Role of Rhizobium endoglucanase CelC2 in cellulose biosynthesis and biofilm formation on plant roots and abiotic surfaces.

BACKGROUND: The synthesis of cellulose is among the most important but poorly understood biochemical processes, especially in bacteria, due to its complexity and high degree of regulation. In this study, we analyzed both the production of cellulose by all known members of the Rhizobiaceae and the diversity of Rhizobium celABC operon predicted to be involved in cellulose biosynthesis. We also investigated the involvement in cellulose production and biofilm formation of celC gene encoding an endoglucanase (CelC2) that is required for canonical symbiotic root hair infection by Rhizobium leguminosarum bv. trifolii. RESULTS: ANU843 celC mutants lacking (ANU843ΔC2) or overproducing cellulase (ANU843C2+) produced greatly increased or reduced amounts of external cellulose micro fibrils, respectively. Calcofluor-stained cellulose micro fibrils were considerably longer when formed by ANU843ΔC2 bacteria rather than by the wild-type strain, in correlation with a significant increase in their flocculation in batch culture. In contrast, neither calcofluor-stained extracellular micro fibrils nor flocculation was detectable in ANU843C2+ cells. To clarify the role of cellulose synthesis in Rhizobium cell aggregation and attachment, we analyzed the ability of these mutants to produce biofilms on different surfaces. Alteration of wild-type CelC2 levels resulted in a reduced ability of bacteria to form biofilms both in abiotic surfaces and in planta. CONCLUSIONS: Our results support a key role of the CelC2 cellulase in cellulose biosynthesis by modulating the length of the cellulose fibrils that mediate firm adhesion among Rhizobium bacteria leading to biofilm formation. Rhizobium cellulose is an essential component of the biofilm polysaccharidic matrix architecture and either an excess or a defect of this "building material" seem to collapse the biofilm structure. These results position cellulose hydrolytic enzymes as excellent anti-biofilm candidates.

Chylous ascytes secondary to acute pancreatitis: a case report and review of literature.

Chylous ascites is an uncommon finding which is due to the presence of thoracic or intestinal lymph in the abdominal cavity. It is usually caused by a chronic disruption of the lymphatic system. The present report is one of the rare cases in the literature of chylous ascites secondary to idiopathic acute pancreatitis, which showed a complete resolution with a combination of low fat enteral nutrition with MCT and somatostatin analogs.

Paraesophageal hiatal hernia. Open vs. laparoscopic surgery.

BACKGROUND: paraesophageal hiatal hernia represents 5-10% of hiatal hernias. Its importance is based on the severe complications it may have, including gastric volvulus, and surgical treatment is recommended when a diagnosis is established. MATERIAL AND METHODS: a retrospective study of all patients who underwent surgery for paresophageal hernia between 1985 and 2007. RESULTS: we studied 90 cases, 68 females and 22 males with a median age of 67.6 years (37-96). Forty-five patients reported pyrosis, 34 epigastric postprandial pain, and 15 dysphagia; eight patients were diagnosed with gastric volvulus. Eighty-one patients underwent elective surgery and 9 emergency surgery. Forty-seven cases underwent an open procedure and 43 a laparoscopic one; 5 (11.6%) of them required conversion. The techniques performed were D Or fundoplication in 35 cases, Nissen in 35, Toupet in 14, simple hiatal closure in 2, Narbona in 1, and Lortat-Jakob in 1; in 10 patients a mesh was placed. The complication rate for open procedure was 10.6 and 9.5% for the laparoscopic one (p > 0.05). Median hospital stay was 9.1 days for the open procedure and 3.4 for the laparoscopic one (p < 0.05). As follow-up, we analyzed 84 patients. After a median follow-up of 12 years (1-19), 15 patients were still symptomatic (17.8%), with recurrence in 8 cases (5 required reoperation). The satisfaction rate was 95.5%. CONCLUSION: equivalent results were observed after laparoscopic and open surgery and a significant shorter hospital stay in the laparoscopic one. Therefore, we think that laparoscopic surgery should be considered as the election procedure for paraesophageal hiatal hernia.

