Soybean Lectin Enhances Biofilm Formation by Bradyrhizobium japonicum in the Absence of Plants

Soybean lectin (SBL) purified from soybean seeds by affinity chromatography strongly bound to Bradyrhizobium japonicum USDA 110 cell surface. This lectin enhanced biofilm formation by B. japonicum in a concentration-dependent manner. Presence of galactose during biofilm formation had different effects in the presence or absence of SBL. Biofilms were completely inhibited in the presence of both SBL and galactose, while in the absence of SBL, galactose was less inhibitory. SBL was very stable, since its agglutinating activity of B. japonicum cells as well as of human group A+ erythrocytes was resistant to preincubation for one week at 60°C. Hence, we propose that plant remnants might constitute a source of this lectin, which might remain active in soil and thus favor B. japonicum biofilm formation in the interval between soybean crop seasons

Distinctive phehotypes of Bradyrhizobium diazoefficiens transcriptional regulator (phaR) mutants grown on different carbon sources.

Resumen del poster presentado en: XVII Congreso Argentino de Microbiología General (SAMIGE). Córdoba, Argentina, 25-28 octubre 2022

A Rhizobium leguminosarum CHDL- (Cadherin-Like-) lectin participates in assembly and remodeling of the biofilm matrix

In natural environments most bacteria live in multicellular structures called biofilms. These cell aggregates are enclosed in a self-produced polymeric extracellular matrix, which protects the cells, provides mechanical stability and mediates cellular cohesion and adhesion to surfaces. Although important advances were made in the identification of the genetic and extracellular factors required for biofilm formation, the mechanisms leading to biofilm matrix assembly, and the roles of extracellular proteins in these processes are still poorly understood. The symbiont Rhizobium leguminosarum requires the synthesis of the acidic exopolysaccharide and the PrsDE secretion system to develop a mature biofilm. PrsDE is responsible for the secretion of the Rap family of proteins that share one or two Ra/CHDL (cadherin-like-) domains. RapA2 is a calcium-dependent lectin with a cadherin-like β sheet structure that specifically recognizes the exopolysaccharide, either as a capsular polysaccharide (CPS) or in its released form [extracellular polysaccharide (EPS)]. In this study, using gain and loss of function approaches combined with phenotypic and microscopic studies we demonstrated that RapA lectins are involved in biofilm matrix development and cellular cohesion. While the absence of any RapA protein increased the compactness of bacterial aggregates, high levels of RapA1 expanded distances between cells and favored the production of a dense matrix network. Whereas endogenous RapA(s) are predominantly located at one bacterial pole, we found that under overproduction conditions, RapA1 surrounded the cell in a way that was reminiscent of the capsule. Accordingly, polysaccharide analyses showed that the RapA lectins promote CPS formation at the expense of lower EPS production. Besides, polysaccharide analysis suggests that RapA modulates the EPS size profile. Collectively, these results show that the interaction of RapA lectins with the polysaccharide is involved in rhizobial biofilm matrix assembly and remodeling.Facultad de Ciencias ExactasInstituto de Biotecnologia y Biologia Molecula

Abundance of the Quorum-Sensing Factor Ax21 in Four Strains of Stenotrophomonas maltophilia Correlates with Mortality Rate in a New Zebrafish Model of Infection

Stenotrophomonas maltophilia is a Gram-negative pathogen with emerging nosocomial incidence. Little is known about its pathogenesis and the genomic diversity exhibited by clinical isolates complicates the study of pathogenicity and virulence factors. Here, we present a strategy to identify such factors in new clinical isolates of S. maltophilia, incorporating an adult-zebrafish model of S. maltophilia infection to evaluate relative virulence coupled to 2D difference gel electrophoresis to explore underlying differences in protein expression. In this study we report upon three recent clinical isolates and use the collection strain ATCC13637 as a reference. The adult-zebrafish model shows discrimination capacity, i.e. from very low to very high mortality rates, with clinical symptoms very similar to those observed in natural S. maltophilia infections in fish. Strain virulence correlates with resistance to human serum, in agreement with previous studies in mouse and rat and therefore supporting zebrafish as a replacement model. Despite its clinical origin, the collection strain ATCC13637 showed obvious signs of attenuation in zebrafish, with null mortality. Multilocus-sequence-typing analysis revealed that the most virulent strains, UV74 and M30, exhibit the strongest genetic similitude. Differential proteomic analysis led to the identification of 38 proteins with significantly different abundance in the three clinical strains relative to the reference strain. Orthologs of several of these proteins have been already reported to have a role in pathogenesis, virulence or resistance mechanisms thus supporting our strategy. Proof of concept is further provided by protein Ax21, whose abundance is shown here to be directly proportional to mortality in the zebrafish infection model. Indeed, recent studies have demonstrated that this protein is a quorum-sensing-related virulence factor

