Temperature-dependent differences in community structure of bacteria involved in degradation of petroleum hydrocarbons under sulfate-reducing conditions.

AIM: The aim of this study was to characterize the microbial community involved in anaerobic degradation of petroleum hydrocarbon under low- and moderate-temperature conditions. METHODS AND RESULTS: Sulfate-reducing enrichment cultures growing on crude oil and p-xylene were established at low and moderate temperatures. Bacterial community structures of the cultures were characterized by 16S rRNA gene-based analysis and organisms responsible for degradation of p-xylene were investigated by analysis of the bamA gene, involved in anaerobic degradation of aromatic compounds. The PCR-denaturing gradient gel electrophoresis analysis indicated significant differences in microbial community structures among the cultures, depending on the temperatures of incubation. Difference depending on the temperatures was also observed in the cloning analysis of the bamA gene performed on the p-xylene-degrading enrichment cultures. Majority of clones detected in the culture of moderate temperature were related to Desulfosarcina ovata, whereas more diverse bamA gene sequences were obtained from the culture incubated at low temperature. CONCLUSIONS: Temperature-dependent differences in microbial community were demonstrated by the analyses of two genes. It was suggested that sulfate-reducing bacteria of phylogenetically different groups might be involved in the degradation of petroleum hydrocarbons in different temperature environments. SIGNIFICANCE AND IMPACT OF THE STUDY: This study is the first report of p-xylene-degrading sulfate-reducing enrichment culture at low temperature. The results of the experiments at low temperature were distinctly different from those reported in previous studies performed at moderate temperatures.

In vitro antileishmanial, antiplasmodial and cytotoxic activities of phenolics and triterpenoids from Baccharis dracunculifolia D. C. (Asteraceae).

Baccharis dracunculifolia (Asteraceae), the most important plant source of the Brazilian green propolis (GPE), displayed in vitro activity against Leishmania donovani, with an IC(50) value of 45 microg/mL, while GPE presented an IC(50) value of 49 microg/mL. Among the isolated compounds of B. dracunculifolia, ursolic acid, and hautriwaic acid lactone showed IC(50) values of 3.7 microg/mL and 7.0 microg/mL, respectively. Uvaol, acacetin, and ermanin displayed moderate antileishmanial activity. Regarding the antiplasmodial assay against Plasmodium falciparum, BdE and GPE gave similar IC(50) values (about 20 microg/mL), while Hautriwaic acid lactone led to an IC(50) value of 0.8 microg/mL (D6 clone).

Microbial community analysis at crude oil-contaminated soils targeting the 16S ribosomal RNA, xylM, C23O, and bcr genes.

AIMS: The analyses targeting multiple functional genes were performed on the samples of crude oil-contaminated soil, to investigate community structures of organisms involved in monoaromatic hydrocarbon degradation. METHODS AND RESULTS: Environmental samples were obtained from two sites that were contaminated with different components of crude oil. The analysis on 16S rRNA gene revealed that bacterial community structures were clearly different between the two sites. The cloning analyses were performed by using primers specific for the catabolic genes involved in the aerobic or anaerobic degradation of monoaromatic hydrocarbons, i.e. xylene monooxygenase (xylM), catechol 2,3-dioxygenase (C23O), and benzoyl-CoA reductase (bcr) genes. From the result of xylM gene, it was suggested that there are lineages specific to the respective sites, reflecting the differences of sampling sites. In the analysis of the C23O gene, the results obtained with two primer sets were distinct from each other. A comparison of these suggested that catabolic types of major bacteria carrying this gene were different between the two sites. As for the bcr gene, no amplicon was obtained from one sample. Phylogenetic analysis revealed that the sequences obtained from the other sample were distinct from the known sequences. CONCLUSIONS: The differences between the two sites were demonstrated in the analyses of all tested genes. As for aerobic cleavage of the aromatic ring, it was also suggested that analysis using two primer sets provide more detailed information about microbial communities in the contaminated site. SIGNIFICANCE AND IMPACT OF THE STUDY: The present study demonstrated that analysis targeting multiple functional genes as molecular markers is practical to examine microbial community in crude oil-contaminated environments.

