The GOODSTEP project: General Object-Oriented Database for Software Engineering Processes

The goal of the GOODSTEP project is to enhance and improve the functionality of a fully object-oriented database management system to yield a platform suited for applications such as software development environments (SDEs). The baseline of the project is the O2 database management system (DBMS). The O2 DBMS already includes many of the features regulated by SDEs. The project has identified enhancements to O2 in order to make it a real software engineering DBMS. These enhancements are essentially upgrades of the existing O2 functionality, and hence require relatively easy extensions to the O2 system. They have been developed in the early stages of the project and are now exploited and validated by a number of software engineering tools built on top of the enhanced O2 DBMS. To ease tool construction, the GOODSTEP platform encompasses tool generation capabilities which allow for generation of integrated graphical and textual tools from high-level specifications. In addition, the GOODSTEP platform provides a software process toolset which enables modeling, analysis and enaction of software processes and is also built on top of the extended O2 database. The GOODSTEP platform is to be validated using two CASE studies carried out to develop an airline application and a business application

Visualization of grapevine root colonization by the Saharan soil isolate Saccharothrix algeriensis NRRL B-24137 using DOPE-FISH microscopy

Background and aim There is currently a gap of knowledge regarding whether some beneficial bacteria isolated from desert soils can colonize epi- and endophytically plants of temperate regions. In this study, the early steps of the colonization process of one of these bacteria, Saccharothrix algeriensis NRRL B-24137, was studied on grapevine roots to determine if this beneficial strain can colonize a non-natural host plant. An improved method of fluorescence in situ hybridization (FISH), the double labeling of oligonucleotide probes (DOPE)-FISH technique was used to visualize the colonization behavior of such bacteria as well as to determine if the method could be used to track microbes on and inside plants. Methods A probe specific to Saccharothrix spp. was firstly designed. Visualization of the colonization behavior of S. algeriensis NRRL B-24137 on and inside roots of grapevine plants was then carried out with DOPE-FISH microscopy. Results The results showed that 10 days after inoculation, the strain could colonize the root hair zone, root elongation zone, as well as root emergence sites by establishing different forms of bacterial structures as revealed by the DOPE-FISH technique. Further observations showed that the strain could be also endophytic inside the endorhiza of grapevine plants. Conclusions Taking into account the natural niches of this beneficial strain, this study exemplifies that, in spite of its isolation from desert soil, the strain can establish populations as well as subpopulations on and inside grapevine plants and that the DOPE-FISH tool can allow to detect it

Detection of quorum sensing molecules in Burkholderia cepacia culture supernatants with enzyme-linked immunosorbent assays.

The Burkholderia cepacia complex (Bcc) employs a quorum sensing (QS) mechanism which is a cell density-dependent bacterial communication system to regulate certain gene expressions. As with many other Gram-negative bacteria, Burkholderia cepacia species use (N-acyl-)homoserine lactones (AHLs or HSLs) as signalling molecules. Because of the essential role of QS in bacterial behavior, the aim of this study was to demonstrate the applicability of our in-house-developed enzyme-linked immunosorbent assays (ELISAs) for the detection of bacterial activities via HSLs in B. cepacia strain LA3 culture supernatants. For this purpose the previously developed monoclonal antibodies (mAbs) HSL1/2-2C10 and HSL1/2-4H5 were exploited. N-3-Oxo-decanoyl-L-homoserine lactone (3-oxo-C10-HSL) was used as main analyte throughout all experiments. With the bacterial culture medium (named ABC medium) a matrix effect in both ELISAs was visible (slight increase in optical density, shift in test midpoints (IC(50)) and working ranges). For example, ELISA with mAb HSL1/2-2C10 and enzyme tracer HSL3-HRP (HSL derivative conjugated to horseradish peroxidase) had an IC(50) of 120 μg L(-1) for 3-oxo-C10-HSL in phosphate-buffered saline versus 372 μg L(-1) in ABC medium. A significant increase of HSLs in B. cepacia strain LA3 culture supernatants after 12 h to 48 h of growth was observed. Although the analytical result of these immunoassays cannot distinguish HSLs from homoserines (HSs), the appearance of these compounds can be easily followed. Hydrolysis and spiking experiments were carried out with these biological samples. According to our knowledge, these are the first immunoassays for the detection of quorum sensing molecules in biological culture supernatants. This study provides a cost-effective, fast, and sensitive analytical method for detection of HSLs/HSs in biological samples without complex sample preparation and will offer a quick idea about B. cepacia activities. The low sample amount requirement (less than 1 mL) constitutes a tremendous advantage for many analytical questions with biological samples

Root colonization by Pseudomonas sp. DSMZ 13134 and impact on the indigenous rhizosphere bacterial community of barley.

