Specific sequence modifications of a cry3B endotoxin gene result in high levels of expression and insect resistance

Solanum melongena (eggplant) cv. Picentia and the wild species Solanum integrifolium were transformed with both a wild type (wt) and four mutagenized versions of Bacillus thuringiensis (Bt) gene Bt43 belonging to the cry3 class. The Bt gene was partly modified in its nucleotide sequence by replacing four target regions (W: +1 to +170; X: +592 to +1057; Y: +1203 to +1376; Z: +1376 to +1984) with synthetic fragments obtained by polymerase chain reaction amplification of crude oligonucleotides. The synthetic Bt genes were designed to avoid, in their modified regions, sequences such as ATTTA sequence, polyadenylation sequences and splicing sites, which might destabilize the messenger RNA. Furthermore, the codon usage was improved for a better expression in the plant system. The amino acid composition was not altered. Four versions of the modified Bt gene were obtained, BtE, BtF, BtH and BtI, with a nucleotide subtitution percentage of 8.2, 8.6, 14, and 16%, respectively, in comparison to the wt gene Bt43. Modified versions contained different subsets of substituted regions: BtE-W + Z, BtF - Y + Z, BtH-X + Y + Z, BtI - W + X + Y + Z. In the final modified version (BtI), overall guanine+cytosine was increased from the 34.1% of the wt gene to 45.5%, and most of the destabilizing sequences were eliminated. Transgenic plants obtained with the more modified versions, BtH and BtI, were fully resistant to Leptinotarsa decemlineata Say first- and third-instar larvae, while Bt43 wt, BtE and BtF genotypes did not cause mortality and did not affect larval development.     

Understanding spaces as places: extending interaction design paradigms

This short paper focuses on some issues involved in the design of ubiquitous technologies and their integration within physical spaces. Current design methodologies and techniques do not explicitly consider the importance of gaining a full understanding of the human experience of space when designing technologies that will pervade and become an integral part of our physical environment. This paper is specifically focused on the importance of considering spaces as places, and of analysing those features of a place that are going to shape users interactions with technologically enhanced environments. It also examines the future implications that this approach may have on theoretical and methodological aspects of interaction design.

Molecular mechanisms driving <Emphasis Type="Italic">Streptococcus mitis</Emphasis> entry into human gingival fibroblasts in presence of chitlac-nAg and saliva

The molecular mechanisms leading to Streptococcus mitis capability of entering oral cells were investigated in a co-culture of S. mitis and Human Gingival Fibroblasts (HGFs) in the presence of saliva. An innovative colloidal solution based on silver nanoparticles (Chitlac-nAg), a promising device for daily oral care, was added to the experimental system in order to study the effects of silver on the bacterial overgrowth and ability to enter non-phagocytic eukaryotic cells. The entry of bacteria into the eukaryotic cells is mediated by a signalling pathway involving FAK, integrin β1, and the two cytoskeleton proteins vinculin and F-actin, and down-regulated by the presence of saliva both at 3 and 48?h of culture, whereas Chitlac-n Ag exposure seems to influence, by incrementing it, the number of bacteria entering the fibroblasts only at 48?h. The formation of fibrillary extrusion from HGFs and the co-localization of bacteria and silver nanoparticles within the fibroblast vacuoles were also recorded. After longer experimental times (72 and 96?h), the number of S. mitis chains inside gingival cells is reduced, mainly in presence of saliva. The results suggest an escape of bacteria from fibroblasts to restore the microbial balance of the oral cavity.

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Saliva improves Streptococcus mitis protective effect on human gingival fibroblasts in presence of 2-hydroxyethyl-methacrylate

This study aimed to investigate the effect of saliva on Streptococcus mitis free cells and on S. mitis/human gingival fibroblasts (HGFs) co-culture model, in presence of 2-hydroxyethyl-methacrylate (HEMA). The bacterial aggregation both in the planktonic phase and on HGFs, as well as the apoptotic and necrotic eukaryotic cells amount were analyzed, in presence of saliva and/or HEMA. The aggregation test revealed a significant saliva aggregation effect on S. mitis strains compared to the untreated sample. No significant differences were recorded in the amount of culturable bacteria in all studied conditions; however, from microscopy images, the saliva/HEMA combining effect induced a significant bacterial aggregation and adhesion on HGFs. HEMA treatment decreased viable eukaryotic cell number with a parallel increment of necrotic cells, but when saliva was added to the co-culture, the viable cells percentage increased to a value comparable to the control sample.     

Comparison of an objective method of measuring bulbar redness to the use of traditional grading scales.

PURPOSE: The primary objective was to compare measures of bulbar redness objectively using a photometric method with standard grading methods. Measures of redness were made on 24 participants wearing a silicone hydrogel contact lens in one eye for overnight wear. This report compares hyperaemia after 1 week of daily wear (baseline) with redness measured after 6 months of overnight wear. METHOD: A new method of objectively measuring bulbar conjunctival redness was performed using the Spectrascan650 Photometer by Photo Research under fixed illumination. Photometric measures in CIEu(*) chromaticity values involve the measurement of chromaticity, a physical analogue of redness, greenness and blueness in the image. This method was validated in Part 1 of the study using repeated measurements on the photographic CCLRU scale. In Part 2 of the study, the photographic grading scale (CCLRU) from 0 (none) to 100 (extreme) was used to make the comparison. RESULTS: Part 1 indicated that the photometer provides a repeatable and reliable measure of bulbar redness (CCC=0.989). A moderately strong and significant correlation was found between the CIEu(*) chromaticity values and the analogue data (R=0.795, p=0.000) at each measurement session (from baseline to 1 day, 1 week, and 1, 3 and 6 months of overnight wear). CONCLUSIONS: This new standardized and objective method of measuring bulbar redness has great potential to replace subjective grading scales, especially with multi-centre studies, where variability between investigators occurs. This method may also detect smaller changes between visits or between eyes.