Endophytic colonization and in planta nitrogen fixation by a Herbaspirillum sp. isolated from wild rice species.

Nitrogen-fixing bacteria were isolated from the stems of wild and cultivated rice on a modified Rennie medium. Based on 16S ribosomal DNA (rDNA) sequences, the diazotrophic isolates were phylogenetically close to four genera: Herbaspirillum, Ideonella, Enterobacter, and Azospirillum. Phenotypic properties and signature sequences of 16S rDNA indicated that three isolates (B65, B501, and B512) belong to the Herbaspirillum genus. To examine whether Herbaspirillum sp. strain B501 isolated from wild rice, Oryza officinalis, endophytically colonizes rice plants, the gfp gene encoding green fluorescent protein (GFP) was introduced into the bacteria. Observations by fluorescence stereomicroscopy showed that the GFP-tagged bacteria colonized shoots and seeds of aseptically grown seedlings of the original wild rice after inoculation of the seeds. Conversely, for cultivated rice Oryza sativa, no GFP fluorescence was observed for shoots and only weak signals were observed for seeds. Observations by fluorescence and electron microscopy revealed that Herbaspirillum sp. strain B501 colonized mainly intercellular spaces in the leaves of wild rice. Colony counts of surface-sterilized rice seedlings inoculated with the GFP-tagged bacteria indicated significantly more bacterial populations inside the original wild rice than in cultivated rice varieties. Moreover, after bacterial inoculation, in planta nitrogen fixation in young seedlings of wild rice, O. officinalis, was detected by the acetylene reduction and (15)N(2) gas incorporation assays. Therefore, we conclude that Herbaspirillum sp. strain B501 is a diazotrophic endophyte compatible with wild rice, particularly O. officinalis.

IS1631 occurrence in Bradyrhizobium japonicum highly reiterated sequence-possessing strains with high copy numbers of repeated sequences RSalpha and RSbeta.

From Bradyrhizobium japonicum highly reiterated sequence-possessing (HRS) strains indigenous to Niigata and Tokachi in Japan with high copy numbers of the repeated sequences RSalpha and RSbeta (K. Minamisawa, T. Isawa, Y. Nakatsuka, and N. Ichikawa, Appl. Environ. Microbiol. 64:1845-1851, 1998), several insertion sequence (IS)-like elements were isolated by using the formation of DNA duplexes by denaturation and renaturation of total DNA, followed by treatment with S1 nuclease. Most of these sequences showed structural features of bacterial IS elements, terminal inverted repeats, and homology with known IS elements and transposase genes. HRS and non-HRS strains of B. japonicum differed markedly in the profiles obtained after hybridization with all the elements tested. In particular, HRS strains of B. japonicum contained many copies of IS1631, whereas non-HRS strains completely lacked this element. This association remained true even when many field isolates of B. japonicum were examined. Consequently, IS1631 occurrence was well correlated with B. japonicum HRS strains possessing high copy numbers of the repeated sequence RSalpha or RSbeta. DNA sequence analysis indicated that IS1631 is 2,712 bp long. In addition, IS1631 belongs to the IS21 family, as evidenced by its two open reading frames, which encode putative proteins homologous to IstA and IstB of IS21, and its terminal inverted repeat sequences with multiple short repeats.

Follow-up of patients with superior vena cava syndrome by functional analysis of radionuclide venography.