The tumor-preventing effect of a mixture of several lactic acid bacteria on 1,2-dimethylhydrazine-induced colon carcinogenesis in mice.

The anti-tumor effect of a dietary supplement obtained from mixed cultures of several lactic acid bacteria was examined in the colon of tumor-inducing ICR male mice by use of a carcinogen, 1,2-dimethylhydrazine (DMH, 20 mg/kg body weight, 1 intra-muscular injection per week for 10 weeks). The animals were sacrificed either 15 weeks or 24-26 weeks after the first carcinogen injection. Macroscopically, the incidence of colon tumors at a 24-26 week period of tumor induction was apparently lower in mice treated with both the DMH and dietary supplement (76%) than in those treated with DMH alone (100%). Histologically, microadenomas were induced predominantly in the anal half of the total colon, and large lymphoid aggregates were often associated with dysplastic crypts in the distal colon. Apoptotic cell masses were shed into the distended lumen of the involved crypts. The statistical analysis at a 15-week period of tumor induction indicated that the incidence of microadenomas per tumor-induced mouse was lowered significantly by use of the dietary supplement. From the present results, it is suggested that the intake of the dietary supplement inhibits the early development of colon adenomas, and the inhibition of microadenomas results in a reduction of subsequent polyp and tumor yield in the mouse colon.

Hypothalamic hamartoma associated with an arachnoid cyst.

A hypothalamic hamartoma associated with an arachnoid cyst in an 8-year-old boy is reported herein. He presented with precocious puberty, and neuroimaging studies demonstrated a solid mass in the prepontine cistern and a huge arachnoid cyst in the left cranial fossa. The mass appeared isointense to the surrounding cerebral cortex on T1-weighted magnetic resonance images, hyperintense on T2-weighted images, and was not enhanced after administration of Gd-DTPA. The patient underwent a left frontotemporal craniotomy and a cyst-peritoneal shunt was inserted. Histological features of the cyst wall and the mass were characteristic of an arachnoid cyst and hamartoma, respectively. While a hypothalamic hamartoma associated with an arachnoid cyst is rare, such a case may help clarify the geneses of both anomalous lesions.

Normal diffusion-weighted imaging in cerebral air embolism complicating angiography.

We report a case of cerebral air embolism resulting from accidental air infection during cerebral angiography. A 60-year-old man was accidentally injected with air via the left subclavian artery. Angiography demonstrated air within the basilar artery. The patient showed signs of posterior circulation ischaemia (confusion, blindness, gaze palsy and hemiparesis). However, MRI, including diffusion-weighted imaging, showed no abnormality 4 h later. The patient was treated with hyperbaric oxygen within 5 h of the embolism. All symptoms and signs resolved completely within a week.

Thiamine-deficient lactic acidosis with brain tumor treatment. Report of three cases.

Lactic acidosis due to thiamine deficiency is known to complicate chemotherapy and radiotherapy treatment of malignant extracranial tumors, but to the authors' knowledge, this complication has not been reported in patients treated for malignant brain tumors. They report three such cases, demonstrating that this complication can occur during treatment of brain tumors. In all patients, consciousness levels deteriorated within 1 to 2 days. Serum lactic acid levels increased to concentrations between 62 and 96.7 mg/dl, resulting in severe metabolic acidosis. A low blood thiamine level (9 ng/ml) was demonstrated at the onset in one case, and high-dose thiamine infusions dramatically improved lactic acidemia as well as impairment of consciousness in two cases. In the other case, hydrocephalus was suspected initially, resulting in a delay in thiamine supplementation. Clinical differentiation of this form of lactic acidosis from hydrocephalus or tumor progression can be very difficult in a patient undergoing treatment for a malignant brain tumor. Demand for thiamine is thought to be increased in patients with malignant brain tumors, and supplemental thiamine during treatment is necessary to prevent lactic acidosis. When this complication occurs, immediate treatment with sufficient thiamine is essential, together with normalization of pH by using sodium bicarbonate. With timely intervention, the level of consciousness can recover to the preacidotic state with no new neurological deficits.

