Comparison of species, virulence genes and clones of Aeromonas isolates from clinical specimens and well water in Okinawa Prefecture, Japan.

AIMS: To reveal the sources of Aeromonas infection in Okinawa Prefecture of Japan, the species, virulence genes and clones of strains isolated from clinical specimens and well water were compared. METHODS AND RESULTS: The properties of both isolates were investigated by sequencing of rpoD, detection of 10 virulence genes using PCR and genotyping with pulsed-field gel electrophoresis. In all, 68 clinical and 146 well water strains of Aeromonas were isolated and the main species were A. caviae, A. dhakensis, A. hydrophila and A. veronii biovar sobria. Aeromonas dhakensis possessed various virulence genes; however, A. caviae possessed only fla. The same or similar clones were distributed in certain areas of Okinawa and one clone had survived several months in the biliary system of two patients, respectively. CONCLUSION: Although the same Aeromonas clone was not isolated from clinical and well water samples, our study revealed the detected patterns of virulence genes in both isolates, the distribution of identical/similar clones in the Okinawan environment and long-time survival in patient's organs. SIGNIFICANCE AND IMPACT OF THE STUDY: We investigated the association between Aeromonas patients and well water exposure. This study provides the properties of species, virulence genes and clones of Aeromonas isolated from samples of these origins.

Serum cytokeratin 19 fragment 21-1 and carcinoembryonic antigen combination assay as a biomarker of tumour progression and treatment response in extramammary Paget disease.

BACKGROUND: Extramammary Paget disease (EMPD) is a rare intraepithelial adenocarcinoma affecting the genitals and axillary regions. As metastasis of these tumours is itself rare, solid disease management strategies have not been established. Serum carcinoembryonic antigen (CEA) and cytokeratin 19 fragment 21-1 (CYFRA 21-1) levels have been identified as candidate biomarkers for tumour progression in EMPD. OBJECTIVES: To determine the accuracy of and the correlation between these markers in patients with EMPD. METHODS: Serum CEA and CYFRA 21-1 levels were examined in 30 patients with EMPD treated at Keio University Hospital, and compared against clinical information. Both assays were performed at the time of diagnosis, during the postoperative observation period, and following systemic treatment in those with confirmed metastasis. Serum levels were then correlated with tumour progression status and treatment responses. RESULTS: Normal levels for both assays were observed in all 11 patients with primary localized disease (100%). In patients with metastatic disease the CEA positivity rate was 79% (15 of 19 patients), with a rate of 63% (12 of 19 patients) for CYFRA 21-1. Changes in CEA and CYFRA 21-1 levels were statistically independent; however, using a combined view, elevated levels of either marker improved the positivity rate to 95% (18 of 19 patients). Use of both markers also correlated well with treatment responses. CONCLUSIONS: The combination of CEA and CYFRA 21-1 is useful for predicting metastasis and treatment response in patients with EMPD, especially in those who only have elevation of a single marker. What's already known about this topic? Serum levels of carcinoembryonic antigen (CEA) and cytokine 19 fragment 21-1 (CYFRA 21-1) have been shown to be elevated in patients with extramammary Paget disease (EMPD). Elevation of serum CEA levels is associated with tumour progression of EMPD. A single small study reported that serum CYFRA 21-1 levels are elevated in patients with EMPD with lymph node metastasis. What does this study add? Serum CEA and CYFRA 21-1 were present in 79% and 63% of 19 cases of metastatic EMPD, respectively. Elevations of CEA and CYFRA 21-1 were statistically independent. CEA and CYFRA 21-1 combination assays were positive in 95% of cases of metastatic EMPD. What is the translational message? Combination assays with CEA and CYFRA 21-1 are useful for monitoring treatment response in patients with metastatic EMPD, particularly in those with elevation of either marker.

