In Memoriam

The Patient Self-Determination Act was implemented in December 1991. Before and after its implementation, we used a structured interview of 302 randomly selected patients to determine their awareness, understanding, and use of advance directives. Implementation of the Act did not have a major effect on these. Although more than 90% of patients were aware of the living will, only about a third selected the correct definition or the correct circumstances in which it applied, and less than 20% of patients had completed one. About a third of patients were aware of a Durable Power of Attorney for Health Care and chose the correct definition, and about half identified the correct circumstances in which it applies; less than 10% had completed such a document. Surprisingly, patients who said they had completed advance directives did not demonstrate better understanding of these documents. Our results indicate that many patients, including some who have completed advance directives, do not fully understand them. It may be unwise to regard these documents as carefully considered, compelling statements of patients'' preferences. Appropriate responses to our findings include increased public education, revising state statutes to bring them into congruence with public perception, and expanding the dialogue between physicians and patients.     

Metal specificity in DNA damage prevention by sulfur antioxidants

Metals such as Cu^I and Fe^II generate hydroxyl radical (OH) by reducing endogenous hydrogen peroxide (H₂O₂). Because antioxidants can ameliorate metal-mediated oxidative damage, we have quantified the ability of glutathione, a primary intracellular antioxidant, and other biological sulfur-containing compounds to inhibit metal-mediated DNA damage caused hydroxyl radical. In the Cu^I/H₂O₂ system, six sulfur compounds, including both reduced and oxidized glutathione, inhibited DNA damage with IC₅₀ values ranging from 3.4 to 12.4 μM. Glutathione and 3-carboxypropyl disulfide also demonstrated significant antioxidant activity with Fe^II and H₂O₂. Additional gel electrophoresis and UV-vis spectroscopy studies confirm that antioxidant activity for sulfur compounds in the Cu^I system is attributed to metal coordination, a previously unexplored mechanism. The antioxidant mechanism for sulfur compounds in the Fe^II system, however, is unlike that of Cu^I. Our results demonstrate that glutathione and other sulfur compounds are potent antioxidants capable of preventing metal-mediated oxidative DNA damage at well below their biological concentrations. This novel metal-binding antioxidant mechanism may play a significant role in the antioxidant behavior of these sulfur compounds and help refine understanding of glutathione function in vivo.     

Partial 9q trisomy associated with a 9,21 translocation

Summary A new case of partial trisomy 9q was found in a child presenting two de novo aberrations: a deletion of the long arms of 9 and a 9,21 translocation. A tentative cytogenetic explanation is put forward.     

Altered sensitivity to colchicine and PHA in human cultured cells

Summary PHA-stimulated lymphocytes cultivated from a pair of human monozygotic twins yielded mostly tetraploid cells when colchicine was not used to arrest the metaphases. The rate of tetraploidy was also enhanced by colchicine in fibroblasts cultured without PHA. In in situ condition, larger than usual cells were observed. Other defects found in parental lymphocyte cultures included C-anaphase cells and increased cell aggregation. These results suggest a membrane mutation resulting in hypersensitivity to PHA and variant response to colchicine.     

Preventing metal-mediated oxidative DNA damage with selenium compounds

Copper and iron are two widely studied transition metals associated with hydroxyl radical (˙OH) generation, oxidative damage, and disease development. Because antioxidants ameliorate metal-mediated DNA damage, DNA gel electrophoresis assays were used to quantify the ability of ten selenium-containing compounds to inhibit metal-mediated DNA damage by hydroxyl radical. In the Cu(I)/H(2)O(2) system, selenocystine, selenomethionine, and methyl-selenocysteine inhibit DNA damage with IC(50) values ranging from 3.34 to 25.1 μM. Four selenium compounds also prevent DNA damage from Fe(II) and H(2)O(2). Additional gel electrophoresis experiments indicate that Cu(I) or Fe(II) coordination is responsible for the selenium antioxidant activity. Mass spectrometry studies show that a 1?:?1 stoichiometry is the most common for iron and copper complexes of the tested compounds, even if no antioxidant activity is observed, suggesting that metal coordination is necessary but not sufficient for selenium antioxidant activity. A majority of the selenium compounds are electroactive, regardless of antioxidant activity, and the glutathione peroxidase activities of the selenium compounds show no correlation to DNA damage inhibition. Thus, metal binding is a primary mechanism of selenium antioxidant activity, and both the chemical functionality of the selenium compound and the metal ion generating damaging hydroxyl radical significantly affect selenium antioxidant behavior.     