Improvement of the pulmonary absorption of (Asu1,7)-eel calcitonin by various absorption enhancers and their pulmonary toxicity in rats.

The effects of absorption enhancers on the pulmonary absorption of (Asu1,7)-eel calcitonin (ECT) and their pulmonary toxicity were examined by means of in situ pulmonary experiments. The absorption of ECT from the lungs was estimated by its hypocalcemic effect. The pulmonary membrane toxicity of absorption enhancers was evaluated by the leakage of Evans Blue from the plasma into the lungs. In the absence of absorption enhancers, a slight hypocalcemic effect was obtained following intrapulmonary administration of ECT. However, we found significant hypocalcemic effects after the ECT administration with 10 mM n-lauryl beta-D-maltopyranoside (LM), 10 mM sodium glycocholate (NaGC), and 10 mM linoleic acid-HCO60 (hydrogenated caster oil) mixed micelle (MM). The plasma calcium levels decreased as the amount of LM coadministered with ECT increased. In contrast, 10 mM EDTA did not improve the pulmonary absorption of ECT. Overall, a correlation between the pulmonary absorption of ECT and local toxicity was observed in the presence of these additives. However, 1 mM LM, 10 mM NaGC, and 10 mM MM improve the pulmonary absorption of ECT with low pulmonary toxicity. These findings suggest that the use of these adjuvants would be a useful approach for improving the pulmonary absorption of ECT.

Thalamic gliomas: a clinicopathologic analysis of 20 cases with reference to patient age.

Twenty patients (M 11, F9; ranging from 1-77 years old) with histologically proven glial tumours in the thalamic region, treated from 1979 until 1994 at Kyushu University Hospital were retrospectively reviewed and analysed in order to elucidate their clinical and neuropathological characteristics. The initial common clinical manifestations were those of increased intracranial pressure or motor weakness. The histological diagnosis of the tumour was pilocytic astrocytoma in 2 patients, fibrillary astrocytoma in 7, anaplastic astrocytoma in 7, and glioblastoma multiforme in 4. The initial treatment was surgery alone in 4 patients, surgery followed by radiation therapy in 5, surgery followed by radiation therapy and chemotherapy in 9, and conventional radiation therapy alone in 2 patients. The 3-year overall actuarial survival rate for all patients was 20% but was related to both the histological type and the age of the patients: As a result, the rate was 44% for patients with low-grade astrocytoma compared to 0% for those with high-grade astrocytoma. While 5 out of 11 patients under the age of 25 years at their initial presentation have survived for from 2-16 years after the diagnosis, all patients presenting after the age of 25 years died within 3 years after treatment. Thalamic glial tumours are not a homogeneous group of tumours in terms of clinical behaviour and histopathological features, and the poor overall results, especially in adult tumours, thus emphasise the need for continued research in the treatment of these tumours.

Degradative capacities and 16S rRNA-targeted whole-cell hybridization of sulfate-reducing bacteria in an anaerobic enrichment culture utilizing alkylbenzenes from crude oil.