Correction to: Common findings of blaCTX-M-55-encoding 104-139 kbp plasmids harbored by extended-spectrum ß-lactamase-producing Escherichia coli in pork meat, wholesale market workers, and patients with urinary tract infection in Vietnam.

The original version of this article unfortunately contained a mistake. The legends of Tables 2 and 3, Fig. 1 are incorrect. The corrected legends are given below.

Common findings of bla CTX-M-55-encoding 104-139 kbp plasmids harbored by extended-spectrum ß-lactamase-producing Escherichia coli in pork meat, wholesale market workers, and patients with urinary tract infection in Vietnam.

Extended-spectrum, ß-lactamase-producing Escherichia coli (ESBL-E) harboring the bla CTX-M-55-encoding plasmid (ESBL-E55) has been reported to be associated with urinary tract infection (UTI). The aims of this study were to clarify the prevalence of ESBL-E55 in pork meats and workers from the same wholesale market, as well as patients with UTI from a nearby hospital in Vietnam; we also investigated the plasmids encoding bla CTX-M-55. Sequencing analysis showed that 66.6% of the ESBL-E isolated from pork meats contained bla CTX-M-55, whereas the gene was present in 25.0% of workers and 12.5% of patients with UTI. Plasmid analysis showed that several sizes of plasmid encoded bla CTX-M-55 in ESBL-E55 isolated from pork meats, whereas ESBL-E55 isolated from workers and patients with UTI contained only 104-139 kbp of bla CTX-M-55-encoding plasmids. This indicates that the 104-139 kbp sizes of bla CTX-M-55-encoding plasmids were commonly disseminated in pork meats, wholesale market workers, and patients with UTI.

Serous cystic neoplasm of the pancreas: a multinational study of 2622 patients under the auspices of the International Association of Pancreatology and European Pancreatic Club (European Study Group on Cystic Tumors of the Pancreas).

OBJECTIVES: Serous cystic neoplasm (SCN) is a cystic neoplasm of the pancreas whose natural history is poorly known. The purpose of the study was to attempt to describe the natural history of SCN, including the specific mortality. DESIGN: Retrospective multinational study including SCN diagnosed between 1990 and 2014. RESULTS: 2622 patients were included. Seventy-four per cent were women, and median age at diagnosis was 58 years (16-99). Patients presented with non-specific abdominal pain (27%), pancreaticobiliary symptoms (9%), diabetes mellitus (5%), other symptoms (4%) and/or were asymptomatic (61%). Fifty-two per cent of patients were operated on during the first year after diagnosis (median size: 40 mm (2-200)), 9% had resection beyond 1 year of follow-up (3 years (1-20), size at diagnosis: 25 mm (4-140)) and 39% had no surgery (3.6 years (1-23), 25.5 mm (1-200)). Surgical indications were (not exclusive) uncertain diagnosis (60%), symptoms (23%), size increase (12%), large size (6%) and adjacent organ compression (5%). In patients followed beyond 1 year (n=1271), size increased in 37% (growth rate: 4 mm/year), was stable in 57% and decreased in 6%. Three serous cystadenocarcinomas were recorded. Postoperative mortality was 0.6% (n=10), and SCN's related mortality was 0.1% (n=1). CONCLUSIONS: After a 3-year follow-up, clinical relevant symptoms occurred in a very small proportion of patients and size slowly increased in less than half. Surgical treatment should be proposed only for diagnosis remaining uncertain after complete workup, significant and related symptoms or exceptionally when exists concern with malignancy. This study supports an initial conservative management in the majority of patients with SCN. TRIAL REGISTRATION NUMBER: IRB 00006477.

Extended-spectrum ß-lactamase-producing Escherichia coli strains in the feces of carriers contribute substantially to urinary tract infections in these patients.

