Regulatory associations between the metabolism of sulfur-containing amino acids and xanthan biosynthesis in Xanthomonas campestris pv. campestris B100.

The γ-proteobacterium Xanthomonas campestris pv. campestris (Xcc) B100 synthesizes the exopolysaccharide xanthan, a commercially relevant thickening agent produced commonly by industrial scale fermentation. This work was inspired by the observation that methionine is an inhibitor of xanthan formation in growth experiments. Therefore, the global effects of methionine supplementation were characterized through cultivation experiments, genome-wide microarray hybridizations and qRT-PCR. Specific pull down of DNA-binding proteins by using the intergenic regions upstream of xanA, gumB and gumD led to the identification of six transcriptional regulators, among them the LysR-family transcriptional regulator CysB. An insertion mutant of this gene was analyzed by growth experiments, microarray experiments and qRT-PCR. Based on our experimental data, we developed a model that describes the methionine-dependent co-regulation of xanthan and sulfur-containing compounds in Xanthomonas. These data substantially contribute to better understand the impact of methionine as a compound in xanthan production media used in industrial fermentations.

Effects and Consequences of Being an OSCE Examiner in Surgery-A Qualitative Study.

OBJECTIVE: Even though objective structured clinical examination (OSCE) is a well-investigated format for competency-based practical examination, only a few studies have explored the motivations of OSCE examiners and their opinions, both positive and negative, toward being an examiner. The aim of this study was to gain insights into the views of OSCE examiners using semi-structured interviews. DESIGN: Surgical OSCE examiners were queried at two medical faculties in Germany via semi-structured interviews. The interviews were transcribed verbatim and analyzed using the techniques of structured qualitative content analysis. SETTING: This study was conducted at the medical faculties of the Goethe University, Frankfurt, Germany and of the Otto-von-Guericke University, Magdeburg, Germany. PARTICIPANTS: All of the study participants were surgeons working at the university hospital of one of the faculties. RESULTS: A total of 29 examiners were queried until a saturation of content was achieved. A critical reflection of one's own teaching was described as a major benefit by most participants. Furthermore, they noted that the standards and competences examined during the OSCE boosted the detail of their teaching sessions in the wards. However, the examiners criticized missed operations due the examination and were not appreciated by superiors for being an examiner. Most of the examiners (22/29) preferred to be an examiner themselves rather than appointing student peer examiners. If they had appointed someone else, that would mean they would miss valuable experiences useful for their own teaching. CONCLUSIONS: Being an OSCE examiner confers several advantages, notably the reflection of one's own teaching, which was described as highly valuable by the examiners.

Prevalence of Helicobacter pylori Infection in Patients with Minimal Hepatic Encephalopathy.

BACKGROUND: Ammonia is a critical factor in the pathogenesis of minimal hepatic encephalopathy (MHE). Most of the ammonia is from bacterial production in the intestinal tract. Non-absorbable antibiotics and non-absorbable disaccharides are therefore the therapy of choice. A few studies have addressed the effect of ammonia produced by H. pylori in the pathogenesis of MHE. METHODS: In this prospective clinical trial, 84 consecutive patients with liver cirrhosis (LC) underwent laboratory, psychometric und neurophysiological testing to determine serological H. pylori status, MHE and blood ammonia levels. Relevant clinical and demographic characteristics were documented. RESULTS: Out of 84 LC patients (83% male), 29% presented with MHE as assessed by critical flicker frequency analysis (CFF). The prevalence of H. pylori infection in the cohort was 21%; 22% of H. pylori-infected patients presented with MHE according to the criterion of a positive CFF result. If the criterion for MHE was a positive CFF and a positive NCT-A result, then 17% of H. pylori positive patients suffered from MHE. The prevalence of MHE in H. pylori-negative patients, based on CFF alone and on the combination, was 30%. A proportion of 19% of the patients with MHE had increased blood ammonia levels. CONCLUSION: The amount of ammonia produced by H. pylori does not affect venous ammonia levels. Therefore, an additional benefit of H. pylori eradication in the treatment of hepatic encephalopathy in patients with LC is unlikely to occur.

Characterization of the pyrophosphate-dependent 6-phosphofructokinase from Xanthomonas campestris pv. campestris.

Xanthomonads are plant pathogenic proteobacteria that produce the polysaccharide xanthan. They are assumed to catabolize glucose mainly via the Entner-Doudoroff pathway. Whereas previous studies have demonstrated no phosphofructokinase (PFK) activity in xanthomonads, detailed genome analysis revealed in Xanthomonas campestris pathovar campestris (Xcc) genes for all Embden-Meyerhof-Parnas pathway (glycolysis) enzymes, including a conserved pfkA gene similar to 6-phosphofructokinase genes. To address this discrepancy between genetic and physiological properties, the pfkA gene of Xcc strain B100 was cloned into the expression vector pET28a+. The 45-kDa pfkA gene product exhibited no conventional PFK activity. Bioinformatic analysis of the Xcc PfkA amino acid sequence suggested utilization of pyrophosphate as an alternative cosubstrate. Pyrophosphate-dependent PFK activity was shown in an in vitro enzyme assay for purified Xcc PfkA, as well as in the Xcc B100 crude protein extract. Kinetic constants were determined for the forward and reverse reactions. Primary structure conservation indicates the global presence of similar enzymes among Xanthomonadaceae.