A mesophilic sulfate-reducing enrichment culture growing anaerobically on crude oil was used as a model system to study which nutritional types of sulfate-reducing bacteria may develop on original petroleum constituents in oil wells, tanks, and pipelines. Chemical analysis of oil hydrocarbons during growth revealed depletion of toluene and o-xylene within 1 month and of m-xylene, o-ethyltoluene, m-ethyltoluene, m-propyltoluene, and m-isopropyltoluene within approximately 2 months. In anaerobic counting series, the highest numbers of CFU (6 x 10(6) to 8 x 10(6) CFU ml-1) were obtained with toluene and benzoate. Almost the same numbers were obtained with lactate, a substrate often used for detection of the vibrio-shaped, incompletely oxidizing Desulfovibrio sp. In the present study, however, lactate yielded mostly colonies of oval to rod-shaped, completely oxidizing, sulfate-reducing bacteria which were able to grow slowly on toluene or crude oil. Desulfovibrio species were detected only at low numbers (3 x 10(5) CFU ml-1). In agreement with this finding, a fluorescently labeled, 16S rRNA-targeted oligonucleotide probe described in the literature as specific for members of the Desulfovibrionaceae (suggested family) hybridized only with a small portion (< 5%) of the cells in the enrichment culture. These results are consistent with the observation that known Desulfovibrio species do not utilize aromatic hydrocarbons, the predominant substrates in the enrichment culture. All known sulfate-reducing bacteria which utilize aromatic compounds belong to a separate branch, the Desulfobacteriaceae (suggested family). Most members of this family are complete oxidizers. For specific hybridization with members of this branch, the probe had to be modified by a nucleotide exchange. Indeed, this modified probe hybridized with more than 95% of the cells in the enrichment culture. The results show that completely oxidizing, alkylbenzene-utilizing sulfate-reducing bacteria rather than Desulfovibrio species have to be considered in attempts to understand the microbiology of sulfide production in oil wells, tanks, and pipelines when no electron donors other than the indigenous oil constituents are available.

Chemotherapy for progressive pilocytic astrocytomas in the chiasmo-hypothalamic regions.

Over the past 25 years, we have treated 17 patients with chiasmo-hypothalamic astrocytomas. Before 1988, the initial treatments consisted of surgery and/or radiotherapy, while since 1989, 4 children (1 male, 3 females, aged 3-8 years) were treated primarily with chemotherapy. None of them was associated with neurofibromatosis. After a biopsy of the tumor, the intravenous administration of ranimustine (MCNU; 30-86 mg/m2) and/or nimustine (ACNU; 30.3-64.1 mg/m2) was given without radiation therapy. Chemotherapy was usually given as an out-patient, with a total of 5-13 courses. The total doses of MCNU and ACNU administered ranged from 150 to 570 mg and from 64.8 mg to 100 mg, respectively. After chemotherapy 2 patients showed clinical improvement and tumor regression on neuro-imaging, while one patient showed clinical improvement and tumor size stabilization on neuro-imaging. The remaining one child, however, showed a clinical worsening and tumor progression on neuro-imaging studies. He was thus treated with a second chemotherapy regimen with carboplatin and etoposide, which brought about tumor regression. The acute and subacute toxicity of chemotherapy was mild. All patients are now leading almost normal lives with a median of 43 months after diagnosis. Although a longer and more careful clinical observation is required, the authors conclude that chemotherapy with MCNU and/or ACNU may benefit patients with unresectable pilocytic astrocytoma requiring treatment. The advantages of this therapy include its mild side effects and the lack of any hospitalization in most patients. It may also delay the need for radiation therapy, which can have a deleterious effect on the young developing brain.

Physiological correlates of abnormal behaviors in magnesium-deficient rats.

In order to elucidate the mechanism of behavioral alterations in magnesium-deficient rats, changes in the electroencephalogram (EEG) and electrocardiogram (ECG) were studied during auditory stimulation and correlated with the behavioral alterations. Weanling rats were fed either a Mg-deficient diet or a control synthetic diet for 2-3 weeks before the experiment. EEGs were recorded from the hippocampus and the sensorimotor and auditory cortices, and ECGs with a telemetry system. White noise with an intensity of 100 dB was given continuously to induce behavioral changes. The Mg-deficient rats developed consistent and graded behavioral changes in response to the stimulation, showing running-jumping behavior (stage 1), followed by tonic limb convulsion (stage 2) and finally by falling down on the floor (stage 3). The EEGs also showed consistent changes with spike activity, initiating in the hippocampus (stage 2) and then spreading to the neocortices bilaterally (stage 3). These findings indicate that the behavioral changes induced by auditory stimulation in the Mg-deficient rats are due to seizures arising in deeper brain structures, particularly in the limbic system, and projecting secondarily to the neocortices. The ECG changes, mainly consisting of marked bradyarrhythmia, occurred as early as the appearance of the EEG spikes, indicating that they were also related to the seizure. We conclude therefore that Mg deficiency in rats causes increased excitability of the central nervous system, resulting in seizures possibly originated in the limbic system, later developing secondary generalization, and also causing cardiac dysfunctions.

