The effects of demographics, functioning, and perceptions on the relationship between self-reported and objective measures of driving exposure and patterns among older adults.

The exploratory study reported here was intended to examine: how strongly subjectively reported driving avoidance behaviors (commonly referred to as self-regulation) and exposure were related to their objectively measured counterparts and whether it depended on the specific behavior; the extent to which gender and age play a role in the association between subjectively reported driving avoidance behaviors and exposure and their objectively measured counterparts; and the extent to which demographics, health and functioning, driving-related perceptions, and cognition influence the association between subjective and objective driving avoidance behaviors overall. The study used data from the Longitudinal Research on Aging Drivers (LongROAD) study, a multisite, prospective cohort study designed to generate empirical data for understanding the role of medical, behavioral, environmental, and technological factors in driving safety during the process of aging. Objective driving measures were derived from GPS/datalogger data from 2131 LongROAD participants' vehicles. The corresponding subjective measures came from a comprehensive questionnaire administered to participants at baseline that asked them to report on their driving exposure, patterns, and other aspects of driving. Several other variables used in the analyses came from the comprehensive questionnaire and an inperson clinical assessment administered to participants at baseline. A series of simple linear and logistic models were fitted to examine the relationship between the subjective and objective driving measures of interest, and a multivariable analysis was conducted to examine the potential role of selected factors in the relationship between objective and subjective driving avoidance behaviors. Results of the models are presented and overall findings are discussed within the context of the existing research literature.

The impact of culture medium on the development and physiology of biofilms of Pseudomonas fluorescens formed on polyurethane paint.

Microbial biofilms cause the deterioration of polymeric coatings such as polyurethanes (PUs). In many cases, microbes have been shown to use the PU as a nutrient source. The interaction between biofilms and nutritive substrata is complex, since both the medium and the substratum can provide nutrients that affect biofilm formation and biodeterioration. Historically, studies of PU biodeterioration have monitored the planktonic cells in the medium surrounding the material, not the biofilm. This study monitored planktonic and biofilm cell counts, and biofilm morphology, in long-term growth experiments conducted with Pseudomonas fluorescens under different nutrient conditions. Nutrients affected planktonic and biofilm cell numbers differently, and neither was representative of the system as a whole. Microscopic examination of the biofilm revealed the presence of intracellular storage granules in biofilms grown in M9 but not yeast extract salts medium. These granules are indicative of nutrient limitation and/or entry into stationary phase, which may impact the biodegradative capability of the biofilm.

Bacterial sunscreen: layer-by-layer deposition of UV-absorbing polymers on whole-cell biosensors.

UV-protective coatings on live bacterial cells were created from the assembly of cationic and UV-absorbing anionic polyelectrolytes using layer-by-layer (LbL) methodology. A cationic polymer (polyallylamine) and three different anionic polymers with varying absorbance in the UV range (poly(vinyl sulfate), poly(4-styrenesulfonic acid), and humic acid) were used to encapsulate Escherichia coli cells with two different green fluorescent protein (GFP) expression systems: constitutive expression of a UV-excitable GFP (GFPuv) and regulated expression of the intensely fluorescent GFP from amphioxus (GFPa1) through a theophylline-inducible riboswitch. Riboswitches activate protein expression after specific ligand-RNA binding events. Hence, they operate as a cellular biosensor that will activate reporter protein synthesis after exposure to a ligand target. E. coli cells coated with UV-absorbing polymers demonstrated enhanced protection of GFP stability, metabolic activity, and viability after prolonged exposure to radiation from a germicidal lamp. The results show the effectiveness of LbL coatings to provide UV protection to living cells for biotechnological applications.

Supramolecular assembly of a biomineralizing antimicrobial peptide in coarse-grained Monte Carlo simulations.