A novel low-cost Safe System-aligned treatment for regional and remote intersections

An intersection active warning system aiming to improve vehicle safety at intersections has been developed and trialled. This system, named RJAWS Lite, is an advisory variation of the existing mandatory-speed based RJAWS and provides two major visual warnings: (i) speed advisory on major approaches and (ii) run-through prevention on the minor approach. A controlled before-after analysis conducted on speed data from a trial at six rural intersections in South Australia indicates that RJAWS Lite can provide substantial safety benefits, albeit to a lesser degree than the full version of the RJAWS. Nonetheless, the RJAWS Lite involves considerably lower equipment and installation costs, combined with less restrictive regulations, than the mandatory full version of the RJAWS and so could be expected to be suitable for more widespread installation, which could therefore effectively increase road safety at a larger number of rural junctions. Future streamlining of the current large billboard-style RJAWS signage may likely contribute to improve driver compliance with the advised speed and therefore further increase the predicted safety benefits.Mongiardini M., Stokes C.S

Acceptance Criteria for Validation Metrics in Roadside Safety Based on Repeated Full-Scale Crash Tests

This paper proposes acceptance criteria for quantitative comparison metrics to be applied in the Verification and Validation (V&V) process of computational models used in roadside safety. Typically, the degree of verification or validation of a numerical model is assessed by qualitatively comparing the shapes of two curves, but qualitative comparisons are subjective and open to interpretation. Using quantitative comparison metrics in the V&V process allows for an objective measure of the reliability of a numerical model. Two comparison metrics were selected from a group of 16 metrics found in the literature. Acceptance criteria suitable to the typical scatter of full-scale crash tests were established by comparing ten essentially identical vehicle redirectional crash tests. Since the tests were as identical as can be achieved experimentally, the values of the quantitative metrics represented the reasonable range for the metric corresponding to matched experiments. Typical residual errors expected in full-scale tests are also discussed

Profile of crashes at intersections in South Australia

Intersections represent one of the most safety critical parts of the road network. They are characterised by a high risk of collisions with large impact angles and high speeds. In 2019, 20% of fatal crashes in metropolitan areas occurred at intersections. Knowledge of both the types and relative incidence of intersection crashes should assist in identifying effective treatments aimed at reducing the risk of casualty crashes (fatal, injury) in this critical part of the road network. Therefore, the objective of this report was to provide a broad overview of the various types of crashes occurring at intersections in South Australia. A top-down analysis was carried out on police reported road crashes that occurred in South Australia between 2013 and 2019 (inclusive). After an initial overview of all crashes at intersections, a more specific analysis was carried out on casualty crashes at intersections. Finally, a specific analysis was conducted on pedestrian-related casualty crashes at intersections. At each level of this top-down approach, a breakdown analysis was conducted to further examine the role of relevant factors such as type of control (signalised, priority, roundabouts), road environment (urban or rural) and speed limit. Results are presented in the form of various graphs and cover three major geographical areas: (i) the entire state, (ii) Adelaide metropolitan area only, (iii) outside of Adelaide metropolitan area. This information can be used to support and inform future research activities as well as the selection of appropriate countermeasures to improve safety at intersections.M Mongiardini, ME Elsegood, CS Stoke

Evaluation of Centreline ATLM along Curves in Mountainous Roads

Centreline Audio Tactile Line Marking (ATLM) was installed on a popular narrow road in the Adelaide foothills. A before-after evaluation was conducted to assess the ATLM potential for influencing lane position/crossing and speed along four trial curves. Vehicle lane position and speed at those curves, and between-site travel times were analysed. . Post treatement, all vehicle types tended to safely shift towards the edgeline along right-hand curves. However, variability along left-hand curves existed amongst the four sites. Generally, the treatment did not appear to reduce lane crossing frequency on curves, except at one site. Centreline ATLM did not appear to induce a speed decrease along curves. Nonetheless, a slight reduction of the average travel speed along the treated route may indicate a generalised speed calming effect. The study suggests merit for centreline ATLM along curves regarding lane positioning, but a limited effect on voluntary lane crossing and speed.Mongiardini, M., Stokes, C., Woolley, J