Padronização da técnica da reação em cadeia da polimerase (PCR) para detecção de micro-organismos patogênicos em amostras de leite bovino in natura oriundas de produtores do Vale do Taquari, RS.

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Padronização da técnica da reação em cadeia da polimerase (PCR) para detecção de micro-organismos patogênicos em amostras de leite bovino in natura oriundas de produtores do Vale do Taquari, RS.

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DETECÇÃO DE STAPHYLOCOCCUS AUREUS ATRAVÉS DA TÉCNICA DE REAÇÃO EM CADEIA DA POLIMERASE (PCR), EM AMOSTRAS DE LEITE BOVINO IN NATURA OBTIDAS DE PRODUTORES NO SUL DO BRASIL

O leite é o alimento mais consumido mundialmente e de elevado valor nutricional. Considerando a comercialização do leite cru e o alto risco de contaminação, o presente estudo investigou a presença de Staphylococcus aureus em amostras de leite bovino in natura, de produtores do estado do Rio Grande do Sul (RS), Brasil. Utilizou- -se a Técnica de Reação em Cadeia da Polimerase (PCR), e compararam-se os dados obtidos com os resultados dos métodos microbiológicos, segundo a Instrução Normativa 62 do Ministério da Agricultura e Abastecimento. Três protocolos distintos de extração de DNA diretamente do leite foram testados. A sensibilidade do protocolo C foi de 104 UFC/mL sendo o escolhido para a extração de DNA das amostras de leite in natura de oito produtores. Os resultados da PCR foram comparados com culturas bacterianas e testes bioquímicos. O fragmento de 132 pb, correspondente a S. aureus, foi identificado em cinco amostras pela técnica de PCR e seis amostras foram positivas através de testes microbiológicos e bioquímicos. Os resultados obtidos pela pesquisa indicam que o protocolo utilizado pode ser eficiente para a extração de DNA diretamente do leite, bem como para a detecção S. aureus em amostras de leite in natura

Oficinas de Biotecnologia para o Ensino Médio: Antioxidantes, a Fonte da juventude?

Theory and practice about biotechnology-related issues have been topics of workshops for students and teachers of the Post Graduate in Biotechnology UNIVATES to secondary education students. We held ten different workshops aimed at the food production and health, among which stands out in this study, the workshop entitled "Antioxidants: the fountain of youth?", which evaluated the antioxidant potential of natural juice orange select and pearl pineapple. Previously, a protocol was structured on the practice, which aimed to evaluate the juice antioxidant potential of the fruit orange select and pearl pineapple, which was compared with the antioxidant reference compound, ascorbic acid, with adaptations according to spectrophotometric method in vitro DPPH. The workshops about antioxidants have 180 minutes duration and have been taught by three monitors responsible for assisting students throughout the activity. The workshops involved 388 students from seven schools in six municipalities of Taquari-RS Valley, and the workshop about antioxidants had 62 participants. This activity aimed to promote interaction and diffusion of knowledge among students of higher education and secondary education.Teoria e prática sobre assuntos relacionados à Biotecnologia foram temas de oficinas realizadas por discentes e docentes do Programa de Pós-Graduação em Biotecnologia da UNIVATES a estudantes de Ensino Médio. Foram ministradas dez diferentes oficinas voltadas à produção de alimentos e à saúde, entre as quais destaca-se, neste estudo, a oficina intitulada “Antioxidantes: a fonte da juventude?”, que avaliou o potencial antioxidante do suco natural da laranja seleta e do abacaxi pérola. Previamente, foi estruturado um protocolo sobre a prática, a qual objetivou avaliar o potencial antioxidante do suco das frutas laranja seleta e abacaxi pérola, o qual foi comparado com o composto antioxidante de referência, ácido ascórbico, com adaptações segundo o método espectrofotométrico in vitro DPPH. As oficinas sobre antioxidantes tiveram duração de 180 minutos e foram ministradas por três monitores responsáveis por auxiliar os alunos durante toda a atividade. As oficinas envolveram 388 alunos de sete escolas de seis municípios do Vale do Taquari-RS, sendo que a oficina sobre antioxidantes contou com 62 participantes. Tal atividade buscou promover a interação e difusão de conhecimento entre alunos de Ensino Superior e de Ensino Médio

Polimorfismos no SNP CGIL4: estudo de associação ao fenótipo de resistência a mastite clinica em vacas holandesas