In addition to imaging, radionuclide venography can be used for studying haemodynamic changes in superior vena cava syndrome (SVCS) by using the indices of transit time (TT), time of half peak count (TH) and peak count ratio (PC ratio). The objective of this study was to examine the utility of applying these indices, alongside images, in the post-therapy assessment of SVCS patients. Radionuclide venography was performed twice on 36 patients with SVCS due to thoracic malignancies, 29 of whom received specific anti-cancer therapy between the two studies (Group I); the other 7 did not receive such therapy (Group II). On the images, 12 patients in Group I showed improvement, one a deterioration and 16 no change; in Group II, 4 showed a deterioration and 3 no change. Using the indices, nearly all patients in Group I had decreased post-therapy values indicating haemodynamic improvement. A significant difference was seen between the mean (+/- S.E.) pre- and post-therapy values for TT (pre vs post: 6.2 +/- 0.7 vs 2.7 +/- 0.3 s) and TH (pre vs post: 16.9 +/- 2.3 vs 7.9 +/- 0.8 s) (P < 0.001). Although the PC ratio showed a non-significant decrement, it related to the status of collateral change. The above indices provide clinically valuable information about the haemodynamic status of patients with SVCS and can be used for their post-therapy evaluation.

New Bradyrhizobium japonicum strains that possess high copy numbers of the repeated sequence RS alpha.

In a survey of DNA fingerprints of indigenous Bradyrhizobium japonicum with the species-specific repeated sequences RS alpha and RS beta, 21 isolates from three field sites showed numerous RS-specific hybridization bands. The isolates were designated highly reiterated sequence-possessing (HRS) isolates, and their DNA hybridization profiles were easily distinguished from the normal patterns. Some HRS isolates from two field sites possessed extremely high numbers of RS alpha copies, ranging from 86 to 175 (average, 128), and showed shifts and duplications of nif- and hup-specific hybridization bands. The HRS isolates exhibited slower growth than normal isolates, although no difference in symbiotic properties was detected between the HRS and normal isolates. Nucleotide sequence analysis of 16S rRNA genes showed that HRS isolates were strains of B. japonicum. There was no difference in the spectra of serological and hydrogenase groupings of normal and HRS isolates. Some HRS isolates possessed a tandem repeat RS alpha dimer that is similar to the structure of (IS30)2, which was shown to cause a burst of transpositional rearrangements in Escherichia coli. The results suggest that HRS isolates are derived from normal isolates in individual fields by genome rearrangements that may be mediated by insertion sequences such as RS alpha.

Chronic solitary pulmonary nodule due to unsuspected pulmonary infarction from silent pulmonary embolism.

A case of pulmonary embolism showing a longstanding solitary pulmonary nodule is presented. An asymptomatic 57 year-old man with a solitary nodule in the right lower lobe was referred to our hospital. A pulmonary perfusion-ventilation scan following a sudden onset of dyspnoea established the diagnosis of recurrent pulmonary embolism. The nodule gradually disappeared after anticoagulant treatment, indicating that the nodule was pulmonary infarction from silent pulmonary embolism. Although the incidence of pulmonary infarction is low in Japan, this case suggests that pulmonary infarction from silent pulmonary embolism should be considered as one important cause of a solitary pulmonary nodule.

Radionuclide venography and its functional analysis in superior vena cava syndrome.

UNLABELLED: In addition to imaging, radionuclide venography maybe used for the functional assessment of superior vena cava (SVC) syndrome by applying the indices of transit time (TT), time of half-peak count (TH) and peak count ratio (PC ratio). METHODS: Ten healthy subjects (Group N) and 107 patients with SVC syndrome (64 symptomatic and 43 asymptomatic) were studied. Images were visually assessed for collaterals or jugular venous reflux and values of the indices were calculated. RESULTS: The 107 patients were subclassified into three groups according to the images obtained. Collateral circulation was seen in 37 patients (Group C). In 20 patients, jugular venous reflux was observed (Group J). Fifty patients who showed neither collaterals nor reflux were included in Group P. In comparison to Group N [3.6 +/- 0.56 see (sem)], TT values were significantly higher (p < 0.05) for Group J (7.13 +/- 1.16 sec) and Group C (7.00 +/- 0.87 sec). Values of TH were significantly prolonged (p < 0.05) for Group J (23.6 +/- 4.8 sec) and Group C (18.8 +/- 2.2 sec) in comparison to Group N (9.2 +/- 1.5 sec). PC ratio values were higher in all patient groups in comparison to Group N (3.4 +/- 0.57). CONCLUSION: These indices are potentially useful in the initial diagnosis and post-therapeutic evaluation of SVC syndrome. In the absence of other causes, appearance of jugular venous reflux may be considered a sign of SVC syndrome.

