Data management in life cycle assessment: a case study of wastewater treatment.

To assess the environmental impact of wastewater treatment, life cycle assessment (LCA) is a frequently applied instrument. However, these studies often require large amounts of data. The complexity and heterogeneity of these data result in the need for a systematic data management approach. Especially the generation of the life cycle inventory (LCI) holds the potential to be facilitated by automation. A case study in the wastewater sector was used to demonstrate the implementation of data management. A database structure was developed to store the raw data of the wastewater plants (WWTPs) and make it accessible through code. The code interacted with the database, implemented calculations, and automatically created the inventory based on the processed data. The database provides a consistent structure for the raw data and can also be used for backup purposes. Because it is machine-readable it can be accessed through the code that enables the automated generation of the LCI. As a proof of concept, a sequence of the code is provided with a user interface and can be tested online. We found that for most use cases, basic programming tools were sufficient for systematic data management, and, therefore, the approach is considered accessible for LCA practitioners.

Diagnostic accuracy of the ROCHE Septifast PCR system for the rapid detection of blood pathogens in neonatal sepsis-A prospective clinical trial.

INTRODUCTION: Diagnosis of neonatal sepsis remains a major challenge in neonatology. Most molecular-based methods are not customized for neonatal requirements. The aim of the present study was to assess the diagnostic accuracy of a modified multiplex PCR protocol for the detection of neonatal sepsis using small blood volumes. METHODS: 212 episodes of suspected neonatal late onset sepsis were analyzed prospectively using the Roche SeptiFast® MGRADE PCR with a modified DNA extraction protocol and software-handling tool. Results were compared to blood culture, laboratory biomarkers and clinical signs of sepsis. RESULTS: Of 212 episodes, 85 (40.1%) were categorized as "not infected". Among these episodes, 1 was false positive by blood culture (1.2%) and 23 were false positive by PCR (27.1%). Of 51 (24.1%) episodes diagnosed as "culture proven sepsis", the same pathogen was detected by blood culture and PCR in 39 episodes (76.5%). In 8 episodes, more pathogens were detected by PCR compared to blood culture, and in 4 episodes the pathogen detected by blood culture was not found by PCR. One of these episodes was caused by Bacillus cereus, a pathogen not included in the PCR panel. In 76/212 (35.8%) episodes, clinical sepsis was diagnosed. Among these, PCR yielded positive results in 39.5% of episodes (30/76 episodes). For culture-positive sepsis, PCR showed a sensitivity of 90.2% (95%CI 86.2-94.2%) and a specificity of 72.9% (95%CI 67.0-79.0%). CONCLUSION: The Roche SeptiFast® MGRADE PCR using a modified DNA extraction protocol showed acceptable results for rapid detection of neonatal sepsis in addition to conventional blood culture. The benefit of rapid pathogen detection has to be balanced against the considerable risk of contamination, loss of information on antibiotic sensitivity pattern and increased costs.

Molecular detection of late-onset neonatal sepsis in premature infants using small blood volumes: proof-of-concept.

BACKGROUND: Conventional blood culture is still the gold standard for sepsis diagnosis but results are not immediately available and pathogens are only detected in approximately 25% of cases. New molecular assays for the detection of blood stream pathogens are promising diagnostic tools. OBJECTIVES: The aim of the study was to adapt and evaluate a multiplex PCR system using 100 µl blood. - METHODS: 46 blood specimens of very low birth weight infants (818 ± 242 g) with suspected sepsis were analyzed using the Roche SeptiFast MGRADE PCR with a modified DNA extraction protocol and software handling tool for decreased blood volume requirements. RESULTS: In the non-infected group, 5/21 infants had a positive PCR result with coagulase-negative staphylococci. All pathogens detected in the blood culture positive group (n = 15) were also detected by PCR. In addition, 4/6 patients had a positive PCR result in the clinical sepsis group (clinical and laboratory signs of sepsis but negative blood culture). Overall, the PCR was demonstrated to have a higher sensitivity (90.5%; 95%CI 68.2-98.3%) in comparison to blood culture (71.4%; 95%CI 47.7-87.8%) including clinical sepsis cases, even though it had a lower specificity (80.0%; 95%CI 58.7-92.4% versus 100.0%; 95%CI 83.4-100.0%). CONCLUSIONS: These first data demonstrate the usability and potential benefit of this multiplex PCR using a modified DNA extraction for the rapid detection of nosocomial sepsis in preterm infants in addition to blood culture.

