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The objective of this study was to identify the relative contribution of tenderness factors for three beef muscles with similar tenderness ratings. Longissimus lumborum (LL), tensor fascia latae (TF) and gastrocnemius (GC) were collected from 10 USDA low Choice beef carcasses and assigned to a 5 or 21 days aging period (n = 60). Sarcomere length, troponin-T degradation, collagen content, mature collagen crosslink density, intramuscular lipid content and trained panel analysis were measured. Correlation and multivariate regression analysis indicated each muscle has a specific tenderness factor that contributed to the overall tenderness evaluated by trained panelists. The equations indicated LL tenderness was driven by lipid content (P .05); TF tenderness was driven by collagen content (P .05). GC tenderness was driven by proteolysis (P .01), and only collagen content can be casually used as an overall tenderness predictor for all three cuts.

We apply a cascaded linear model analysis to a micro-channel plate x-ray framing camera. We establish a theoretical expression of the Noise Power Spectrum (NPS) at the detector's output and assess its accuracy by comparing it to the NPS of Monte Carlo simulations of the detector's response to a uniform illumination. We also demonstrate that fitting the NPS of experimental data against a parametric model based on this expression can yield valuable information on the imaging ability of framing cameras, offering an alternative approach to the usual method employed to measure their modulation transfer functions.

ObjectivesWoody breast is a myopathy observed in chicken breast meat (Pectoralis major) characterized by its tough and rubbery texture. However, the exact causation of woody breast texture is still unknown. We hypothesize that sarcoplasmic reticulum (SR) dysfunctionality early postmortem results in rapid leakage of intracellular calcium may partially contribute to the abnormal meat texture observed in woody breast meat. The objective of this preliminary study was to investigate this hypothesis.Materials and MethodsFourteen Ross line broiler breast fillets (7 severe woody breast and 7 normal) were collected at 3 h postmortem from a commercial processing plant located in the southeast United States. The 7 woody breast samples also exhibited moderate to severe white striping. The 7 normal samples did not exhibit any signs of white striping or woody breast. Each sample was trimmed, weighed, vacuum packaged and frozen at –20°C at approximately 8 h postmortem. One 1.9 cm strip across the cranial end of each fillet was fabricated and pulverized in liquid nitrogen to measure sarcomere length (Laser Scan Confocal Microscope with a 100x/NA 1.4 objective), calpain activity (immunoblotting for µ-calpain autolysis), proteolysis (immunoblotting for troponin-T degradation) and collagen content (hydroxyproline content). Purge was also collected from each sample to evaluate protein (bicinchoninic acid assay) and free calcium concentration (atomic absorption).ResultsWoody breast fillets were heavier than normal chicken breast fillets (522.9 vs. 446.9g; P < 0.05). Woody breast samples tended to have shorter sarcomeres (1.70 vs. 2.02 µm; P = 0.0543) and less intact troponin-T compared to normal breast samples (relative intact troponin-T band density: 49.98 vs. 56.97%; P = 0.0515) at 8 h postmortem. It was interesting to note that no µ-calpain band was detected through immunoblotting for both the woody breast and normal samples at 8 h postmortem. Other studies have found similar results as poultry µ-calpain autolyzed at a much rapid rate than µ-calpain in mammalian species. In addition, the purge from woody breast samples also had higher levels of free calcium compared to normal samples (6.2 vs. 4.2 nmol calcium/mg protein; P < 0.05). Lastly, there was more collagen present in the woody breast samples compared to normal chicken breast samples (3.89 vs. 2.08 mg collagen/g muscle tissue; P < 0.05).ConclusionThe results indicated that the cause of texture abnormality of woody breast may be the combined effects of more calcium being released from the SR early postmortem resulting in shorter sarcomere length and more collagen being deposited in the chicken breast meat. Additional research with the focus on SR integrity and functionality as well as collagen crosslinks are needed to further elucidate the basic mechanism of woody breast texture formation.