Jejunal and bladder perforation: complication of intravesical Nd-YAG laser irradiation of bladder tumour.

Nd-YAG laser coagulation is a therapeutic method for the treatment of refractory superficial bladder cancer. Complications resulting from its use are uncommon. We report a case of jejunal and bladder perforation 24 h after Nd-YAG bladder irradiation of a recurrent bladder tumour.

[Multidisciplinary approach to the treatment of rectal cancer: the benefits of neoadjuvant therapy]. / Abordaje multidisciplinar en el tratamiento del cáncer de recto: eficacia del tratamiento neoadyuvante.

OBJECTIVE: the aim of this study was to assess the impact of neoadjuvant treatment on rectal cancer following involvement of a multidisciplinary team (MDT). MATERIALS AND METHODS: between January 2000 and December 2005, 90 patients with rectal adenocarcinoma were evaluated by a MDT and operated on after receiving neoadjuvant treatment with radiochemotherapy (RTCT) -67% were men and 33% were women, with a mean age of 65.04 years (21-83 years). Surgery was low anterior resection in 50% and abdominoperineal amputation in 42.2%. RESULTS: the rate of complications associated with neoadjuvant treatment was 54.4%, with gastrointestinal complications being most frequent. However, this toxicity was tolerated by most patients. It was severe in two cases (2.2%), leading to chemotherapy discontinuation. A histological analysis of specimens showed a complete pathologic response in 10 cases (11.1%) and a partial response (downstaging of T) in 32 cases (35.6%), hence overall response to neoadjuvant treatment was 46.6%. Postoperative complications included anastomotic leakage in 8.3%, perineal wound complications in 34.2%, and urinary disease in 12.2%. The surgical mortality rate was 0%. Local recurrence occurred in 4.4%, and distant metastases were found in 22.2%. Both overall and disease-free survivals were 80 and 64%, respectively. CONCLUSIONS: neoadjuvant treatment results in low local recurrence rates and optimal survival rates, with no increase in morbidity or mortality. A systematic evaluation by a MDT in the context of a clinical protocol offers better cure rates.

Pancreatic neuroendocrine tumours.

INTRODUCTION: Pancreatic neuroendocrine tumours (PNT) are infrequent epithelial neoplasms associated with a better outcome than pancreatic adenocarcinoma. MATERIALS AND METHODS: We analysed our 22 years of experience in managing PNT. Forty-nine patients (27 women and 22 men) with a mean age of 49 years were studied. There were 28 insulinomas, eight glucagonomas, three gastrinomas, one VIPoma and one carcinoid. Eight patients presented with nonfunctional tumours. Enucleation was performed in 20 patients, distal pancreatectomy in 16, middle pancreatic resection in four, cephalic pancreatoduodenectomy in two and total pancreatoduodenectomy in one. In six patients, the tumour was not resected. RESULTS: Postoperative complication rate was 22%: six pancreatic fistulas, three intra-abdominal collections, one remnant pancreatitis and one pancreatic pseudocyst. There was no mortality. 39 cases showed benign histologic features and ten malignant ones. Symptomatic palliation was achieved in 94% of the cases. Five patients presented recurrences: three liver metastases and two pancreatic recurrences. Actuarial mean survival was 163 months and was longer in insulinomas, in those tumours completely resected and in tumours with benign histological features. CONCLUSION: Conservative surgery of the pancreas is preferred, but aggressive surgery is indicated when the primary tumour can be controlled. Despite of minimising pancreatic resection, there is a high complication rate, mainly pancreatic fistulas, though they can often be conservatively managed. Insulinomas are the PNT with better outcome; those completely resected also associate a better prognosis.