Ureterorenoscopic biopsy and urinary cytology according to the 2004 WHO classification underestimate tumor grading in upper urinary tract urothelial carcinoma.

OBJECTIVES: To determine accuracy of upper tract cytology and ureteroscopic biopsy according to the 2004 World Health Organization (WHO) classification in predicting the correct tumor grade in patients with urothelial cancer of the upper urinary tract (UUT-UC). METHODS: Pathology reports of 77 nephroureterectomy specimens were retrospectively analyzed for tumor grade and compared with preoperatively gained cytology and ureteroscopic biopsy results. For analysis, the 2004 WHO classification was used. RESULTS: Overall sensitivity of cytology and biopsy in diagnosis of UUT-UC was 64% and 74%, respectively. Accuracy of cytology and biopsy in predicting high grade cancer was 53% and 58%, respectively. Combination of cytology and biopsy could improve sensitivity (84%) and accuracy (68%), but even for this combination, 15% of high grade tumors were misinterpreted as low grade cancer. CONCLUSION: Our results show only limited accuracy for preoperative cytology and ureterorenoscopically performed biopsies in the prediction of the correct tumor grading of an UUT-UC. Therefore, we suggest the use of additional diagnostic procedures before the decision for definitive surgical treatment in patients with UUT-UC is made.

Nosocomial rhinovirus infection in preterm infants.

During 11 months, all preterm infants admitted to our neonatal care facility with suspected respiratory tract infection were screened for respiratory viruses by polymerase chain reaction. Rhinovirus infection was identified in 16 infants, leading to severe respiratory compromise in most cases. Distribution of rhinovirus infections during the year showed a strong clustering trend, suggesting a major role for nosocomial transmission.

Staging of muscle-invasive bladder cancer: can computerized tomography help us to decide on local treatment?

OBJECTIVES: To assess the power of multi-detector row computerized tomography (MDCT) in daily routine as a basic staging procedure for the decision on local treatment of patients with bladder cancer. PATIENTS AND METHODS: We retrospectively analysed 276 patients who had undergone radical cystectomy between 2004 and 2008 and correlated the MDCT findings with pathological findings, number of removed lymph nodes and type of urinary diversion. RESULTS: Accuracy of MDCT in predicting pathological tumour stage was 49% (kappa coefficient, 0.23; P < 0.001). Overstaging occurred in 23.4%, and understaging occurred in 24.7%. Accuracy in predicting lymph node metastases was 54% (kappa coefficient, 0.04; P = 0.297). Overstaging and understaging occurred in 8.3 and 29.4%, respectively. Significantly more ileal conduits were performed in patients with high postoperative pathological tumour stages (P = 0.04) and positive lymph nodes (P = 0.013). In contrast, there was no correlation between preoperative CT tumour/nodal stage and the number of removed lymph nodes (P = 0.44 and P = 0.732, respectively), and between preoperative tumour stage and type of urinary diversion (P = 0.126). CONCLUSIONS: MDCT as a preoperative staging procedure has a low accuracy in predicting the correct tumour and nodal stage, and therefore, it has little impact on decision-making for local treatment of muscle-invasive bladder cancer during radical cystectomy.

Individual learning curve reduces the clinical value of urinary cytology.

UNLABELLED: The aim of the study was to define the learning curve of a single cytologist as a limitation of urinary cytology. A total of 1034 cytologic and histologic findings of patients undergoing transurethral resection of the bladder for suspicion of bladder cancer were reviewed, and cytologic evaluations of a single cytologist from the beginning of his learning period were compared with the results of a cytologist at a national reference center. Our results showed that the individual learning curve has a significant impact on the quality of urine cytology. BACKGROUND: To define the learning curve of a single cytologist as a limitation of urinary cytology and to compare his results with a reference center for urinary cytology. PATIENTS AND METHODS: Cytologic and histologic findings of patients undergoing transurethral resection of the bladder for suspicion of bladder cancer were reviewed. Cytologic evaluations from a single local cytologist during his learning period and of a cytologist of a national reference center were compared. Changes of sensitivity and specificity of cytology from the local cytologist over the time period were assessed. Differences were estimated by using kappa statistics. RESULTS: The local cytologist evaluated 1034 cytologic findings. Sensitivity and specificity of cytology estimated by the local cytologist were 86% and 66% at the beginning of the learning period, but 68% and 84% at the end of it (P ≤ .05). In high-grade carcinomas, sensitivity did not decrease over the years (P > .05). In contrast, the sensitivity in the diagnosis of low-grade tumors decreased from 86% to 56% (P ≤ .05). The reference center estimated a constant sensitivity and specificity of 59% and 97%, respectively. Agreement of findings between local and national cytologists increased significantly over the learning period (P < .05). CONCLUSION: The individual learning curve has a significant impact on the quality of urinary cytology. Both specificity of cytology and sensitivity for low-grade tumors changed significantly when done by a local cytologist at the beginning of learning period. These findings emphasize the impact of the individual learning curve on the clinical value of urinary cytology in diagnosis of bladder cancer.

