Abstract The objective of this work was to evaluate microfluidic chip electrophoresis, known as lab-on-a-chip technique, for the detection of milk adulteration using cheese whey in comparison with SDS-PAGE. Raw, pasteurized, processed at an ultra-high temperature (UHT), and powdered milk samples received increasing concentrations of cheese whey (0, 1, 2.5, 5, 10, 20, 30, 50, and 100% v/v), and were subjected to lab-on-a-chip electrophoresis and SDS-PAGE to detect their mixtures. The lab-on-a-chip methodology was able to separate and quantify milk proteins. In addition, the tested technique is easy, rapid, sensitive, and can detect the addition of cheese whey in milk from the lowest level tested (1%) for milk proteins α-casein and β-casein.

Resumo O objetivo deste trabalho foi avaliar a eletroforese com dispositivo de microfluidos, conhecida como técnica lab-on-a-chip, para detecção de adulteração de leite com soro de queijo, em comparação ao SDS-PAGE. Amostras de leite cru, pasteurizado, processado em temperatura ultra-alta (UHT) e em pó receberam adição de soro de queijo em concentrações crescentes (0, 1, 2,5, 5, 10, 20, 30, 50 e 100% v/v) e foram submetidas a eletroforese lab-on-a-chip e SDS-PAGE para detectar suas misturas. A metodologia lab-on-a-chip foi capaz de separar e quantificar as proteínas do leite. Além disso, a técnica lab-on-a-chip é fácil, rápida, sensível e pode detectar adição de soro de queijo no leite do nível mais baixo testado (1%) para as proteínas do leite α-caseína e β-caseína.

Abstract Nutritionally, cheese and other dairy products have been highlighted especially as a calcium source. Access to minerals is dependent on variables related to diet, lifestyle, and also the health status. This study aimed to evaluate the bioaccessibility and calcium partition in Minas Padrão cheese over its ripening period for experimentally obtained samples. The cheese of the study has been manufactured using two different technologies (A and B). Also to evaluate the evolution of pH over cheese ripening and the influence of calcium partitions in its bioaccessibility along the cheese ripening period. The mean values found for total calcium content were (549 mg⋅100 g-1 – A and 644 mg⋅100 g-1 – B). For calcium in the aqueous phase on average 460 mg⋅100 g-1 – A and 404 mg⋅100 g-1 – B. These and other changes found are possibly due to physicochemical and biochemical changes, especially with respect to pH variations. Calcium bioaccessibility decreased in the beginning and then incresed in the end of the ripening period. The results obtained in this work hopefully may allow further studies and the development of products in which calcium bioaccessibility might be indicated on the product`s label accordingly to its ripening time.

Abstract Real-time PCR (qPCR) has been used for rapid identification of Salmonella Typhimurium and Staphylococcus aureus in dairy foods, but is unable to differentiate viable and unviable pathogens. Ethidium bromide monoazide (EMA), a DNA-intercalating agent, can detect only viable cells because selectively enter cells considered unviable and bind to their DNA, inhibiting its amplification during qPCR. The objective was to establish a protocol for detection of viable Salmonella Typhimurium and S. aureus, experimentally inoculated in coalho cheese, by the use of EMA combined with qPCR. The protocol was effective for the identification of viable Salmonella Typhimurium in coalho cheese but not for the S. aureus cells. Concentrations of viable Salmonella Typhimurium cells of 1 CFU/10 g of coalho cheese could be detected. The monoplex protocol enables the rapid and specific identification of viable Salmonella Typhimurium in coalho cheese, making it an alternative method for the quality and safety control of cheeses.

Resumo: O objetivo deste trabalho foi caracterizar bacteriófagos autóctones e determinar sua atividade lítica em Staphylococcus aureus. Seis fagos foram isolados de água de lavagem de pisos de estábulos por meio do enriquecimento de cultura com três estirpes de S. aureus. Todos os fagos foram caracterizados pela digestão do DNA com enzimas de restrição e pelo sequenciamento do fragmento de DNA que codifica a endolisina. Cada fago foi testado contra 100 estirpes de S. aureus isoladas de casos de mastite bovina e de produtos lácteos pelo método de lise em placa. Sequências do gene de endolisina apresentaram alta conservação, com mais de 99% de similaridade a nível do nucleotídeo entre os fagos isolados. Foram identificados três domínios envolvidos no reconhecimento e na lise da parede celular bacteriana. Dois bacteriófagos isolados de estábulos apresentam alta atividade lítica em S. aureus, em ampla gama de estirpes, o que indica seu potencial para estudos de fagoterapia em gado leiteiro ou como agente de controle biológico para produtos lácteos.