To transform undergraduate biology education, faculty need to provide opportunities for students to engage in the process of science. The rise of research approaches using next-generation (NextGen) sequencing has been impressive, but incorporation of such approaches into the undergraduate curriculum remains a major challenge. In this paper, we report proceedings of a National Science Foundation–funded workshop held July 11–14, 2011, at Juniata College. The purpose of the workshop was to develop a regional research coordination network for undergraduate biology education (RCN/UBE). The network is collaborating with a genome-sequencing core facility located at Pennsylvania State University (University Park) to enable undergraduate students and faculty at small colleges to access state-of-the-art sequencing technology. We aim to create a database of references, protocols, and raw data related to NextGen sequencing, and to find innovative ways to reduce costs related to sequencing and bioinformatics analysis. It was agreed that our regional network for NextGen sequencing could operate more effectively if it were partnered with the Genome Consortium for Active Teaching (GCAT) as a new arm of that consortium, entitled GCAT-SEEK(quence). This step would also permit the approach to be replicated elsewhere.

The immunoglobulin degrading enzyme of Streptococcus pyogenes, IdeS, is an unusual cysteine protease produced by group A streptococci for which the only known substrate is immunoglobulin G (IgG). To date, IdeS has not been found to cleave any of the known synthetic substrates that other cysteine proteases hydrolyse, thus making the development of an IdeS detection assay difficult. Furthermore, at high doses of substrate, product generation is inhibited potentially due to the need for a dimeric enzyme complex with IgG. In this study we have developed a mass spectral assay for IdeS activity based on the detection of an Mr approximately 25,300 Fc fragment that retains the ability to bind streptococcal protein G. Using this assay procedure, evidence for a multimeric enzyme-substrate complex was obtained as well as identifying isolated heavy chains as a non-substrate inhibitor of IdeS activity. Under appropriate experimental conditions the assay could be used to detect IdeS activity in bacterial culture media or in human plasma without a requirement for purified reactants. The availability of a rapid and sensitive assay for IdeS should facilitate the detailed biochemical characterization of this unusual bacterial cysteine protease.

Surface-enhanced laser desorption/ionization time-of-flight analysis was used to monitor both the kinetics and heterogeneity of product formation during the biotinylation of a number of model proteins and peptide targets. The selected molecules were the IgG-binding protein, protein A, human serum albumin, and a synthetic peptide corresponding to the N terminus of a streptococcal M1 protein. The extent of biotinylation was determined by kinetic analysis of the shift in molecular mass from the native material. Each residue modified by reaction with N-hydroxysuccinimide biotin resulted in an addition of approximately 341 amu to the native protein or polypeptide. The novelty of the method was in the ability to determine the molecular mass shift, without first separating the targeted molecule from the biotinylating reagent. The analysis was rapid, simple, and provided information on the average number of biotin molecules added and the homogeneity of the resulting product.

Binding proteins that have high affinities for mammalian plasma proteins that are expressed on the surface of bacteria have proven valuable for the purification and detection of several biologically important molecules from human and animal plasma or serum. In this study, we have isolated a high affinity albumin-binding molecule from a group G streptococcal isolate of bovine origin and have demonstrated that the isolated protein can be biotinylated without loss of binding activity and can be used as a tracer for quantification of human serum albumin (HSA). The binding protein can be immobilized and used as a selective capture reagent in a competitive ELISA format using a biotinylated HSA tracer. In this assay format, the sensitivity of detection for 50% inhibition of binding of HSA was less than 1 microg/ml. When attached to the bacterial surface, this binding protein can be used to deplete albumin from human plasma, as analyzed by surface-enhanced laser desorption ionization time of flight mass spectrometry.

Batrachochytrium dendrobatidis is a major pathogen of frogs worldwide, associated with declines in amphibian populations. Diagnosis of chytridiomycosis to date has largely relied upon histological and immunohistochemical examination of toe clips. This technique is invasive and insensitive particularly at early stages of infection when treatment may be possible. We have developed a real-time PCR Taqman assay that can accurately detect and quantify one zoospore in a diagnostic sample. This assay will assist the early detection of B. dendrobatidis in both captive and wild populations, with a high degree of sensitivity and specificity, thus facilitating treatment and protection of endangered populations, monitoring of pristine environments and preventing further global spread via amphibian trade.

