Incisional hernia following closure of loop ileostomy: The main predictor is the patient, not the surgeon.

BACKGROUND: Incisional hernia at the ileostomy site occurs in 0-48% of patients undergoing loop ileostomy closure. Risk factors for ileostomy-site hernia are not currently well understood. We explored the predictive value of patient and clinical factors for ileostomy-site hernias. METHOD: Loop ileostomy reversals undertaken between 1st January 2009 and 31st December 2013 were retrospectively evaluated. Preoperative patient data (BMI, age, gender, blood pressure, diabetes), surgical variables (preoperative ileostomy marking, intraoperative management (suture type, closure method), postoperative complications (≤30 days), approach, urgency, and chemotherapy, hospital stay, stoma closure interval, follow-up duration) were collected. Patients were followed up by clinical examination and postoperative imaging. RESULTS: 193 loop ileostomy reversals were identified. Operative indications included: colorectal cancer (n = 102, 52.8%); inflammatory bowel disease (n = 47, 24.3%); diverticulosis (n = 20, 10.4%); assorted indications (n = 19, 9.8%); and inflammatory fistulae (n = 5, 2.6%). Median duration of clinical follow-up was 20.5 months (0-69). Hernia occurred in 26 patients (13.5%), detected at a median of eight months post-reversal. Radiological follow-up occurred in 72% of patients and, as a reference standard, in 100% of patients diagnosed with a hernia. Concordance between clinical and radiological findings was 88.5%. Postoperative complications predicted higher hernia risk. BMI and preoperative blood pressure were significant hernia predictors. Differences in the type of suture material to close the defect (absorbable vs. non-absorbable) and stoma skin closure method (primary vs. secondary intention healing) were non-predictive of hernia. CONCLUSION: Whilst BMI and patient comorbidity are the major hernia predictors, variability in surgical practice does not constitute a significant risk factor for ileostomy-site incisional hernia.

Visualization of cellulose synthases in Arabidopsis secondary cell walls.

Cellulose biosynthesis in plant secondary cell walls forms the basis of vascular development in land plants, with xylem tissues constituting the vast majority of terrestrial biomass. We used plant lines that contained an inducible master transcription factor controlling xylem cell fate to quantitatively image fluorescently tagged cellulose synthase enzymes during cellulose deposition in living protoxylem cells. The formation of secondary cell wall thickenings was associated with a redistribution and enrichment of CESA7-containing cellulose synthase complexes (CSCs) into narrow membrane domains. The velocities of secondary cell wall-specific CSCs were faster than those of primary cell wall CSCs during abundant cellulose production. Dynamic intracellular of endomembranes, in combination with increased velocity and high density of CSCs, enables cellulose to be synthesized rapidly in secondary cell walls.

Recent Y chromosome divergence despite ancient origin of dioecy in poplars (Populus).

All species of the genus Populus (poplar, aspen) are dioecious, suggesting an ancient origin of this trait. Despite some empirical counter examples, theory suggests that nonrecombining sex-linked regions should quickly spread, eventually becoming heteromorphic chromosomes. In contrast, we show using whole-genome scans that the sex-associated region in Populus trichocarpa is small and much younger than the age of the genus. This indicates that sex determination is highly labile in poplar, consistent with recent evidence of 'turnover' of sex-determination regions in animals. We performed whole-genome resequencing of 52 P. trichocarpa (black cottonwood) and 34 Populus balsamifera (balsam poplar) individuals of known sex. Genomewide association studies in these unstructured populations identified 650 SNPs significantly associated with sex. We estimate the size of the sex-linked region to be ~100 kbp. All SNPs significantly associated with sex were in strong linkage disequilibrium despite the fact that they were mapped to six different chromosomes (plus 3 unmapped scaffolds) in version 2.2 of the reference genome. We show that this is likely due to genome misassembly. The segregation pattern of sex-associated SNPs revealed this to be an XY sex-determining system. Estimated divergence times of X and Y haplotype sequences (6-7 Ma) are much more recent than the divergence of P. trichocarpa (poplar) and Populus tremuloides (aspen). Consistent with this, in P. tremuloides, we found no XY haplotype divergence within the P. trichocarpa sex-determining region. These two species therefore have a different genomic architecture of sex, suggestive of at least one turnover event in the recent past.

