Cultivation of three color strains of Gracilaria domingensis in an integrated organic system

Species of the genus Gracilaria are widely distributed around the world, and many are wild harvested and cultivated for phycocolloid production (agar) and human food. This study investigated the growth performance of three color strains of Gracilaria domingensis in an integrated organic system. The three chromatic variants (red, green, and brown) were cultured in a shrimp farm at two depths (surface and 20 cm). The highest relative growth rates were obtained in the first week of cultivation for all three strains studied. Although the growth rates and biomass were slightly higher at a depth of 20 cm, significant differences were not observed at different depths (p < 0.05). The growth of the three strains varied significantly over time (p < 0.001). The maximal growth values obtained were 11.25 ± 1.50% day−1 (red), 12.76 ± 3.96% day−1 (green), and 7.86 ± 1.75% day−1 (brown). The mean growth rates for the whole period were 4.63 ± 6.97, 3.38 ± 9.09, and 2.16 ± 6.61% day−1 for the red, green, and brown strains, respectively. The pigment content (chlorophyll a, phycocyanin, and phycoerythrin) of the three strains studied was higher at a depth of 20 cm, while the maximum electron transport rate (ETRmax) and photosynthetic efficiency were higher at the surface. The high growth rates obtained in this study demonstrate the great potential of this species for integrated organic farming. Moreover, the ecological success shown by the red strain suggests that this strain is the most suitable for integrated cultivation.     

Manipulating the pathway of ammonia assimilation in transgenic non-legumes and legumes

In this paper we discuss the ways in which our understanding of the nature of the molecular controls of nitrogen assimilation has been increased by the use of non-leguminous and leguminous plants with genetically-altered capacities for ammonia assimilation. Using tobacco or Lotus as model plants, Glutamine synthetase (GS) and glutamate synthase (GOGAT) activities have been altered by stimulating or inhibiting in an organ- or tissue-specific manner the expression of the corresponding genes. In a few selected examples, the physiological impact of these genetic manipulations has been studied on plants grown under different nitrogen regimes. The use of such genetically-modified plants will allow us to better understand the molecular control of this metabolic pathway. It is also potentially of great importance in agriculture if such internal and stable modifications are beneficial in terms of nitrogen use efficiency, thus avoiding an excessive utilization of fertilizers or herbicides (GS inhibitors). Our current knowledge and prospects for future development are explored.     

<Emphasis Type="Italic">Eucalyptus</Emphasis> agroforestry system for small farms: 2-year experiment with rice and beans in Minas Gerais,Brazil

In Minas Gerais, Brazil, Eucalyptus plantation agroforestry techniques for small farms appear to be an adequate alternative to small-scale plantations, and by integrating timber and food production, they can meet local demands for aliment and wood. Intercropping of Eucalyptus camaldulensis with rice and beans was studied to compare yields of intercropping and monocultures over 2 years. In the first year, no differences were found in the productivity between the intercropped and monoculture systems of E. camaldulensis, rice and beans in both seasons. In the second year, E. camaldulensis in monoculture showed the lowest productivity. If compared with E. camaldulensis intercropped with agricultural crops, during the rainy season, beans intercropped with E. camaldulensis showed similar productivity when compared to monoculture. In the dry season, the productivity of beans in monoculture was higher than intercropping. In the second year, rice productivity was very low for both systems due to the lack of precipitation during the fructification phase. The equivalence area index (EAI) confirmed the advantages of the intercropping system over the monoculture system for E. camaldulensis and beans and rice, at least in the first 2 years.     

Generation of equine TSLP-specific antibodies and their use for detection of TSLP produced by equine keratinocytes and leukocytes

