Taxonomic characterization of Sphaerotilus microaerophilus sp. nov., a sheath-forming microaerophilic bacterium of activated sludge origin.

A microaerophilic Gram-stain-negative bacilliform bacterial strain, FB-5 T, was isolated from activated sludge in Yokohama, Japan, that exhibited filamentous growth and formed a microtube (sheath). Cells were motile using a single polar flagellum. The optimum growth temperature and pH were 30 °C and 7.5, respectively. Strain FB-5 T was catalase-negative. Peptides and amino acids were utilized as energy and carbon sources. Sugars and organic acids were not utilized. Vitamin B12 enhanced the growth of strain FB-5 T. Sulfur-dependent lithotrophic growth was possible. Major respiratory quinone was UQ-8. Major fatty acids were C16:1ω7 and C16:0. The genomic DNA G + C content was 69.16%. Phylogenetic analysis of the 16S rRNA gene suggested that strain FB-5 T belongs to the genus Sphaerotilus. The close relatives were S. natans subsup. sulfidivorans and S. natans subsup. natans with 98.0% and 97.8% similarity based on the 16S rRNA gene analysis, respectively. The genome size (6.06 Mbp) was larger than that (4.39-5.07 Mbp) of the Sphaerotilus strains. The AAI values against the related strains ranged from 71.0 to 72.5%. The range of ANI values was 81.7 - 82.5%. In addition to these distinguishable features of the genome, the core genome and dDDH analyses suggested that this strain is a novel member of the genus Sphaerotilus. Based on its physiological properties and genomic features, strain FB-5 T is considered as a novel species of the genus Sphaerotilus, for which the name S. microaerophilus sp. nov. is proposed. The type strain is FB-5 T (= JCM 35424 T = KACC 23146 T).

Enzymatic degradation of glucosaminoglucan and cellulase resistance of cellulose nanofiber coated with glucosaminoglucan.

AIMS: Enzymatic degradation of ß-1,4-linked glucose and glucosamine (glucosaminoglucan, GG), which is prepared from Thiothrix nivea and can act as a cellulose-aminating agent with a strong affinity to cellulose, was attempted. METHODS AND RESULTS: A chitosanase-secreting fungal strain was isolated as a GG-degrading microbe. GG was found to be degraded by not only chitosanases but also cellulases. Based on nuclear magnetic resonance spectroscopy, both enzymes were found to produce GlcN-Glc from GG. The cellulases also produced GlcN-Glc-GlcN-Glc as an additional final digest. Furthermore, aminated (GG-coated) cellulose nanofibers exhibited cellulase resistance. The flexibility of GG adsorbed onto a cellulose crystal was almost identical to that of cellulose, as estimated via the molecular dynamics calculations. CONCLUSIONS: The chitosanase and cellulase hydrolyzed the ß-1,4-linkage from Glc to GlcN and were expected to recognize the tetramer and hexamer units of GG depending on their final products. The cellulose nanofibers acquired cellulase resistance via amination with GG, probably because of the lower activity of cellulase to GG than cellulose.

A heterodimeric hyaluronate lyase secreted by the activated sludge bacterium Haliscomenobacter hydrossis.

Haliscomenobacter hydrossis is a filamentous bacterium common in activated sludge. The bacterium was found to utilize hyaluronic acid, and hyaluronate lyase activity was detected in its culture. However, no hyaluronate lyase gene was found in the genome, suggesting the bacterium secretes a novel hyaluronate lyase. The purified enzyme exhibited two bands on SDS-PAGE and a single peak on gel filtration chromatography, suggesting a heterodimeric composition. N-terminal amino acid sequence and mass spectrometric analyses suggested that the subunits are molybdopterin-binding and [2Fe-2S]-binding subunits of a xanthine oxidase family protein. The presence of the cofactors was confirmed using spectrometric analysis. Oxidase activity was not detected, revealing that the enzyme is not an oxidase but a hyaluronate lyase. Nuclear magnetic resonance analysis of the enzymatic digest revealed that the enzyme breaks hyaluronic acid to 3-(4-deoxy-ß-d-gluc-4-enuronosyl)-N-acetyl-d-glucosamine. As hyaluronate lyases (EC 4.2.2.1) are monomeric or trimeric, the enzyme is the first heterodimeric hyaluronate lyase.

[Significance and Treatment Strategy of Glasgow Prognostic Score in High Risk Stage â ¡ Colorectal Cancer].

