Biofertilization with Azospirillum brasilense improves in vitro culture of Handroanthus ochraceus, a forestry, ornamental and medicinal plant.

Biofertilization with plant growth-promoting rhizobacteria is a potential alternative to plant productivity. Here, in vitro propagation of Handroanthus ochraceus (yellow lapacho), a forest crop with high economic and environmental value, was developed using the Azospirillum brasilense strains Cd and Az39 during rhizogenesis. Epicotiles of in vitro plantlets were multiplied in Woody Plant Medium (WPM). For rooting, elongated shoots were transferred to auxin-free Murashige-Skoog medium with Gamborg's vitamins and WPM, both at half salt concentration (½MSG and ½WPM), and inoculated with Cd or Az39 at the base of each shoot. Anatomical studies were performed using leaves cleared and stained with safranin for optical microscopy and leaves and roots metalized with gold-palladium for scanning electron microscopy (SEM). In ½WPM auxin-free medium, A. brasilense Cd inoculation produced 55% of rooting, increased root fresh and dry weight (45% and 77%, respectively), and led to lower stomata size and density with similar proportion of open and closed stomata. Both strains selectively increased the size or density of glandular trichomes in ½MSG. Moreover, bacteria were detected on the root surface by SEM. In conclusion, the difference in H. ochraceus response to A. brasilense inoculation depends on the strain and the plant culture media. Cd strain enhanced rooting in auxin-free ½WPM and produced plantlets with features similar to those expected in ex vitro plants. This work presents an innovative in vitro approach using beneficial plant-microorganism interaction as an ecologically compatible strategy in plant biotechnology.

Use of artichoke (Cynara scolymus) flower extract as a substitute for bovine rennet in the manufacture of Gouda-type cheese: characterization of aspartic proteases.

Artichoke (Cynara scolymus L.) flower extract was assayed with the aim of replacing animal rennet in the manufacture of Gouda-type cheeses from bovine milk. Floral extract coagulated milk within a suitable time for use on an industrial scale, while the yield of cheese obtained was equal to that achieved with bovine abomasum. Five proteolytic fractions with milk-clotting activity were isolated in a two-step purification protocol, three belonging to the cardosin group. Cheeses made with C. scolymus proteases must be brined for a longer period (40 h) to prevent overproteolysis and avoid the development of a background flavor. The type of coagulant (bovine or vegetable) had no significant effect on the cheeses' chemical parameters analyzed throughout ripening, and no significant organoleptic differences were detected between those manufactured with C. scolymus or animal rennet. The results indicate that C. scolymus flower extract is suitable for replacing animal rennet in the production of Gouda-type cheeses.

In vitro propagation of jojoba.

Jojoba (Simmondsia chinensis (Link) Schn.) is a nontraditional crop in arid and semi-arid areas. Vegetative propagation can be achieved by layering, grafting, or rooting semi-hardwood cuttings, but the highest number of possible propagules is limited by the size of the plants and time of the year. Micropropagation is highly recommended strategy for obtaining jojoba elite clones. For culture initiation, single-node explants are cultivated on Murashige and Skoog medium (MS) supplemented with Gamborg's vitamins (B5), 11.1 µM BA (N(6)-benzyl-adenine), 0.5 µM IBA (indole-3-butyric acid), and 1.4 µM GA(3) (gibberellic acid). Internodal and apical cuttings proliferate on MS medium containing B5 vitamins and 4.4 µM BA. Rooting is achieved on MS medium (half strength mineral salt) amended with B5 vitamins and 14.7 µM IBA during 7 days and transferred to develop in auxin-free rooting medium. Plantlets are acclimatized using a graduated humidity regime on soil: peat: perlite (5:1:1) substrate. This micropagation protocol produces large numbers of uniform plants from selected genotypes of jojoba.

In vitro propagation of fraser photinia using Azospirillum-mediated root development.

