Comparative sensitivity to the fungicide tebuconazole of biofilm and plankton microbial communities in freshwater ecosystems.

Stream and lake ecosystems in agricultural watersheds are exposed to fungicide inputs that can threaten the structure and functioning of aquatic microbial communities. This research analyzes the impact of the triazole fungicide tebuconazole (TBZ) on natural biofilm and plankton microbial communities from sites presenting different degrees of agricultural contamination. Biofilm and plankton communities from less-polluted (LP) and polluted (P) sites were exposed to nominal concentrations of 0 (control), 2 and 20 µg TBZ L(-1) in 3-week microcosm experiments. Descriptors of microbial community structure (bacterial density and chlorophyll-a concentration) and function (bacterial respiration and production and photosynthesis) were analyzed to chart the effects of TBZ and the kinetics of TBZ attenuation in water during the experiments. The results showed TBZ-induced effects on biofilm function (inhibition of substrate-induced respiration and photosynthetic activity), especially in LP-site communities, whereas plankton communities experienced a transitory stimulation of bacterial densities in communities from both LP and P sites. TBZ attenuation was stronger in biofilm (60-75%) than plankton (15-18%) experiments, probably due to greater adsorption on biofilms. The differences between biofilm and plankton responses to TBZ were likely explained by differences in community structure (presence of extracellular polymeric substances (EPS) matrix) and microbial composition. Biofilm communities also exhibited different sensitivity levels according to their in-field pre-exposure to fungicide, with P-site communities demonstrating adaptation capacities to TBZ. This study indicates that TBZ toxicity to non-targeted aquatic microbial communities essentially composed by microalgae and bacteria was moderate, and that its effects varied between stream and lake microbial communities.

Enhanced solid-liquid separation of dairy manure with natural flocculants.

The aim of this study was to determine the effectiveness of natural flocculants to reduce solids and nutrient loads in dairy cow wastewater using solid-liquid separation; chitosan was used as a model. Its use efficiency and optimum application rate were determined using flushed dairy cow manure of varied strengths - 0.4%, 0.8%, 1.6%, and 3.2% total solids (TS) content. Treatments consisted of nine rates of chitosan. The flocculated manure was dewatered using 1-mm and 0.25-mm screens. Separation by screening alone was not effective; average efficiencies were about 60% for total suspended solids (TSS), 22% for total Kjeldahl nitrogen (TKN), and 26% for total phosphorus (TP). Mixing with chitosan before screening substantially increased separation. At optimum chitosan rate (0.5g/L for the highest strength effluent), separation efficiencies were >95% for TSS, >73% for TKN, and >54% for TP. The results of this study indicate that natural flocculants such as chitosan are useful for the solid-liquid separation treatment of livestock wastewater.

Learning from what went wrong--two case studies.

Safer design of process equipment can protect against unexpected events. Two case studies involving the design of a process vessel and the subsequent events will be reviewed. One case study will show how the original design minimized equipment damage from an operational error, and how additional safeguards will prevent recurrence. The second case study will show that over time small process changes can lead to an unexpected chemical reaction that results in a vessel rupture. We will also cover the additional safeguards added to prevent recurrence.

HIV lipodystrophy: review of the syndrome and report of a case treated with liposuction.

BACKGROUND: A syndrome characterized by loss of fat on the face and limbs, localized fatty deposits on the trunk, and metabolic disturbances is becoming increasingly recognized in the human immunodeficiency virus (HIV) patient population. OBJECTIVE: To increase awareness of this syndrome among dermatologists and dermatologic surgeons and to review its various treatment options, including liposuction. METHODS: We present a patient with HIV lipodystrophy syndrome who underwent tumescent liposuction. We also describe our experience with liposuction in the management of this condition and review the treatment options that have been proposed in the literature. RESULTS: In the medical management of HIV lipodystrophy, various agents have been utilized but most have yielded disappointing results. Preliminary evidence on the use of tumescent liposuction in these patients suggests that significant improvement in the cosmetic disfigurement can be achieved. CONCLUSION: This syndrome is common among HIV-infected patients and remains difficult to treat. Although medical therapy may be preferable in most patients, liposuction represents a viable option in selected individuals.

Propylthiouracil hypersensitivity: report of two patients with vasculitis and review of the literature.

