How to juggle priorities? An interactive tool to provide quantitative support for strategic patient-mix decisions: an ophthalmology case

An interactive tool was developed for the ophthalmology department of the Academic Medical Center to quantitatively support management with strategic patient-mix decisions. The tool enables management to alter the number of patients in various patient groups and to see the consequences in terms of key performance indicators. In our case study, we focused on the bottleneck: the operating room. First, we performed a literature review to identify all factors that influence an operating room's utilization rate. Next, we decided which factors were relevant to our study. For these relevant factors, two quantitative methods were applied to quantify the impact of an individual factor: regression analysis and computer simulation. Finally, the average duration of an operation, the number of cancellations due to overrun of previous surgeries, and the waiting time target for elective patients all turned out to have significant impact. Accordingly, for the case study, the interactive tool was shown to offer management quantitative decision support to act proactively to expected alterations in patient-mix. Hence, management can anticipate the future situation, and either alter the expected patient-mix or expand capacity to ensure that the key performance indicators will be met in the future

A broader role for AmyR in Aspergillus niger: regulation of the utilisation of d-glucose or d-galactose containing oligo- and polysaccharides

AmyR is commonly considered a regulator of starch degradation whose activity is induced by the presence of maltose, the disaccharide building block of starch. In this study, we demonstrate that the role of AmyR extends beyond starch degradation. Enzyme activity assays, genes expression analysis and growth profiling on d-glucose- and d-galactose-containing oligo- and polysaccharides showed that AmyR regulates the expression of some of the Aspergillus niger genes encoding α- and β-glucosidases, α- and β- galactosidases, as well as genes encoding α-amlyases and glucoamylases. In addition, we provide evidence that d-glucose or a metabolic product thereof may be the inducer of the AmyR system in A. niger and not maltose, as is commonly assumed

Expression, Purification and Characterization of Arginase from Helicobacter pylori in Its Apo Form

Arginase, a manganese-dependent enzyme that widely distributed in almost all creatures, is a urea cycle enzyme that catalyzes the hydrolysis of L-arginine to generate L-ornithine and urea. Compared with the well-studied arginases from animals and yeast, only a few eubacterial arginases have been characterized, such as those from H. pylori and B. anthracis. However, these enzymes used for arginase activity assay were all expressed with LB medium, as low concentration of Mn2+ was detectable in the medium, protein obtained were partially Mn2+ bonded, which may affect the results of arginase activity assay. In the present study, H. pylori arginase (RocF) was expressed in a Mn2+ and Co2+ free minimal medium, the resulting protein was purified through affinity and gel filtration chromatography and the apo-form of RocF was confirmed by flame photometry analysis. Gel filtration indicates that the enzyme exists as monomer in solution, which was unique as compared with homologous enzymes. Arginase activity assay revealed that apo-RocF had an acidic pH optimum of 6.4 and exhibited metal preference of Co2+>Ni2+>Mn2+. We also confirmed that heat-activation and reducing regents have significant impact on arginase activity of RocF, and inhibits S-(2-boronoethyl)-L-Cysteine (BEC) and Nω-hydroxy-nor-Arginine (nor-NOHA) inhibit the activity of RocF in a dose-dependent manner

Membrane TNF confers protection to acute mycobacterial infection

BACKGROUND: Tumour necrosis factor (TNF) is crucial for the control of mycobacterial infection as TNF deficient (KO) die rapidly of uncontrolled infection with necrotic pneumonia. Here we investigated the role of membrane TNF for host resistance in knock-in mice with a non-cleavable and regulated allele (mem-TNF). METHODS: C57BL/6, TNF KO and mem-TNF mice were infected with M. tuberculosis H37Rv (Mtb at 100 CFU by intranasal administration) and the survival, bacterial load, lung pathology and immunological parameters were investigated. Bone marrow and lymphocytes transfers were used to test the role of membrane TNF to confer resistance to TNF KO mice. RESULTS: While TNF-KO mice succumbed to infection within 4–5 weeks, mem-TNF mice recruited normally T cells and macrophages, developed mature granuloma in the lung and controlled acute Mtb infection. However, during the chronic phase of infection mem-TNF mice succumbed to disseminated infection with necrotic pneumonia at about 150 days. Reconstitution of irradiated TNF-KO mice with mem-TNF derived bone marrow cells, but not with lymphocytes, conferred host resistance to Mtb infection in TNF-KO mice. CONCLUSION: Membrane expressed TNF is sufficient to allow cell-cell signalling and control of acute Mtb infection. Bone marrow cells, but not lymphocytes from mem-TNF mice confer resistance to infection in TNF-KO mice. Long-term infection control with chronic inflammation likely disrupting TNF mediated cell-cell signalling, additionally requires soluble TNF