Voronoi tessellation captures very early clustering of single primary cells as induced by interactions in nascent biofilms

Biofilms dominate microbial life in numerous aquatic ecosystems, and in engineered and medical systems, as well. The formation of biofilms is initiated by single primary cells colonizing surfaces from the bulk liquid. The next steps from primary cells towards the first cell clusters as the initial step of biofilm formation remain relatively poorly studied. Clonal growth and random migration of primary cells are traditionally considered as the dominant processes leading to organized microcolonies in laboratory grown monocultures. Using Voronoi tessellation, we show that the spatial distribution of primary cells colonizing initially sterile surfaces from natural streamwater community deviates from uniform randomness already during the very early colonisation. The deviation from uniform randomness increased with colonisation--despite the absence of cell reproduction--and was even more pronounced when the flow of water above biofilms was multidirectional and shear stress elevated. We propose a simple mechanistic model that captures interactions, such as cell-to-cell signalling or chemical surface conditioning, to simulate the observed distribution patterns. Model predictions match empirical observations reasonably well, highlighting the role of biotic interactions even already during very early biofilm formation despite few and distant cells. The transition from single primary cells to clustering accelerated by biotic interactions rather than by reproduction may be particularly advantageous in harsh environments--the rule rather than the exception outside the laboratory.     

Looking inside the box: using Raman microspectroscopy to deconstruct microbial biomass stoichiometry one cell at a time

Stoichiometry of microbial biomass is a key determinant of nutrient recycling in a wide variety of ecosystems. However, little is known about the underlying causes of variance in microbial biomass stoichiometry. This is primarily because of technological constraints limiting the analysis of macromolecular composition to large quantities of microbial biomass. Here, we use Raman microspectroscopy (MS), to analyze the macromolecular composition of single cells of two species of bacteria grown on minimal media over a wide range of resource stoichiometry. We show that macromolecular composition, determined from a subset of identified peaks within the Raman spectra, was consistent with macromolecular composition determined using traditional analytical methods. In addition, macromolecular composition determined by Raman MS correlated with total biomass stoichiometry, indicating that analysis with Raman MS included a large proportion of a cell''s total macromolecular composition. Growth phase (logarithmic or stationary), resource stoichiometry and species identity each influenced each organism''s macromolecular composition and thus biomass stoichiometry. Interestingly, the least variable peaks in the Raman spectra were those responsible for differentiation between species, suggesting a phylogenetically specific cellular architecture. As Raman MS has been previously shown to be applicable to cells sampled directly from complex environments, our results suggest Raman MS is an extremely useful application for evaluating the biomass stoichiometry of environmental microorganisms. This includes the ability to partition microbial biomass into its constituent macromolecules and increase our understanding of how microorganisms in the environment respond to resource heterogeneity.     

How infectious diseases got left out--and what this omission might have meant for bioethics

In this article, we first document the virtually complete absence of infectious disease examples and concerns at the time bioethics emerged as a field. We then argue that this oversight was not benign by considering two central issues in the field, informed consent and distributive justice, and showing how they might have been framed differently had infectiousness been at the forefront of concern. The solution to this omission might be to apply standard approaches in liberal bioethics, such as autonomy and the harm principle, to infectious examples. We argue that this is insufficient, however. Taking infectious disease into account requires understanding the patient as victim and as vector. Infectiousness reminds us that as autonomous agents we are both embodied and vulnerable in our relationships with others. We conclude by applying this reunderstanding of agency to the examples of informed consent and distributive justice in health care.     

Phylogenetic and Functional Heterogeneity of Sediment Biofilms along Environmental Gradients in a Glacial Stream

We used in situ hybridization with fluorescently labeled rRNA-targeted oligonucleotide probes concurrently with measurements of bacterial carbon production, biomass, and extracellular polymeric substances (EPS) to describe the bacterial community in sediments along a glacial stream. The abundance of sediment-associated Archaea, as detected with the ARCH915 probe, decreased downstream of the glacier snout, and a major storm increased their relative abundance by a factor of 5.5 to 7.9. Bacteria of the Cytophaga-Flavobacterium group were also sixfold to eightfold more abundant in the storm aftermath. Furthermore, elevated numbers of Archaea and members of the Cytophaga-Flavobacterium group characterized the phylogenetic composition of the supraglacial ice community. We postulate that glacial meltwaters constitute a possible source of allochthonous bacteria to the stream biofilms. Although stream water temperature increased dramatically from the glacier snout along the stream (3.5 km), sediment chlorophyll a was the best predictor for bacterial carbon production and specific growth rates along the stream. Concomitant with an increase in sediment chlorophyll a, the EPS carbohydrate-to-bacterial-cell ratio declined 11- to 15-fold along the stream prior to the storm, which is indicative of a larger biofilm matrix in upstream reaches. We assume that a larger biofilm matrix is required to assure prolonged transient storage and enzymatic processing of allochthonous macromolecules, which are likely the major substrate for microbial heterotrophs. Bacteria of the Cytophaga-Flavobacterium cluster, which are well known to degrade complex macromolecules, were most abundant in these stream reaches. Downstream, higher algal biomass continuously supplies heterotrophs with easily available exudates, therefore making a larger matrix unnecessary. As a result, bacterial carbon production and specific growth rates were higher in downstream reaches.