Bovine mastitis is the most important udder pathology and have a significant economic impact in the dairy industry. Its etiology is associated to problems with health and milking management. However, there are animals with resistance or susceptibility to mastitis, even when environmental factors are controlled. Recently, some molecular markers were associated to the phenotype of resistance to mastitis. The present study aims to verify if, based on clinical history of cows of second and third lactation, the SNP CGIL4 is associated to the phenotype of resistance to mastitis in Holstein cows in a herd in southern Brazil. Genomic DNA was obtained from blood samples of 160 Holstein cows (second and third lactation) from dairy herds from Rio Grande do Sul state, Brazil. Phenotype of resistance or susceptibility to clinical mastitis was defined based on animal’s clinical history, and the animal was classified as susceptible if presented at least 3 episodes of clinical mastitis during the last lactation. The identification of the CGIL4 SNP was performed using a PCR-RFLP. The association between genotipes and phenotipes was acessed by the χ 2 test. To detect polymorphism, Polymerase Chain Reaction (PCR) was performed using a touch-down PCR reaction producing an amplicon of 399 bp. The Restriction Fragment Length Polymorfism (RFLP) analysis was performed using TaqI restriction enzyme to identify the SNP (G→A). The association between genotipes and phenotipes was acessed by the χ 2 test. The most frequent genotype observed was GG (47.5%), AG with 45% and AA with 7.5%. Other results point AG as the most frequent genotype. Allele frequencies were 69.8% and 30.2% for the alleles G and A, respectively. No association was found between genotypes and phenotype of resistance and susceptibility to clinical mastitis, based in clinical records.A mastite bovina é a principal patologia da glândula mamária e a maior causadora de prejuízos na produção leiteira. Sua etiologia é quase sempre relacionada a problemas de manejo sanitário e de ordenha. Entretanto, observa-se a existência de animais com maior ou menor resistência a mastite, mesmo quando fatores ambientais são controlados. Recentemente, alguns marcadores moleculares foram associados ao fenótipo de resistência a mastite. O presente estudo teve como objetivo verificar se com base no histórico de mastites clinicas em vacas de 2a e 3a lactação, o SNP CGIL4 está relacionado ao fenótipo de resistência a mastite em vacas holandesas em um rebanho no Sul do Brasil. Para obtenção do DNA genômico, foram utilizadas amostras de sangue de 160 vacas de segunda e terceira lactação de rebanhos tecnificados no Vale do Taquari, RS. O fenótipo de resistência ou susceptibilidade à mastite clínica foi determinado com base no histórico clínico dos animais, sendo classificada como susceptível a vaca com mais de 3 episódios da doença durante a lactação anterior. A identificação do SNP CGIL4 foi realizada através da técnica de PCR-RFLP. Foi utilizada uma PCR touch down para obter o amplicon de 399 pb. A analise de RFLP foi feita utilizando a enzima TaqI para identificar o SNP (G→A). A relação com o fenótipo e genótipo foi analisada pelo teste Qui-quadrado. O genótipo mais frequente observado foi GG (47,5%), seguido por AG com 45% e AA com 7,5%. O presente resultado difere de outros estudos que apontaram AG como o genótipo mais encontrado. As frequências alélicas foram 69,8% para o alelo G e 30,2% para o alelo A. Não foi observada associação entre os genótipos testados e fenótipos de resistência e susceptibilidade à mastite clínica baseada em registros clínicos

Molecular screening of bovine raw milk for the presence of Shiga toxin-producing Escherichia coli (STEC) on dairy farms

Milkborne transmission of Shiga toxin- producing Escherichia coli (STEC) has raised considerable concern due to recent outbreaks worldwide and poses a threat to public health. The aim of this study was to develop a sensitive and specific multiplex PCR assay to detect the presence of STEC in bovine raw milk. To identify E. coli (ATCC 25922) contamination, the gene uspA was used, and PCR sensitivity and specificity were accessed by testing diluted samples ranging from 2 to 2.0 × 10(6) CFU/mL. To detect STEC, the stx1 and stx2 genes were selected as targets. After reaction standardization, the multiplex assay was tested in raw milk collected from 101 cows on dairy farms. PCR assay for E. coli detection had a specificity of 100% and sensitivity of 79% (P<0.0001), with a lower detection limit of 2 CFU/mL. Multiplex PCR assay had 100% sensitivity for E. coli positive raw milk samples, and 31.1% were contaminated with STEC, 28.3% of stx2, and 1.9% of stx1. The multiplex PCR assay described in the present study can be employed to identify and screen E. coli harboring stx1 and stx2 genes in raw milk on dairy farms and in industries

