Growth of Pseudomonas taiwanensis VLB120∆C biofilms in the presence of n-butanol.

Biocatalytic processes often encounter problems due to toxic reactants and products, which reduce biocatalyst viability. Thus, robust organisms capable of tolerating or adapting towards such compounds are of high importance. This study systematically investigated the physiological response of Pseudomonas taiwanensis VLB120∆C biofilms when exposed to n-butanol, one of the potential next generation biofuels as well as a toxic substance using microscopic and biochemical methods. Initially P. taiwanensis VLB120∆C biofilms did not show any observable growth in the presence of 3% butanol. Prolonged cultivation of 10 days led to biofilm adaptation, glucose and oxygen uptake doubled and consequently it was possible to quantify biomass. Complementing the medium with yeast extract and presumably reducing the metabolic burden caused by butanol exposure further increased the biomass yield. In course of cultivation cells reduced their size in the presence of n-butanol which results in an enlarged surface-to-volume ratio and thus increased nutrient uptake. Finally, biofilm enhanced its extracellular polymeric substances (EPS) production when exposed to n-butanol. The predominant response of these biofilms under n-butanol stress are higher energy demand, increased biomass yield upon medium complements, larger surface-to-volume ratio and enhanced EPS production. Although we observed a distinct increase in biomass in the presence of 3% butanol it was not possible to cultivate P. taiwanensis VLB120∆C biofilms at higher n-butanol concentrations. Thereby this study shows that biofilms are not per se tolerant against solvents, and need to adapt to toxic n-butanol concentrations.

Comparison of Single-Level and Multiple-Level Outcomes of Total Disc Arthroplasty: 24-Month Results.

BACKGROUND: Low back pain is one of the most prevalent problems in industrialized countries, affecting as many as 80% of all adults at some time in their lives. Among the significant contributors to low back pain is degenerative disc disease (DDD). Although fusion has been well accepted for treatment of DDD, high rates of complications and stress to adjacent segments remain a concern. Lumbar total disc replacement (TDR) was developed with a goal of preserving motion and avoiding various fusion-related complications, but the relative merits of single vs. multiple level arthroplasty remain unclear. METHODS: This is a multi-center, single arm, prospective post-market registry of the M6-L, consisting of consecutive patients presenting with lumbar DDD who agreed to participate. This paper reports on those patients who have completed at least 24 months of followup to date. Clinical outcome measures include the Oswestry Disability Index (ODI) and back and leg Visual Analogue Scales (VAS). Radiographic analysis of disc angle and range of motion (ROM) was also performed. RESULTS: Results for 83 patients comprising 121 implants in two cohorts (49 single level (SL), 34 multiple levels (ML)) are reported. Both cohorts experienced significant improvement at 24 months including significant decreases in ODI and VAS. Relative to SL procedures, ML procedures demonstrated either comparable results, or results that trended favorably towards the ML procedures. Index and global ROM at 24 months were not significantly different between the two cohorts, while the disc angles were larger in the SL cohort regardless of index level. CONCLUSIONS: This is the first study to report clinical and radiographic outcomes of TDR with the M6-L in SL vs ML procedures with two years of followup. The results suggest initial device safety and effectiveness when used for the treatment of lumbar degenerative disc disease at one or more levels.

Maverick total disc replacement in a real-world patient population: a prospective, multicentre, observational study.

PURPOSE: Controlled trials have shown that total disc replacement (TDR) can provide pain and disability relief to patients with degenerative disc disease; however, whether these outcomes can also be achieved for patients treated in normal surgical practice has not been well documented. METHODS: This prospective, international study observed changes in disability and back pain in 134 patients who were implanted with Maverick TDR within the framework of routine clinical practice and followed for 2 years post-surgery. Primary and secondary outcomes were the differences from baseline to 6 months post-surgery in the means of the Oswestry Disability Index and the change in back pain intensity assessed on a 10-cm visual analogue scale, respectively. Mean patient age at surgery was 43 years, but ranged up to 65 years. RESULTS: One hundred twenty-three patients had an implant at one level, 10 patients at two levels, and one patient at three levels. Statistically significant improvements in mean disability (-25.4) and low back pain intensity (-4.0) scores were observed at 6 months postoperatively (P < 0.0001 for both) in the hands of experienced surgeons (>10 TDRs per centre). During the study, 56 patients (42 %) experienced a complication or adverse event. CONCLUSIONS: This is the first international observational study to report outcomes of TDR in real-world clinical settings. We showed statistically significant improvements in disability and pain scores at 6 months following Maverick TDR, which were maintained for 2 years alongside an acceptable rate of perioperative complications. The safety and tolerability shown in this observational study were comparable to those from controlled trials.

Metabolic engineering of Pseudomonas sp. strain VLB120 as platform biocatalyst for the production of isobutyric acid and other secondary metabolites.

BACKGROUND: Over the recent years the production of Ehrlich pathway derived chemicals was shown in a variety of hosts such as Escherichia coli, Corynebacterium glutamicum, and yeast. Exemplarily the production of isobutyric acid was demonstrated in Escherichia coli with remarkable titers and yields. However, these examples suffer from byproduct formation due to the fermentative growth mode of the respective organism. We aim at establishing a new aerobic, chassis for the synthesis of isobutyric acid and other interesting metabolites using Pseudomonas sp. strain VLB120, an obligate aerobe organism, as host strain. RESULTS: The overexpression of kivd, coding for a 2-ketoacid decarboxylase from Lactococcus lactis in Ps. sp. strain VLB120 enabled for the production of isobutyric acid and isobutanol via the valine synthesis route (Ehrlich pathway). This indicates the existence of chromosomally encoded alcohol and aldehyde dehydrogenases catalyzing the reduction and oxidation of isobutyraldehyde. In addition we showed that the strain possesses a complete pathway for isobutyric acid metabolization, channeling the compound via isobutyryl-CoA into valine degradation. Three key issues were addressed to allow and optimize isobutyric acid synthesis: i) minimizing isobutyric acid degradation by host intrinsic enzymes, ii) construction of suitable expression systems and iii) streamlining of central carbon metabolism finally leading to production of up to 26.8 ± 1.5 mM isobutyric acid with a carbon yield of 0.12 ± 0.01 g g(glc)⁻¹. CONCLUSION: The combination of an increased flux towards isobutyric acid using a tailor-made expression system and the prevention of precursor and product degradation allowed efficient production of isobutyric acid in Ps. sp. strain VLB120. This will be the basis for the development of a continuous reaction process for this bulk chemicals.

Characterization of a biofilm membrane reactor and its prospects for fine chemical synthesis.

Biofilms are known to be robust biocatalysts. Conventionally, they have been mainly applied for wastewater treatment, however recent reports about their employment for chemical synthesis are increasingly attracting attention. Engineered Pseudomonas sp. strain VLB120 Delta C biofilm growing in a tubular membrane reactor was utilized for the continuous production of (S)-styrene oxide. A biofilm specific morphotype appeared in the effluent during cultivation, accounting for 60-80% of the total biofilm irrespective of inoculation conditions but with similar specific activities as the original morphotype. Mass transfer of the substrate styrene and the product styrene oxide was found to be dependent on the flow rate but was not limiting the epoxidation rate. Oxygen was identified as one of the main parameters influencing the biotransformation rate. Productivity was linearly dependent on the specific membrane area and on the tube wall thickness. On average volumetric productivities of 24 g L(aq) (-1) day(-1) with a maximum of 70 g L(aq) (-1) day(-1) and biomass concentrations of 45 g (BDW) L(aq) (-1) have been achieved over long continuous process periods (>or=50 days) without reactor downtimes.