In this study, we investigated the usefulness of Glasgow prognostic score(GPS)as a prognostic factor for Stage Ⅱ colorectal cancer, and the treatment strategy by individualizing adjuvant chemotherapy. We enrolled 86 patients with Stage Ⅱ primary colorectal cancer who underwent curative resection. This study examines the prognostic significance of clinicopathological factors and GPS, NLR, LMR, PLR. Multivariate analyses was performed to evaluate the factors affecting recurrence free survival. The 5-year OS was 92.5%, and the RFS was 86% in Stage Ⅱ colorectal cancer. The recurrence rate was 12.8%. In multivariate analysis, GPS(HR: 13.66, p=0.005)was extracted as an independent poor prognosis factor. In comparison of survival rates, RFS of GPS 0, 1 was 95.2% and that of GPS 2 43.8%, and GPS 2 had a significantly poor prognosis(p< 0.01). GPS 2 is an independent high risk factor for recurrence of Stage Ⅱ colorectal cancer. In order to improve the prognosis of Stage Ⅱ colorectal cancer, individualized adjuvant chemotherapy is important.

Diagnosis and treatment of gastric hamartomatous inverted polyp (GHIP) using a modified combination of laparoscopic and endoscopic approaches to neoplasia with a non-exposure technique (modified CLEAN-NET): a case report.

BACKGROUND: Gastric hamartomatous inverted polyp (GHIP) is a pathological condition where enlarged gastric glands with cystic dilatation grow in the submucosa. It is difficult to excise the tissue due to its location. In addition, even if the tissue is taken correctly, making an accurate diagnosis is difficult due to foveolar epithelium in the tissue, which can be misdiagnosed as gastric mucosal epithelium. Thus, an accurate diagnosis of GHIP is rarely established from a biopsy alone preoperatively. We here report a case of GHIP with a central dimple, which was diagnosed and treated using a modified combination of laparoscopic and endoscopic approaches to neoplasia with a non-exposure technique (modified CLEAN-NET). CASE PRESENTATION: A 60-year-old man with a submucosal tumor (SMT) in the stomach was referred to our hospital by a primary care doctor. On examination, a gastrointestinal stromal tumor was suspected. Modified CLEAN-NET was performed for diagnostic and therapeutic purposes. The histopathological examination of the resected specimen showed an enlarged gland duct in the submucosal layer. This finding, along with immunostaining results, led to the diagnosis of GHIP. The postoperative course was uneventful without any symptoms. CONCLUSIONS: GHIP should be considered among the differential diagnoses of SMT of the stomach. Modified CLEAN-NET may be beneficial in the removal of SMTs such as GHIP with a central dimple because it can avoid stomach deformation of the stomach and tumor dissemination.

Prognostic role of FUT8 expression in relation to p53 status in stage II and III colorectal cancer.

The expression of fucosyltransferase 8, an enzyme responsible for core fucosylation encoded by FUT8, influences tumor biology and correlates with patient prognosis in several solid cancers. We hypothesized that p53 alteration modifies prognostic associations of FUT8 expression in colorectal cancer (CRC), since FUT8 has recently been identified as a direct transcriptional target of wild-type p53. Utilizing multiple datasets of microarray and RNA sequence of CRC, FUT8 mRNA was found to be highly expressed in wild-type p53 tumors (n = 382) compared to those of mutant p53 (n = 437). Prognostic values of FUT8 expression in conjunction with the p53 status for disease-free survival (DFS) were analyzed using two independent cohorts of stage II and III CRC after curative surgery, including the immunohistochemistry (IHC) cohort (n = 123) and the microarray cohort (n = 357). In both cohorts, neither FUT8 expression nor the p53 status was associated with DFS. Strikingly, positive expression of FUT8 protein was significantly associated with better DFS only in tumors with negative p53, while it had no prognostic impact in tumors with positive p53 in the IHC cohort. Although not statistically significant, a similar prognostic trend was observed in the microarray cohort when patients were stratified by the p53 status. Our results suggest that the prognostic values of FUT8 expression on DFS may be modified by the p53 status, and the expression of FUT8 protein can be a prognostic biomarker for patients with stage II and III CRC.

[A Case of Simultaneous Recurrence of Colon Cancer at the Drain Placement and Anastomotic Sites Postoperatively].

A woman in her 70s presented to our hospital with abdominal pain and right lower abdominal mass. Colonoscopy revealed circumferential ascending colon cancer. She underwent right hemicolectomy, D3 lymphadenectomy, and ileocolonic functional end-to-end anastomosis. The tumorwas pathologically diagnosed as T4aN1M0, Stage Ⅲa. Nine months afterthe first surgery, tumor marker levels increased, and detailed examination yielded a diagnosis of isolated recurrence at the site of drain removal in the abdominal wall. The tumor was resected without exposure. Four months after the second surgery, the tumor recurred in the abdominal wall. Furthermore, colonoscopy revealed anastomotic recurrence. Both recurrent tumors at the anastomotic site and in the abdominal wall were resected. No more recurrence has been detected to date. In this case, a possible cause of recurrence is implantation of cancer cells. Sufficient consideration should be given to innovations in intraoperative maneuvers and surgical wound protection, among others.