Neuroendocrine differentiation in pancreatic duct carcinoma special emphasis on duct-endocrine cell carcinoma of the pancreas.

To evaluate the significance of neuroendocrine differentiation in duct carcinoma of the pancreas, we investigated 79 pancreatic carcinomas, applying histochemistry and immunohistochemistry (chromogranin A, Leu-7, synaptophysin and neuron-specific enolase (NSE), and correlated the morphologic differentiation pattern with clinicopathological characteristics. There were two types of neuroendocrine differentiation: scattered (n = 23) and diffuse (n = 3). The scattered type of pancreatic duct carcinoma contained scattered endocrine cells amounting to less than 10% of the neoplastic cells and was seen more frequently in well-differentiated carcinomas. There was no characteristic clinical feature found in the scattered type when compared with the tumors devoid of endocrine cells (n = 53). In contrast, the diffuse type showed diffuse immunostaining with NSE and synaptophysin in tumor cells and dense core granules ultrastructurally. These tumors showed a greater hypervascularity in angiography (p < 0.01) and the patients had relatively longer survival (33.3 months, p < 0.05) than unresectable cases of other histological types of pancreatic cancer. Two types of neuroendocrine differentiation (scattered and diffuse) existed in pancreatic ductal carcinoma. The diffuse type (Duct-Endocrine Cell Carcinoma of the Pancreas) showed synchronous duct and endocrine differentiation and particular clinical features.

Treatment of T3 and T4 carcinomas of the gallbladder.

Forty-nine patients with T3 and T4 carcinoma of the gallbladder were treated by three different regimens. Group 1 consisted of 26 patients treated with combined resection alone; group 2 was twelve patients whose tumors had spread to the hepatoduodenal ligament treated with combined resection plus intraoperative radiation therapy; group 3 comprised eleven patients with nonresectable tumors treated with hyperthermia in combination with chemoradiation therapy (HCRT). The difference in the survival rates between group 1 and 2 were statistically significant (p < 0.001); however, the difference between group 2 and 3 was not significant. In group 1, there was a significant difference between patients with and without lymph node involvement in the 3-year survival rate (p < 0.01). Thus, the only patients without involvement of regional lymph nodes and the hepatoduodenal ligament have the best potential for long-term survival through aggressive surgical approaches. HCRT may provide an alternative palliation for patients with advanced carcinoma showing obstructive jaundice.

High-resolution electron microscopy of technegas and pertechnegas.

Once inhaled, technegas remains in the lungs for a long period of time, whereas pertechnegas rapidly disappears. To investigate this difference, the morphology of technegas and of pertechnegas was investigated using high-resolution transmission electron microscopy. The technegas and pertechnegas were generated in an atmosphere of pure argon and in 3% oxygen in argon, respectively, using a commercially available technegas generator. For the technegas, the technetium crystals were observed to be covered with carbon, whereas they were found to have no carbon coating with the pertechnegas. Whether or not the technetium is coated with carbon appears to be responsible for the differences in behaviour of technegas and pertechnegas after deposition on the lung epithelium following inhalation.

Hematogenous metastases of pancreatic ductal carcinoma.