Monte Carlo simulations are used to model the self-organizing behavior of the biomineralizing peptide KSL (KKVVFKVKFK) in the presence of phosphate. Originally identified as an antimicrobial peptide, KSL also directs the formation of biosilica through a hypothetical supramolecular template that requires phosphate for assembly. Specificity of each residue and the interactions between the peptide and phosphate are considered in a coarse-grained model. Both local and global physical quantities are calculated as the constituents execute their stochastic motion in the presence and absence of phosphate. Ordered peptide aggregates develop after simulations reach thermodynamic equilibrium, wherein phosphates form bridging ligands with lysines and are found interdigitated between peptide molecules. Results demonstrate that interactions between the lysines and phosphate drive self-organization into lower energy conformations of interconnected peptide scaffolds that resemble the supramolecular structures of polypeptide- and polyamine-mediated silica condensation systems. Furthermore, the specific phosphate-peptide organization appears to mimic the zwitterionic structure of native silaffins (scaffold proteins of diatom shells), suggesting a similar template organization for silica deposition between the in vitro KSL and silaffin systems.

Encapsulation of bacterial spores in nanoorganized polyelectrolyte shells.

Layer-by-layer assembly uses alternating charged layers of polyionic polymers to coat materials sequentially in a sheath of functionalized nanofilms. Bacterial spores were encapsulated in organized ultrathin shells using layer-by-layer assembly in order to assess the biomaterial as a suitable core and determine the physiological effects of the coating. The shells were constructed on Bacillus subtilis spores using biocompatible polymers polyglutamic acid, polylysine, albumin, lysozyme, gelatin A, protamine sulfate, and chondroitin sulfate. The assembly process was monitored by measuring the electrical surface potential (zeta-potential) of the particles at each stage of assembly. Fluorescent laser confocal microscopy and scanning electron microscopy confirmed the formation of uniform coatings on the spores. The coating surface charge and thickness (20-100 nm) could be selectively tuned by using appropriate polymers and the number of bilayers assembled. The effect of each coating type on germination was assessed and compared to native spores. The coated spores were viable, but the kinetics and extent of germination were changed from control spores in all instances. The results and insight gained from the experiments may be used to design various bioinspired systems. The spores can be made dormant for a desired amount of time using the LbL encapsulation technique and can be made active when appropriate.

Lysozyme catalyzes the formation of antimicrobial silver nanoparticles.

Hen egg white lysozyme acted as the sole reducing agent and catalyzed the formation of silver nanoparticles in the presence of light. Stable silver colloids formed after mixing lysozyme and silver acetate in methanol and the resulting nanoparticles were concentrated and transferred to aqueous solution without any significant changes in physical properties. Activity and antimicrobial assays demonstrated lysozyme-silver nanoparticles retained the hydrolase function of the enzyme and were effective in inhibiting growth of Escherichia coli, Staphylococcus aureus, Bacillus anthracis, and Candida albicans. Remarkably, lysozyme-silver nanoparticles demonstrated a strong antimicrobial effect against silver-resistant Proteus mirabilis strains and a recombinant E. coli strain containing the multiple antibiotic- and silver-resistant plasmid, pMG101. Results of toxicological studies using human epidermal keratinocytes revealed that lysozyme-silver nanoparticles are nontoxic at concentrations sufficient to inhibit microbial growth. Overall, the ability of lysozyme to assemble silver nanoparticles in a one-step reaction offers a simple and environmentally friendly approach to form stable colloids of nontoxic silver nanoparticles that combine the antimicrobial properties of lysozyme and silver. The results expand the functionality of nanomaterials for biological systems and represent a novel antimicrobial composite for potential aseptics and therapeutic use in the future.

Hybrid antimicrobial enzyme and silver nanoparticle coatings for medical instruments.

