Towards a genome-scale metabolic model for the Kluyveromyces lactis yeast

The interest in Kluyveromyces lactis (K. lactis) has begun in academia due to its ability to metabolize the betaglycoside (1). Since then, this yeast has been considered a model organism for studies in genetics and physiology (2). This yeast had its genome sequenced back in 2004 (3) and recently we have published a full metabolic re-annotation of its genome (4). This re-annotation can be used, among other applications, to reconstruct genome-scale metabolic models. These models allow anticipating a given organism's phenotype from its genome sequence. The reconstruction of biochemical networks is, currently, a valid alternative to microorganisms modelling as the output provided by the in silico simulations permits focusing on experiments with promising results. Thus, we propose a new fully compartmentalised genome-scale metabolic model for K. lactis, the iOD1759 which comprises 1759 metabolic genes

Reconstructing genome-scale metabolic models with Merlin

The reconstruction of genome-scale metabolic models is based on the well-known stoichiometry of biochemical reactions. Usually the main objective of a reconstruction is the in silico simulation of the phenotypic behaviour of a microorganism, under different environmental and genetic conditions, thus representing an important tool in Metabolic Engineering. The genome of the yeast Kluyveromyces lactis was used as a case study for this method, providing information for the first stage of the reconstruction of this eukaryote. Given and input of 5085 gene sequences, Merlin identified more than 4200 distinct organisms and approximately 394.000 genes with sequence similarities to the K. lactis genome. This information, after user appraisal, will be used to assemble a metabolic model with the reactions catalysed by the enzymes encoded in the genome. Such model, in the SBML format, can be used as a first raw approach to the study of the K. lactis metabolism

Genome-wide metabolic (re-) annotation of Kluyveromyces lactis

Even before having its genome sequence published in 2004, Kluyveromyces lactis had long been considered a model organism for studies in genetics and physiology. Research on Kluyveromyces lactis is quite advanced and this yeast species is one of the few with which it is possible to perform formal genetic analysis. Nevertheless, until now, no complete metabolic functional annotation has been performed to the proteins encoded in the Kluyveromyces lactis genome. Results In this work, a new metabolic genome-wide functional re-annotation of the proteins encoded in the Kluyveromyces lactis genome was performed, resulting in the annotation of 1759 genes with metabolic functions, and the development of a methodology supported by merlin (software developed in-house). The new annotation includes novelties, such as the assignment of transporter superfamily numbers to genes identified as transporter proteins. Thus, the genes annotated with metabolic functions could be exclusively enzymatic (1410 genes), transporter proteins encoding genes (301 genes) or have both metabolic activities (48 genes). The new annotation produced by this work largely surpassed the Kluyveromyces lactis currently available annotations. A comparison with KEGG's annotation revealed a match with 844 (~90%) of the genes annotated by KEGG, while adding 850 new gene annotations. Moreover, there are 32 genes with annotations different from KEGG. Conclusions The methodology developed throughout this work can be used to re-annotate any yeast or, with a little tweak of the reference organism, the proteins encoded in any sequenced genome. The new annotation provided by this study offers basic knowledge which might be useful for the scientific community working on this model yeast, because new functions have been identified for the so-called metabolic genes. Furthermore, it served as the basis for the reconstruction of a compartmentalized, genome-scale metabolic model of Kluyveromyces lactis, which is currently being finished.This work was partially supported by the MIT-Portugal Program in Bioengineering (MIT-Pt/BS-BB/0082/2008) and a PhD grant (SFRH / BD / 47307 / 2008) from Portuguese FCT (Fundacao para a Ciencia e Tecnologia)

Esophageal schwannoma mimicking non-functional parathyroid adenoma on ^99mTc-sestamibi imaging: a case report.