Monolignol ferulate transferase introduces chemically labile linkages into the lignin backbone.

Redesigning lignin, the aromatic polymer fortifying plant cell walls, to be more amenable to chemical depolymerization can lower the energy required for industrial processing. We have engineered poplar trees to introduce ester linkages into the lignin polymer backbone by augmenting the monomer pool with monolignol ferulate conjugates. Herein, we describe the isolation of a transferase gene capable of forming these conjugates and its xylem-specific introduction into poplar. Enzyme kinetics, in planta expression, lignin structural analysis, and improved cell wall digestibility after mild alkaline pretreatment demonstrate that these trees produce the monolignol ferulate conjugates, export them to the wall, and use them during lignification. Tailoring plants to use such conjugates during cell wall biosynthesis is a promising way to produce plants that are designed for deconstruction.

A 34K SNP genotyping array for Populus trichocarpa: design, application to the study of natural populations and transferability to other Populus species.

Genetic mapping of quantitative traits requires genotypic data for large numbers of markers in many individuals. For such studies, the use of large single nucleotide polymorphism (SNP) genotyping arrays still offers the most cost-effective solution. Herein we report on the design and performance of a SNP genotyping array for Populus trichocarpa (black cottonwood). This genotyping array was designed with SNPs pre-ascertained in 34 wild accessions covering most of the species latitudinal range. We adopted a candidate gene approach to the array design that resulted in the selection of 34 131 SNPs, the majority of which are located in, or within 2 kb of, 3543 candidate genes. A subset of the SNPs on the array (539) was selected based on patterns of variation among the SNP discovery accessions. We show that more than 95% of the loci produce high quality genotypes and that the genotyping error rate for these is likely below 2%. We demonstrate that even among small numbers of samples (n = 10) from local populations over 84% of loci are polymorphic. We also tested the applicability of the array to other species in the genus and found that the number of polymorphic loci decreases rapidly with genetic distance, with the largest numbers detected in other species in section Tacamahaca. Finally, we provide evidence for the utility of the array to address evolutionary questions such as intraspecific studies of genetic differentiation, species assignment and the detection of natural hybrids.

Fast-track management of patients undergoing proximal pancreatic resection.

INTRODUCTION: To avoid the risk of complications of biliary drainage, a feasibility study was carried out to determine whether it might be possible to fast-track surgical treatment, with resection before biliary drainage, in jaundiced patients with proximal pancreatic/peri-ampullary malignancy. PATIENTS AND METHODS: Over an 18-month period, based on their presenting bilirubin levels and other logistical factors, all jaundiced patients who might be suitable for fast-track management were identified. Data on complications and hospital stay were compared with those patients in whom a conventional pathway (with biliary drainage) was used during the same time period. Data were also compared with a group of patients from the preceding 6 months. RESULTS: Nine patients were fast-tracked and 49 patients treated in the conventional pathway. Fast-track patients mean (SD) serum bilirubin level was 265 micromol/l (81.6) at the time of the operation compared to 43 micromol/l (51.3; P > or = 0.0001) in conventional patients. Mean (SD) of time from referral to operation, 14 days (9) versus 59 days (36.9), was significantly shorter in fast-track patients than conventional patients (P < or = 0.0001). Length of hospital stay mean (SD) at 17 (6) days versus 22 days (19.6; P = 0.2114), surgical complications and mortality in fast-track patients were similar to conventional patients. Prior to surgery, the 49 conventional patients underwent a total of 73 biliary drainage procedures resulting in seven major complications. Comparison with the group of patients from the previous 6 months indicated that the conventional group were not disadvantaged. CONCLUSIONS: Fast-track management by resection without biliary drainage of selected patients with distal biliary strictures is safe and has the potential to reduce the waiting time to surgery, overall numbers of biliary drainage procedures and the complications thereof.

Comparison of multislice computed tomography and endoscopic ultrasonography with operative and histological findings in suspected pancreatic and periampullary malignancy.