Fraser photinia (Photinia × fraseri Dress.) is a woody plant of high ornamental value. The traditional propagation system for photinia is by rooting apical cuttings using highly concentrated auxin treatments. However, photinia micropropagation is an effective alternative to traditional in vivo propagation which is affected by the seasonal supply of cuttings, the long time required to obtain new plants, and the difficulties in rooting some clones.A protocol for in vitro propagation of fraser photinia using the plant growth-promoting ability of some rhizobacteria is described here. Bacterial inoculation is a new tool in micropropagation protocols that improves plant development in in vitro culture. Shoots culture on a medium containing MS macro- and microelements, Gamborg's vitamins (BM), N (6)-benzyladenine (BA, 11.1 µM), and gibberellic acid (1.3 µM) produce well-established explants. Proliferation on BM medium supplemented with 4.4 µM BA results in four times the number of shoots per initial shoot that develops monthly. Consequently, there is a continuous supply of plant material since shoot production is independent of season. Azospirillum brasilense inoculation, after 49.2 µM indole-3-butyric acid pulse treatment, stimulates early rooting of photinia shoots and produces significant increase in root fresh and dry weights, root surface area, and shoot fresh and dry weights in comparison with controls. Furthermore, inoculated in vitro photinia plants show anatomical and morphological changes that might lead to better adaptation in ex vitro conditions after transplanting, compared with the control plants.

Micropropagation of photinia employing rhizobacteria to promote root development.

An alternative protocol was developed for in vitro propagation of photinia (Photinia x fraseri Dress), an ornamental shrub, using the plant growth-promoting rhizobacteria (PGPR) Azospirillum brasilense and Azotobacter chroococcum during rhizogenesis. Shoot tips from four-year-old mature plants, cut in spring and summer, were used as initial explants. They were cultured on Murashige-Skoog (MS) medium with Gamborg's vitamins, N(6)-benzyladenine (BA: 11.1 microM) and gibberellic acid (GA(3): 1.3 microM), obtaining 63% of established explants. The highest shoot length (22.9 mm) and multiplication rate (4.3) was achieved by cultivating for four weeks in the same basal medium supplemented with 4.4 microM BA. Both auxin induction and bacterial inoculation were used for rooting. Elongated shoots were treated with two concentrations of indole-3-butyric acid (IBA: 4.9 or 49.2 microM) during 6 days for auxin induction. Then, the shoots were transferred to an auxin-free medium and inoculated with A. brasilense Cd, Sp7 or A. chroococcum (local strain). Bacterial inoculation induced earlier rooting of photinia shoots. A. brasilense Cd with 49.2 microM IBA pulse showed a significant increase (P

Purification and characterization of a milk-clotting aspartic proteinase from globe artichoke (Cynara scolymus L.).

The study of proteinase expression in crude extracts from different organs of the globe artichoke (Cynara scolymus L.) disclosed that enzymes with proteolytic and milk-clotting activity are mainly located in mature flowers. Maximum proteolytic activity was recorded at pH 5.0, and inhibition studies showed that only pepstatin, specific for aspartic proteinases, presented a significant inhibitory effect. Such properties, in addition to easy enzyme inactivation by moderate heating, make this crude protease extract potentially useful for cheese production. Adsorption with activated carbon, together with anion exchange and affinity chromatography, led to the isolation of a heterodimeric milk-clotting proteinase consisting of 30- and 15-kDa subunits. MALDI-TOF MS of the 15-kDa chain determined a 15.358-Da mass, and the terminal amino sequence presented 96% homology with the smaller cardosin A subunit. The amino terminal sequence of the 30-kDa chain proved to be identical to the larger cardosin A subunit. Electrophoresis evidenced proteinase self-processing that was confirmed by immunoblots presenting 62-, 30-, and 15-kDa bands.

La funcion hepatica a traves del espectro clinico de la hanseniasis / The hepatic function through the clinical phantom of the leprosy

Se estudio la funcion hepatica em 100 pacientes hansenianos agrupados seguin la classificacion especial y la presencia o no de episodios reacionales del seguiente modo: pacientes quiescentes: 20 LL, 13 BL, 7 BB, 9 BT, 12 TT. Pacientes reaccionales: 12 LL, 10 BL, 7 BB, 5 TT. Como grupo testigo se estudiaron 10 individuos sanos no contactantes com pacientes hansenianos y sin antecedentes de patologia hepatica. Los resultados obtenidos muestran una hiperproteinemia a expensas de las globulinas sin disminucion de albumina en todas las formas clinicas de la hanseniase reaccionales y quiescentes, y un aumento de la actividad enzimatica serica de GPT, GOT y fosfatasa alcalina en los pacientes reaccionales. Estos incrementos de la actividade enzimatica caracterizan un duno hepatocelular leve, en que la hilirruhinemia y la prueba de turbidez del timol no presentan alteraciones. Se discuten los mecanismos fisiopatologicos causantes de estas alteraciones, postulindos su vinculacion com fenomenos de hipersensibilidad a tipo III y IV.