Over the last few decades, the ability of rhizosphere bacteria to promote plant growth has been considered to be of scientific, ecological, and economic interest. The properties and mechanisms of interaction of these root-colonizing bacteria have been extensively investigated, and plant protection agents that are based on these bacterial strains have been developed for agricultural applications. In the present study, the root colonization of barley by Pseudomonas sp. DSMZ 13134, that is contained in the commercially available plant protection agent Proradix, was examined using the fluorescence in situ hybridization method with oligonucleotide probes and specific gfp-tagging of the inoculant strain in combination with confocal laser scanning microscopy. In the first phase of root colonization, the inoculant strain competed successfully with seed and soil-borne bacteria (including Pseudomonads) for the colonization of the rhizoplane. Pseudomonas sp. DSMZ 13134 could be detected in all parts of the roots, although it did not belong to the dominant members of the root-associated bacterial community. Gfp-tagged cells were localized particularly in the root hair zone, and high cell densities were apparent on the root hair surface. To investigate the impact of the application of Proradix on the structure of the dominant root-associated bacterial community of barley, T-RFLP analyses were performed. Only a transient community effect was found until 3 weeks post-application

Determination of phosphorus in small amounts of protein samples by ICP-MS

Inductively coupled plasma mass spectrometry (ICP-MS) is used for phosphorus determination in protein samples. A small amount of solid protein sample (down to 1 micro g) or digest (1-10 micro L) protein solution was denatured in nitric acid and hydrogen peroxide by closed-microvessel microwave digestion. Phosphorus determination was performed with an optimized analytical method using a double-focusing sector field inductively coupled plasma mass spectrometer (ICP-SFMS) and quadrupole-based ICP-MS (ICP-QMS). For quality control of phosphorus determination a certified reference material (CRM), single cell proteins (BCR 273) with a high phosphorus content of 26.8+/-0.4 mg g(-1), was analyzed. For studies on phosphorus determination in proteins while reducing the sample amount as low as possible the homogeneity of CRM BCR 273 was investigated. Relative standard deviation and measurement accuracy in ICP-QMS was within 2%, 3.5%, 11% and 12% when using CRM BCR 273 sample weights of 40 mg, 5 mg, 1 mg and 0.3 mg, respectively. The lowest possible sample weight for an accurate phosphorus analysis in protein samples by ICP-MS is discussed. The analytical method developed was applied for the analysis of homogeneous protein samples in very low amounts [1-100 micro g of solid protein sample, e.g. beta-casein or down to 1 micro L of protein or digest in solution (e.g., tau protein)]. A further reduction of the diluted protein solution volume was achieved by the application of flow injection in ICP-SFMS, which is discussed with reference to real protein digests after protein separation using 2D gel electrophoresis.The detection limits for phosphorus in biological samples were determined by ICP-SFMS down to the ng g(-1) level. The present work discusses the figure of merit for the determination of phosphorus in a small amount of protein sample with ICP-SFMS in comparison to ICP-QMS

Effects of glyphosate on the bacterial community associated with roots of transgenic Roundup Ready^® soybean.

Introduction of glyphosate-resistant soybean plants into agricultural systems has greatly increased the application frequency of glyphosate. Because glyphosate is able to inhibit 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) not only in plants but also in different microorganisms, its application could lead to shifts in rhizosphere microbial communities in farming soils. In this study, greenhouse experiments were conducted with the objective to evaluate the effects of glyphosate on the composition and diversity of rhizosphere bacterial communities of transgenic soybean. This was especially relevant, because foliar applied glyphosate is transported down to the roots and exuded into the rhizosphere. After two foliar herbicide applications, root samples of treated and untreated plants were analysed by 16S rRNA gene T-RFLP analysis. Multivariate statistical analysis of the data and diversity indices were used to assess changes in the microbial populations in response to glyphosate applications. A comparison of rhizosphere communities revealed that the abundance of a T-RF representing microbes related to Burkholderia sp. significantly decreased under glyphosate application, while the abundance of a T-RF representing uncultured Gemmatimonadetes significantly increased. Interestingly, the bacterial community associated with soybean roots after glyphosate application not only demonstrated effective resilience after the disturbance but in addition, T-RF diversity also increased in comparison to the untreated control samples. The results suggest that bacterial diversity was even stimulated in the rhizosphere after glyphosate application