BACKGROUND: This study compared multislice computed tomography (MSCT) with endoscopic ultrasonography (EUS) in the diagnosis and staging of pancreatic and periampullary malignancy. METHODS: Data were collected prospectively on patients having MSCT and EUS for suspected pancreatic and periampullary malignancy. RESULTS: Eighty-four patients had MSCT and EUS, of whom 35 underwent operative assessment (29 resections). In assessing malignancy, there was no significant difference between MSCT and EUS, and agreement was good (82 per cent, kappa = 0.49); the sensitivity and specificity of MSCT were 97 and 87 per cent, compared with 95 and 52 per cent respectively for EUS (P = 0.264). For portal vein/superior mesenteric vein invasion, MSCT was superior (P = 0.017) and agreement was moderate (72 per cent, kappa = 0.42); the sensitivity and specificity were 88 and 92 per cent for MSCT, and 50 and 83 per cent for EUS. For resectability, there was no significant difference and agreement was good (78 per cent, kappa = 0.51). EUS had an impact on the management of 14 patients in whom MSCT suggested benign disease or equivocal resectability. CONCLUSION: MSCT is the imaging method of choice for pancreatic and periampullary tumours. Routine EUS should be reserved for those with borderline resectability on MSCT.

Complications of loop ileostomy closure: a retrospective cohort analysis of 123 patients.

BACKGROUND: Loop ileostomies are often formed in order to defunction distal anastomoses. The aim of this study was to review the complications following closure of loop ileostomies. METHODS: This is a retrospective case note analysis of all loop ileostomy closures performed in the Northumbria Healthcare NHS Trust (population over 500,000) over a 5-year period between 2001 and 2005. RESULTS: A total of 123 case records were reviewed. Complications occurred in 41 patients (33.3%), with 9 patients (7.3%) requiring further intervention. There were 4 (3.3%) postoperative deaths. Complications were more common in patients with increased comorbidity (p = 0.0007) and postoperative death was more frequent among the elderly (p = 0.0006). Postoperative death was more common in those patients who had their stomas created during surgery (elective or emergency) for diverticular disease (3 patients, p = 0.006). Patients with diverticular disease had significantly higher comorbidity and peritoneal contamination at the time of primary surgery. Ileostomy reversal after anterior resection for cancer was associated with a lower complication rate than the rest of the cohort (26%, p = 0.0003) but there was no significant difference in mortality. Neither the grade of the surgeon, the case volume, or the anastomotic technique affected postoperative morbidity. Reoperation was more common in patients whose closure procedure took less time (p = 0.002) and in those who had a shorter wait from creation to reversal of the stoma (p < 0.0001). CONCLUSIONS: Reversal of loop ileostomy may be associated with significant morbidity and mortality. Increasing the delay from creation to closure may result in fewer complications.There is an increased risk in older patients with more comorbidity, particularly when the primary procedure is for diverticular disease with significant peritoneal contamination.

Increase in serum bilirubin levels in obstructive jaundice secondary to pancreatic and periampullary malignancy--implications for timing of resectional surgery and use of biliary drainage.

BACKGROUND: Routine preoperative biliary drainage in cases of jaundice secondary to pancreatobiliary malignancy is associated with a significant risk of complications, failure and stent occlusion. It may be possible to avoid biliary drainage in those patients who are not deeply jaundiced. AIMS: To measure presenting serum bilirubin and its rate of increase in patients with malignant obstructive jaundice. To predict the urgency with which surgery should be performed to avoid preoperative biliary drainage. PATIENTS AND METHODS: Prospective data collection for all pancreatic and periampullary malignancies over a period of 18 months was carried out. Serum bilirubin levels before successful drainage were recorded. Rates of increase in bilirubin and the number of days for bilirubin to reach different thresholds were calculated. RESULTS: Of 111 patients, 66 (59%) had resectable disease on imaging investigations. Median serum bilirubin on presentation was 160 micromol/l. Median increase was 13.1 micromol/l/day or approximately 100 micromol/l/week. The predicted number of days for bilirubin levels to reach a variety of thresholds varied significantly. For a patient presenting with a serum bilirubin of 160 micromol/l, the mean number of days for it to rise to 200 micromol/l, 300 micromol/l, 400 micromol/l and 500 micromol/l was 3, 13, 22 and 31 days, respectively. CONCLUSIONS: There is a variable window of opportunity in jaundiced patients with pancreatic and periampullary malignancy during which surgery may be performed to avoid biliary drainage procedures, depending on the threshold for operating on the jaundiced patient.

