Engineering and characterization of Xanthomonas campestris pv. campestris towards an enhanced production of the exopolysaccharide xanthan

Steffens T. Engineering and characterization of Xanthomonas campestris pv. campestris towards an enhanced production of the exopolysaccharide xanthan. Bielefeld: Universität Bielefeld; 2017.Xanthan is the industrially most important bacterial polysaccharide, with broad fields of application (Hublik 2012). It is produced by the Gram negative phytopathogen Xanthomonas campestris pv. campestris (Xcc). Presented in this work are three systematical, mutational approaches in order to optimize xanthan production and gain new insights in the Xcc metabolism. In a first approach three mutant strains, carrying mutations in genes involved in the lipopolysaccharide (LPS) O-antigen biosynthesis, were used. The mutants Xcc H21012 (wxcB), Xcc H28110 (wxcK) and Xcc H20110 (wxcN) were constructed by Barbara Hötte and colleagues (1990) and in this thesis the phenotypic analyses are described. The mutant Xcc H21012 (wxcB) was characterized by the inhibition of the entire O-antigen. Moreover, the mutant phenotype was displayed through an increased xanthan production. In Xcc H28110 (wxcK) and Xcc H20110 (wxcN), no increase in xanthan production was detected. However, a novel LPS phenotype with an extended O-antigen main chain and no O-antigen branches could be detected. For the second approach, in order to construct a Xcc strain towards enhanced xanthan production, two flagellar genes were mutated to inhibit the motility. First the structural gene fliC was mutated, then the basal flagellar gene fliM. Mutations were performed in laboratory strain Xcc B100 and in an industrial production strain, Xcc JBL007. Phenotypic analyses revealed the inhibition of motility. Furthermore, both mutational approaches resulted in enhanced xanthan producer strains. Remarkably, even the industrial production strain could be improved. Not only in production, but also in xanthan quality. The xanthan from Xcc JBL007 fliC – and Xcc JBL007 fliM – showed enhanced rheological properties. Both was shown under distinct industrial cultivation conditions. While the chemical composition did not change between xanthan by the initial or by a mutant strain, notable differences in persistence length could be measured via atomic force microscopy. During a study with the purpose to identify sucrose related regulators, two novel regulators encoded by xcc-b100_2791 (crt1) and xcc-b100_2861 (crt2) were identified. Since they are putatively involved in Xcc carbohydrate regulation, mutants were constructed by Tobias Loka. As third approach of this thesis the mutant strains Xcc B100 crt1 – and Xcc B100 crt2 – were tested towards their cultivation characteristics and production abilities. Both mutants showed an increased xanthan yield, as compared to the initial strain Xcc B100. During the course of this work, both mutations were transferred into the production strain Xcc JBL007, after the corresponding genes in this strain were identified and sequence homologies were ensured. The xanthan production abilities of the mutant strains were then tested under industrial cultivation conditions. Both mutant strains exceeded the production of the initial strain Xcc JBL007. Results presented in this study demonstrate the possibility to further improve the xanthan production by Xcc, including the industrial xanthan production, through rational strain design

On the AAVenue to success: Advances in technologies for AAV production

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Leading ‘us’ to be active: A two-wave test of relationships between identity leadership, group identification, and attendance

Although physical activity participation has numerous physiological and psychological benefits, inactivity rates remain high, and a greater understanding of the factors that drive participation is needed. Growing evidence indicates that (1) the strength of individuals’ social identification as a member of a particular physical activity group (e.g., an exercise group or sports team) is positively associated with their group-relevant participation, and (2) physical activity leaders (e.g., exercise group leaders, coaches, and captains) can foster members’ identification, and thus their greater group-relevant participation. Extending previous cross-sectional research, we examined relationships over time between sports group members’ perceptions of their leaders’ engagement in identity leadership, their group identification, and attendance. Participants (N = 186) from amateur sports teams completed measures of identity leadership, group identification, and attendance on two occasions, eight weeks apart. Lagged regressions indicated that perceptions of leaders’ engagement in identity leadership at Time 1 predicted members’ group identification at Time 2, controlling for their group identification at Time 1; and members’ group identification at Time 2 was associated with their attendance at Time 2, controlling for their attendance at Time 1. Mediation analysis demonstrated a significant indirect effect of perceptions of leaders’ engagement in identity leadership on group members’ attendance through greater group identification. Findings provide evidence of the participation-related benefits of forming, and maintaining, strong social identities in physical activity settings, and point to the role leaders can play in fostering members' sustained identification and participation