Two patients with a hypersensitivity vasculitis in association with propylthiouracil (PTU) administration are described. Although both patients presented with a cutaneous eruption, our first patient suffered severe systemic manifestations and the second patient's involvement was primarily limited to the skin. Patients with a vascular hypersensitivity reaction to PTU typically present with constitutional symptoms, acral purpuric skin lesions, and variable involvement of multiple organ systems. The reaction is treated by urgent withdrawal of PTU and implementation of supportive measures and immunosuppressive agents, as necessary. Prompt recognition of this condition and initiation of appropriate therapy lead to complete recovery in most cases.

Sensitive determination of erythromycin in human plasma by LC-MS/MS.

A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the analysis of erythromycin in human plasma (EDTA as anticoagulant) was developed and validated. The concentration ranges were 0.5-50 and 50-5000 ng ml-1. The procedure involved alkalization of 0.5 ml of plasma, one step liquid-liquid extraction, dryness of the extract and reconstitution in 80:20 water:acetonitrile. An Inertsil ODS-2 5 microns, 3.0 x 50 mm column (Metachem) with a C8 guard column and isocratic mobile phase were used for liquid chromatography. The mobile phase consisted of 1:1 acetonitrile:water with 2 mM NH4OAc and 0.1% HOAc. A flow rate of 0.7 ml min-1 was used. The analysis time on LC-MS/MS for one sample was approximately 2 min. A Turbo-Ionspray source was interfaced between the HPLC and triple quadrupole mass spectrometer (Sciex API III Plus). MS/MS analysis used Multi-Reaction Monitoring (MRM) mode. The lowest limit of quantitation (LOQ) was 0.5 ng ml-1 with all Quality Control (QC) sample recoveries varying between 88 and 105%. Nine intraday and interday calibration curves were generated yielding correlation coefficients ranging from 0.995 to 1.000. Average recovery for erythromycin at 1 ng ml-1 was 105% (+/- 4.5%). Average recovery for the internal standard was 83-103%. Short-term and long-term stability in the freezer (-20 degrees C), bench stability, and stability after 3 freeze/thaw cycles at -20 and -80 degrees C were conducted. The samples were found to be stable under all conditions. The method developed and validated proved useful for clinical pharmacokinetic study sample analysis with high throughput due to its high sensitivity and very short analysis time.

Translational suppression by trinucleotide repeat expansion at FMR1.

Fragile X syndrome is the result of the unstable expansion of a trinucleotide repeat in the 5'-untranslated region of the FMR1 gene. Fibroblast subclones from a mildly affected patient, each containing stable FMR1 alleles with 57 to 285 CGG repeats, were shown to exhibit normal steady-state levels of FMR1 messenger RNA. However, FMR protein was markedly diminished from transcript with more than 200 repeats. Such transcripts were associated with stalled 40S ribosomal subunits. These results suggest that a structural RNA transition beyond 200 repeats impedes the linear 40S migration along the 5'-untranslated region. This results in translational inhibition by trinucleotide repeat expansion.

Human genes containing polymorphic trinucleotide repeats.

Expansions of trinucleotide repeats within gene transcripts are responsible for fragile X syndrome, myotonic dystrophy and spinal and bulbar muscular atrophy. To identify other human genes with similar features as candidates for triplet repeat expansion mutations, we screened human cDNA libraries with repeat probes and searched databases for transcribed genes with repeats. From both strategies, 40 genes were identified and 14 characterized. Five were found to contain repeats which are highly polymorphic including the N-cadherin, BCR, glutathione-S-transferase and Na+/K+ ATPase (beta-subunit) genes. These data demonstrate the occurrence of other human loci which may undergo this novel mechanism of mutagenesis giving rise to genetic disease.

Abnormalities in protein synthesis and degradation induced by extracellular pH in BC3H1 myocytes.

Metabolic acidosis impairs protein and amino acid metabolism in rat muscle. To examine how extracellular acidification affects cellular protein turnover, we studied the BC3H1 myocyte. At pH 7.1 vs. 7.4, intracellular pH was lower; the decrease was greater in cells incubated in N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid-tris(hydroxymethyl)aminomethane compared with bicarbonate buffer. We monitored degradation of proteins labeled with L-[14C]phenylalanine by measuring radioactivity released into media containing an excess of unlabeled phenylalanine. Extracellular acidification increased degradation compared with incubation at pH 7.4. Adding a physiological concentration of insulin (1 nM) decreased protein degradation at pH 7.1 and 7.4; a supraphysiological (71 nM) insulin concentration decreased degradation at pH 7.1 to the same rate as cells incubated at pH 7.4 without insulin. Compared with pH 7.4, protein synthesis decreased 29% at pH 7.2; at pH 7.6 it increased 129%. Insulin stimulated protein synthesis at all pHs, but at pH 7.4 the insulin-induced increase was less than the rate at pH 7.6 without insulin. Dexamethasone did not change protein breakdown regardless of the pH; it had variable effects on protein synthesis. Thus extracellular acidification causes marked changes in protein turnover in BC3H1 myocytes.