The archaeal Lsm protein binds to small RNAs.

Proteins of the Lsm family, including eukaryotic Sm proteins and bacterial Hfq, are key players in RNA metabolism. Little is known about the archaeal homologues of these proteins. Therefore, we characterized the Lsm protein from the haloarchaeon Haloferax volcanii using in vitro and in vivo approaches. H. volcanii encodes a single Lsm protein, which belongs to the Lsm1 subfamily. The lsm gene is co-transcribed and overlaps with the gene for the ribosomal protein L37e. Northern blot analysis shows that the lsm gene is differentially transcribed. The Lsm protein forms homoheptameric complexes and has a copy number of 4000 molecules/cell. In vitro analyses using electrophoretic mobility shift assays and ultrasoft mass spectrometry (laser-induced liquid bead ion desorption) showed a complex formation of the recombinant Lsm protein with oligo(U)-RNA, tRNAs, and an small RNA. Co-immunoprecipitation with a FLAG-tagged Lsm protein produced in vivo confirmed that the protein binds to small RNAs. Furthermore, the co-immunoprecipitation revealed several protein interaction partners, suggesting its involvement in different cellular pathways. The deletion of the lsm gene is viable, resulting in a pleiotropic phenotype, indicating that the haloarchaeal Lsm is involved in many cellular processes, which is in congruence with the number of protein interaction partners.

Urinary cytology in era of fluorescence endoscopy: redefining the role of an established method with a new reference standard.

OBJECTIVES: To assess whether the use of fluorescence endoscopy (FE) decreases the clinical value of urinary cytology compared with the use of white light endoscopy (WLE). METHODS: The endoscopic, cytologic, and histologic findings of patients who had undergone transurethral resection of the bladder with or without FE were reviewed. The number and characteristics of the tumors that had been overlooked by WLE or FE but detected by cytology were analyzed. An assessment of whether the sensitivity and specificity of urinary cytology changed according to the use of FE or WLE was conducted. RESULTS: The data from 1705 patients were included. Histologic findings were obtained from 238 patients using WLE and from 1467 patients using FE. Histologically confirmed malignancy was found in 641 patients, and the disease of 1064 was classified as benign. FE was superior to WLE in sensitivity in the detection of bladder cancer (94.3% vs 86.3%, P <.05). Cytology detected 53 of 88 tumors that were not detected by WLE (47 high-grade tumors); 31 tumors were overlooked by FE, of which 20 were detected by cytology (12 were high-grade tumors). The sensitivity and specificity of cytology was 66.0% and 78.4%, respectively. The specificity of cytology using WLE and FE was 75% and 79.1% (P >.05) and the sensitivity was 61.4% and 67.4% (P >.05), respectively. CONCLUSIONS: Although FE demonstrated a great sensitivity, cytology still has a role even when using FE, because a small group of patients with high-grade tumors were detected by urinary cytology only.

Influence of clinical information on the interpretation of urinary cytology in bladder cancer: how suggestible is a cytologist?

OBJECTIVE: to determine the influence of the knowledge of the endoscopic findings and the influence of the patient's history on the cytologist's judgement, as urinary cytology is known to be subjective and has several limitations, in particular a high inter- and intra-observer variability. PATIENTS AND METHODS: we analysed the cytological and histological findings of patients who underwent transurethral resection of a bladder tumour, and determined whether the cytologist was aware of the endoscopic findings or not. The sensitivity and specificity of cytology were calculated with or without this knowledge, and that of the patients' bladder cancer history. RESULTS: the findings of 1705 patients were reviewed; in 641 the histological examination confirmed a malignant tumour and 1046 were classified as benign. The sensitivity of cytology was 66.0% and the specificity was 78.4%. The cytologist was aware of the endoscopic finding and patient history in 742 cases, and unaware of the endoscopic findings in 963. The specificity was higher in the latter group (80.2% vs 73.0%; P= 0.006). The specificity in patients with the endoscopic findings described as 'negative', 'inflammation', 'scar tissue', 'flat lesion', 'suspicious for tumour', and 'exophytic tumour' was 89.8%, 89.9%, 85.0%, 77.1%, 63.2% and 48.6%, respectively (P < 0.001). In 898 patients the history was negative for bladder tumours. Among these patients the sensitivity and specificity of cytology was 67.3% and 79.7%; the sensitivity and specificity was 65.4% and 74.8% for the 807 patients with a positive history of bladder cancer (P= 0.054). CONCLUSION: both being aware of the endoscopic findings and a positive patient history for bladder cancer lowers the specificity of cytology. Consequently, the cytologist should be unaware of the endoscopic findings.