Management of hilar cholangiocarcinoma in the North of England: pathology, treatment, and outcome.

AIM: To assess the management and outcome of hilar cholangiocarcinoma (Klatskin tumor) in a single tertiary referral center. METHODS: The notes of all patients with a diagnosis of hilar cholangiocarcinoma referred to our unit for over an 8-year period were identified and retrospectively reviewed. Presentation, management and outcome were assessed. RESULTS: Seventy-five patients were identified. The median age was 64 years (range 34-84 years). Male to female ratio was 1:1. Eighty-nine percent of patients presented with jaundice. Most patients referred were under Bismuth classification 3a, 3b or 4. Seventy patients required biliary drainage, 65 patients required 152 percutaneous drainage procedures, and 25 had other complications. Forty-one patients had 51 endoscopic drainage procedures performed (15 failed). Of these, 36 subsequently required percutaneous drainage. The median number of drainage procedures for all patients was three, 18 patients underwent resection (24%), nine had major complications and three died post-operatively. The 5-year survival rate was 4.2% for all patients, 21% for resected patients and 0% for those who did not undergo resection (P = 0.0021). The median number of admissions after diagnosis in resected patients was two and three in non-resected patients (P<0.05). Twelve patients had external-beam radiotherapy, seven brachytherapy, and eight chemotherapy. There was no significant benefit in terms of survival (P = 0.46) or hospital admissions. CONCLUSION: Resection increases survival but carries the risk of significant morbidity and mortality. Percutaneous biliary drainage is almost always necessary and endoscopic drainage should be avoided if possible.

An ethanologenic yeast exhibiting unusual metabolism in the fermentation of lignocellulosic hexose sugars.

Three lignocellulosic substrate mixtures [liquid fraction of acid-catalyzed steam-exploded softwood, softwood spent sulfite liquor (SSL) and hardwood SSL] were separately fermented by the industrially employed SSL-adapted strain Tembec T1 and a natural galactose-assimilating isolate (Y-1528) of Saccharomyces cerevisiae to compare fermentative efficacy. Both strains were confirmed as S. cerevisiae via molecular genotyping. The performance of strain Y-1528 exceeded that of Tembec T1 on all three substrate mixtures, with complete hexose sugar consumption ranging from 10 to 18 h for Y-1528, vs 24 to 28 h for T1. Furthermore, Y-1528 consumed galactose prior to glucose and mannose, in contrast to Tembec T1, which exhibited catabolite repression of galactose metabolism. Ethanol yields were comparable regardless of the substrate utilized. Strains T1 and Y-1528 were also combined in mixed culture to determine the effects of integrating their distinct metabolic capabilities during defined hexose sugar and SSL fermentations. Sugar consumption in the defined mixture was accelerated, with complete exhaustion of hexose sugars occurring in just over 6 h. Galactose was consumed first, followed by glucose and mannose. Ethanol yields were slightly reduced relative to pure cultures of Y-1528, but normal growth kinetics was not impeded. Sugar consumption in the SSLs was also accelerated, with complete utilization of softwood- and hardwood-derived hexose sugars occurring in 6 and 8 h, respectively. Catabolite repression was absent in both SSL fermentations.

Cellular machinery of wood production: differentiation of secondary xylem in Pinus contorta var. latifolia.