To investigate the heterogeneity of hematogenous metastases of pancreatic ductal carcinoma, we investigated carcinomatous spread in 130 autopsy cases. Hepatic metastases occurred most frequently, in 81 cases (62%), which may be explained by the fact that all veins draining the pancreas flow into the portal system. We closely examined the 49 cases without hepatic metastases. Sixteen patients had pulmonary metastases without hepatic metastases, whereas seven had peculiar hematogenous metastases without hepatic or pulmonary metastases. Fifteen of these 23 patients had pancreatic body carcinomas. The unusual patterns of spread might be due to (a) hepatofugal portosystemic shunting induced by splenic vein obstruction, (b) retrograde lymphatic infiltration from metastatic tracheobronchial lymph nodes, or (c) aggressive characteristics of the tumors indicated by peculiar histologic features such as pleomorphic or mucoepidermoid carcinoma, etc. Sixteen patients showed only lymph node metastases and 10 had no distant metastases. Seventeen of these 26 cases had pancreatic head carcinoma. Histologically, two patients had mucinous cystadenocarcinomas, and six had adenocarcinomas producing rich mucin. The average age of the group with no distant metastases was higher.

[Pulmonary nuclear medicine].

Pulmonary nuclear medicine dates back to Knipping and West in the late 1950's but practically starts with the successful production of 131I-MAA by GV Taplin in 1963. Not only is the diagnosis of pulmonary embolism greatly facilitated by using 131I-MAA but also studies of regional lung function have made rapid progress. Radioactive gas and aerosol inhalation have been used to study ventilation distribution in the lungs. Using nuclear medicine technology regional hypoxic vasoconstriction was found to play a great role in regulating regional perfusion distribution in the lungs. Ventilation and perfusion mismatch and match indicate organic lung diseases and pulmonary vascular diseases, respectively. Aerosol deposition patterns in the lungs are helpful in the differential diagnosis of chronic obstructive lung diseases (COPD). Technigas is an ultrafine aerosol which is probably more useful for ventilation studies than conventional aerosols produced by either a jet or an ultrasonic nebulizer. Besides respiratory lung function pulmonary nuclear medicine techniques have made it possible to study nonrespiratory lung function. One is mucociliary clearance mechanisms. They can be studied by using a nonabsorbable aerosol like 99mTc-albumin. Dynamic mucociliary clearance function can be visualized in vivo by radioaerosol inhalation lung cine-scintigraphy. Four abnormal mucociliary transport patterns were discernible in COPD. An objective evaluation of a drug effect on mucociliary transport is feasible. Detailed quantitative analysis of mucociliary clearance is also possible by computer techniques. Pulmonary epithelial permeability is studied following inhalation of 99mTc-DTPA aerosol. Inhaled 99mTc-DTPA disappears from the lungs more rapidly in smokers and patients with interstitial lung diseases. Nuclear medicine has great potential to elucidate other functions of the lung which are still not defined yet by the present knowledge of lung function.

Inhalation of pertechnegas: similar clearance from the lungs to that of inhaled pertechnetate aerosol.

It has been reported that the inhalation of pertechnegas (TGO3) generated under an atmosphere of 3% oxygen (O2) in argon (Ar) instead of 100% Ar without O2 in the technegas generator, offers a simple but accurate quantification of clearance from the lungs, permitting it to be used in place of 99Tcm-diethylenetriamine pentaacetic acid (DTPA). The disappearance from the lungs of inhaled TGO3 was so similar to that of inhaled pertechnetate aerosol (TcO4) in this study, that there was neither a statistically significant difference in the clearance half-time (t1/2), nor a difference in the lung images between them. Neither TGO3 nor TcO4 inhalation could distinguish smokers from non-smokers using t1/2. When inhaled, TGO3 appears to behave in a similar manner to TcO4 in the lungs and hence cannot be used in place of 99Tcm-DTPA in clinical practice.

A case of pleomorphic carcinoma of the pancreas showing sequential histological change by immunohistochemical study.

Pleomorphic carcinoma of the pancreas is a rare, histologically characterized pancreatic tumor with a rapid and fatal course. We report a case of a resected pleomorphic carcinoma located in the body of the pancreas in a 61-yr-old male. Histological analysis of the resected specimen revealed the coexistence of pleomorphic carcinoma with adenocarcinoma, but the recurrent tumors at autopsy 20 mo later were only of the adenocarcinomatous type. Cells in the adenocarcinomatous component showed a diffuse reactivity for CA19-9, CEA, and cytokeratin, and a focal reactivity for vimentin. In contrast, vimentin was diffusely expressed in pleomorphic lesion. Adenocarcinoma at autopsy expressed CA19-9, CEA, and cytokeratin, but not vimentin. These findings suggest that the recurrent adenocarcinoma may have developed as a consequence of sequential change in the nature of the tumor.

