Risk factors associated with the bovine subclinical mastitis in an Amazon micro-region.

In this study, we evaluated the main risk factors for the occurrence of bovine mastitis, in the southeastern of Pará, in the Brazilian Amazon. We surveyed 91 dairy farmers to identify management practices and bovine breed characteristics. From each farm, 50 mL of milk sample was collected for microbiological analysis and somatic cell count (SCC). Depending on the management practices and breed, a logit model was used to determine the odds ratio of subclinical mastitis (SCM) occurrence. In irrigated pastures, an SCM-associated risk factor, the occurrence of SCM was 5.03 times higher than that in the non-irrigated pastures. Similarly, in Girolando breed herds, the occurrence of SCM increased by 5.8 times compared to the crossbred herds. Moreover, the occurrence of mastitis was 33 times higher in farms using common cloths for drying teats than in farms using paper towels. Therefore, adoption of better management practices can lead to SCC reduction, milk quality improvement and a guarantee to contain SCC within prescribed Brazilian limits for the Amazon region.

Novel Microsatellite Markers Used for Determining Genetic Diversity and Tracing of Wild and Farmed Populations of the Amazonian Giant Fish Arapaima gigas.

The Amazonian symbol fish Arapaima gigas is the only living representative of the Arapamidae family. Environmental pressures and illegal fishing threaten the species' survival. To protect wild populations, a national regulation must be developed for the management of A. gigas throughout the Amazon basin. Moreover, the reproductive genetic management and recruitment of additional founders by aquaculture farms are needed to mitigate the damage caused by domestication. To contribute to the sustainable development, we investigated the genetic diversity of wild and cultivated populations of A. gigas and developed a panel composed by 12 microsatellite markers for individual and population genetic tracing. We analyzed 368 samples from three wild and four farmed populations. The results revealed low rates of genetic diversity in all populations, loss of genetic diversity and high inbreeding rates in farmed populations, and genetic structuring among wild and farmed populations. Genetic tracing using the 12 microsatellite markers was effective, and presented a better performance in identifying samples at the population level. The 12-microsatellite panel is appliable to the legal aspects of the trade of the A. gigas, such as origin discrimination, reproductive genetic management by DNA profiling, and evaluation and monitoring of genetic diversity.

Braga nasuta (Cymothoidae): an ectoparasite of the Giant Amazonian fish Arapaima gigas (Osteoglossidae) fingerlings cultured in the Amazon region in Northern Brazil / Braga nasuta (Cymothoidae): um ectoparasito de alevinos do peixe gigante da Amazônia Arapaima gigas (Osteoglossidae), cultivado na região Amazônica, no Norte do Brasil

Isopods Cymothoidae are organisms that parasitize several fish species, both marine and freshwater, provoking important physiological alterations and secondary infections. The genus Braga was proposed to harbor three species of parasitic isopods in freshwater fish from South America: B. brasiliensis, B. cichlae and B. nasuta. Posteriorly, other four species were included: B. patagonica, B. amapaensis, B. fluviatilis and B. bachmanni. Regarding the geographical distribution of the genus, there are registers in Brazil, Argentina, Suriname and Paraguay. In this study, 3,625 fingerlings of pirarucu Arapaima gigas were examined from a commercial fish farm in the Amazon region, Pará State, to observe and identify possible parasites. A total of eleven ectoparasitic isopods were carefully removed from the body surface of the hosts and fixed in alcohol 70%. They were processed and identified as Braga nasuta. Parasitological indexes were prevalence of 0.303%, mean intensity of 1.000±0.000 and mean abundance of 0.003±0.055. This is the first report of B. nasuta in pirarucu fingerlings.

Isópodes Cymothoidae são organismos que parasitam ampla variedade de espécies de peixes, tanto marinhos quanto dulcícolas, provocando importantes alterações fisiológicas e infecções secundárias. O gênero Braga foi proposto inicialmente para abrigar três espécies de isópodes parasitos de peixes de água doce da América do Sul: B. brasiliensis, B. cichlae e B. nasuta; posteriormente, outras quatro espécies foram inseridas: B. patagônica, B. amapaensis, B. fluviatilis e B. bachmanni. Em relação à distribuição geográfica das espécies deste gênero, há registros no Brasil, na Argentina, em Suriname e no Paraguai. Neste estudo, 3.625 alevinos de pirarucu Arapaima gigas de uma piscicultura comercial na região amazônica, Estado do Pará, foram examinados para observar e identificar possíveis parasitos. Ao total, foram encontrados onze isópodes ectoparasitos, cuidadosamente retirados da superfície corporal de hospedeiros e fixados em álcool 70%. Eles foram processados e identificados como Braga nasuta. Os índices parasitológicos tiveram prevalência de 0,303%, intensidade media de 1.000±0.000 e abundância média de 0,003±0,055. Este é o primeiro relato de B. nasuta em alevinos de pirarucu.