We report a method for the synthesis of antimicrobial coatings on medical instruments that combines the bacteriolytic activity of lysozyme and the biocidal properties of silver nanoparticles. Colloidal suspensions of lysozyme and silver nanoparticles were electrophoretically deposited onto the surface of stainless steel surgical blades and needles. Electrodeposited films firmly adhered to stainless steel surfaces even after extensive washing and retained the hydrolytic properties of lysozyme. The antimicrobial efficacy of coatings was tested by using blades and needles in an in vitro lytic assay designed to mimic the normal application of the instruments. Coated blades and needles were used to make incisions and punctures, respectively, into agarose infused with bacterial cells. Cell lysis was seen at the contact sites, demonstrating that antimicrobial activity is transferred into the media, as well as retained on the surface of the blades and needles. Blade coatings also exhibited antimicrobial activity against a range of bacterial species. In particular, coated blades demonstrated potent bactericidal activity, reducing cell viability by at least 3 log within 1.5 h for Klebsiella pneumoniae, Bacillus anthracis Sterne, and Bacillus subtilis and within 3 h for Staphylococcus aureus and Acinetobacter baylyi. The results confirmed that complex antimicrobial coatings can be created using facile methods for silver nanoparticle synthesis and electrodeposition.

Synthesis of bioinorganic antimicrobial peptide nanoparticles with potential therapeutic properties.

Amphiphilicity and cationicity are properties shared between antimicrobial peptides and proteins that catalyze biomineralization reactions. Merging these two functionalities, we demonstrate a reaction where a cationic antimicrobial peptide catalyzes self-biomineralization within inorganic matrices. The resultant antimicrobial peptide nanoparticles retain biocidal activity, protect the peptide from proteolytic degradation, and facilitate a continuous release of the antibiotic over time. Taken together, these properties demonstrate the therapeutic potential of self-synthesizing biomaterials that retain the biocidal properties of antimicrobial peptides.

Histidine ligand protonation and redox potential in the rieske dioxygenases: role of a conserved aspartate in anthranilate 1,2-dioxygenase.

The Rieske dioxygenase, anthranilate 1,2-dioxygenase, catalyzes the 1,2-dihydroxylation of anthranilate (2-aminobenzoate). As in all characterized Rieske dioxygenases, the catalytic conversion to the diol occurs within the dioxygenase component, AntAB, at a mononuclear iron site which accepts electrons from a proximal Rieske [2Fe-2S] center. In the related naphthalene dioxygenase (NDO), a conserved aspartate residue lies between the mononuclear and Rieske iron centers, and is hydrogen-bonded to a histidine ligand of the Rieske center. Engineered substitutions of this aspartate residue led to complete inactivation, which was proposed to arise from elimination of a productive intersite electron transfer pathway [Parales, R. E., Parales, J. V., and Gibson, D. T. (1999) J. Bacteriol. 181, 1831-1837]. Substitutions of the corresponding aspartate, D218, in AntAB with alanine, asparagine, or glutamate also resulted in enzymes that were completely inactive over a wide pH range despite retention of the hexameric quaternary structure and iron center occupancy. The Rieske center reduction potential of this variant was measured to be approximately 100 mV more negative than that for the wild-type enzyme at neutral pH. The wild-type AntAB became completely inactive at pH 9 and exhibited an altered Rieske center absorption spectrum which resembled that of the D218 variants at neutral pH. These results support a role for this aspartate in maintaining the protonated state and reduction potential of the Rieske center. Both the wild-type and D218A variant AntABs exhibited substrate-dependent rapid phases of Rieske center oxidations in stopped-flow time courses. This observation does not support a role for this aspartate in a facile intersite electron transfer pathway or in productive substrate gating of the Rieske center reduction potential. However, since the single turnovers resulted in anthranilate dihydroxylation by the wild-type enzyme but not by the D218A variant, this aspartate must also play a crucial role in substrate dihydroxylation at or near the mononuclear iron site.

Redox-dependent structural changes in archaeal and bacterial Rieske-type [2Fe-2S] clusters.