[Causes of conversion in laparoscopic surgery for gastroesophageal reflux disease: an analysis of our experience]. / Causas de conversión en la cirugía laparoscópica de la enfermedad por reflujo gastroesofágico: análisis de nuestra experiencia.

BACKGROUND: since its introduction in 1991 laparoscopic antireflux surgery has gained great success and popularity among surgeons, and now it is the gold standard for the treatment of gastroesophageal reflux disease (GERD). AIM: to identify and evaluate the causes of conversion in the laparoscopic surgery of GERD and hiatus hernia. MATERIAL AND METHODS: since January 1993 to August 2007 606 laparoscopic antireflux procedures were performed in our hospital. There were 296 women and 310 men with a median age of 53.5 years. The main indication for surgery was evidence of intractable or recurrent GERD symptoms after adequate medical treatment with associated hiatal hernia. The preoperative workup included manometry, pH-metry, oral endoscopy, and barium swallow. The surgical technique was mainly the Nissen-Rossetti procedure. RESULTS: mean postoperative hospital stay was 2.7 days. The operation had to be converted to an open procedure in 43 cases (7%). Conversions were more frequent in the first decade of the learning curve (26 vs. 17, p < 0.016), and fewer among the group of experts in advanced laparoscopic surgery (15 vs. 28, p < 0.017). In 17 cases conversions were due to an intraoperative complication whereas in 26 cases a conversion was done because of technical difficulties. Esophageal perforation and pneumothorax rates were 0.8 and 1%, respectively, and mortality and morbidity rates were 0.1 and 12%. CONCLUSION: the rate of conversion is acceptable and significantly decreases with surgeon experience.

Rhizobium cellulase CelC2 is essential for primary symbiotic infection of legume host roots.

The rhizobia-legume, root-nodule symbiosis provides the most efficient source of biologically fixed ammonia fertilizer for agricultural crops. Its development involves pathways of specificity, infectivity, and effectivity resulting from expressed traits of the bacterium and host plant. A key event of the infection process required for development of this root-nodule symbiosis is a highly localized, complete erosion of the plant cell wall through which the bacterial symbiont penetrates to establish a nitrogen-fixing, intracellular endosymbiotic state within the host. This process of wall degradation must be delicately balanced to avoid lysis and destruction of the host cell. Here, we describe the purification, biochemical characterization, molecular genetic analysis, biological activity, and symbiotic function of a cell-bound bacterial cellulase (CelC2) enzyme from Rhizobium leguminosarum bv. trifolii, the clover-nodulating endosymbiont. The purified enzyme can erode the noncrystalline tip of the white clover host root hair wall, making a localized hole of sufficient size to allow wild-type microsymbiont penetration. This CelC2 enzyme is not active on root hairs of the nonhost legume alfalfa. Microscopy analysis of the symbiotic phenotypes of the ANU843 wild type and CelC2 knockout mutant derivative revealed that this enzyme fulfils an essential role in the primary infection process required for development of the canonical nitrogen-fixing R. leguminosarum bv. trifolii-white clover symbiosis.

Pseudomyxoma peritonei secondary to epithelial appendicular neoplasms. Experience in a non-specialised centre.