INTRODUCTION: The current increase in the incidence of urinary tract infections (UTIs) worldwide caused by extended-spectrum ß-lactamase (ESBL)-producing Escherichia coli may be due to the high number of ESBL-producing Enterobacteriaceae carriers in the community. However, whether ESBL-producing bacteria can cause UTIs in carriers remains uncertain. MATERIALS AND METHODS: In this study, 21 fecal carriers of ESBL-producing Enterobacteriaceae were assessed for UTIs caused by ESBL-producing E. coli. Bacterial isolates obtained from patients' urine and stool specimens were phenotypically and genotypically examined. Clonal similarities of isolates were assessed by multilocus sequence typing (MLST) and random amplified polymorphic DNA (RAPD) fingerprinting. RESULTS: The study revealed that 9 of 21 carriers developed UTIs, and genetic analysis showed that 44% of the UTIs developed were caused by the same ESBL-producing E. coli as that found in the feces of the patients. CONCLUSIONS: The ESBL-producing E. coli in carriers can cause UTIs.

High expression of heat shock protein 60 in follicular cells of Hashimoto's thyroiditis.

Heat shock protein 60 expression in the thyrocytes of Hashimoto's thyroiditis and Graves' disease was studied immunohistochemically. Thyrocytes of Hashimoto's thyroiditis showed high expression of heat shock protein 60 not only in tall eosinophilic-cells but also in low and flattened cells, although the former was stained with moderate to strong staining intensity and the latter weakly to moderately. Follicular cells around lymphoid cell follicles were stained more intensely, whereas cells apart from lymphoid cell follicles were stained weakly to moderately. In Graves' disease, only follicular cells around lymphoid cell follicles were stained with varying intensities. Since the pattern in all positive staining was granular, it was thought that heat shock protein 60 overexpressed in thyrocytes located on mitochondria. To investigate the immunological role of overexpression of self heat shock protein 60 in the thyrocytes of Hashimoto's thyroiditis, gamma delta TCR+ T-cells in the tissue and the IgG class of anti-self heat shock protein 60 antibodies were studied. gamma delta TCR+ T-cells were detected among lymphoid cells scattered in interfollicular connective tissue. No difference in antibody level was seen among subjects with Hashimoto's thyroiditis, Graves' disease and normal subjects. Self heat shock protein 60 overexpression in the thyrocytes of Hashimoto's thyroiditis may play a disease-modifying role, although it does not influence the anti-self heat shock protein 60 antibody level.

Evaluation of effect of lung resection on lobar ventilation and perfusion using intrabronchial capnography.

Intrabronchial capnography was applied in 11 lung cancer patients to investigate the effects of lobectomy on regional lung function. Spirometry and intrabronchial capnography were performed before surgery (PRE), and during the early (POST1,19 +/- 5 POD) and late (POST2, 184 +/- 98 POD) postoperative periods. End-tidal carbon dioxide concentration (EtCO2) and Smidt's velocity profile index (V-index) were calculated from each lobar capnogram obtained bronchoscopically. The V-index of lobes without cancer on the operated-on side increased after surgery (PRE, 10.7 +/- 5.0%; POST1, 14.3 +/- 9.4%, NS; POST2, 16.8 +/- 8.6%, p < 0.05), while the V-index on the unoperated-on side decreased after surgery (PRE, 10.5 +/- 5.3%; POST1, 7.9 +/- 3.5%, p < 0.05; POST2, 7.2 +/- 2.9%, p < 0.05). EtCO2 after surgery was lower on the operated-on side (POST1, 5.1 +/- 1.1%; POST2, 4.6 +/- 1.1%) than on the unoperated-on side (POST1, 5.4 +/- 0.9%, p < 0.05; POST2, 5.0 +/- 0.9%, p < 0.01). Since the V-index and EtCO2 are compatible with the expiratory flow rate and the perfusion/ventilation ratio, respectively, we concluded that the air flow decreased on the operated-on side and increased on the unoperated-on side postoperatively and that perfusion on the operated-on side was more severely reduced than ventilation. These findings suggest that intrabronchial capnography is useful for assessing the ventilation and perfusion of the individual lobes as single units.