The influence of a modified lipopolysaccharide O-antigen on the biosynthesis of xanthan in Xanthomonas campestris pv. campestris B100

Steffens T, Vorhölter F-J, Giampa M, Hublik G, Pühler A, Niehaus K. The influence of a modified lipopolysaccharide O-antigen on the biosynthesis of xanthan in Xanthomonas campestris pv. campestris B100. BMC Microbiology. 2016;16(1): 93.Background The exopolysaccharide xanthan is a natural product which is extensively used in industry. It is a thickening agent in many fields, from oil recovery to the food sector. Xanthan is produced by the Gram negative bacterium Xanthomonas campestris pv. campestris (Xcc). We analyzed the lipopolysaccharide (LPS) of three mutant strains of the Xcc wild type B100 to distinguish if the xanthan production can be increased when LPS biosynthesis is affected. Results The Xcc B100 O-antigen (OA) is composed of a linear main chain of rhamnose residues with N-acetylfucosamine (FucNAc) side branches at every second rhamnose. It is the major LPS constituent. The O-antigen was missing completely in the mutant strain H21012 (deficient in wxcB), since neither rhamnose nor FucNAc could be detected as part of the LPS by MALDI-TOF-MS, and only a slight amount of rhamnose and no FucNAc was found by GC analysis. The LPS of two other mutants was analyzed, Xcc H28110 (deficient in wxcK) and H20110 (wxcN). In both of them no FucNAc could be detected in the LPS fraction, while the rhamnose moieties were more abundant than in wild type LPS. The measurements were carried out by GC and confirmed by MALDI-TOF-MS analyses that indicated an altered OA in which the branches are missing, while the rhamnan main chain seemed longer than in the wild type. Quantification of xanthan confirmed our hypothesis that a missing OA can lead to an increased production of the extracellular polysaccharide. About 6.3 g xanthan per g biomass were produced by the Xcc mutant H21012 (wxcB), as compared to the wild type production of approximately 5 g xanthan per g biomass. In the two mutant strains with modified OA however, Xcc H28110 (wxcK) and Xcc H20110 (wxcN), the xanthan production of 5.5 g and 5.3 g, respectively, was not significantly increased. Conclusions Mutations affecting LPS biosynthesis can be beneficial for the production of the extracellular polysaccharide xanthan. However, only complete inhibition of the OA resulted in increased xanthan production. The inhibition of the FucNAc side branches did not lead to increased production, but provoked a novel LPS phenotype. The data suggests an elongation of the linear rhamnan main chain of the LPS OA in both the Xcc H28110 (wxcK) and Xcc H20110 (wxcN) mutant strains

Leaders’ creation of shared identity impacts group members’ effort and performance: Evidence from an exercise task

There is growing evidence that leaders’ effectiveness derives in part from their creation of a sense of identity that is shared by members of a group they are attempting to lead (i.e., their identity entrepreneurship). Little is known, however, about the impact of identity entrepreneurship in sport and exercise settings, particularly in relation to its effect on group members’ effort and performance. Using a pre-post between subjects experimental design, we examined the effect of leaders’ identity entrepreneurship on group members’ effort and performance during 5km cycling time trials. Following a baseline session (in which time trials were completed individually), participants (N=72) were randomly allocated to either a high or low identity entrepreneurship condition, and further randomly divided into groups of five (including a leader who was a confederate). In the subsequent test sessions (which participants attended with their fellow group members), leaders displayed either high or low identity entrepreneurship behaviors. Results indicated that, compared to participants in the low identity entrepreneurship condition, those in the high identity entrepreneurship condition maintained greater effort (maximum heart rate), and demonstrated improved (rather than poorer) performance (average power output in the first 60 seconds of time trials). Examination of pacing showed that the largest increases in participants’ average power output occurred in theearly stages of their second time trials for those in the high identity entrepreneurship condition only. Results provide causal evidence that leaders who create a shared sense of identity among team members are able to inspire greater participant effort and performance