Abstract: The objective of this work was to characterize autochthonous bacteriophages and to determine their lytic activity on Staphylococcus aureus. Six phages were isolated from dairy barn flush water through enrichment cultures with three S. aureus strains. All phages were characterized by DNA digestion by restriction enzymes and sequencing of the DNA fragment encoding endolysin. Each phage was tested against 100 S. aureus strains isolated from bovine mastitis and from dairy products using the lysis-plate method. The sequences of the endolysin gene were highly conserved, with nucleotide similarity higher than 99% among the isolated phages. Three domains involved in the recognition and lysis of the bacterial cell wall were identified. Two bacteriophages isolated from the dairy barns present high lytic activity on S. aureus, on a wide range of host strains, indicating their potential for studies on phage therapy in dairy cattle or as a biological control agent for dairy products.

RESUMO Em rebanhos leiteiros, um dos objetivos mais importantes da seleção é a melhoria da produção e composição do leite. Vários estudos demonstraram que os genes candidatos da kappa-caseína (CSN3) e da β-lactoglobulina (β-LG) estão associados a produção de leite, qualidade do leite e características de saúde em animais leiteiros. Portanto, o objetivo deste estudo foi detectar polimorfismos nos genes CSN3 e β-LG e avaliar possíveis associaçoes desses polimorfismos com a produção de leite em até 305 dias (305MY) e a capacidade de transmissão prevista (PTA) de leite em bovinos da raça Girolando. No total, 138 touros e 729 vacas (n = 867) foram amostrados. A genotipagem foi realizada pelo método PCR-RFLP utilizando as enzimas HinfI e HaeIII para os genes CSN3 e β-LG, respectivamente. Os resultados estatísticos revelaram dois alelos A e B para ambos os genes. Os genotipos e alelos mais frequentes para os genes CSN3 e β-LG foram respectivamente: AA (0,7324) e A (0,8558) e AB (0,4827) e A (0,5017). O teste x2 revelou que os dois loci estavam em equilibrio de Hardy-Weinberg (p <0,001). Os efeitos de substituiçao alélica para as variantes nao foram significativos para as características 305 MY e PTA para 305MY (p> 0,05). Portanto, as variantes alélicas identificadas nos genes β-LG e CSN3 devem ser mais investigadas antes de serem incluidas nos programas de melhoramento desenhados para bovinos leiteiros Girolando objetivando melhorar as características do leite analisadas no presente estudo.

SUMMARY In dairy farm animals, one the most important goal of the selection is the improvement of milk yield and composition. Several studies have demonstrated that the candidate genes of the kappa-casein (CSN3) and β - lactoglobulin (β-LG) are associated with milk yield, milk quality and health traits in dairy animals. Therefore the aim of this study was to detect polymorphisms in CSN3 and β-LG genes and its association with milk yield in up to 305 days (305MY) and predicted transmission capacity (PTA) for 305MY in Girolando cattle. Totally, 138 bulls and 729 cows (n=867) were sampled. The genotypes of both genes were obtained by the PCR-RFLP method using HinfI and HaeIII enzymes for CSN3 and β-LG genes, respectively. Statistical results revealed two alleles A and B for both genes. The genotypes and alleles more frequents for CSN3 and β-LG genes were respectively: AA (0.7324) and A (0.8558), and AB (0.4827) and A (0.5017). The x2 test revealed that the two loci were at Hardy-Weinberg equilibrium (p<0.001). The allele substitution effects for the variants were not significant on 305MY and PTA for 305MY (p>0.05). The allele variants of β-LG and CSN3 might be more investigated before include them into future breeding schemes designed for Girolando dairy cattle with objective of improving milk traits as milk yield in up to 305 days (305MY) and predicted transmission capacity (PTA) for 305MY..

ABSTRACT Due to their ecological and economic importance, honey bees have attracted much scientific attention, which has intensified due to the recent population decline of these insects in the several parts of the world. Among the factors related to these patterns, infection by pathogens are the most relevant, mainly because of the easy dissemination of these microorganisms. Although no zoonotic diseases are associated with these insects, the presence of infectious agents in bee products should still be considered because they play a role as disease dispersers, increasing the risk to animal health. Because of the possibility of dispersion of pathogens via bee products, this work aimed to identify the presence of spores of the pathogens Paenibacillus larvae, Ascosphaera apis and Nosema spp. in samples of honey, pollen and royal jelly that are registered with Brazil's Federal Inspection Service (S.I.F.) and commercially available in the state of São Paulo. Of the 41 samples of bee products analyzed, only one showed no contamination by any of these pathogens. N. ceranae and P. larvae had the highest prevalence considering all the samples analyzed (present in 87.80% and 85.37% of the total, respectively), with N. apis present in 26.83% and A. apis present in 73.17% of the samples. These results provide support for the formulation of government regulations for sanitary control of exotic diseases by preventing dispersion of pathogens, including through illegal importation, since local and international trade and the transfer of colonies between regions play important roles in the dispersion of these microorganisms.