Higher Vulnerability of Menadione-Exposed Cortical Astrocytes of Glutaryl-CoA Dehydrogenase Deficient Mice to Oxidative Stress, Mitochondrial Dysfunction, and Cell Death: Implications for the Neurodegeneration in Glutaric Aciduria Type I.

Patients affected by glutaric aciduria type I (GA-I) show progressive cortical leukoencephalopathy whose pathogenesis is poorly known. In the present work, we exposed cortical astrocytes of wild-type (Gcdh +/+ ) and glutaryl-CoA dehydrogenase knockout (Gcdh -/- ) mice to the oxidative stress inducer menadione and measured mitochondrial bioenergetics, redox homeostasis, and cell viability. Mitochondrial function (MTT and JC1-mitochondrial membrane potential assays), redox homeostasis (DCFH oxidation, nitrate and nitrite production, GSH concentrations and activities of the antioxidant enzymes SOD and GPx), and cell death (propidium iodide incorporation) were evaluated in primary cortical astrocyte cultures of Gcdh +/+ and Gcdh -/- mice unstimulated and stimulated by menadione. We also measured the pro-inflammatory response (TNFα levels, IL1-ß and NF-ƙB) in unstimulated astrocytes obtained from these mice. Gcdh -/- mice astrocytes were more vulnerable to menadione-induced oxidative stress (decreased GSH concentrations and altered activities of the antioxidant enzymes), mitochondrial dysfunction (decrease of MTT reduction and JC1 values), and cell death as compared with Gcdh +/+ astrocytes. A higher inflammatory response (TNFα, IL1-ß and NF-ƙB) was also observed in Gcdh -/- mice astrocytes. These data indicate a higher susceptibility of Gcdh -/- cortical astrocytes to oxidative stress and mitochondrial dysfunction, probably leading to cell death. It is presumed that these pathomechanisms may contribute to the cortical leukodystrophy observed in GA-I patients.

Ornithine and Homocitrulline Impair Mitochondrial Function, Decrease Antioxidant Defenses and Induce Cell Death in Menadione-Stressed Rat Cortical Astrocytes: Potential Mechanisms of Neurological Dysfunction in HHH Syndrome.

Hyperornithinemia-hyperammonemia-homocitrullinuria (HHH) syndrome is caused by deficiency of ornithine translocase leading to predominant tissue accumulation and high urinary excretion of ornithine (Orn), homocitrulline (Hcit) and ammonia. Although affected patients commonly present neurological dysfunction manifested by cognitive deficit, spastic paraplegia, pyramidal and extrapyramidal signs, stroke-like episodes, hypotonia and ataxia, its pathogenesis is still poorly known. Although astrocytes are necessary for neuronal protection. Therefore, in the present study we investigated the effects of Orn and Hcit on cell viability (propidium iodide incorporation), mitochondrial function (thiazolyl blue tetrazolium bromide-MTT-reduction and mitochondrial membrane potential-ΔΨm), antioxidant defenses (GSH) and pro-inflammatory response (NFkB, IL-1ß, IL-6 and TNF-α) in unstimulated and menadione-stressed cortical astrocytes that were previously shown to be susceptible to damage by neurotoxins. We first observed that Orn decreased MTT reduction, whereas both amino acids decreased GSH levels, without altering cell viability and the pro-inflammatory factors in unstimulated astrocytes. Furthermore, Orn and Hcit decreased cell viability and ΔΨm in menadione-treated astrocytes. The present data indicate that the major compounds accumulating in HHH syndrome impair mitochondrial function and reduce cell viability and the antioxidant defenses in cultured astrocytes especially when stressed by menadione. It is presumed that these mechanisms may be involved in the neuropathology of this disease.

Induction of a Proinflammatory Response in Cortical Astrocytes by the Major Metabolites Accumulating in HMG-CoA Lyase Deficiency: the Role of ERK Signaling Pathway in Cytokine Release.