INTRODUCTION: Pseudomyxoma peritonei is an infrequent entity, defined by collections of gelatinous material in the abdomen and pelvis and mucinous implants on peritoneum, secondary to the rupture of a mucinous lesion, usually of ovarian or appendiceal origin. MATERIALS AND METHODS: We present our experience of 11 cases (6 males and 5 females) diagnosed with pseudomyxoma peritonei secondary to epithelial appendicular neoplasms over 27 years. The mean age of the patients was 68 years. Clinical manifestations were abdominal distension (55%), right lower quadrant pain (45%) suggesting acute appendicitis and constitutional syndrome (36%). An abdominal mass was detected at physical examination in 4 patients. CT scan revealed a tumour in right iliac fossa in 4 patients, peritoneal enlargement in 1 and a liquid collection in 1. Preoperative diagnosis was acute abdomen in 5 patients, peritoneal carcinomatosis in 3 and undetermined abdominal mass in 3. RESULTS: Surgical findings suggested pseudomyxoma peritonei in 8 patients and peritoneal carcinomatosis in 3. Appendicectomy was performed in 9 patients, and in 3 of them bilateral anexectomy was also performed. One patient underwent ileocaecal resection and another a right hemicolectomy. In all the cases, mucinous material was eliminated as much as possible. Pathology revealed mucinous cystoadenoma in 6 cases, mucinous cystoadenocarcinoma in 3 and epithelial hyperplasia in 2 patients. Median survival was 54 months, with a 5- year survival rate of 40%. The last case we treated was sent to a reference centre for the treatment of pseudomyxoma peritonei. CONCLUSIONS: There is no consensus on the best treatment for pseudomyxoma peritonei. We recommend avoiding incomplete surgical resections in non-reference centres and submitting patients to a reference centre to undergo adequate treatment.

Peritoneal carcinomatosis secondary to carcinoid tumour.

Carcinoid tumours are neuroendocrine neoplasms that can appear in every location of the digestive tract. They are low aggressive tumours, although they often produce local invasion and hepatic metastases, whose resection allows long-term survival. We report a case of a 64-year-old man with ileal carcinoid tumour, that underwent ileal resection and metastasectomy of one lesion in liver segment II. Surgical findings indicated peritoneal carcinomatosis. Carcinoid dissemination as peritoneal carcinomatosis has been rarely described in the literature. Cytoreductive surgery, always when complete resection is aimed, achieves asymptomatic long-term survivals.

[Diagnosis and treatment of pancreatic insulinoma]. / Manejo diagnóstico y terapéutico del insulinoma pancreático.

INTRODUCTION: insulinoma is the most frequent pancreatic endocrine tumor. Its preoperative diagnosis has been controversial for many years. The aim of this study was to evaluate the experience in the management and treatment of this kind of tumor at Hospital Ramón y Cajal. MATERIAL AND METHODS: between January 1999 and July 2006, 12 patients were operated on in our hospital (9 females and 3 males) who had been diagnosed with insulinoma, with a mean age of 56 years (16-72 years). RESULTS: octreotide scintigraphy allowed a diagnosis in 33.3% of cases, abdominal CT in 83.3%, and echoendoscopy in 100%. Intraoperative ultrasonography confirmed the presence of an insulinoma in 100% of cases. In all cases a complete excision was possible, with 9 enucleations and 3 distal pancreatectomies. Two patients developed a pancreatic fistula, and one a pancreatic pseudocyst that healed spontaneously without surgery. The overall cure rate was 100%. After a mean follow-up of 48 months no recurrences have been reported. CONCLUSION: in our experience, we consider advisable that abdominal CT and echoendoscopy be performed before surgery. However, the initial procedure of choice would be pancreas palpation and intraoperative ultrasonography. Surgery cured 100% of cases, and the procedure selected depends on size, location, distance from the main pancreatic duct, and relation to multiple endocrine neoplasm 1 (MEN-1).

Strains of Mesorhizobium amorphae and Mesorhizobium tianshanense, carrying symbiotic genes of common chickpea endosymbiotic species, constitute a novel biovar (ciceri) capable of nodulating Cicer arietinum.

