Hydrothermal impacts on trace element and isotope ocean biogeochemistry.

Hydrothermal activity occurs in all ocean basins, releasing high concentrations of key trace elements and isotopes (TEIs) into the oceans. Importantly, the calculated rate of entrainment of the entire ocean volume through turbulently mixing buoyant hydrothermal plumes is so vigorous as to be comparable to that of deep-ocean thermohaline circulation. Consequently, biogeochemical processes active within deep-ocean hydrothermal plumes have long been known to have the potential to impact global-scale biogeochemical cycles. More recently, new results from GEOTRACES have revealed that plumes rich in dissolved Fe, an important micronutrient that is limiting to productivity in some areas, are widespread above mid-ocean ridges and extend out into the deep-ocean interior. While Fe is only one element among the full suite of TEIs of interest to GEOTRACES, these preliminary results are important because they illustrate how inputs from seafloor venting might impact the global biogeochemical budgets of many other TEIs. To determine the global impact of seafloor venting, however, requires two key questions to be addressed: (i) What processes are active close to vent sites that regulate the initial high-temperature hydrothermal fluxes for the full suite of TEIs that are dispersed through non-buoyant hydrothermal plumes? (ii) How do those processes vary, globally, in response to changing geologic settings at the seafloor and/or the geochemistry of the overlying ocean water? In this paper, we review key findings from recent work in this realm, highlight a series of key hypotheses arising from that research and propose a series of new GEOTRACES modelling, section and process studies that could be implemented, nationally and internationally, to address these issues.This article is part of the themed issue 'Biological and climatic impacts of ocean trace element chemistry'.

Expansion in size of a terminal deletion: a paradigm shift for parental follow-up studies.

BACKGROUND: Parental studies are often necessary subsequent to the identification of a chromosome abnormality. The recommended studies are based on assumptions about how chromosome rearrangements occur. One such assumption is that deletion size is stable through generations. RESULTS: We have identified a family where a small subtelomeric deletion in a phenotypically and cytogenetically normal mother expanded nearly 10-fold into a clinically consequential and cytogenetically visible deletion in her affected daughter. CONCLUSION: Traditional parental follow-up studies would have not identified this expansion, but would have rather classified the deletion in the daughter as either de novo or benign. Only by sizing the deletion by array comparative genomic hybridisation in both the mother and the daughter was the expansion recognised. Previous assumptions about chromosome behaviour suggest that this phenomenon may have been easily missed in other cases of chromosomal deletions. Therefore, this case illustrates the need for more comprehensive analyses of parental chromosome structure when characterising an abnormality in a child.

Comparison of targeted and whole genome analysis of postnatal specimens using a commercially available array based comparative genomic hybridisation (aCGH) microarray platform.

PURPOSE: The University of Utah Comparative Genomic Hybridization Microarray Laboratory was one of the first US laboratories to offer comparative genomic hybridisation (CGH) microarray testing using a commercial platform in a clinical setting. Results for 1076 patients (1598 chips) are presented. METHODS: The Spectral Genomics/PerkinElmer Constitutional Chip (targeted array), SpectralChip 2600 (whole genome array) and a "Combo" chip (both arrays run simultaneously) were the tests offered. Abnormal results were confirmed by an alternative method, most often fluorescence in situ hybridisation. RESULTS: In 669 cases with known normal cytogenetics, an abnormal detection rate of 10.8% was observed, (5.3%, 12.2% and 14.1% for the Constitutional Chip, SpectralChip 2600 and Combo assay, respectively). Known copy number variants and single clone abnormalities are not included in these rates. Single clone abnormalities are reported separately. For 1076 total cases, we report an average abnormal rate of 16.9% (8.7%, 23.7% and 18.6% for the three assays). This rate includes characterisation of some abnormalities previously identified by cytogenetics. CONCLUSIONS: CGH microarray provides a likely aetiology for the clinical phenotype in many cytogenetically normal cases, and a whole genome array generally identifies copy number changes more effectively than a targeted chip alone.

Benign copy number changes in clinical cytogenetic diagnostics by array CGH.