Immunogenic tumor cell death induced by chemotherapy in patients with breast cancer and esophageal squamous cell carcinoma.

It has been reported that chemo-radiotherapy can induce immunogenic tumor cell death (ICD), which triggers T-cell immunity mainly mediated by high-mobility group box 1 protein (HMGB1) and calreticulin. However, there is still limited information to support this theory relating to chemotherapy alone. In the present study, the expression of HMGB1 and calreticulin was evaluated by immunohistochemistry in pre-treatment biopsy specimens and surgically resected specimens, which were obtained from patients with breast cancer (n=52) and esophageal squamous cell carcinoma (ESCC) (n=8) who had been treated with neoadjuvant chemotherapy (NAC). We also analyzed HMGB1 and calreticulin expression in breast cancer cell lines treated with chemotherapeutic drugs. As a result, both HMGB1 and calreticulin expression levels were significantly upregulated after NAC in both breast cancer and ESCC tissues. However, no significant correlation was observed between HMGB1 expression and pathological response after NAC or between HMGB1 expression and patient survival. Furthermore, although overall survival in the high infiltration group of CD8-positive T cells was significantly superior to that in the low infiltration group in breast cancer patients, there were no correlations between the number of CD8-positive T cells and HMGB1 or calreticulin expression levels. In addition, chemotherapeutic drugs induced upregulation of HMGB1 and calreticulin in all tested cell lines. Our findings indicate that chemotherapy alone can significantly induce ICD regardless of the degree of pathological response after chemotherapy.

Association between circulating galectin-3 levels and the immunological, inflammatory and nutritional parameters in patients with colorectal cancer.

Galectin-3, a ß-galactoside-binding lectin, exhibits pleiotropic biological functions and has a role as one of the immunological modulators. However, the associations between circulating galectin-3 and immunological, inflammatory and nutritional parameters have not yet been fully elucidated. The serum concentration of galectin-3 was examined in association with interleukin-10 (IL-10), IL-12 and IL17 production, lymphocyte stimulation, neutrophil/lymphocyte ratio (NLR), white blood cell count (WBC), C-reactive protein (CRP) and rapid turnover proteins, including retinol-binding protein (RBP), prealbumin (PA) and transferrin (TF) in 50 patients with untreated colorectal cancers. Significant increases (P<0.05) were observed in the serum galectin-3 levels in patients with untreated colorectal cancer (9.6±4.5 ng/ml) compared with the normal controls (3.2±1.6 ng/ml). Higher serum galectin-3 concentrations were observed in patients with colon cancer (11.5±4.4 ng/ml) compared to in patients with rectal cancer (8.0±4.0 ng/ml) (P=0.005). The levels of circulating galectin-3 inversely correlated with the production of IL-10 (r=-0.59, P<0.001), and IL-12 (r=-0.69, P<0.001). Galectin-3 concentration also inversely correlated with the lymphocyte stimulation assay stimulation index (r=-0.42, P=0.021). However, the level of serum galectin-3 correlated with IL-17 production (r=0.67, P<0.001). Serum galectin-3 levels exhibited significant correlations with NLR (r=0.41, P=0.009), WBC (r=0.32, P=0.035), and CRP (r=0.63, P<0.001), and statistically significant inverse correlations with RBP (r=-0.45, P=0.002), PA (r=-0.46, P=0.001) and TF (r=-0.72, P<0.001). Galectin-3 may be one of the key factors in the regulation of immunological, inflammatory and nutritional conditions.

Stromal VCAN expression as a potential prognostic biomarker for disease recurrence in stage II-III colon cancer.