Allergic horses react to innocuous environmental substances by activation of Th2 cells and production of allergen-specific IgE antibodies. The mechanisms leading to Th2 differentiation are not well understood. In humans and mice, epithelial cell-derived thymic stromal lymphopoietin (TSLP) plays a central role in this process. Little is known about equine TSLP (eqTSLP) and its role in allergic diseases and our current knowledge is limited to the assessment of TSLP mRNA expression. In order to be able to study eqTSLP at the protein level, the aim of the present study was to produce recombinant eqTSLP protein and generate TSLP-specific antibodies. EqTSLP was cloned from a skin biopsy sample from a horse with chronic urticaria and eqTSLP protein was expressed in E.coli and in mammalian cells. Recombinant proteins were designed to include C-terminal Histag, which allowed subsequent purification and detection by Histag-specific Ab. Polyclonal and monoclonal eqTSLP-specific Ab were generated after immunization of mice with E.coli-expressed TSLP. Their specificity was tested by western blotting and ELISA. In addition, a commercially available polyclonal human TSLP-specific antibody was tested for cross-reactivity with eqTSLP. Expression of TSLP protein was confirmed by western blotting using Histag-specific Ab. E.coli-expressed TSLP appears as a band of ～13 kDa, whereas mammalian cell-expressed TSLP showed several bands at 20-25 kDa, probably representing several glycosylation forms. Polyclonal and monoclonal antibodies generated in this study, as well as commercially available human TSLP-specific Ab reacted with both E.coli- and mammalian cell-expressed TSLP in western blotting and ELISA. A capture ELISA was established to quantitate TSLP in cell supernatants and validated using supernatants from primary equine keratinocytes and peripheral blood leukocytes (PBL). Increased TSLP concentrations were found after stimulation of keratinocytes with PMA+ionomycine and with Culicoides extract. Similarly, increased TSLP concentrations were detected in PBL after stimulation with ConA, Culicoides extract, or IgE cross-linking. In conclusion, recombinant TSLP proteins and TSLP-specific antibodies produced in this study will allow further studies of the role of TSLP in equine allergic diseases.     

Characterization of the inflammatory infiltrate and cytokine expression in the skin of horses with recurrent urticaria

Background – Recurrent urticaria (RU) is a common skin disease of horses, but little is known about its pathogenesis. Hypothesis/Objective – The aim of this study was to characterize the inflammatory cell infiltrate and cytokine expression pattern in the skin of horses with RU. Animals – Biopsies of lesional and nonlesional skin of horses with RU (n = 8) and of skin from healthy control horses (n = 8) were evaluated. Methods – The inflammatory cell infiltrate was analysed by routine histology. Immunohistochemistry was used to identify T cells (CD3), B cells (CD79), macrophages (MAC387) and mast cells (tryptase). Expression of T‐helper 2 cytokines (interleukins IL‐4, IL‐5 and IL‐13), a T‐helper 1 cytokine (interferon‐γ), IL‐4 receptor α and thymic stromal lymphopoietin was assessed by quantitative RT‐PCR. Results – In subepidermal lesional skin of RU‐affected horses, increased numbers of eosinophils (P ≤ 0.01), CD79‐positive (P ≤ 0.01), MAC387‐positive (P ≤ 0.01) and tryptase‐positive cells (P ≤ 0.05) were found compared with healthy horses. Subepidermal lesional skin of RU‐affected horses contained more eosinophils (P ≤ 0.05) and tryptase‐positive cells (P ≤ 0.05) compared with nonlesional skin. There was no significant difference in infiltrating cells between nonlesional skin and skin of healthy horses. Expression of IL‐4 (P ≤ 0.01), IL‐13 (P ≤ 0.05), thymic stromal lymphopoietin (P ≤ 0.05) and IL‐4 receptor α (P ≤ 0.05) was increased in lesional skin of RU‐affected horses compared with control horses. Expression of IL‐4 was higher (P ≤ 0.05) in lesional compared with nonlesional RU skin. Conclusions and clinical importance – Analysis of cytokine expression and inflammatory infiltrate suggests that T‐helper 2 cytokines, eosinophils, mast cells and presumptive macrophages play a role in the pathogenesis of equine RU.     

In vitro analysis of expression vectors for DNA vaccination of horses: the effect of a Kozak sequence

One of the prerequisite for developing DNA vaccines for horses are vectors that are efficiently expressed in horse cells.We have analysed the ectopic expression of the human serum albumin gene in primary horse cells from different tissues. The vectors used are of pcDNA and pUC origin and include the cytomegalovirus (CMV) promoter. The pUC vectors contain CMV intron A whereas the pcDNA vectors do not.Insertion of intron A diminished the expression from the pcDNA vectors whereas insertion of a Kozak sequence upstream of the gene in two types of pUC vectors increased significantly the in vitro expression in primary horse cells derived from skin, lung, duodenum and kidney.We report for the first time the significance of full consensus Kozak sequences for protein expression in horse cells in vitro.