoblasts and some myotubes in the ILI(4) subgroup. Conclusion: During the acute inflammatory phase, the AlGaInP laser treatment was found to have anti-inflammatory effects, reducing the number of leukocytes at the injury site and accelerating the regeneration of connective tissue.

Survival and Neuronal Differentiation of Mesenchymal Stem Cells Transplanted into the Rodent Brain Are Dependent upon Microenvironment

Introduction: The successful integration of stem cells in adult brain has become a central issue in modern neuroscience. In this study we sought to test the hypothesis that survival and neurodifferentiation of mesenchymal stem cells (MSCs) may be dependent upon microenvironmental conditions according to the site of implant in the brain. Methods: MSCs were isolated from adult rats and labeled with enhanced-green fluorescent protein (eGFP) lentivirus. A cell suspension was implanted stereotactically into the brain of 50 young rats, into one neurogenic area (hippocampus), and into another nonneurogenic area (striatum). Animals were sacrificed 6 or 12 weeks after surgery, and brains were stained for mature neuronal markers. Cells coexpressing NeuN (neuronal specific nuclear protein) and GFP (green fluorescent protein) were counted stereologically at both targets. Results: The isolated cell population was able to generate neurons positive for microtubule-associated protein 2 (MAP2), neuronal-specific nuclear protein (NeuN), and neurofilament 200 (NF200) in vitro. Electrophysiology confirmed expression of voltage-gated ionic channels. Once implanted into the hippocampus, cells survived for up to 12 weeks, migrated away from the graft, and gave rise to mature neurons able to synthesize neurotransmitters. By contrast, massive cell degeneration was seen in the striatum, with no significant migration. Induction of neuronal differentiation with increased cyclic adenosine monophosphate in the culture medium before implantation favored differentiation in vivo. Conclusions: Our data demonstrated that survival and differentiation of MSCs is strongly dependent upon a permissive microenvironment. Identification of the pro-neurogenic factors present in the hippocampus could subsequently allow for the integration of stem cells into nonpermissive areas of the central nervous system.

Mast cell repopulation of the peritoneal cavity: contribution of mast cell progenitors versus bone marrow derived committed mast cell precursors

Background: Mast cells have recently gained new importance as immunoregulatory cells that are involved in numerous pathological processes. One result of these processes is an increase in mast cell numbers at peripheral sites. This study was undertaken to determine the mast cell response in the peritoneal cavity and bone marrow during repopulation of the peritoneal cavity in rats. Results: Two mast cell specific antibodies, mAb AA4 and mAb BGD6, were used to distinguish the committed mast cell precursor from more mature mast cells. The peritoneal cavity was depleted of mast cells using distilled water. Twelve hours after distilled water injection, very immature mast cells could be isolated from the blood and by 48 hours were present in the peritoneal cavity. At this same time the percentage of mast cells in mitosis increased fourfold. Mast cell depletion of the peritoneal cavity also reduced the total number of mast cells in the bone marrow, but increased the number of mast cell committed precursors. Conclusions: In response to mast cell depletion of the peritoneal cavity, a mast cell progenitor is released into the circulation and participates in repopulation of the peritoneal cavity, while the committed mast cell precursor is retained in the bone marrow.

The Lectin ArtinM Induces Recruitment of Rat Mast Cells from the Bone Marrow to the Peritoneal Cavity

Background: The D-mannose binding lectin ArtinM is known to recruit neutrophils, to degranulate mast cells and may have potential therapeutic applications. However, the effect of ArtinM on mast cell recruitment has not been investigated. Methodology: Male Wistar rats were injected i.p. with ArtinM or ConA (control). The ability of the lectin to degranulate peritoneal and mesenteric mast cells was examined. Recruitment of mast cells to the peritoneal cavity and mesentery after ArtinM injection was examined with or without depletion of peritoneal mast cells by distilled water. Results: ArtinM degranulated both peritoneal and mesentery mast cells in vitro. Three days after i.p. injection of the lectin there were reduced numbers of mast cells in the peritoneal lavage, while at 7 days post injection of ArtinM, the number of peritoneal mast cells was close to control values. Since immature mast cells are recruited from the bone marrow, the effect of the lectin on bone marrow mast cells was examined. Injection of ArtinM resulted in an increased number of mast cells in the bone marrow. To determine if degranulation of mast cells in the peritoneal cavity was required for the increase in bone marrow mast cells, the peritoneal cavity was depleted of mast cells with ultrapure water. Exposure to ArtinM increased the number of mast cells in the bone marrow of rats depleted of peritoneal mast cells. Conclusions: The ArtinM induced recruitment of mast cells from the bone marrow to the peritoneal cavity may partially explain the therapeutic actions of ArtinM.