New TMJ clinical data and the implication on diagnosis and treatment.

A review of the literature and the presentation of 220 patients with acute temporomandibular disorders and a control group are presented. Condylar position in the fossae is related to clinical symptoms. Ten important clinical factors were recorded and the data compared without reference to specific condylar position in the fossae and then in relation to condylar displacement groups. The significantly high incidence of posterior condylar displacement and joint pain in the patients described in this report indicates that elimination of the term joint in our definition of this disorder may be premature.

Effects of microdialysis on brain metabolism in normal and seizure states.

The effect of intracranial microdialysis on brain glucose metabolism in control and kainic acid-treated rats was assessed by semi-quantitative [14C]2-deoxyglucose autoradiography. A dialysis fiber loop was implanted into the piriform cortex or a horizontal Vita fiber into the hippocampus, and 24 h later, fibers were perfused with Krebs-Ringer bicarbonate solution before and after injection of kainic acid (16 mg/kg, i.p.) [14C]2-Deoxyglucose was injected i.p. 3 h after the injection of kainic acid. Rats injected with kainic acid were initially lethargic and then proceeded through behavioral phases of staring, "wet-dog shakes", Straub tail, rearing, forepaw clonus, and, in some cases, tonic-clonic convulsions. Three hours after kainic acid, the fiber presence in the piriform cortex enhanced kainic acid-induced metabolic activity in areas adjacent to the fiber assembly, whereas the fiber in hippocampus attenuated kainic acid-induced metabolic activity in areas adjacent to the fiber assembly. The results indicate that intracranial microdialysis alters the already abnormal brain metabolism in a kainic acid-induced seizure state, but has no significant effect in the non-seizure control state.

Kainic acid-induced seizures: changes in brain extracellular ions as assessed by intracranial microdialysis.

The effect of kainic acid on extracellular [K+], [Ca2+], and [Na+] in the rat piriform cortex and hippocampus was studied by means of intracranial microdialysis. Either a dialysis fiber loop or horizontal Vita fiber were stereotaxically implanted within the piriform cortex or hippocampus, respectively. About 24 h later, fibers were perfused (1 ml/min) with Krebs-Ringer bicarbonate solution. Effluent samples were collected before (four at 30 min intervals), and after (six at 30 min intervals) administration of kainic acid (16 mg/kg, i.p.) or kainic acid vehicle. Kainic acid induced sequential signs of lethargy, staring, "wet-dog shakes," forepaw clonus, and tonic-clonic convulsions. In these awake free-moving rats, kainic acid induced a rapid and prolonged increase in extracellular [K+] and an apparent, but not statistically significant, decrease in extracellular [Ca2+] within the hippocampus. In the piriform cortex, kainic acid induced increases in extracellular [K+] and [Na+], which were associated with early pre-convulsive signs. In contrast to the pronounced ion changes commonly seen when the brain is activated by factors such as local application of excitatory substances or when the brain is made ischemic or hypoxic, extracellular ion concentrations are relatively well maintained during parenteral kainic acid-induced seizures.

Characterization of a new flavin metabolite from human urine.

A new flavin metabolite comprising approximately 5% of the total flavin of human urine was isolated and characterized using absorption and fluorescence spectra, oxidation-reduction and hydrolysis data, and ninhydrin reactions. The flavin is a derivative associated with a peptide residue in ester linkage from an amino acid carboxyl to the ribityl chain of riboflavin, probably at the 5'-terminus.

Flavin catabolites: identification and quantitation in human urine.

Riboflavin is the primary flavin excreted in human urine but significant amounts of 7 alpha-hydroxyriboflavin and lesser amounts of 8 alpha-hydroxyriboflavin are present and reflect tissue microsomal oxidations. A newly found flavin catabolite of an 8 alpha-sulfonyl type may reflect intake and/or turnover of such thioether-linked flavin as occurs in monoamine oxidase. Additionally, lesser amounts of 10-hydroxyethylflavin (indicative of intestinal microbial action on the vitamin) and traces of lumiflavin (arising from photodecomposition) constitute part of the remaining flavin, which acutely reflects level of intake.

Attenuation of cerebral glucose use in kainic acid-treated rats by diazepam.