The Characterization of Helicobacter pylori DNA Associated with Ancient Human Remains Recovered from a Canadian Glacier

Helicobacter pylori is a gram-negative bacterium that colonizes the stomach of nearly half of the world's population. Genotypic characterization of H. pylori strains involves the analysis of virulence-associated genes, such as vacA, which has multiple alleles. Previous phylogenetic analyses have revealed a connection between modern H. pylori strains and the movement of ancient human populations. In this study, H. pylori DNA was amplified from the stomach tissue of the Kwäday Dän Ts'ìnchi individual. This ancient individual was recovered from the Samuel Glacier in Tatshenshini-Alsek Park, British Columbia, Canada on the traditional territory of the Champagne and Aishihik First Nations and radiocarbon dated to a timeframe of approximately AD 1670 to 1850. This is the first ancient H. pylori strain to be characterized with vacA sequence data. The Tatshenshini H. pylori strain has a potential hybrid vacA m2a/m1d middle (m) region allele and a vacA s2 signal (s) region allele. A vacA s2 allele is more commonly identified with Western strains, and this suggests that European strains were present in northwestern Canada during the ancient individual's time. Phylogenetic analysis indicated that the vacA m1d region of the ancient strain clusters with previously published novel Native American strains that are closely related to Asian strains. This indicates a past connection between the Kwäday Dän Ts'ìnchi individual and the ancestors who arrived in the New World thousands of years ago

Multifaceted SlyD from Helicobacter pylori: implication in [NiFe] hydrogenase maturation

SlyD belongs to the FK506-binding protein (FKBP) family with both peptidylprolyl isomerase (PPIase) and chaperone activities, and is considered to be a ubiquitous cytosolic protein-folding facilitator in bacteria. It possesses a histidine- and cysteine-rich C-terminus binding to selected divalent metal ions (e.g., Ni2+, Zn2+), which is important for its involvement in the maturation processes of metalloenzymes. We have determined the solution structure of C-terminus-truncated SlyD from Helicobacter pylori (HpSlyDΔC). HpSlyDΔC folds into two well-separated, orientation-independent domains: the PPIase-active FKBP domain and the chaperone-active insert-in-flap (IF) domain. The FKBP domain consists of a four-stranded antiparallel β-sheet with an α-helix on one side, whereas the IF domain folds into a four-stranded antiparallel β-sheet accompanied by a short α-helix. Intact H. pylori SlyD binds both Ni2+ and Zn2+, with dissociation constants of 2.74 and 3.79 μM respectively. Intriguingly, binding of Ni2+ instead of Zn2+ induces protein conformational changes around the active sites of the FKBP domain, implicating a regulatory role of nickel. The twin-arginine translocation (Tat) signal peptide from the small subunit of [NiFe] hydrogenase (HydA) binds the protein at the IF domain. Nickel binding and the recognition of the Tat signal peptide by the protein suggest that SlyD participates in [NiFe] hydrogenase maturation processes

Persistent Gastric Colonization with Burkholderia pseudomallei and Dissemination from the Gastrointestinal Tract following Mucosal Inoculation of Mice

Melioidosis is a disease of humans caused by opportunistic infection with the soil and water bacterium Burkholderia pseudomallei. Melioidosis can manifest as an acute, overwhelming infection or as a chronic, recurrent infection. At present, it is not clear where B. pseudomallei resides in the mammalian host during the chronic, recurrent phase of infection. To address this question, we developed a mouse low-dose mucosal challenge model of chronic B. pseudomallei infection and investigated sites of bacterial persistence over 60 days. Sensitive culture techniques and selective media were used to quantitate bacterial burden in major organs, including the gastrointestinal (GI) tract. We found that the GI tract was the primary site of bacterial persistence during the chronic infection phase, and was the only site from which the organism could be consistently cultured during a 60-day infection period. The organism could be repeatedly recovered from all levels of the GI tract, and chronic infection was accompanied by sustained low-level fecal shedding. The stomach was identified as the primary site of GI colonization as determined by fluorescent in situ hybridization. Organisms in the stomach were associated with the gastric mucosal surface, and the propensity to colonize the gastric mucosa was observed with 4 different B. pseudomallei isolates. In contrast, B. pseudomallei organisms were present at low numbers within luminal contents in the small and large intestine and cecum relative to the stomach. Notably, inflammatory lesions were not detected in any GI tissue examined in chronically-infected mice. Only low-dose oral or intranasal inoculation led to GI colonization and development of chronic infection of the spleen and liver. Thus, we concluded that in a mouse model of melioidosis B. pseudomallei preferentially colonizes the stomach following oral inoculation, and that the chronically colonized GI tract likely serves as a reservoir for dissemination of infection to extra-intestinal sites