Small RNAs in haloarchaea: identification, differential expression and biological function.

To elucidate the role of small noncoding RNAs (sRNAs) in archaea we applied RNomics to identify sRNAs in the halophilic archaeon Haloferax volcanii. Using a size-selected cDNA library, 39 different previously uncharacterized sRNAs were identified ranging in size from 130 to 460 nucleotides. Twenty-one of these sRNAs are located in intergenic regions and 18 in antisense orientation. One of the intergenic sRNAs codes for a peptide. Only a minor fraction of sRNA genes were preceded by promoter elements (15 of 39), indicating that the majority might be generated by processing from larger precursors. Northern blot analyses of the intergenic sRNAs revealed differential expression for several sRNAs. Deletion mutants of two sRNAs were constructed, demonstrating that this approach is suitable to elucidate their biological function. Both mutant strains showed a defined phenotype: sRNA(30) gene deletion mutant was less resistant to higher temperatures and sRNA(63) gene deletion mutant resulted in a severe growth defect at low salt concentrations. Proteome analyses revealed clear differences between wildtype and deletion strains. These results represent the first reported examples of experimentally characterizing the function of sRNAs, excepting snoRNAs, in archaea. Taken together, we showed that haloarchaea encode sRNAs, some of which are differentially expressed and which have the potential to fulfil important biological functions in vivo.

Small RNAs of the halophilic archaeon Haloferax volcanii.

In recent years, sRNAs (small non-coding RNAs) have been found to be abundant in eukaryotes and bacteria and have been recognized as a novel class of gene expression regulators. In contrast, much less is known about sRNAs in archaea, except for snoRNAs (small nucleolar RNAs) that are involved in the modification of bases in stable RNAs. Therefore bioinformatic and experimental RNomics approaches were undertaken to search for the presence of sRNAs in the model archaeon Haloferax volcanii, resulting in more than 150 putative sRNA genes being identified. Northern blot analyses were used to study (differential) expression of sRNA genes. Several chromosomal deletion mutants of sRNA genes were generated and compared with the wild-type. It turned out that two sRNAs are essential for growth at low salt concentrations and high temperatures respectively, and one is involved in the regulation of carbon metabolism. Taken together, it could be shown that sRNAs are as abundant in H. volcanii as they are in well-studied bacterial species and that they fulfil important biological roles under specific conditions.

Induced release of membrane vesicles from rat adipocytes containing glycosylphosphatidylinositol-anchored microdomain and lipid droplet signalling proteins.

Synthesis and degradation of lipids in mammalian adipocytes are tightly and coordinatedly regulated by insulin, fatty acids, reactive oxygen species and drugs. Conversely, the lipogenic or lipolytic state of adipocytes is communicated to other tissues by the secretion of soluble adipocytokines. Here we report that insulin, palmitate, H(2)O(2) and the antidiabetic sulfonylurea drug glimepiride induce the release of the typical lipid droplet (LD) protein, perilipin-A, as well as typical plasma membrane microdomain (DIGs) proteins, such as caveolin-1 and the glycosylphosphatidylinositol (GPI)-anchored proteins, Gce1 and CD73 from rat adipocytes. According to biochemical and morphological criteria these LD and GPI-proteins are embedded within two different types of phospholipid-containing membrane vesicles, collectively called adiposomes. Adiposome release was not found to be causally related to cell lysis or apoptosis. The interaction of Gce1 and CD73 with the adiposomes apparently depends on their intact GPI anchor. Pull-down of caveolin-1, perilipin-A and CD73 together with phospholipids (via binding to annexin-V) as well as mutually of caveolin-1 with CD73 or perilipin-A (via coimmunoprecipitation) argues for their colocalization within the same adiposome vesicle. Taken together, certain lipogenic and anti-lipolytic agents induce the specific release of a subset of LD and DIGs proteins, including certain GPI-proteins, in adiposomes from primary rat adipocytes. Given the (c)AMP-degrading activities of Gce1 and CD73 and LD-forming function of perilipin-A and caveolin-1, the physiological relevance of the release of adiposomes from adipocytes may rely on the intercellular transfer of lipogenic and anti-lipolytic information.