To develop prognostic biomarkers that can discriminate stage II-III colorectal cancer patients with high risk of postoperative recurrence, we conducted a genome-wide screening of relapse-related genes utilizing multiple microarray cohorts. Among differentially expressed genes between tumor and nontumor, we identified eight candidate genes associated with relapse in two datasets of stage II-III patients (n = 94 and 145, respectively, P < 0.05). Using datasets of laser-microdissected samples and FACS-purified cell populations, the localization of candidate genes, including COL4A2, COL4A1, VCAN and SERPINE1, were found predominantly in cancer stroma rather than epithelial components. Among those relapse-related stromal genes, VCAN mRNA, specifically expressed in cancer-associated fibroblasts, was further validated to be a prognostic factor in two additional independent datasets, consisting of 453 (P = 0.0334) and 89 (P = 0.0041) stage II-III patients. Furthermore, in our large set of formalin-fixed paraffin-embedded cohort (n = 338), VCAN protein was detected exclusively in cancer stroma by immunohistochemistry, demonstrating a stepwise increase of stromal VCAN from normal tissues through stage 0 to stage IV tumors. Stromal VCAN protein was associated with shorter relapse-free survival (RFS) in stage II-III colon cancer, independent of other clinical factors by multivariate analysis (P = 0.004). Stratified analyses revealed that stromal VCAN was a strong prognostic indicator particularly in stage II colon cancer (P = 0.0029). In all five analyzed cohorts, the expression of VCAN, in transcript or protein levels, was associated with poor RFS in stage II-III patients. We conclude that VCAN is a promising biomarker to identify stage II-III patients at high risk of relapse who may benefit from intensive postoperative management.

[A Case of Port Site Recurrence after Laparoscopic Distal Gastrectomy for Advanced Gastric Cancer].

It has been suggested that port site recurrence is a potential complication after laparoscopic distal gastrectomy for gastric cancer, especially considering the increased number of laparoscopic surgeries being performed. We encountered a case of an 84-year-old man who was diagnosed with 2 port site recurrences at the navel and right hypochondrium after laparoscopic distal gastrectomy(D2). Pathological diagnosis for the original tumor was tub2, pT4a, pN1(1/38), M0, pStage III A, and HER2(0). As first-line chemotherapy with S-1 plus CDDP for the port site recurrence failed, second-line chemotherapy with ramucirumab plus paclitaxel(RAM plus PTX)was administered. Although RAM plus PTX therapy induced shrinkage of the port site recurrence, liver metastasis was detected as a new lesion. RAM mono-therapy maintained good QOL for 18 months after surgery.

[Risk Evaluation and Prognostic Prediction of Colorectal Cancer in Elderly Patients Over 80 Years of Age].

This study aimed to investigate risk factors and prognosis ofcolorectal cancer in patients over 80 years ofage. Surgical risk was evaluated by colorectal POSSUM(CR-POSSUM)and prognosis by Glasgow prognostic score(GPS). The analysis included 56 patients aged over 80 years with colorectal cancer during 2002-2012. Mean operation time, blood loss, and period ofhospitalization were 130 min, 111 mL, and 19.9 days, respectively. Postoperative complications occurred in 26 patients (46.4%; complications group). The 5-year overall survival rate for patients with complication scores above 2 was 51.1%, compared to 82.3% in a control group, and patients in the complication group also exhibited a poorer prognosis. CR-POS SUM scores were significantly higher in the complication group than in the control group in PS, OS, and PMR. Further analysis revealed that patients with GPS 0 or 1 had a significantly higher 5-year survival rate(84.9%)than those with GPS2(38.9%, p <0.05).

[A Case of Gastrointestinal Perforation in a Patient with Colon Cancer and Stent Placement during Bevacizumab Treatment, Alleviated with Conservative Therapy].

The case involved a 52-year-old man. He was diagnosed with sigmoid colon cancer and multiple liver and lung metastases. After placement of a metallic stent for circumferential stricture, 5-fluorouracil/Leucovorin/oxaliplatin (FOLFOX4) plus bevacizumab therapy was introduced. After 17 courses, gastrointestinal perforation was noted at the site of stent placement, but this was alleviated with conservative therapy. Although gastrointestinal perforation is known to be a serious complication that occurs during chemotherapy combined with bevacizumab, further studies are needed to investigate the incidence of gastrointestinal perforation after stent placement more fully.

[A case of metastatic liver carcinoma treated with radiofrequency ablation and hepatectomy].

We report a case of metastatic liver carcinoma treated by radiofrequency ablation (RFA) and hepatectomy. A 53-year-old man suffering from sigmoid colon carcinoma and unresectable multiple liver metastases was treated with capecitabine and oxaliplatin and bevacizumab. After the seventh course, tumor reduction was confirmed but liver dysfunction meant that it was difficult to continue the same regimen. Because preoperative evaluation of liver function showed a high risk of postoperative liver failure, a combination of partial hepatectomy and RFA was planned in order to reduce the amount of liver resection. High anterior resection, partial hepatectomy and RFA were performed as planned. He underwent S-1 therapy following 7 courses of irinotecan, S-1, and bevacizumab therapy. To date, no recurrence has been observed 18 months after the operation. A combination of hepatectomy and RFA is expected to be an effective local treatment for multiple liver metastases of colorectal cancer, although the evidence is currently insufficient.