AIMS: To identify several strains of Mesorhizobium amorphae and Mesorhizobium tianshanense nodulating Cicer arietinum in Spain and Portugal, and to study the symbiotic genes carried by these strains. METHODS AND RESULTS: The sequences of 16S-23S intergenic spacer (ITS), 16S rRNA gene and symbiotic genes nodC and nifH were analysed. According to their 16S rRNA gene and ITS sequences, the strains from this study were identified as M. amorphae and M. tianshanense. The type strains of these species were isolated in China from Glycyrrhiza pallidiflora and Amorpha fruticosa nodules, respectively, and are not capable of nodulating chickpea. These strains carry symbiotic genes, phylogenetically divergent from those of the chickpea isolates, whose nodC and nifH genes showed more than 99% similarity with respect to those from Mesorhizobium ciceri and Mesorhizobium mediterraneum, the two common chickpea nodulating species in Spain and Portugal. CONCLUSIONS: The results from this study showed that different symbiotic genes have been acquired by strains from the same species during their coevolution with different legumes in distinct geographical locations. SIGNIFICANCE AND IMPACT OF THE STUDY: A new infrasubspecific division named biovar ciceri is proposed within M. amorphae and M. tianshanense to include the strains able to effectively nodulate Cicer arietinum.

Characterization of xylanolytic bacteria present in the bract phyllosphere of the date palm Phoenix dactylifera.

AIMS: Despite the interest of phyllosphere microbiology, no studies have addressed the bacteria present in bract phyllosphere, an ecosystem that has special characteristics in palm trees because the dry bracts remain on the plant until pruning and may contain polymer-degrading bacteria involved in plant degradation. Therefore, the aim of this work was to characterize xylanolytic bacteria isolated from palm bract phyllosphere. METHODS AND RESULTS: Twelve xylanolytic strains were isolated and characterized by phenotypic features and complete sequencing of 16S rRNA gene. The results showed that the isolates were phenotypically and genotypically diverse. Gram-positive isolates were classified into genus Paenibacillus some of them belonging to hitherto undescribed species of this genus. Gram-negative isolates were classified into genera Pseudomonas and Acinetobacter. CONCLUSIONS: The results of this work confirm the complexity of the bacterial populations present in phyllospheric ecosystems and suggest that bacteria involved in plant degradation are present at the early degradation steps of this process in dry palm tree bracts. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first study on bract phyllospheric bacteria able to hydrolyse vegetal polymers and offers a new perspective in the search of unexplored sources of xylanase-producing strains.

Evolution of the therapeutic approach of visceral artery aneurysms.

INTRODUCTION: Visceral artery aneurysms are an infrequent entity. Most of them are asymptomatic, but when they present as a rupture, a high mortality is associated. MATERIAL AND METHODS: We review our experience of 18 cases between 1988 and 2006. RESULTS: 9 males and 9 females with a mean age of 66,5 years are analyzed. Aneurysms were located in splenic artery (9), hepatic artery (2), superior mesenteric artery (2), celiac trunk (3), inferior mesenteric artery (1) and gastroduodenal artery (1). Three of them were associated with abdominal aorta aneurysms. Fourteen patients were asymptomatic, three presented abdominal pain and one case presented with rupture and intraperitoneal bleeding. Surgical treatment was performed in thirteen of the patients and endovascular in five. Two cases of endovascular treatment failed and surgery was necessary. Postsurgical mortality was 0 and complications appeared in 2 patients. Mean hospital stay after surgical treatment was 11 days and 3 days after endovascular one. None of the patients presented recurrences, and one has chronic mesenteric ischemia as sequelae of surgical treatment of a superior mesenteric artery aneurysm. CONCLUSION: Visceral artery aneurysms must be treated if it is feasible, due to the potential risk of rupture. Endovascular treatment associates lower morbimortality rates and shorter hospital stay than surgical one, but nowadays many aneurysms are not suitable for this management. Endovascular treatment is a technically difficult approach, that requires a specific training and the first cases represent a learning curve.

Cellulomonas xylanilytica sp. nov., a cellulolytic and xylanolytic bacterium isolated from a decayed elm tree.

A Gram-positive, aerobic, non-motile bacterium was isolated from a decayed elm tree. Phylogenetic analysis based on 16S rDNA sequences revealed 99.0 % similarity to Cellulomonas humilata. Chemotaxonomic data that were determined for this isolate included cell-wall composition, fatty acid profiles and polar lipids; the results supported the placement of strain XIL11(T) in the genus Cellulomonas. The DNA G+C content was 73 mol%. The results of DNA-DNA hybridization with C. humilata ATCC 25174(T), in combination with chemotaxonomic and physiological data, demonstrated that isolate XIL11(T) should be classified as a novel Cellulomonas species. The name Cellulomonas xylanilytica sp. nov. is proposed, with strain XIL11(T) (=LMG 21723(T)=CECT 5729(T)) as the type strain.