The objectives of this study were to define cell structure during pine secondary xylem development and to integrate this information with current knowledge of the biochemistry and physiology of secondary cell wall biosynthesis in gymnosperms. Lodgepole pine (Pinus contorta var. latifolia Englem.) cambium and secondary xylem were cryofixed using high pressure freezing and freeze-substitution which allowed excellent preservation of the cell structure of developing secondary xylem and enabled high-resolution transmission electron microscopic viewing of these cells for the first time. In contrast to their precursors in the adjacent cambial zone, developing tracheids were active in secondary wall deposition, with abundant cortical microtubules and developing bordered pits. These cells were also characterized by unusual Golgi structures: the trans-Golgi network was highly developed and the associated vesicles were large and darkly stained. These unusual Golgi structures persisted throughout the period of xylem maturation until programmed cell death occurred. Immuno-cytochemistry and enzyme-gold probes were used to investigate the distribution of key secretory products (mannans) and a lignification-associated enzyme (coniferin beta-glucosidase) during xylogenesis. Mannans were localized to the secondary cell wall, the trans-Golgi cisternae and trans-Golgi network vesicles of developing xylem. Coniferin beta-glucosidase was found only in the secondary cell wall. The cell wall localization of coniferin beta-glucosidase, the enzyme responsible for cleaving glucose from coniferin to generate free coniferyl alcohol, provides a mechanism to de-glucosylate monolignols in muro. A two-step model of lignification of conifer tracheids is proposed. First, Golgi-mediated secretion deposits monolignols into the cell wall, where they polymerize in cell corners and middle lamella. Secondly, cell lysis releases stored, vacuolar monolignol glucosides into the wall where they are deglucosylated and their polymerization is influenced by the wall environment including the lignin deposited earlier.

Degradation of trilinolein by laccase enzymes.

Laccase enzymes were investigated for their potential to catalyze the oxidation of trilinolein and methyl linoleate. This study demonstrates that laccase enzymes can oxidize unsaturated fatty acid esters and their associated lipids. The reaction products resulting from laccase-catalyzed reactions with trilinolein were analyzed using combined reversed-phase high-performance liquid chromatography and mass spectrometry via an atmospheric pressure chemical ionization source. The dominant oxidation products detected were monohydroperoxides, bishydroperoxides, and epoxides. This paper presents the first detailed investigation into the interaction between laccase enzymes and lipids containing unsaturated fatty acids.

The influence of bark on the fermentation of Douglas-fir whitewood pre-hydrolysates.

Douglas-fir ( Pseudotsuga menziesii) whitewood was supplemented with increasing concentrations of bark (0-30%) and was pretreated using SO(2)-catalysed steam explosion. The presence of bark in the feedstock resulted in the decreased recovery of total sugars, furfural and 5-hydroxymethylfurfural in the resultant pre-hydrolysate. No detrimental impact on monomer sugar recovery was observed. The concentration of lipophilic extractives present in the pre-hydrolysate increased with increasing bark loading, to a maximum of 0.43 g x l(-1). The water-soluble pre-hydrolysates were fermented by Saccharomyces cerevisiae to determine the impact of bark on sugar consumption and ethanol production. Despite the inclusion of bark, fermentation of all pre-hydrolysates resulted in the complete consumption of hexose sugars within 48 h. Ethanol yields were greater than 0.43 g x g(-1) for all pre-hydrolysates regardless of bark content, indicating that, up to a content of 30%, bark had a negligible impact on the fermentation of the pre-hydrolysates to ethanol.

Do enzymatic hydrolyzability and Simons' stain reflect the changes in the accessibility of lignocellulosic substrates to cellulase enzymes?

In an attempt to elucidate the impact of substrate accessibility to cellulases on the susceptibility of lignocellulosic substrates to enzymatic hydrolysis, a hydrogen peroxide treated, Douglas fir kraft pulp was dried using several methods with varying levels of intensity. Oven-drying at 50 and 100 degrees C, air-drying, and freeze-drying methods were employed to remove the interfibrillar water from the pulp samples. Subsequently, the never-dried and variably dried pulps were hydrolyzed using a commercial cellulase preparation supplemented with additional beta-glucosidase. Drying reduced the susceptibility of the substrates to enzymatic hydrolysis, which can be attributed to the hornifying effect that drying has on fibers. This effect was more pronounced for the fibers that were oven-dried at 100 degrees C (23% reduction) and 50 degrees C (15% reduction), and there was a good correlation between the Simons's stain results and the enzymatic digestibility of the dried pulps. These observations indicated that drying significantly reduced the population of larger pores and that the partial closure of larger pores created a large number of smaller pores that were not accessible to the displacement dye molecules (orange dye). The inaccessibility of the cellulose to the enzymes, due to the collapse or closure of the large pores, appears to be the primary reason for the lower susceptibility of the dried pulps to enzymatic hydrolysis.