RESUMO A geléia real pode conter grãos de pólen e sua presença pode ser usada para determinar a origem fitogeográfica do produto. Este estudo analisou a origem fitogeográfica de amostras comerciais de geléia real do Estado de São Paulo, com registro no Sistema de Inspeção Federal (SIF) brasileiro, que estavam contaminadas com os esporos da bactéria Paenibacillus larvae, causadora da doença Cria Pútrida Americana. Os grãos de pólen de Castanea apresentaram a maior porcentagem total, com menores percentuais de Cirsium/Carduus, Cistus, Parthenocissus, Prunus, Quercus, Robinia, Scrophulariaceae, Taraxacum, Tilia, dentre outros. Este espectro polínico é incompatível com amostras de geléia real produzidas no Brasil. O espectro polínico assemelhou- se ao de um produto importado, compatível com a origem do Hemisfério Norte. A legislação brasileira não exige que a origem fitogeográfica dos produtos apícolas importados seja analisada por procedimentos palinológicos, mas é obrigatório ter um certificado atestando a ausência de patógenos emitido pelo país de origem, monitorado com o objetivo de impedir que doenças exóticas penetrem no Brasil. A palinologia, portanto, provou ser fundamental na detecção de importações desses lotes contaminados.

ABSTRACT Royal jelly may contain pollen grains and their presence can be used to determine the phytogeographical origin of the product. This study analyzed the phytogeographical origin of commercial royal jelly samples from São Paulo State, tested as part of the Brazilian Federal Inspection System (SIF), found to be contaminated with spores of the bacterium Paenibacillus larvae, that causes the American Foulbrood Disease. The pollen grains of Castanea had the highest total percentage, with lower percentages of Cirsium/Carduus, Cistus, Parthenocissus, Prunus, Quercus, Robinia, Scrophulariaceae, Taraxacum, Tilia, among others. This pollen spectrum is incompatible with royal jelly samples produced in Brazil. The pollen spectrum resembled that of an imported product, compatible with the Northern Hemisphere origin. Brazilian legislation does not require the phytogeographic origin of imported bee products to be analyzed by palynological procedures, but it is mandatory to have a certificate issued by the country of origin attesting the absence of pathogens, monitored with the objective of preventing the exotic diseases from entering Brazil. Palynology, therefore, proved to be fundamental in detecting imports of this contaminated batches.

O objetivo neste estudo foi avaliar a estrutura genética da população de bovinos da raça Girolando no Brasil. Analisou-se o arquivo de pedigree de 26.969 animais, composto de 3.031 machos e 23.938 fêmeas. O nível de conteúdo de informação do pedigree na geração atual foi 61%, mostrando ser de qualidade moderada. O coeficiente de endogamia médio e o coeficiente de relação médio da população Girolando foram 0,11 e 0,13%, respectivamente. O tamanho efetivo da população, considerando a geração completa traçada, foi 188, acima do nível crítico. Do total de 9.457 ancestrais que contribuíram para a população de referência, 457 explicaram 50% da variabilidade genética da população. O número efetivo de fundadores foi 551 e o de ancestrais 393. O intervalo médio de geração foi de 5,26 anos, sendo ligeiramente maior nas trilhas gaméticas mãe-filho e pai-filha. A partir dos coeficientes estimados, pode-se concluir que a endogamia nos rebanhos da raça Girolando foi de pequena magnitude e que as práticas de acasalamento foram adequadas durante o período avaliado. No entanto, é importante continuar com o monitoramento desses coeficientes a fim de prevenir perda de variabilidade genética

The aim of this study was to evaluate the population structure of Girolando cattle in Brazil. The pedigree file contained 26,969 individuals, from which 3,031 were males and 23,938 were females. The average level of completeness of the pedigree in the current generation was of reasonable quality (61%). Inbreeding and average relatedness coefficients were low: 0.11 and 0.13%, respectively. Estimates of effective population size considering the full generations traced was 188, which is above the critical level range. The number of ancestors that contributed to the reference population was 9,457 animals, from which 457 explained 50% of the genetic variability of the population. The effective number of founders and the effective number of ancestors in this population were, respectively, 551 and 393. The average generation interval was 5.26 years, slightly higher in genetic pathways dam-son and sire-daughter. The inbreeding in the Girolando breed was of small magnitude, indicating that the current practices of mating were adequate during the study period. However, it is important to continue monitoring these coefficients in order to prevent loss of genetic variability