Technetium-99m sestamibi single-photon emission computed tomography/computed tomography ( <sup>99m</sup> Tc-sestamibi SPECT/CT) is a mainstay of the pre-operative localization of parathyroid lesions. We report here the case of a 30 year-old woman with a fortuitously discovered 2 cm cervical mass for which a parathyroid origin was originally suspected due to its retro-thyroidal localization and a personal history of nephrolithiasis. Normal serum calcium and parathyroid hormone (PTH) levels excluded primary hyperparathyroidism, raising suspicion of a non-functional parathyroid adenoma, and SPECT/CT imaging showed that the mass was <sup>99m</sup> Tc-sestamibi-avid. Fine-needle aspiration (FNA) was performed; cytology was non-diagnostic but the needle washout was negative for thyroglobulin, calcitonin and PTH, arguing against a thyroidal or parathyroidal origin of the mass. Core needle biopsy revealed a schwannoma, ostensibly originating from the recurrent laryngeal nerve; upon surgical resection, it was finally found to arise from the esophageal submucosa. This case illustrates the fact that endocrinologists, radiologists, nuclear medicine, head and neck, and other specialists investigating patients with cervical masses should be aware that schwannomas need to be considered in the differential diagnosis of focal <sup>99m</sup> Tc-sestamibi uptake in the neck region

Contrôle du Phomopsis, maladie émergente, en culture de carotte porte-graîne

The first triangular browning umbels caused by Diaporthe angelicae on carrot seed crops were observed in France in 2007. This fungal phytopathogen negatively impacted carrot seed production. Little is known about this fungus. In order to control it in carrot seed production, a better understanding of its life cycle and its epidemiology is required. So, this is the reason why a three-years research project named DIAPOCAR was initiated in 2012. It appears that flower is the main sensitive organ and flowering the most sensitive stage of the umbel. In the field, contaminations are particularly observed during flowering of the umbels of order I and II if the weather is favourable to disease. Field fungicide trials show significant efficacy differences between fungicides and only three of them are efficient against carrot phomopsis. Finally, model simulations with the sunflower phomopsis model are promising and this tool to aid decision-making can be used to apply fungicides only when the weather is favourable to the disease. It should help carrot seed growers to determine the best timing for fungicide applications. Thus, a greater understanding of the biology of this fungus has led to a better control of phomopsis on carrot seed crops

Heterologous expression of glucose oxidase in the yeast Kluyveromyces marxianus

<p>Abstract</p> <p>Background</p> <p>In spite of its advantageous physiological properties for bioprocess applications, the use of the yeast <it>Kluyveromyces marxianus </it>as a host for heterologous protein production has been very limited, in constrast to its close relative <it>Kluyveromyces </it><it>lactis</it>. In the present work, the model protein glucose oxidase (GOX) from <it>Aspergillus niger </it>was cloned into <it>K. marxianus </it>CBS 6556 and into <it>K. lactis </it>CBS 2359 using three different expression systems. We aimed at verifying how each expression system would affect protein expression, secretion/localization, post-translational modification, and biochemical properties.</p> <p>Results</p> <p>The highest GOX expression levels (1552 units of secreted protein per gram dry cell weight) were achieved using an episomal system, in which the <it>INU1 </it>promoter and terminator were used to drive heterologous gene expression, together with the <it>INU1 </it>prepro sequence, which was employed to drive secretion of the enzyme. In all cases, GOX was mainly secreted, remaining either in the periplasmic space or in the culture supernatant. Whereas the use of genetic elements from <it>Saccharomyces cerevisiae </it>to drive heterologous protein expression led to higher expression levels in <it>K. lactis </it>than in <it>K. marxianus</it>, the use of <it>INU1 </it>genetic elements clearly led to the opposite result. The biochemical characterization of GOX confirmed the correct expression of the protein and showed that <it>K. marxianus </it>has a tendency to hyperglycosylate the protein, in a similar way as already observed for other yeasts, although this tendency seems to be smaller than the one of e.g. <it>K. lactis </it>and <it>S. cerevisiae</it>. Hyperglycosylation of GOX does not seem to affect its affinity for the substrate, nor its activity.</p> <p>Conclusions</p> <p>Taken together, our results indicate that <it>K. marxianus </it>is indeed a good host for the expression of heterologous proteins, not only for its physiological properties, but also because it correctly secretes and folds these proteins.</p