Establishment of a cell line from an adenocarcinoma of the lung producing carcinoembryonic antigen (CEA) and CA19-9.

An adenocarcinoma cell line was established from pleural effusion of a 67-year-old woman and designated as TK-SA. The population doubling time of the cells was 100.4 hr. Chromosomal analysis revealed the TK-SA to have human aneuploid chromosomes with a near-triploid mode. Ultrastructurally, the TK-SA was compatible with adenocarcinoma. The cell line was highly tumorigenic. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide dye assay demonstrated resistance of the cell line to Cis-platin (CDDP), Cyclophosphamide (CPM) and Tegafur/Uracil (UFT). High carcinoembryonic antigen (CEA) and CA19-9 levels were similar in patient's serum and conditioned medium.

Annular pancreas associated with carcinoma in the dorsal part of pancreas divisum.

A carcinoma in the dorsal part of the pancreas divisum with an annular pancreas in the anterior part is reported. A 79-yr-old female was admitted in our hospital complaining of epigastralgia. Computed tomography (CT) and ultrasound (US) showed an irregular mass in the pancreatic body. A pancreatogram obtained through the major duodenal papilla demonstrated only the ventral pancreatic duct that encircled the duodenum. Contrast medium injected from the minor duodenal papilla showed Santorini's duct obstruction at the neck portion of the pancreas without communication with the ventral pancreatic duct. The patient died with liver metastases. Autopsy confirmed annular pancreas and a 6-cm tumor in the pancreatic body extending to the pancreatic head and pancreas divisum. Pancreatic carcinoma; histologically a moderately differentiated adenocarcinoma; originated from the dorsal part of pancreas divisum. To our knowledge this is the first report of pancreatic carcinoma associated with annular pancreas coexistent with pancreas divisum.

Technegas versus krypton-81m gas as an inhalation agent. Comparison of pulmonary distribution at total lung capacity.

Exactly how the pulmonary distribution of inhaled radioactive gas and Technegas, or ultra-small aerosol particulates, differs from each other, is still uncertain. The authors compared the distribution of inhaled Kr-81 m gas and Technegas in the lungs at total lung capacity in 13 control subjects with no clinical conditions and 13 patients with various chest diseases. In normal lungs, there was no difference in the distribution ratios in the right and left lungs between inhaled Kr-81m gas and Technegas. However, there was a significant difference in the lungs of patients with pulmonary disease. Technegas tended to deposit more in the lung bases than did Kr-81m gas. Despite these statistical differences, they were visually, or qualitatively, similar. From a practical and clinical standpoint, Technegas seems to be useful as an inhalation agent, unless quantitative analyses are required.

Intraoperative radiation therapy for pancreatic carcinoma. The choice of treatment modality.

Ninety patients with carcinoma of the pancreas treated between 1976 and 1990 were reviewed retrospectively. Intraoperative radiation therapy (IORT) in combination with external beam radiation therapy (EBRT) for localized but unresectable tumors (n = 29) prolonged survival significantly more than IORT alone (n = 16) (p < 0.01); it seems EBRT enhanced or contributed to the better results obtained with IORT plus EBRT. Moreover, IORT, alone or in combination, relieved pain. Adjuvant IORT for residual tumors (n = 20) might not effectively prolong survival, because the difference in survival rate between noncurative resection plus IORT and nonresection plus IORT in combination with EBRT was not significant. Curative tumor resection of stage III disease in combination with IORT (n = 9) resulted in significantly longer survival as compared with curative tumor resection alone (n = 8) (p < 0.05). It may be advisable to administer IORT in combination with EBRT to patients with advanced pancreatic carcinoma, avoiding aggressive tumor resection, when curative tumor resection cannot be performed.