Germ cell tumor in the hypothalamo-neurohypophysial region: clinical features and treatment.

Twenty-one patients with germ cell tumors (17 germinomas and 4 teratomas) involving the hypothalamic-neurohypophysial (HN) region were reviewed retrospectively. Eleven patients were males and 10 females, and their ages ranged from 7 to 45 years (average 18.5 years). Diabetes insipidus was the initial and the most prominent symptom in most germinomas; in teratomas the most prominent symptom was visual disturbance. Fifteen patients with germinomas were treated by radiotherapy, and 4 with teratomas were treated by surgical resection alone. Two recent germinoma patients with extensive CSF dissemination were treated with systemic chemotherapy consisting of anticancer platinum drugs and etoposide, which resulted in a complete disappearance of the tumors. Patients with germinoma treated after the introduction of CT scanning had a greatly improved mortality rate, and their actual survival rate was 87.5% over 10 years. On the basis of this review, the authors consider that diagnosis at an early stage of the disease and chemotherapy, which can be an effective therapeutic alternative to radiation therapy, may improve not only the mortality rate but also the quality of life of patients with HN germ cell tumors.

[MR imaging of stereotactic thalamotomy using radiofrequency methods].

Sixty-eight stereotactic radiofrequency (rf) thalamic lesions in 57 patients with movement disorders were evaluated by magnetic resonance (MR) imaging. Postoperative periods ranged from 5 days to 4 years and 9 months. All 68 rf lesions were clearly detected on T2-weighted images (T2WI). Changes in signal intensity on T2WI were classified into five patterns, as follows: Pattern I: lesions with three concentric zones consisting of an inner hypointense, middle hyperintense and outer hypointense zone (31 lesions); Pattern II: lesions consisting of an inner hypointense and outer hyperintense zone (4 lesions); Pattern III: lesions consisting of an inner hyperintense and outer hypointense zone (27 lesion); Pattern IV: lesions of a hyperintense area alone (2 lesions); Pattern V: lesions of a hypointense area alone (4 lesions). The outer hypointense rim in Patterns I and III is thought to represent hemosiderin deposition. The abnormal signal intensity on T2WI caused by rf ranged from 2 to 12 mm in diameter, and lesions in the late phase were smaller than lesions in the early phase.

Localization of stereotactic targets by microrecordings of thalamic somatosensory evoked potentials.

To improve the localization of stereotactic targets, somatosensory evoked potentials (SEPs) were recorded from the thalamus and subthalamic area using a specially designed semimicroelectrode in 61 patients and a conventional "macroelectrode" in 17 patients. By means of the semimicroelectrode, median nerve stimulation evoked two distinct SEPs, consisting of a diphasic wave with a huge positivity restricted to the nucleus ventrocaudalis (Vc) and a triphasic wave of lower amplitude with a major negativity in the ventral part of the nucleus ventrointermedius (Vim) and nucleus ventrooralis posterior (Vop) as well as the subthalamic lemniscal pathway. The Vim-Vc junction could thus be clearly delineated by an abrupt transition of SEPs from one type to the other with a precision of 1 mm. The parvicellular part of the Vc (Vcpc), situated in its basal region, was distinguishable from the Vc proper by a significant reduction of the positivity elicited by stimulation of the median nerve and by a rapid growth of a diphasic SEPs to stimulation of the posterior tibial nerve. In the other thalamic nuclei, stimulation of the median nerve elicited triphasic SEPs of a very small amplitude, suggesting a volume conduction current from the lemniscal pathway. With the macroelectrode, the positivity in the Vc was sensitive to electrode manipulation and the thalamic nuclei could not be distinctly outlined. SEP monitoring using the semimicroelectrode significantly improved the precision of target localization, which allowed minimizing of the volume of the therapeutic lesion without losing surgical effectiveness, while avoiding complications associated with increased penetration of the coagulating electrode. It is suggested that recording serial thalamic SEPs with the semimicroelectrode is a practical method to refine stereotactic targets in the thalamus.