A database of apparently benign copy number variants (bCNVs) detected by a Spectral Genomics Inc./PerkinElmer BAC array platform has been maintained through the University of Utah Comparative Genomic Hybridization laboratory since 2005. The target population for this database represents 1,275 patients with abnormal phenotypes, primarily children referred for developmental delay and mental retardation. These bCNVs are independent of any identified copy number abnormality detected. The most common 35 bCNVs observed and their frequencies are reported here, and a subset of ten of the patients studied was evaluated on a new oligonucleotide CNV array set designed by Agilent Technologies. There was a 76% concordance of calls for these 35 bCNVs detected by both array platforms in the same patients. The higher resolution of the Agilent oligonucleotide array compared to the BAC array allowed determination of the precise breakpoints of the observed CNVs, in addition to documentation of additional CNVs of smaller sizes. As expected, observed CNVs and their frequencies were generally consistent with those of other previously published and available databases, including the Database of Genomic Variants (http://projects.tcag.ca/variation/). The availability of these data should assist other clinical laboratories in the evaluation of CNVs of unknown clinical significance.

Rapid detection of rabies and rabies-related viruses by RT-PCR and enzyme-linked immunosorbent assay.

A rapid detection method for the six established genotypes of rabies and rabies-related viral RNA using RT-PCR-ELISA is described. The detection of digoxigenin-labelled amplified products is performed by solution hybridization to two specific, biotin-labelled, capture probes, which are complementary to the inner region of the amplification products. The capture probe and amplified product hybrid are then immobilised on a streptavidin-coated microtitre plate, bound products are detected by an anti-DIG Fab fragment conjugated to peroxidase, and colorimetric reaction automatically measured. This method was up to 100-fold more sensitive than Southern blot hybridization, detecting 0.00002 TCID50/ml of a genotype 1, classical rabies virus strain. The complete detection methodology from RT-PCR to PCR-ELISA detection could be completed within 10 h. Using this procedure, we were 100% successful in detecting 60 isolates from a representative selection of the six established genotypes from all over the world. This test is a useful additional tool for the detection of the rabies and rabies-related viruses, which is easy to perform, rapid and highly sensitive.

Rapid detection of viruses of the tick-borne encephalitis virus complex by RT-PCR of viral RNA.

Studies were performed to identify a pair of primers, specific for the tick-borne encephalitis (TBE) virus complex of the Flaviviridae, with which to develop a rapid and specific identification system based on reverse transcription and the polymerase chain reaction (RT-PCR). The specificity of a putative primer pair was examined by RT-PCR of representative viruses from other antigenic complexes of the Flaviviridae and by computer sequence homology checks. All viruses of the TBE complex tested, with a single exception, were identified by RT-PCR using the identified primer pair. Accumulated data suggest that one of the putative primers identified in these studies may have flavivirus group specificity. The advantages of such a primer in the development of identification systems for all virus complexes of the Flaviviridae is discussed.

Renal disease and occupational exposure to organic solvents: a case referent approach.

Several recent studies have suggested that a relation may exist between exposure to occupational organic solvents and diseases of the kidney--particularly malignancy and glomerulonephritis. Two case referent studies were undertaken in the West Midlands to investigate these possibilities. In the case of renal cancer 54 live cases of biopsy proved adenocarcinoma of the kidney were compared with an equal number of community based healthy referents matched for age, sex, place of residence, and socioeconomic and ethnic grouping. For glomerulonephritis, 50 biopsy proved cases were matched in the same manner with 50 referents. Fourteen other patients were also reviewed who, on biopsy, proved not to have glomerulonephritis. For both sets of cases and their referents each individual was interviewed and a detailed account obtained of medical history and environmental exposures. Exposure to solvents was assessed independently and "blind" in a semiquantitative way by an experienced occupational hygienist. Past exposure was estimated for 10 different solvent types and 17 material types. No relation was found between exposure to solvents and renal cancer or glomerulonephritis. In the case of renal cancer the numbers studied only precluded a fourfold excess risk. For glomerulonephritis, the study, although methodologically superior to most other published studies and of similar size, was of similar power to the renal cancer investigation.

Investigation of the mechanism of hepatotoxicity of N-methylformamide in mice: effects on calcium sequestration in hepatic microsomes and mitochondria and on hepatic plasma membrane potential.

N-Methylformamide is an antitumour drug with hepatotoxic properties. Three potential targets for hepatocellular toxic lesions caused by N-methylformamide were investigated: the mitochondrial and microsomal Ca2+ pumps and the functional integrity of the plasma membrane. The administration of N-methylformamide to mice caused a dramatic decrease in the ability of the liver mitochondria to sequester [45Ca2+]. This effect was dose-dependent and was not caused by dimethylformamide, N-hydroxymethylformamide or formamide. The microsomal Ca2+ pump was not affected by N-methylformamide. Incubations of isolated mitochondria with N-methylformamide for 1 hr also led to the inhibition of the Ca2+ sequestration. Incubation of isolated mouse hepatocytes with N-methylformamide did not cause changes in plasma membrane potential as measured using the lipophilic cation triphenylmethylphosphonium. Of the three targets studied, the mitochondrial Ca2+ pump may be the one through which N-methylformamide triggers the events leading ultimately to hepatic necrosis.