Application of an air-filled tube for measuring intraesophageal pressure.

We devised a new method for measuring esophageal pressure (Pes) with use of a flexible tube without a balloon at a constant rate of airflow through the tube into the esophagus (balloonless method). A study with 133Xe showed that the air that accumulated in the esophagus did not interfere with the measurement of Pes. We measured dynamic compliance (Cdyn) and pulmonary resistance (RL) with the balloonless method in 19 subjects and obtained a static deflation pressure-volume curve (P-V curve) in 10 other subjects. Cdyn was 0.243 +/- 0.099 l/cmH2O and RL was 1.52 +/- 0.42 cmH2O.l-1.s. In 6 of the 10 subjects, a P-V curve was also obtained with the balloon tube (balloon method). K, the index of compliance in the exponential function V = V0(1-e-KP) where V0 is volume at infinite pressure, was 0.136 +/- 0.040 cmH2O-1 with the balloonless method and 0.153 +/- 0.023 cmH2O-1 with the balloon method. No statistically significant difference was found between these two values. In conclusion, Cdyn, RL, and the P-V curve can be obtained precisely with the balloonless method.

Difference in inhaled aerosol deposition patterns in the lungs due to three different sized aerosols.

Deposition patterns of inhaled aerosol in the lungs were studied in five normal subjects and 20 patients with lung disease by inhaling radioaerosols with three different particle size distributions. These aerosols were generated from BARC, UltraVent, and Mistogen-EN-142. Particle size distributions generated by these three nebulizers were 0.84, 1.04 and 1.93 microns in activity median aerodynamic diameter (AMAD) with its geometric standard deviation (sigma g) of 1.73, 1.71 and 1.52, respectively. Deposition patterns of inhaled aerosols were compared qualitatively and quantitatively by studying six different parameters: alveolar deposition ratio (ALDR), Xmax, Xmean, standard deviation (S.D.), skewness and kurtosis of the radioactive distribution in the lungs following inhalation. It has been found that aerosol deposition patterns varied with particle size. The unevenness of aerosol deposition, Xmax, Xmean and the number of 'hot spots' became more prominent with the increase in particle size, whereas values of ALDR and S.D. decreased as particle size increased. Knowing these deposition characteristics would facilitate a judicious application of aerosol inhalation to medical use.

[Pulmonary epithelial permeability in rats with bleomycin-induced pneumonitis].

This study was performed to investigate the mechanism by which 99mTc-DTPA molecules pass through the pulmonary epithelium following inhalation of 99mTc-DTPA aerosol. Interstitial pneumonitis was induced in 6-week-old male rats by instilling 1 mg/kg of bleomycin into the trachea. Disappearance of radioactivity from the lungs was measured with a gamma camera every 2 weeks to estimate pulmonary epithelial permeability, and light- and electron-microscopic histopathologic examinations were performed at the same intervals. There was a statistically significant increase in the pulmonary epithelial permeability at 2 weeks after the instillation of bleomycin. However, subsequent changes in pulmonary epithelial permeability were not uniform; some animals showed recovery and some showed further increase and/or partial recovery. Microscopically, increase in the capillary bed, round cell infiltration, and widening of the interstitial space were observed in addition to the presence of macrophages in the alveolar spaces at 2 weeks. Electron microscopic examination revealed vacuolization, thinning and detachment of the alveolar epithelium, and denudation of the basement membrane. Prominent fibrosis, honeycombing, thinning of the pulmonary epithelium, and increase in collagen fibers were observed after 18 weeks. We consider that vacuolization, thinning, and detachment of the pulmonary epithelium and denudation of the basement membrane are related to the increase in pulmonary epithelial permeability in bleomycin-induced interstitial pneumonitis.