The biosynthesis of amylase and collagenase, produced by A. acidocaldarius and A. sendaiensis respectively, were studied, and different matrices evaluated for the microorganisms immobilization and enzymes production optimization, such as loofa sponge, alginate-sponge and alginate, followed by concentration through ultrafiltration. Using a wheat bran substrate, the amylase enzyme displayed enzymatic activity of 0.45 U mL-1 and optimum temperature and pH conditions of 75 °C and pH 3.0, respectively. Thermal stability was in the range of 55 to 60 °C. The apparent Km and Vmax were 3.2 mg mL-1 and 0.5 U mL-1, respectively. The production of collagenase by A. sendaiensis was carried out with potato dextrose broth substrate and the activity obtained was 7.2 U mL-1. For amylase, the best results were obtained from immobilization in loofa sponge and the use of ultrafiltration (0.67 U mL-1) and for the collagenase extract, from the free biomass and ultrafiltration (13.6 U mL-1). The use of an ultrafiltration system enabled an average increase of 54% in the activity of both enzymes. Therefore, Alicyclobacillus are capable of producing enzymes of industrial interest, with the possibility of economically viable application of the substrate, and the use of immobilization and ultrafiltration produced positive results.
The objective of this study was to determinate the seroprevalence of caseous lymphadenitis (CL) in commercial sheep herds in the Federal District, Brazil. One thousand and twenty-eight serum samples were collected, between March and June 2004, from all the properties (32) with at least 20 adult females in the flock. The seroprevalence of caseous lymphadenitis was determined by ELISA with secreted proteins of Corynebacterium pseudotuberculosis. The real prevalence value for the animals was 44.0% (CI 95: 41.0; 47.0), and 50.0% of the 32 herds had at least one animal seropositive for CL. Therefore, these data show that CL is fairly widespread in commercial flocks of sheep in the Federal District.
O objetivo deste trabalho foi determinar a prevalência de anticorpos contra linfadenite caseosa (LC) em rebanhos ovinos comerciais do Distrito Federal (DF). Foram coletadas 1.028 amostras de soro entre março e junho de 2004, de todas as propriedades (32) do Distrito Federal com pelo menos 20 fêmeas adultas no rebanho. A soroprevalência da linfadenite caseosa foi determinada por ELISA com proteínas secretadas de Corynebacterium pseudotuberculosis. Cinquenta por cento das 32 propriedades apresentaram pelo menos um animal soropositivo para o LC e a prevalência real para animais foi de 44,0% (IC 95: 41,0; 47,0), portanto, esses dados sugerem que a LC está presente em rebanhos ovinos comerciais no Distrito Federal.
Lactococcus lactis, the model lactic acid bacterium, is a good candidate for heterologous protein production in both foodstuffs and the digestive tract. We attempted to produce Streptomyces tendae antifungal protein 1 (Afp1) in L. lactis with the objective of constructing a strain able to limit fungal growth. Since Afp1 activity requires disulfide bond (DSB) formation and since intracellular redox conditions are reportedly unfavorable for DSB formation in prokaryotes, Afp1 was produced as a secreted form. An inducible expression-secretion system was used to drive Afp1 secretion by L. lactis; Afp1 was fused or not with LEISSTCDA, a synthetic propeptide (LEISS) that has been described to be a secretion enhancer. Production of Afp1 alone was not achieved, but production of LEISS-Afp1 was confirmed by Western blot and immunodetection with anti-Afp1 antibodies. This protein (molecular mass: 9.8 kDa) is the smallest non-bacteriocin heterologous protein ever reported to be secreted in L. lactis via the Sec-dependent pathway. However, no anti-fungal activity was detected, even in concentrated samples of induced supernatant. This could be due to a too low secretion yield of Afp1 in L. lactis, to the absence of DSB formation, or to an improper DSB formation involving the additional cysteine residue included in LEISS propeptide. This raises questions about size limits, conformation problems, and protein secretion yields in L. lactis.
The aim of this paper is to report the morphology and distribution of microspines in diplopods pylorus, as these are important structures present along the alimentary tract of arthropods. The morphology of the internal surface of the pylorus of Pseudonannolene tricolor Brolemann, 1901 and Rhinocricus padbergi Verhoeff, 1938 was analyzed by SEM. Pseudonannolene tricolor presents two morphologically distinct pyloric regions: anterior and posterior. The first region is characterized by the presence of thin microspines that increase in number and size towards the posterior portion; the second region presents smaller and triangular-shaped microspines distributed throughout small plates. The pylorus of R. padbergi does not present differentiated regions; the anterior portion is characterized by microspines grouped in plates that decrease in number and increase in size towards the ileum.
Este trabalho relata a morfologia e distribuição de microespinhos no piloro de diplópodos, uma vez que estas são estruturas importantes presentes ao longo do trato digestório dos artrópodos. A morfologia da superfície interna do piloro de Pseudonannolene tricolor Brolemann, 1901 e Rhinocricus padbergi Verhoeff, 1938 foi analisada por meio de MEV. Pseudonannolene tricolor apresenta duas regiões pilóricas morfologicamente distintas: anterior e posterior. A primeira região caracteriza-se pela presença de finos microespinhos que aumentam em número e tamanho em direção a porção posterior; a segunda região apresenta microespinhos pequenos, de formato triangular, distribuído em pequenas placas. O piloro de R. padbergi não apresenta regiões diferenciadas; a porção anterior caracteriza-se por microespinhos agrupados em placas, os quais decrescem em número e aumentam em tamanho, em direção ao íleo.
DNA-based immunization has initiated a new era of vaccine research. One of the main goals of gene vaccine development is the control of the levels of expression in vivo for efficient immunization. Modifying the vector to modulate expression or immunogenicity is of critical importance for the improvement of DNA vaccines. The most frequently used vectors for genetic immunization are plasmids. In this article, we review some of the main elements relevant to their design such as strong promoter/enhancer region, introns, genes encoding antigens of interest from the pathogen (how to choose and modify them), polyadenylation termination sequence, origin of replication for plasmid production in Escherichia coli, antibiotic resistance gene as selectable marker, convenient cloning sites, and the presence of immunostimulatory sequences (ISS) that can be added to the plasmid to enhance adjuvanticity and to activate the immune system. In this review, the specific modifications that can increase overall expression as well as the potential of DNA-based vaccination are also discussed.