Proteins containing Rieske-type [2Fe-2S] clusters play important roles in many biological electron transfer reactions. Typically, [2Fe-2S] clusters are not directly involved in the catalytic transformation of substrate, but rather supply electrons to the active site. We report herein X-ray absorption spectroscopic (XAS) data that directly demonstrate an average increase in the iron-histidine bond length of at least 0.1 A upon reduction of two distantly related Rieske-type clusters in archaeal Rieske ferredoxin from Sulfolobus solfataricus strain P-1 and bacterial anthranilate dioxygenases from Acinetobacter sp. strain ADP1. This localized redox-dependent structural change may fine tune the protein-protein interaction (in the case of ARF) or the interdomain interaction (in AntDO) to facilitate rapid electron transfer between a lower potential Rieske-type cluster and its redox partners, thereby regulating overall oxygenase reactions in the cells.

X-ray crystal structure of benzoate 1,2-dioxygenase reductase from Acinetobacter sp. strain ADP1.

One of the major processes for aerobic biodegradation of aromatic compounds is initiated by Rieske dioxygenases. Benzoate dioxygenase contains a reductase component, BenC, that is responsible for the two-electron transfer from NADH via FAD and an iron-sulfur cluster to the terminal oxygenase component. Here, we present the structure of BenC from Acinetobacter sp. strain ADP1 at 1.5 A resolution. BenC contains three domains, each binding a redox cofactor: iron-sulfur, FAD and NADH, respectively. The [2Fe-2S] domain is similar to that of plant ferredoxins, and the FAD and NADH domains are similar to members of the ferredoxin:NADPH reductase superfamily. In phthalate dioxygenase reductase, the only other Rieske dioxygenase reductase for which a crystal structure is available, the ferredoxin-like and flavin binding domains are sequentially reversed compared to BenC. The BenC structure shows significant differences in the location of the ferredoxin domain relative to the other domains, compared to phthalate dioxygenase reductase and other known systems containing these three domains. In BenC, the ferredoxin domain interacts with both the flavin and NAD(P)H domains. The iron-sulfur center and the flavin are about 9 A apart, which allows a fast electron transfer. The BenC structure is the first determined for a reductase from the class IB Rieske dioxygenases, whose reductases transfer electrons directly to their oxygenase components. Based on sequence similarities, a very similar structure was modeled for the class III naphthalene dioxygenase reductase, which transfers electrons to an intermediary ferredoxin, rather than the oxygenase component.

Cloning and expression of the benzoate dioxygenase genes from Rhodococcus sp. strain 19070.

The bopXYZ genes from the gram-positive bacterium Rhodococcus sp. strain 19070 encode a broad-substrate-specific benzoate dioxygenase. Expression of the BopXY terminal oxygenase enabled Escherichia coli to convert benzoate or anthranilate (2-aminobenzoate) to a nonaromatic cis-diol or catechol, respectively. This expression system also rapidly transformed m-toluate (3-methylbenzoate) to an unidentified product. In contrast, 2-chlorobenzoate was not a good substrate. The BopXYZ dioxygenase was homologous to the chromosomally encoded benzoate dioxygenase (BenABC) and the plasmid-encoded toluate dioxygenase (XylXYZ) of gram-negative acinetobacters and pseudomonads. Pulsed-field gel electrophoresis failed to identify any plasmid in Rhodococcus sp. strain 19070. Catechol 1,2- and 2,3-dioxygenase activity indicated that strain 19070 possesses both meta- and ortho-cleavage degradative pathways, which are associated in pseudomonads with the xyl and ben genes, respectively. Open reading frames downstream of bopXYZ, designated bopL and bopK, resembled genes encoding cis-diol dehydrogenases and benzoate transporters, respectively. The bop genes were in the same order as the chromosomal ben genes of P. putida PRS2000. The deduced sequences of BopXY were 50 to 60% identical to the corresponding proteins of benzoate and toluate dioxygenases. The reductase components of these latter dioxygenases, BenC and XylZ, are 201 residues shorter than the deduced BopZ sequence. As predicted from the sequence, expression of BopZ in E. coli yielded an approximately 60-kDa protein whose presence corresponded to increased cytochrome c reductase activity. While the N-terminal region of BopZ was approximately 50% identical in sequence to the entire BenC or XylZ reductases, the C terminus was unlike other known protein sequences.