Exercise capacity of thoracotomy patients in the early postoperative period.

OBJECTIVE: We investigated the mechanism involved with the initial drop and subsequent recovery of exercise capacity in the early postoperative period of thoracotomy patients. METHODS: Sixteen patients (13 who had undergone lobectomy, 3 who had undergone pneumonectomy) underwent a routine pulmonary function test (PFT) and a cardiopulmonary exercise test preoperatively, within 14 postoperative days (POD; post-1; mean +/- SD, 9 +/- 2 POD), and after 14 POD (post-2; mean, 26 +/- 12 POD). RESULTS: After surgery on post-1, PFT results of FVC, FEV(1), and maximum ventilatory volume (MVV) significantly decreased. Oxygen uptake (VO(2)) at a venous blood lactate level of 2.2 mmol/L (La-2. 2), which was adopted as the empirical anaerobic threshold, and maximum V O(2) (VO(2)max) decreased significantly to 88.2 +/- 7.9% and 73.1 +/- 15.4% of the preoperative values, respectively. La-2.2 min ventilation (VE)/ MVV and maximum VEmax)/MVV increased significantly from 0.36 +/- 0.08 to 0. 66 +/- 0.20 and from 0.58 +/- 0.14 to 0.80 +/- 0.09, respectively. On post-2, though La-2.2 VO(2) did not change, VO(2)max improved significantly to 81.5 +/- 19.7% of the preoperative values, in association with significant increases in maximal tidal volume and VEmax, which were produced by significant increases in the PFT results. La-2.2 VE/MVV also decreased significantly to 0.49 +/- 0.13, which indicated a sufficient recovery of respiratory reserve at submaximal exercise. CONCLUSIONS: The initial drop of exercise capacity after lung resection seems to be derived from both circulatory and ventilatory limitations. Further, the subsequent recovery within 1 month seems to be produced by an improvement in ventilatory limitation, which was caused by the surgical injury to the chest wall.

Human CD8 and CD4 T cell epitopes of epithelial cancer antigens.

Recent human tumor immunology research has identified several genes coding immunogenic peptides recognized by CD8 cytotoxic T lymphocytes (CTLs) in melanoma tumors. Very recently, CD4 T cell antigenic epitopes were also determined in certain melanoma tumors. The use of these peptides in conjunction with human immunotherapy could prove to be of great benefit. However, such peptides in clinically common tumors of epithelial cell origin, such as of the stomach, colon, lung, etc., have not yet been determined extensively. We describe for the first time an HLA-A31 (A*31012)-restricted natural antigenic peptide recognized by the CD8 CTL TcHST-2 of gastric signet ring cell carcinoma cell line HST-2. We also identified the HLA-DRB1*08032-restricted peptide recognized by the CD4 T cell line TcOSC-20 of squamous cell carcinoma OSC-20 derived from the oral cavity. The antigenic peptide of HST-2, designated F4.2, is composed of 10 amino acid residues with two anchor motif residues necessary for binding to HLA-A31 molecules. The synthetic F4.2 peptide enhanced the reactivity of TcHST-2 against HST-2 cells. Furthermore, introduction of an expression minigene coding F4.2 peptide to HLA-A31(+) cells conferred cytotoxic susceptibility to TcHST-2 on the cells. Some stomach cancer lines into which the HLA-A31 gene had been introduced, such as MKN28-A31-2, were lysed by TcHST-2, suggesting the presence of F4.2 peptide in at least some HLA-A31(+) stomach cancers. Furthermore, F4.2 peptide induced an F4.2 peptide-specific CTL response in at least 30-40% of HLA-A31(+) peripheral blood lymphocytes from gastric cancer patients, suggesting that F4.2 peptide could be used as a cancer vaccine for gastric tumors. The natural antigenic peptide of OSC-20 was also determined using acid extraction and biochemical separation and by mass spectrometry. Consequently, OSC-20 peptide was designated as the 6-1-5 peptide, an HLA-DRB1*08032-restricted 16-mer peptide with two possible anchor motifs. It has an amino acid sequence identical to that of human alpha-enolase, suggesting that it was derived from the processed parental alpha-enolase protein. We are presently attempting to determine the genes that code tumor rejection antigens recognized by HLA-A24- and A26-restricted T cells, including those of pulmonary and pancreatic carcinomas. The search for these antigenic peptides may lead to the identification of immunogenic peptide antigens that would be suitable for clinical use in commonly occurring epithelial cancers.