Neuromodulatory effects of Calyptranthes grandifolia extracts against 6-hydroxydopamine-induced neurotoxicity in SH-SY5Y cells

Alzheimer’s and Parkinson's diseases are neurodegenerative disorders characterized by progressive neuronal dysfunction. Previous studies revealed that some natural products have neuroprotective properties, including species of the Myrtaceae family. However, the neuromodulatory potential of Calyptranthes grandifolia is not clear. In the present study, we examined the ability of the ethanol and hexane leaf extracts of C. grandifolia to prevent 6-hydroxydopamine (6-OHDA)-induced neurotoxicity in vitro. Initially, we investigated the potential of the extracts to inhibit the neurodegenerative-related enzymes c-Jun N-terminal kinase 3 (JNK3) and acetylcholinesterase (AChE). In addition, SH-SY5Y cell viability was assessed by MTT assay after 100 mM 6-OHDA-induced cell damage. In order to verify the possible effects of both extracts on 6-OHDA-induced cell death, hydrogen peroxide generation, mitochondrial potential and caspases-3 activity were assessed. Our findings revealed that ethanol extract exhibited inhibitory activity against JNK3 and AChE. In addition, when co-treating SH-SY5Y cells with 6-OHDA and the extracts, oxidative stress was inhibited by both extracts through a decrease of mitochondrial depolarization and caspases-3 activity. In summary, ethanol and hexane extracts of C. grandifolia have some suppressive property against neurotoxicity induced by 6-OHDA.info:eu-repo/semantics/publishedVersio

Adjunctive role of Calyptranthes tricona extract with probiotic Kluyveromyces marxianus on colorectal adenocarcinoma Caco-2 cells

The composition of microbiota may influence the development of colorectal cancer (CRC). In addition, probiotics can decrease the chance of developing cancer or its progress. For that reason, it is encouraging to assess the influence of plant extracts as adjuvants of the health-promoting effects of probiotics. Thus, this study aimed to investigate the in vitro beneficial properties of Calyptranthes tricona leaf ethanol extract in association or not with the lactic yeast Kluyveromyces marxianus on colon adenocarcinoma Caco-2 cells. C. tricona extract inhibited the in vitro p38α MAPK activity and exhibited immunomodulation on isolated human lymphocytes. Further, the extract did not induce cytotoxicity towards K. marxianus or Caco-2 cells; leading to an increased yeast adhesion to cells in a dose-dependent manner. In conclusion, this preliminary study demonstrates that C. tricona extract has effects on enzyme inhibition and immune function. In addition, when associated with K. marxianus, C. tricona may possess beneficial properties for application as enhancer of probiotic’s protective role on CRC cells. However, further studies are necessary in order to elucidate the mechanisms involved.info:eu-repo/semantics/publishedVersio

Lymphocyte genotoxicity and protective effect of Calyptranthes tricona (Myrtaceae) against H 2 O 2-induced cell death in MCF-7 cells

Calyptranthes tricona is a species (Myrtaceae) native to South Brazil. Plants belonging to this family are folkloric used for analgesia, inflammation, and infectious diseases. However, little is known about the toxic potential of C. tricona. The present study aimed to evaluate the antioxidant activity of C. tricona ethanol and hexane leaf extracts, as well as verify their effect on human lymphocytes and MCF-7 cells. The extracts were subjected to preliminary phytochemical screening, antioxidant activity using DPPH and ORAC methods. Genotoxic and mutagenic effects in cultured human lymphocytes were assessed using the comet assay and the micronucleus assay, respectively. In addition, cell viability by MTT assay and fluorometric analysis of mitochondrial potential and caspases-9 activity were performed in order to verify the possible effects of both extracts on H2O2-induced cell death of MCF-7 cells. Our findings revealed that the phenol content and the antioxidant activity were only present in the ethanol extract. Also, the phytochemical screening presented steroids, triterpenoids, condensed tannins, and flavones as the main compounds. However, both extracts were capable of inducing concentration-dependent DNA damage in human lymphocytes. When treating MCF-7 cells with the extracts, both of them inhibited MCF-7 cell death in response to oxidative stress through a decrease of mitochondrial depolarization and caspases-9 activity. Thus, our results need to be considered in future in vitro and in vivo studies of C. tricona effects. In the meanwhile, we recommend caution in the acute/chronic use of this homemade preparation for medicinal purpose.info:eu-repo/semantics/publishedVersio