Enzymatic treatment of mechanical pulp fibers for improving papermaking properties.

Three enzyme preparations (crude cellulase, laccase, and proteinase) were evaluated for their potential to improve the papermaking properties of mechanical pulp. After treating a long fibre-rich fraction of the pulp with enzyme, the fibres were recombined with untreated fines for handsheet making and testing. None of the enzymes altered the retention of fines or the consolidation of the furnish mix during handsheet formation. All three enzymes increased tensile stiffness index, which is a measure of the initial resistance of the handsheets to strain. Only the laccase preparation, an enzyme that modifies pulp lignin, consistently increased fibre bonding to enhance other strength properties of the handsheets.

Analysis of molecular size distributions of cellulose molecules during hydrolysis of cellulose by recombinant Cellulomonas fimi beta-1,4-glucanases.

Four beta-1,4-glucanases (cellulases) of the cellulolytic bacterium Cellulomonas fimi were purified from Escherichia coli cells transformed with recombinant plasmids. Previous analyses using soluble substrates had suggested that CenA and CenC were endoglucanases while CbhA and CbhB resembled the exo-acting cellobiohydrolases produced by cellulolytic fungi. Analysis of molecular size distributions during cellulose hydrolysis by the individual enzymes confirmed these preliminary findings and provided further evidence that endoglucanase CenC has a more processive hydrolytic activity than CenA. The significant differences between the size distributions obtained during hydrolysis of bacterial microcrystalline cellulose and acid-swollen cellulose can be explained in terms of the accessibility of beta-1,4-glucan chains to enzyme attack. Endoglucanases and cellobiohydrolases were much more easily distinguished when the acid-swollen substrate was used.

Characterization and cloning of a 37.6-kb plasmid carried by Legionella pneumophila recovered from patients and hospital water over a 12-year period.

For 12 years, strains of Legionella pneumophila serogroup 1 harbouring a 37.6-kb (23 MDa) plasmid have predominated among patient and potable water isolates at the Victoria General Hospital, Halifax, N.S. Plasmid DNA recovered from 24 strains isolated between 1983 and 1995 was digested with the restriction endonucleases EcoRI, HindIII, KpnI, PvuII, XbaI, and BamHI. The distribution of cutting sites indicated that the 23-MDa size group had remained essentially unchanged during this period, suggesting the persistence of a single plasmid type. Further fragmentation pattern analysis permitted the construction of a physical map of the prototype 23-MDA plasmid, pLp4269. Double digestion with BamHI-HindIII enabled the cloning of 94.4% of pLp4269 into pBluescript vector. A 2.1-kb fragment was not clonable. Plasmid pLp4269 is the first of the smaller Legionella extrachromosomal DNAs to be characterized in this way.

Cellobiose dehydrogenase, an active agent in cellulose depolymerization.

The ability of cellobiose dehydrogenase purified from Phanerochaete chrysosporium to modify a Douglas fir kraft pulp was assessed. Although the addition of cellobiose dehydrogenase alone had little effect, supplementation with cellobiose and iron resulted in a substantial reduction in the degree of polymerization of the pulp cellulose. When the reaction was monitored over time, a progressive depolymerization of the cellulose was apparent with the concomitant production of cellobiono-1,5-lactone. Analysis of the reaction filtrates indicated that glucose and arabinose were the only neutral sugars generated. These sugars are derived from the degradation of the cellobiose rather than resulting from modifications of the pulp. These results suggest that the action of cellobiose dehydrogenase results in the generation of hydroxyl radicals via Fenton's chemistry which subsequently results in the depolymerization of cellulose. This appears to be the mechanism whereby a substantial reduction in the degree of polymerization of the cellulose can be achieved without a significant release of sugar.