Restraint use patterns for older child passengers in Michigan.

The purpose of the present study was to conduct the first statewide direct observation survey of restraint use designed specifically for older child passengers (4-15 years of age). We also sought to determine the factors that were related to belt use by older child passengers in Michigan so that effective programs could be developed to promote use of restraint devices in this age group. The study found that older child restraint use was about 58% statewide. Restraint use was highest in regions where overall belt use was higher, when the driver was using a safety belt, in sport utility vehicles and vans/minivans, and in the front-right seating position. No difference in restraint use was found for the day of week, the sex of the child, or the type of site where data was collected. The study provides the largest scale analysis to date on older child occupant restraint use patterns. The data provide some much needed empirical data on the restraint use patterns of older child passengers for development of theory to better understand and predict lack of restraint use in this age group.

Characterization and evolution of anthranilate 1,2-dioxygenase from Acinetobacter sp. strain ADP1.

The two-component anthranilate 1,2-dioxygenase of the bacterium Acinetobacter sp. strain ADP1 was expressed in Escherichia coli and purified to homogeneity. This enzyme converts anthranilate (2-aminobenzoate) to catechol with insertion of both atoms of O(2) and consumption of one NADH. The terminal oxygenase component formed an alpha(3)beta(3) hexamer of 54- and 19-kDa subunits. Biochemical analyses demonstrated one Rieske-type [2Fe-2S] center and one mononuclear nonheme iron center in each large oxygenase subunit. The reductase component, which transfers electrons from NADH to the oxygenase component, was found to contain approximately one flavin adenine dinucleotide and one ferredoxin-type [2Fe-2S] center per 39-kDa monomer. Activities of the combined components were measured as rates and quantities of NADH oxidation, substrate disappearance, product appearance, and O(2) consumption. Anthranilate conversion to catechol was stoichiometrically coupled to NADH oxidation and O(2) consumption. The substrate analog benzoate was converted to a nonaromatic benzoate 1,2-diol with similarly tight coupling. This latter activity is identical to that of the related benzoate 1, 2-dioxygenase. A variant anthranilate 1,2-dioxygenase, previously found to convey temperature sensitivity in vivo because of a methionine-to-lysine change in the large oxygenase subunit, was purified and characterized. The purified M43K variant, however, did not hydroxylate anthranilate or benzoate at either the permissive (23 degrees C) or nonpermissive (39 degrees C) growth temperatures. The wild-type anthranilate 1,2-dioxygenase did not efficiently hydroxylate methylated or halogenated benzoates, despite its sequence similarity to broad-substrate specific dioxygenases that do. Phylogenetic trees of the alpha and beta subunits of these terminal dioxygenases that act on natural and xenobiotic substrates indicated that the subunits of each terminal oxygenase evolved from a common ancestral two-subunit component.

Trends in driver and front-right passenger safety belt use in Michigan: 1984-1998.

As part of Michigan's effort to track trends in safety belt use within the state over time, the University of Michigan Transportation Research Institute conducted 20 statewide surveys of safety belt use between 1984 and 1998. Results indicate that Michigan safety belt use trends for drivers and front-right passengers are similar to other statewide and national trends. Belt use in Michigan increased dramatically immediately after the state implemented its mandatory belt use law (secondary enforcement) followed by a smaller decline that leveled off at a rate more than 20 percentage points higher than before the law. Belt use was consistently higher among drivers than front-right passengers, older than younger front-outboard occupants, females than males, and front-outboard occupants exiting freeways than those stopped at local intersections. Examination of belt use trends in Michigan provides useful information for continued efforts to increase belt use in our state and for all states interested in meeting national goals for safety belt use for the year 2000 and beyond.

A statewide analysis of child safety seat use and misuse in Michigan.