An investigation of the mechanism of hepatotoxicity of the antitumour agent N-methylformamide in mice.

N-Methylformamide (NMF) has been reported to cause liver damage in animals and man. This hepatotoxicity was characterized in BALB/c mice by the release of liver enzymes into the plasma and by histopathological examination of livers after single and repeated administration of NMF. Whereas plasma levels of sorbitol dehydrogenase were elevated dramatically 24 hr after 400 mg/kg given as a single dose, the glutathione content of the livers was not different from controls even after repeated administration. Liver damage was apparent on gross inspection and was defined as periacinar necrosis on histopathology. A dose of 100 mg/kg did not cause damage even after repeated injections on five consecutive days. The hypothesis that NMF is metabolized to a chemically reactive species was tested. Incubation of mouse hepatocytes with 7 mM NMF for 80 min produced a decrease in intracellular glutathione. Exposure of hepatocytes to NMF for 240 min led to the production of breakdown products of lipid peroxides at levels significantly above controls. However, incubation of microsomes or mitochondria with NMF and NADPH did not lead to raised levels of lipid peroxides. The effects described were specific to NMF as incubation of N,N-dimethylformamide, N-hydroxymethylformamide or formamide with hepatocytes did not result in glutathione depletion or increased lipid peroxidation. NMF undergoes extensive metabolism in vivo and the results indicate that NMF forms a chemically reactive metabolite, even though incubation of the drug with liver fractions or hepatocytes did not lead to metabolites at levels which were analytically identifiable.

Studies of the pharmacology of N-methylformamide in mice.

When 400 mg/kg of 14C-methyl-labeled N-methylformamide (NMF) was injected ip into mice, the curve for plasma concentration of radioactivity versus time was superimposable on the curve obtained by measuring unmetabolized NMF with gas-liquid chromatography during the first 24 hrs. Radioactivity in plasma was measurable for 8 days after NMF administration, but NMF was not measurable by gas chromatography beyond 24 hrs after administration. Radioactivity was eliminated from the plasma after 60 hrs, with an apparent half-life of 71.1 hrs. Of the radioactivity injected with NMF, 73.6% was recovered in the urine in 24 hrs; 26.4% of this was unchanged NMF. Three percent of the administered radioactivity was exhaled as 14CO2 in 7 hrs at a constant rate of 0.007% per min. One urinary metablite was a stable precursor of formaldehyde, which decomposed to formaldehyde only after alkaline hydrolysis and may well be N-(hydroxymethyl)-formamide. The areas under the plasma concentration versus time curve were estimated after ip, iv, and oral administration of NMF. The bioavailability of NMF was 1.01 after oral administration and 1.10 after ip administration.

Assessment of renal function during high-dose cis-platinum therapy in patients with ovarian carcinoma.

Five courses of cis-dichlorodiammine platinum (II) (100 mg/m2) were given to 22 patients with advanced stage III and IV ovarian cancer. Renal function was assessed by measurement of creatinine clearance, urinary osmolality and urinary B2-microglobulin (B2MG) in all patients, and by urinary alanine aminopeptidase (AAP) and N-acetyl-B-glucosaminidase (NAG) excretion in seven patients. Serum creatinine, creatinine clearance, urinary osmolality, and urinary B2-microglobulin were within the reference ranges and did not change significantly after five courses of cis-platinum in any patient. There was a significant increase in the urinary excretion of both enzymes (AAP and NAG) within 2 days of cis-platinum administration (NAG P less than 0.05 and AAP P less than 0.07). There was evidence of a cumulative effect during treatment for AAP (P less than 0.025).

Detection of virus particles by electron microscopy with polyacrylamide hydrogel.

The use of lyphogel to concentrate the number of virus particles in specimens for electron microscopic examination was studied in parallel with ultracentrifugation. One hundred faecal and urine samples were compared. Both methods had a similar sensitivity. Lyphogel was economical, simple, and rapid in use; in contrast to ultracentrifugation, it required relatively little material. The procedure could be done within a safety cabinet, and virus particles were morphologically undamaged by the process.