3-Hydroxy-3-methylglutaric aciduria (HMGA) is an inherited metabolic disorder caused by 3-hydroxy-3-methylglutaryl-CoA lyase deficiency. It is biochemically characterized by predominant tissue accumulation and high urinary excretion of 3-hydroxy-3-methylglutarate (HMG) and 3-methylglutarate (MGA). Affected patients commonly present acute symptoms during metabolic decompensation, including vomiting, seizures, and lethargy/coma accompanied by metabolic acidosis and hypoketotic hypoglycemia. Although neurological manifestations are common, the pathogenesis of brain injury in this disease is poorly known. Astrocytes are important for neuronal protection and are susceptible to damage by neurotoxins. In the present study, we investigated the effects of HMG and MGA on important parameters of redox homeostasis and cytokine production in cortical cultured astrocytes. The role of the metabolites on astrocyte mitochondrial function (thiazolyl blue tetrazolium bromide (MTT) reduction) and viability (propidium iodide incorporation) was also studied. Both organic acids decreased astrocytic mitochondrial function and the concentrations of reduced glutathione without altering cell viability. In contrast, they increased reactive species formation (2'-7'-dichlorofluorescein diacetate (DCFHDA) oxidation), as well as IL-1ß, IL-6, and TNF α release through the ERK signaling pathway. Taken together, the data indicate that the principal compounds accumulating in HMGA induce a proinflammatory response in cultured astrocytes that may possibly be involved in the neuropathology of this disease.

Oxidative Stress, Disrupted Energy Metabolism, and Altered Signaling Pathways in Glutaryl-CoA Dehydrogenase Knockout Mice: Potential Implications of Quinolinic Acid Toxicity in the Neuropathology of Glutaric Acidemia Type I.

We investigated the effects of an acute intrastriatal QUIN administration on cellular redox and bioenergetics homeostasis, as well as on important signaling pathways in the striatum of wild-type (Gcdh +/+ , WT) and knockout mice for glutaryl-CoA dehydrogenase (Gcdh -/- ) fed a high lysine (Lys, 4.7 %) chow. QUIN increased lactate release in both Gcdh +/+ and Gcdh -/- mice and reduced the activities of complex IV and creatine kinase only in the striatum of Gcdh -/- mice. QUIN also induced lipid and protein oxidative damage and increased the generation of reactive nitrogen species, as well as the activities of the antioxidant enzymes glutathione peroxidase, superoxide dismutase 2, and glutathione-S-transferase in WT and Gcdh -/- animals. Furthermore, QUIN induced DCFH oxidation (reactive oxygen species production) and reduced GSH concentrations (antioxidant defenses) in Gcdh -/- . An early increase of Akt and phospho-Erk 1/2 in the cytosol and Nrf2 in the nucleus was also observed, as well as a decrease of cytosolic Keap1caused by QUIN, indicating activation of the Nrf2 pathway mediated by Akt and phospho-Erk 1/2, possibly as a compensatory protective mechanism against the ongoing QUIN-induced toxicity. Finally, QUIN increased NF-κB and diminished IκBα expression, evidencing a pro-inflammatory response. Our data show a disruption of energy and redox homeostasis associated to inflammation induced by QUIN in the striatum of Gcdh -/- mice submitted to a high Lys diet. Therefore, it is presumed that QUIN may possibly contribute to the pathophysiology of striatal degeneration in children with glutaric aciduria type I during inflammatory processes triggered by infections or vaccinations.

Experimental evidence that overexpression of NR2B glutamate receptor subunit is associated with brain vacuolation in adult glutaryl-CoA dehydrogenase deficient mice: A potential role for glutamatergic-induced excitotoxicity in GA I neuropathology.

Glutaric aciduria type I (GA I) is biochemically characterized by accumulation of glutaric and 3-hydroxyglutaric acids in body fluids and tissues, particularly in the brain. Affected patients show progressive cortical leukoencephalopathy and chronic degeneration of the basal ganglia whose pathogenesis is still unclear. In the present work we investigated parameters of bioenergetics and redox homeostasis in various cerebral structures (cerebral cortex, striatum and hippocampus) and heart of adult wild type (Gcdh(+/+)) and glutaryl-CoA dehydrogenase deficient knockout (Gcdh(-/-)) mice fed a baseline chow. Oxidative stress parameters were also measured after acute lysine overload. Finally, mRNA expression of NMDA subunits and GLT1 transporter was determined in cerebral cortex and striatum of these animals fed a baseline or high lysine (4.7%) chow. No significant alterations of bioenergetics or redox status were observed in these mice. In contrast, mRNA expression of the NR2B glutamate receptor subunit and of the GLT1 glutamate transporter was higher in cerebral cortex of Gcdh(-/-) mice. Furthermore, NR2B expression was markedly elevated in striatum of Gcdh(-/-) animals receiving chronic Lys overload. These data indicate higher susceptibility of Gcdh(-/-) mice to excitotoxic damage, implying that this pathomechanism may contribute to the cortical and striatum alterations observed in GA I patients.