Diazepam's impact on kainic acid seizure-induced local cerebral glucose utilization (LCGU) was assessed by a quantitative [14C]2-deoxyglucose method. Male rats were injected i.p. with either kainic acid (12 mg/kg) or its vehicle, 3 or 48 h before LCGU determination. Diazepam (3.2 mg/kg) or its vehicle were injected i.m. 15 min before, 1 and 2.5 h after kainic acid. Diazepam blocked kainic acid-induced overt convulsions, attenuated LCGU increases at 3 h and prevented 48 h LCGU decreases in piriform cortex and amygdala. LCGU in (% of vehicle): CA3 (438%), CA4 (537%) and CA1-ventral (340%) of hippocampus, interpeduncular nucleus (200%) and lateral lemniscus (213%) were still significantly above vehicle levels in the 3 h diazepam-kainic acid group. These results suggest that diazepam suppresses the spread of kainic acid-induced seizure activity from the proposed CA3 epileptogenic focus. In addition, diazepam reduces, but does not abolish, hypermetabolic activity at the foci itself.

Clarification and quantitation of primary (tissue) and secondary (microbial) catabolites of riboflavin that are excreted in mammalian (rat) urine.

Riboflavin derivatives were quantitated and identified in urine of rats fed 0, 2 and 6 micrograms riboflavin/g diet per day both with and without added succinyl sulfathiazole for 6 wk. Two rats from each dietary group were placed in metabolic cages and urine was collected in the dark for 24 h. On the fourth week, a third animal from each group received an i.p. injection of [2-14C]riboflavin before being placed in a metabolic cage and urine collected in the dark for 48 h. Urine samples were extracted with phenol for flavin components and with chloroform for lumichrome and derivatives. Riboflavin was the predominant flavin excreted by rats in all dietary groups, followed by hydroxymethylriboflavins and smaller amounts of flavin mononucleotide (FMN), lumiflavin and 10-hydroxyethylflavin. Carboxylumichromes accounted for 5-10% of the total flavin-derived fluorescence in urine of rats fed 2 and 6 micrograms riboflavin/g diet and were reduced to approximately 3% when sulfathiazole was added to the base diets. Carboxylumichromes were absent from urine of riboflavin-deficient rats. Riboflavin accounted for 85-90% of the recovered radioactivity of all radioactive urine extracts; no radioactively labeled carboxylumichromes were detected. These results indicate that hydroxymethylriboflavins are primary catabolites of riboflavin derived from tissue microsomal oxidations, whereas carboxylumichromes reflect the continued oxidation of ring hydroxymethyl functions plus gut microbial cleavage of the side chain of flavin.

High-performance liquid chromatography of biotin and analogues.

Biotin, analogues, and chemical intermediates were separated by high-performance liquid chromatography (HPLC) using reversed-phase and anion-exchange chromatographic conditions. Reversed-phase separations provided a wide range of retention times and resolution of nearly all the biotin compounds from mixtures of the analogues. Anion-exchange separations gave generally shorter retention times as compared to reversed-phase separations and greater resolution between biotin l- and d-sulfoxide. However, fewer analogues were resolved from mixtures of the compounds with anion-exchange HPLC.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD)-induced immunotoxicity.

The selective toxicity of TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin) for the thymus, consisting primarily of immature T-cells, led us to search for an analogous selective toxicity for the immature B-lymphocytes in the bone marrow. In the dose-response study C57B1/6 male mice were injected with either vehicle alone (corn oil), 30, 60, or 120 micrograms/kg of TCDD i.p. The mice were killed by cervical dislocation 7 days later. In the time-response study, mice were injected with either saline or 120 micrograms/kg i.p. TCDD, 3, 7, 14, or 21 days before killing. In both studies, the following were analyzed: change in body weight, thymus weight, spleen and bone marrow cellularity, and spleen and marrow B-lymphocyte function, measured using the in vitro B-lymphocyte colony forming unit in culture assay, with the mitogen lipopolysaccharide (LPS) from Salmonella typhosa, and the in vitro plaque forming cell assay, with the thymus independent antigen, TNP-LPS. In the dose-response study there was a reduction in thymic weight, spleen B-cell functional response (per spleen), and bone marrow B-cell functional response to 14%, 35-54%, and 20-32% of control, respectively, at a dosage of 120 micrograms/kg. In the time-response study, thymic weight and bone marrow B-cell functional response (per femur) were reduced to 6% and 18% of control, respectively, at day 21. The results indicate that TCDD was selectively more toxic to the immature B-cells in the bone marrow than the more mature B-cells in the spleen. This immunotoxicity was dose-dependent.