Coordinated regulation of esterification and lipolysis by palmitate, H2O2 and the anti-diabetic sulfonylurea drug, glimepiride, in rat adipocytes.

Inhibition of lipolysis by palmitate, H2O2 and the anti-diabetic sulfonylurea drug, glimepiride, in isolated rat adipocytes has previously been shown to rely on the degradation of cyclic adenosine monophosphate by the phosphodiesterase, Gce1, and the 5'-nucleotidase, CD73. These glycosylphosphatidylinositol (GPI)-anchored proteins are translocated from plasma membrane lipid rafts to intracellular lipid droplets upon H2O2-induced activation of a GPI-specific phospholipase C (GPI-PLC) in response to palmitate and glimepiride in intact adipocytes and, as demonstrated here, in cell-free systems as well. The same agents are also known to stimulate the incorporation of fatty acids into triacylglycerol. Here the involvement of H2O2 production, GPI-PLC activation and translocation of Gce1 and CD73 in the agent-induced esterification and accompanying lipid droplet formation was tested in rat adipocytes using relevant inhibitors. The results demonstrate that upregulation of the esterification and accumulation of triacylglycerol by glimepiride depends on the sequential H2O2-induced GPI-PLC activation and GPI-protein translocation as does inhibition of lipolysis. In contrast, stimulation of the esterification and triacylglycerol accumulation by palmitate relies on insulin-independent tyrosine phosphorylation and thus differs from its anti-lipolytic mechanism. As expected, insulin regulates lipid metabolism via typical insulin signalling independent of H2O2 production, GPI-PLC activation and GPI-protein translocation, albeit these processes are moderately stimulated by insulin. In conclusion, triacylglycerol and lipid droplet formation in response to glimepiride and H2O2 may involve the hydrolysis of cyclic adenosine monophosphate by lipid droplet-associated Gce1 and CD73 which may regulate lipid droplet-associated triacylglycerol-synthesizing and hydrolyzing enzymes in coordinated and inverse fashion.

Influencing factors for regional lymph node recurrence of breast cancer.

INTRODUCTION: We investigated whether the risk of developing regional lymph node recurrence was dependant on the number of lymph nodes removed. OBJECT: We followed 2,961 patients of whom 50 (1.69%) developed regional recurrent disease during a median period of 73 months (4-192 months). RESULT: For those women who had involved lymph nodes at initial surgery we were able to establish an inverse correlation between the development of local recurrence and the number of lymph nodes removed in multivariate analysis. For women who had no affected lymph nodes, the number of lymph nodes removed did not influence the incidence of local recurrence. CONCLUSION: We conclude from these data that women who have negative nodal status at surgical staging do not benefit from further systematic axillary resection with regard to regional lymph node recurrence. For those women, however, who have confirmed axillary metastasis, systematic axillary resection lowers the risk of regional lymph node recurrence depending on the number of lymph nodes removed.

Semen quality in men with malignant diseases before and after therapy and the role of cryopreservation.

OBJECTIVE: To review the literature and answer questions about semen quality in young cancer patients before and after therapy and the importance of sperm cryopreservation. METHODS: All aspects of sperm cryopreservation and effects of therapies on semen quality were examined on the basis of MedLine database searches. RESULTS: Chemotherapy, radiation, or their combination results in a significant reduction of sperm quality and as a consequence an indefinite time of infertility follows. The type of cancer and the pretreatment sperm concentrations were the most significant factors governing posttreatment semen quality and recovery of spermatogenesis. Due to their age, fertility and sexual functioning are key issues for these patients. Yet there is no medical protection of the germinal epithelium available. Male germ cell transplantation is in its infancy and still there are no therapeutical options to improve spermatogenesis after damage has occurred. Consequently, cryopreservation represents the only preemptive measure for conserving fertility. CONCLUSION: This manuscript updates the current knowledge of diverse chemotherapeutic treatment regimens and their gonadotoxic effects as well as the development of posttreatment fertility in cancer patients. The importance and rationale of sperm cryopreservation are discussed and possible future options are highlighted.