Treatment of blunt thoracic aortic injury in Germany—Assessment of the TraumaRegister DGU®

Purpose Using the data delivered by the German Trauma Register DGU® from 2002 till 2013, the value of different therapies of blunt thoracic aortic injury (BTAI) in Germany was analyzed. Methods Prospectively collected data of patients suffering from BTAI were retrospectively analyzed with focus on the different treatment modalities for grade I–IV injuries. Results 821 patients suffering from BTAI were identified: 51.6% (424) grade I injury, 35.4% (291) grade II or III injury and 12.9% (106) grade IV injury (77.5% men [44.94 ± 20.6 years]). The main patterns of injury were high- speed accidents and falls (78.0% [n = 640], 21.8% [n = 171] respectively). Significant differences between grade I and grade II/III as well as IV injuries could be assessed for the incidence of cardiopulmonary resuscitation, a Glasgow Coma Scale score below 8 and a systolic blood pressure below 90 mmHg (p-value: <0.001). In the primary admission subgroup, 44.1% (197/447) of the patients received best medical treatment, 55.9% received surgical intervention (250/447): Thereof 37.2% (93/250) received open surgery and 62.8% (147/250) had been treated by endovascular means. Significantly lower 24-h- and in-hospital-mortality rates were encountered after endovascular treatment for all gradings of BTAI (p-value: <0.001). Yet this subgroup of patients showed the lowest incidence of further severe injuries and cardiac arrest. Conclusion Endovascular therapy became the treatment of choice for BTAI in Germany. Patients who have been treated by surgical means showed the highest survival rate, especially endovascular therapy showed a favorable low mortality rate

Imaging features of retroperitoneal extra-adrenal paragangliomas in 10 dogs

Retroperitoneal paragangliomas are rare tumors of the neuroendocrine system. Only a few canine case reports are available with rare descriptions of their imaging features. The objectives of this multi-center, retrospective case series study were to describe the diagnostic imaging features of confirmed retroperitoneal paragangliomas and specify their location. Medical records and imaging studies of 10 affected dogs with cytological or histopathologic results concordant with retroperitoneal paragangliomas were evaluated. Dogs had a median age of 9 years. Four of them had clinical signs and laboratory reports compatible with excessive production of catecholamines. Six ultrasound, four CT, four radiographic, and one MRI studies were included. The paragangliomas did not have a specific location along the aorta. They were of various sizes (median 33&nbsp;mm, range: 9–85&nbsp;mm of length). Masses had heterogeneous parenchyma in six of 10 dogs, regardless of the imaging modality. Strong contrast enhancement was found in all CT studies. Encircling of at least one vessel was detected in six of 10 masses, clear invasion of a vessel was identified in one of 10 masses. In five of 10 cases, the masses were initially misconstrued as lymph nodes by the on-site radiologist. Retroperitoneal paragangliomas appear along the abdominal aorta, often presenting heterogeneous parenchyma, possibly affecting the local vasculature, and displaying strong contrast enhancement on CT. Clinical signs can be secondary to mass effects or excessive catecholamine production. Underdiagnosis and misdiagnosis of this tumor are suspected as they can be silent, of small size, or confused with other structures

De l’angine simple à l’abcès périamygdalien [From tonsillitis to peritonsillar abscess]

Acute tonsillitis is a common disease. The tonsillar infection can be uni- or bilateral with a high rate of spontaneous recovery. In some cases, a peritonsillar cellulitis or abscess can occur, characterized by an inflammatory reaction of the peritonsillar space. The red flags consist of a gradually worsening odynodysphagia, an ipsilateral swelling of the soft palate, a trismus (infrequent in children) and in some cases a change of voice (« hot potato voice »). A needle puncture of the swollen soft palate is then required. If pus is found, a surgical drainage by either hot tonsillectomy or incision under local anesthesia with an appropriate antibiotic therapy must be carried out