Origin and distribution of thalamic somatosensory evoked potentials in humans.

The distribution and generator sources of somatosensory evoked potentials (SEPs) in the thalamus and subthalamic area were studied, using a 'semi-microelectrode' during stereotaxic surgery on 34 patients with involuntary movements or intractable pain. Electrical stimulation was given to the median nerve at the wrist. Two distinct SEPs were evoked by contralateral stimulation. A high voltage (160 microV) positive SEP with a peak latency of 15.5 msec was strictly confined to the ventral part of the sensory relay nucleus (nucleus ventro-caudalis, V.c). A much lower voltage, positive-negative-positive triphasic SEP showed peak latencies of the initial positivity and the major negativity of 13.3 msec and 16.0 msec, respectively, and had maximal voltage (16 microV) in the ventralmost parts of the nucleus ventro-intermedius (V.im) and radiatio praelemniscalis (Ra.prl), and substantial potentials in the lemniscus medialis (L.m) and nucleus ventro-oralis posterior (V.o.p). The potential field of the triphasic SEP spread farther across the different thalamic nuclei and subthalamic region with identical configurations and peak latencies, but with decreasing amplitude. These findings suggest that the high voltage positive SEP reflects a postsynaptic potential generated by the V.c neurons, and the smaller triphasic SEP a presynaptic axonal potential generated in the rostral part of the lemniscal pathway, extending by means of volume conduction.

A case of ocular reticulum cell sarcoma responded to radiation therapy.

We report a case of reticulum cell sarcoma with involvement both in the brain and the eyes. The patient was a 49-year-old man. In 1985, he suffered steroid-resistant uveitis of unknown etiology in his right eye. After that, in 1986, he was aware of a weakness in the right arm and leg and of right sensory disturbances. Histopathological study of a biopsy specimen revealed a malignant lymphoma in the left thalamus, and the lesion was effectively treated by radiation to the brain. In 1987, he complained of visual disturbances in the left eye. According to the history and ocular manifestation, the disease was diagnosed as ocular reticulum cell sarcoma. Radiation therapy to his left orbital cavity was effective and inflammatory signs in the left eye subsided.

Effects of thalamic parafascicular stimulation on the periaqueductal gray and adjacent reticular formation neurons. A possible contribution to pain control mechanisms.

To investigate the mechanism of analgesic effect of electrical stimulation of the thalamic parafascicular nucleus (Pf), we studied modulations of neuronal activities in the periaqueductal gray (PAG) and the adjacent reticular formation (RF) in response to Pf electrical and peripheral noxious stimulations in the rat. Extracellular single unit activities were recorded from 129 neurons in the PAG and adjacent RF under light halothane anesthesia. Pf stimulation caused neuronal responses in approximately 80% of the PAG and adjacent RF neurons, and noxious stimulation in 75%, with predominant excitatory responses to either stimulation. When the responses to the two stimuli were tested in the same neurons (n = 69), 91% responding to noxious stimuli also responded to Pf stimuli, again with predominant excitatory responses to either stimulation. The PAG and adjacent RF neurons that were verified antidromically to project to the nucleus raphe magnus (NRM), showed a similar pattern of response (n = 20). These results suggest that a sizeable population of neurons in the PAG and adjacent RF receives excitatory effects from the Pf and noxious afferents, and that part of these neurons projects to the NRM, which inhibits the dorsal horn cells of the spinal cord (the descending pain suppression system). Thus, part of the mechanism of the analgesic effects of Pf stimulation is due to activation of the descending pain suppression system by exciting the PAG and adjacent RF neurons. A possible role of noxious afferents on the negative feedback to pain mediation through this descending system also has to be considered.