This study was designed to determine frequency of child safety seat use for children under the age of 4 years in Michigan, as well as to determine types of misuse. No state has previously conducted such a statewide survey. A two-stage sampling plan was used with data collected through direct observation at a sample of 88 child-care centers and pediatric clinics throughout the state. Misuse data were collected at a subset of the sites through driver interview and visual and hands-on inspection. The study results indicate that 74.5% of children under 4 years of age in Michigan are in safety seats with use highest in vehicles driven by belted drivers, females, and drivers under 60 years of age. Some degree of seat misuse was found in 88.5% of the inspections. The most common types of misuse were related to snugness of fit (both in installing the seat into the vehicle and in securing the child), use of the safety belt locking clip, and the harness positioning clip. Drivers who had a high occurrence of misuse, as compared with drivers with lower misuse, had lower educational levels, removed the seat frequently from their vehicle, were not the parent of the child, and had children who were both younger and smaller. The results show that efforts to encourage people to use child safety seats have been successful.

Traditional healing and allopathic medicine: issues at the interface.

There is increasing interest in Native traditional healers and the possibility of their working in some form of relationship with the allopathic medical system. It represents a resurgence of effort in an area of great potential benefit to the Native community, but is rife with issues that could destroy the effort at any number of stages in the process. Issues related to professional and institutional responsibility, the power of medicalization, physical and philosophical interactions of the systems of healing, measures of effectiveness, issues of reimbursement, and many more must be dealt with in an intentional and thorough manner if the process is to be successful.

Integrated primary care.

The delivery of medical and health-related services tends to be compartmentalized and fragmented. If the goal is health and if an important part in achieving that goal is increasing the ability of the person, family, and community to care for themselves while providing cost-effective care, then it is imperative to integrate efforts across community and primary care arenas. The Southcentral Foundation is an Alaska Native organization focused on the delivery of community and primary care activities that continues to work at the challenges of integration and coordination. This paper discusses some of the efforts to date and hopes for the future.

Measures of perceived linear size, sagittal motion, and visual angle from optical expansions and contractions.

Using monocular observation, open-loop measurements were obtained of the perceptions of linear size, angular size, and sagittal motion associated with the terminal (largest or smallest) stimuli of repetitive optical expansions and contractions using 1-D or 2-D displays produced on a video monitor at a constant distance from the observer. The perceptions from these dynamic conditions were compared with those from static conditions in which the stimuli were of the same physical size and at the same physical distance as the terminal dynamic stimuli, but that were not part of the optical expansions or contractions. One result, as expected, was that the measures of perceived linear and angular size differed, but also, unexpectedly, some substantial errors were associated with the measures of perceived angular size. Another result was that the amount of size constancy was considerably less than was expected from the obtained amount of perceived motion in depth. Consistent with the latter result, it was found that the size-distance invariance hypothesis (SDIH), using the physical visual angles of the terminal stimuli, predicted only about half of the perceived motion in depth obtained with the dynamic changes. Using the obtained measures of perceived visual angles in the SDIH increased rather than decreased the error in predicting the amount of motion in depth as perceived. An additional experiment suggests that at least some of the error in the measurement of the perceived visual angle is a consequence of error in the perceived origin of the visual angles. The absence of the expected relation between size constancy and perceived motion in depth in the dynamic conditions is hypothesized to be due to cognitive processes associated with off-sized perceptions of the stimuli.

A comparison of two direct-observation methods for measuring daytime safety belt use.

This study compared two methods for measuring front-outboard shoulder-belt use: looking into vehicles when they stopped at a traffic control device (SVDO) and looking into vehicles as they traveled along a traffic corridor (MVDO). The reliability of the latter method has been questioned and certain surveys, such as the National Occupant Protection Use Survey (NOPUS), use both methods to estimate safety belt use. In one experiment, the methods were compared on overall belt use rates and reliability. A second experiment investigated the effect that vehicle speed had on an observer's ability to measure accurately belt use using the MVDO method. The results showed that daytime belt use rates between methods were nearly identical and inter-method reliability was quite high, indicating that front-outboard shoulder-belt use can be measured identically with either method. The second experiment showed that measurement accuracy was not affected by vehicle speeds of up to 60 mph and that overall accuracy was above 95%.