Acquisition of multidrug resistance in recurrent breast cancer demonstrated by the histoculture drug response assay.

Recurrent breast cancer has a very poor response rate to chemotherapy. To understand the degree of acquisition of multidrug resistance in recurrent disease, 24 recurrent breast tumors and 127 primary tumors were evaluated and compared for chemosensitivity in the histoculture drug response assay (HDRA). The evaluation rate was 98.8%. The HDRA utilizes 3-dimensional culture of human tumors on collagen-gel rafts. Doxorubicin (DXR), 5-fluorouracil (5-FU) and mitomycin C (MMC) were tested as standard agents and cisplatin (CDDP) as a candidate agent on surgical specimen of breast cancer in the HDRA. In vitro drug exposure in the HDRA was for 7 days. At the end of the assay, tumor response was assessed by the reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). The mean inhibition rates of primary tumors vs. recurrent tumors were 57.9% and 38.6% for DXR (p<0.0005); 59.9% and 42.8% for MMC (p<0.01); 49.0% and 33.4% for 5-FU (p<0.01); and 34.5% and 16.0% for CDDP (p<0.005), respectively. The recurrent cases were pretreated clinically with CAF (cyclophosphamide, DXR and 5-FU), CEF (cyclophosphamide, epirubicin and 5-FU) or CMF (cyclophosphamide, methotrexate and 5-FU). In the CAF and CEF group, the HDRA sensitivity to CDDP was significantly lower in recurrent disease (p<0.005) than that of primary breast cancer suggesting that one agent can induce resistance to another. This is further suggested by the fact that 64.7% of the recurrent cases were resistant to all 4 agents tested as opposed to 27% of the primary cases and that only 5.9% of the recurrent cases were sensitive to three or more agents as opposed to 18% of the primary cases. The correlation of the HDRA results to clinical outcome in the study was 80.0% with 15 cases evaluated consisting of 5 true positives, 3 false positives, 7 true negatives and no false negatives. Thus, the HDRA gives useful clinical information, in particular for the specific individualized treatment design necessary to overcome the multidrug resistance problem of recurrent breast cancer.

Differential gene expression screening between parental and highly metastatic pancreatic cancer variants using a DNA microarray.

To clarify the difference in genes expressed in hematogenous metastasis and peritoneal dissemination, a broad analysis of differential gene expression analysis between parental cell lines and established metastatic sublines was performed. Using an oligonucleotide array (Gene Chip, Affymetrix), approximately 2,000 genes involved in cancer were analyzed for each of the cell lines. HPC-4H4 (highly metastatic lines to the liver) compared with HPC-4 (low metastatic parental lines), in which 20 overexpressed genes and 5 underexpressed genes were recognized. HPC-4P4a (highly metastatic to the peritoneum) compared with HPC-4, in which 12 overexpressed genes and 15 underexpressed genes were also recognized. Analysis of HPC-4H4 and HPC-4P4a showed comparative up-regulation of 20 genes and down-regulation of 13 in the former, HPC-4H4. Further studies are needed to validate our hypothesis that some of the resulting differentially expressed genes might be implicated in the development of metastasis in pancreatic cancer. In conclusion, this genome-wide expression analysis will help to clarify the molecular mechanisms of cancer metastasis and of the different levels of gene expression in a variety of metastatic potentials in pancreatic cancer.