Parallels between Pathogens and Gluten Peptides in Celiac Sprue

Pathogens are exogenous agents capable of causing disease in susceptible organisms. In celiac sprue, a disease triggered by partially hydrolyzed gluten peptides in the small intestine, the offending immunotoxins cannot replicate, but otherwise have many hallmarks of classical pathogens. First, dietary gluten and its peptide metabolites are ubiquitous components of the modern diet, yet only a small, genetically susceptible fraction of the human population contracts celiac sprue. Second, immunotoxic gluten peptides have certain unusual structural features that allow them to survive the harsh proteolytic conditions of the gastrointestinal tract and thereby interact extensively with the mucosal lining of the small intestine. Third, they invade across epithelial barriers intact to access the underlying gut-associated lymphoid tissue. Fourth, they possess recognition sequences for selective modification by an endogenous enzyme, transglutaminase 2, allowing for in situ activation to a more immunotoxic form via host subversion. Fifth, they precipitate a T cell–mediated immune reaction comprising both innate and adaptive responses that causes chronic inflammation of the small intestine. Sixth, complete elimination of immunotoxic gluten peptides from the celiac diet results in remission, whereas reintroduction of gluten in the diet causes relapse. Therefore, in analogy with antibiotics, orally administered proteases that reduce the host's exposure to the immunotoxin by accelerating gluten peptide destruction have considerable therapeutic potential. Last but not least, notwithstanding the power of in vitro methods to reconstitute the essence of the immune response to gluten in a celiac patient, animal models for the disease, while elusive, are likely to yield fundamentally new systems-level insights

Induction chemotherapy followed by chemoradiotherapy versus chemoradiotherapy alone as neoadjuvant treatment for locally recurrent rectal cancer: study protocol of a multicentre, open-label, parallel-arms, randomized controlled study (PelvEx II)

Background A resection with clear margins (R0 resection) is the most important prognostic factor in patients with locally recurrent rectal cancer (LRRC). However, this is achieved in only 60 per cent of patients. The aim of this study is to investigate whether the addition of induction chemotherapy to neoadjuvant chemo(re)irradiation improves the R0 resection rate in LRRC. Methods This multicentre, international, open-label, phase III, parallel-arms study will enrol 364 patients with resectable LRRC after previous partial or total mesorectal resection without synchronous distant metastases or recent chemo- and/or radiotherapy treatment. Patients will be randomized to receive either induction chemotherapy (three 3-week cycles of CAPOX (capecitabine, oxaliplatin), four 2-week cycles of FOLFOX (5-fluorouracil, leucovorin, oxaliplatin) or FOLFORI (5-fluorouracil, leucovorin, irinotecan)) followed by neoadjuvant chemoradiotherapy and surgery (experimental arm) or neoadjuvant chemoradiotherapy and surgery alone (control arm). Tumours will be restaged using MRI and, in the experimental arm, a further cycle of CAPOX or two cycles of FOLFOX/FOLFIRI will be administered before chemoradiotherapy in case of stable or responsive disease. The radiotherapy dose will be 25 × 2.0 Gy or 28 × 1.8 Gy in radiotherapy-naive patients, and 15 × 2.0 Gy in previously irradiated patients. The concomitant chemotherapy agent will be capecitabine administered twice daily at a dose of 825 mg/m2 on radiotherapy days. The primary endpoint of the study is the R0 resection rate. Secondary endpoints are long-term oncological outcomes, radiological and pathological response, toxicity, postoperative complications, costs, and quality of life. Discussion This trial protocol describes the PelvEx II study. PelvEx II, designed as a multicentre, open-label, phase III, parallel-arms study, is the first randomized study to compare induction chemotherapy followed by neoadjuvant chemo(re)irradiation and surgery with neoadjuvant chemo(re)irradiation and surgery alone in patients with locally recurrent rectal cancer, with the aim of improving the number of R0 resections