Xylanibacterium ulmi gen. nov., sp. nov., a novel xylanolytic member of the family Promicromonosporaceae.

A bacterial strain designated XIL08(T) was isolated from an elm tree affected by Dutch elm disease. Strain XIL08(T) is Gram-positive, aerobic, rod-shaped and non-motile. The complete 16S rDNA sequence of this micro-organism was obtained and phylogenetic analysis based on the neighbour-joining method indicated that the closest related organism belongs to the genus Xylanimonas of the family Promicromonosporaceae, suborder Micrococcineae. Cell-wall analyses revealed the presence of type A4alpha, L-lys-L-ala-D-Glu peptidoglycan. The cell-wall sugars found were rhamnose in large amounts, fucose, mannose and galactose and traces of arabinose and glucose. HPLC analysis of menaquinones revealed two peaks, the main peak corresponding to MK-9(H(4)) and the smaller one to MK-8(H(4)). The major fatty acid found was anteiso-C(15 : 0). Mycolic acids were absent. The polar lipids detected were phosphatidylglycerol, diphosphatidylglycerol, phosphatidylinositol and phosphatidylinositol mannosides. The G+C content of the DNA was 72 mol%. Isolate XIL08(T) hydrolysed xylan but not cellulose. Growth was observed with many carbohydrates including acetate and xylan as the only carbon source. Catalase activity was not detected. The data from this polyphasic study suggest that this bacterium belongs to a novel genus of the family Promicromonosporaceae. It is proposed that isolate XIL08(T) (=LMG 21721(T)=CECT 5731(T)) be classified in a new genus, Xylanibacterium gen. nov., as the type strain of Xylanibacterium ulmi sp. nov.

[Intraductal papillary mucinous tumors of the pancreas: diagnostic and therapeutic approach]. / Neoplasia papilar intraductal mucinosa de páncreas: actitud diagnóstica y terapéutica.

OBJECTIVES: Literature review of the diagnostic and therapeutic approach to intraductal papillary mucinous tumors of the pancreas and report of a new case. CASE REPORT: We present the case of an 85-year-old man treated in our center for steatorrhea and weight loss. Abdominal computerized tomography (CT) showed a heterogeneous pancreatic mass, affecting the hilus, surrounding the superior mesenteric artery and producing dilatation of the biliary tract. Endoscopic retrograde cholangiopancreatography (ERCP) revealed a polypoid neoformation in the lower part of the duodenum and leakage of mucus through the papilla. Biopsy revealed pancreatic adenoma. CONCLUSION: Intraductal papillary mucinous tumor of the pancreas is potentially curable with an uncertain prognosis. Correct diagnosis is required for optimal treatment. One of the greatest problems in this entity continues to be the difficulty of predicting the presence of an infiltrating component, which significantly worsens prognosis. The various diagnostic methods used are abdominal CT, echoendoscopy, ERCP, endoscopy of the Wirsung duct, intraoperative ultrasonography and study of the intraoperative pancreatic resection margin. Treatment requires complete resection of the lesion, although if this provokes pancreatic failure, only tumors with invasive components should be extirpated.

An effective, rapid and simple method for total RNA extraction from bacteria and yeast.

In this work, we describe a rapid and simple method for total RNA extraction from bacteria and yeast. The method allows for the acquirement of high RNA yields while avoiding the use of phenol or other toxic reagents and is less expensive than other methods previously described. The extracted RNA is suitable for applications such as RT-PCR, Northern blot hybridization and low molecular weight RNA (LMW RNA) electrophoresis.