Long descending lymphatic pathway from the pancreaticoduodenal region to the para-aortic nodes: its laterality and topographical relationship with the celiac plexus.

In 6 of 15 postmortem-treated cadaveric specimens, we found macroscopically thick lymphatic collecting vessels that originated from not only the nodes along the common hepatic artery (No. 8 nodes) but also from the pancreaticoduodenal region, and which drained directly into the para-aortic nodes immediately below the left renal vein (No. 16b1-inter or -latero nodes). The collecting vessels, if they originated from the ventral (dorsal) visceral side, passed to the left (right) of the superior mesenteric and celiac arteries. Moreover, the right-side vessels (5 specimens) were classified into superficial and deep courses to the celiac plexus, whereas they were superficial in the left side (2 specimens). One of the deep (right) courses continued to the thoracic duct without any intercalated nodes. In addition, another deep route drained into the para-aortic node immediately above the left renal vein (No. 16a2-inter node). We consider that these collecting vessels form "direct descending pathways" from the relatively peripheral lymphatics in the upper abdomen toward the thoracic duct origin. The pathway seems to be a collateral, or even major drainage route, and it appears responsible for skipped metastasis of primary cancer. Since the classical, limited entity of the intestinal lymph trunk does not coincide with our pathway, it should be reconsidered. The proposed entity of the direct, long descending pathway will influence the selection and modification of lymphadenectomy methods in cancer surgery in the pancreaticoduodenal region.

Ramification pattern and topographical relationship between the portal and hepatic veins in the left anatomical lobe of the human liver.

Sixty-one human livers obtained from donated Japanese adult cadavers were dissected to reveal the ramification pattern of the portal and hepatic veins, and their topographical relationship in the left anatomical lobe. The segmental portal vein supplying S2 (P2) tended to form a single stem, whereas that of S3 (P3) was usually double. An intermediate branch between P2 and P3 was observed in 23.0% of livers. In spite of variation between livers, definite P2 and P3 were identified in 47 specimens. One tributary of the left hepatic vein (LHV) was usually present for drainage of S2, and two tributaries were present for S3 (sometimes also for S2 and/or S4). The latter two tributaries of the LHV and the two subsegmental branches of S3 showed three patterns of three-dimensional interdigitations. From these results, the portal vein system did not seem to have a two segmental composition (i.e., S2 and S3) in 23.0% of specimens, whereas the hepatic vein system did not have an intersegmental course in 23.4%. Thus, there were obvious limitations in using each system to determine the liver segment. Taking the overlapping cases into consideration, the left anatomical lobe of 41.0% of specimens did not seem to fit the definition of Couinaud's liver segment. In addition, four patterns of fissure vein (or scissural vein), > 5 mm in diameter at its terminal, were identified: (1) middle hepatic vein type (left median vein, 9.8%); (2) LHV type (left medial vein, 41.0%); (3) true fissure vein (3.3%); and (4) absent cases (45.9%). The former two types also suggested limitations of the hepatic vein system as an indicator of the segmental border.

[Thoracoscopic partial resection of the lung for peripheral lung tumor].

We report a thoracoscopic partial resection of the lung for the peripheral lung tumor in a 66-year-old woman. Her chest roentgenogram and computed tomographic scan showed a single tumor 1.9 cm in diameter within the periphery of the right lower lobe. A solitary metastatic lung tumor of colon cancer, which was resected one year ago, was suspected. Partial resection of the lung involving this tumor was performed using the MULTIFIRE ENDO GIA 30 disposable surgical stapler under thoracoscopy. There was neither air leak nor bleeding at the resection line. Pathological examination of the tumor revealed benign localized pleural mesothelioma. Her postoperative course was uneventful and she was discharged from the hospital on the eighth postoperative day. Thoracoscopic pulmonary resection using EndoGIA is a safe technique for a peripheral lung tumor.