A Social Identity Approach to Understanding and Promoting Physical Activity

Against the backdrop of a global physical inactivity crisis, attempts to both understand and positively influence physical activity behaviours are characterized by a focus on individual-level factors (e.g., cognitions, attitudes, motivation). Drawn from an emerging body of work that has explored the applicability of social identity and self-categorization theories to domains of sport and health, we outline a new perspective from which to understand and address this pervasive problem. This social identity approach suggests that the groups to which people belong can be, and often are, incorporated into their sense of self and, through this, are powerful determinants of physical activity-related behaviour. We start by reviewing the current state of physical activity research and highlighting the potential for the social identity approach to help understand how social factors influence these behaviours. Next, we outline the theoretical underpinnings of the social identity approach, and provide three key examples that speak to the analytical and practical value of the social identity approach in physical activity settings. Specifically, we argue (1) that social identity can be harnessed to promote engagement in physical activity, (2) that social identity underpins exercise group behaviour, and (3) that social identity underpins effective leadership in exercise settings. We conclude by identifying prospects for a range of theory-informed research developments

Gradual emergence of superconductivity in underdoped LSCO

We present triple-axis neutron scattering studies of low-energy magnetic fluctuations in strongly underdoped La$_{2-x}$Sr$_{x}$CuO$_{4}$ with $x=0.05$, $0.06$ and $0.07$, providing quantitative evidence for a direct competition between these fluctuations and superconductivity. At dopings $x=0.06$ and $x=0.07$, three-dimensional superconductivity is found, while only a very weak signature of two-dimensional superconductivity residing in the CuO$_2$ planes is detectable for $x=0.05$. We find a surprising suppression of the low-energy fluctuations by an external magnetic field at all three dopings. This implies that the response of two-dimensional superconductivity to a magnetic field is similar to that of a bulk superconductor. Our results provide direct evidence of a very gradual onset of superconductivity in cuprates.Comment: 5 pages, 4 figures, and supplementary materia

Putting the ‘we’ into workout: The association of identity leadership with exercise class attendance and effort, and the mediating role of group identification and comfort

This research examined how identity leadership displayed by group exercise instructors is associated with exercisers’ class attendance and in-class effort. Group exercise participants assessed their instructors’ engagement in identity leadership at baseline before indicating their comfort in the exercise environment, identification with the exercise group, class attendance, and in-class effort four weeks later. Results indicated positive associations between instructors’ identity leadership and exercisers’ group identification and comfort in the exercise environment four weeks later. Furthermore, results provided evidence of indirect effects. First, identity leadership was associated with members’ more frequent class attendance through their stronger group identification. Second, identity leadership was associated with members’ greater in-class effort through (a) their stronger group identification, and (b) their greater comfort. These relationships remained significant when accounting for the effect of established motivational predictors (i.e., competence and autonomy). These results point to the role that instructors’ leadership plays in promoting physical activity by suggesting that instructors’ engagement in identity leadership is associated with exercisers’ group-related experiences which, in turn, are a basis for group exercise participation

Genome-wide association analysis of genetic generalized epilepsies implicates susceptibility loci at 1q43, 2p16.1, 2q22.3 and 17q21.32

Genetic generalized epilepsies (GGEs) have a lifetime prevalence of 0.3% and account for 20-30% of all epilepsies. Despite their high heritability of 80%, the genetic factors predisposing to GGEs remain elusive. To identify susceptibility variants shared across common GGE syndromes, we carried out a two-stage genome-wide association study (GWAS) including 3020 patients with GGEs and 3954 controls of European ancestry. To dissect out syndrome-related variants, we also explored two distinct GGE subgroups comprising 1434 patients with genetic absence epilepsies (GAEs) and 1134 patients with juvenile myoclonic epilepsy (JME). Joint Stage-1 and 2 analyses revealed genome-wide significant associations for GGEs at 2p16.1 (rs13026414, Pmeta = 2.5 × 10−9, OR[T] = 0.81) and 17q21.32 (rs72823592, Pmeta = 9.3 × 10−9, OR[A] = 0.77). The search for syndrome-related susceptibility alleles identified significant associations for GAEs at 2q22.3 (rs10496964, Pmeta = 9.1 × 10−9, OR[T] = 0.68) and at 1q43 for JME (rs12059546, Pmeta = 4.1 × 10−8, OR[G] = 1.42). Suggestive evidence for an association with GGEs was found in the region 2q24.3 (rs11890028, Pmeta = 4.0 × 10−6) nearby the SCN1A gene, which is currently the gene with the largest number of known epilepsy-related mutations. The associated regions harbor high-ranking candidate genes: CHRM3 at 1q43, VRK2 at 2p16.1, ZEB2 at 2q22.3, SCN1A at 2q24.3 and PNPO at 17q21.32. Further replication efforts are necessary to elucidate whether these positional candidate genes contribute to the heritability of the common GGE syndrome