Streptococcus pyogenes protein H ( sph ) is an immunoglobulin-binding protein present in the Mga regulon of certain M1 serotype isolates. Although sph is present in many strains, it is frequently not expressed. In this paper we show that protein H was highly expressed after bacteria were injected into the skin of mice and were recovered from the blood, kidney, or spleen at various times postinfection. The percentage of protein H-positive colonies increased with time, reaching 100% in the spleen and kidney within 24 to 72 h postinfection. The up-regulation of sph expression was also observed in a mga mutant.

Protein H is an immunoglobulin-binding protein expressed by certain M1 serotypes of Streptococcus pyogenes. In a recent study of invasive group A isolates, it was found that none of the 16 M1 serotype isolates analyzed expressed protein H on their surface despite the presence of the protein H gene (sph) in approximately one-third of the isolates. Selection of stable protein H-expressing variants could be achieved by infection of prtH(+) non-expressing strains into a mouse skin and recovering bacteria from the spleen. This effect was independent of the transcription regulator Mga, since a similar effect was noted in an mga(-) mutant. Thus, host passage of S. pyogenes can lead to stable high level expression of Protein H.

Rice flour-based batter (RFBB) formulations for chicken drumstick coating were developed as an alternative for traditional wheat flour-based batter (WFBB). Physicochemical properties and storage stability of selected RFBB were evaluated and compared to WFBB. Batter pickup of RFBB formulated in combination with oxidized corn starch and methylcellulose (MC) was not significantly different from that of WFBB. In contrast, batters with only rice and corn flour (60:40% flour weight) exhibited significantly higher pickup. Rice flour batter with 15% oxidized corn starch had the lowest batter pickup. All RFBB exhibited (P0.05) lower oil absorption than WFBB. The TBA values of RFBB and WFBB increased (P0.05) with increased frozen storage time at -40 C for 90 d. The RFBB with MC exhibited the lowest TBA values, whereas WFBB had the highest values. Microstructural analysis revealed that freezing caused structural deterioration of all batters, but the RFBB with MC exhibited less freezing tolerance than other samples. The total plate counts of immediately fried or frozen fried chicken stored for 90 d were less than 1 log cfu/g sample. The RFBB with 5% oxidized corn starch and MC can replace WFBB on fried drumsticks. Additionally, RFBB results in a healthier product due to lower fat absorption.

The convenience and appeal of battered or breaded products have resulted in a sales increase of 100% since 1980. Because of the rapid growth of the Asian-American population and increasing consumption of rice and rice products, rice flour is a logical alternative for wheat flour in traditional batter formulation. The effects of ingredients used in rice flour-based batters on adhesion characteristic for deep-fat fried chicken drumsticks were studied by laser scanning confocal microscopy (LSCM) and texture analysis. Raw chicken drumsticks were predusted with egg albumin powder before dipping into batters prepared from combinations of rice flour, yellow corn flour, oxidized cornstarch, methylcellulose, or xanthan gum. The drumsticks were fried at 175 +/- 5 C until the internal temperature reached at least 71 C. For LSCM, samples were fixed overnight and were sectioned by vibratome (200 micrometer) before viewing. Batter adhesion was determined using an attachment specifically designed for chicken drumsticks. Microstructural analysis showed that batter formulated with a 50:50 mixture of rice and corn flours adhered better to drumsticks than batter with other rice flour ratios. Xanthan gum (0.2%) or methylcellulose (0.3%) alone had poor adhesion to chicken skin. However, when combined with other ingredients, xanthan gum increased the amount of batter pick-up before frying by increasing viscosity. Egg albumin significantly facilitated batter adhesion. The results from texture analysis supported the microstructural studies. As rice flour ratio increased from 50 to 70%, the binding force decreased. Rice flour showed potential as an alternative to wheat flour for batter formulas when the appropriate levels of oxidized starch, xanthan gum, and methylcellulose were included in the formulation.

We have used new gel pore-water profilers and conventional seepage meters to determine the advective flux of water from an underwater saline seep into Lake Kinneret. The gel probes sampled pore waters from medium to coarse sands that could not be sampled by conventional coring methods. The anions Cl, Br, and SO4 were constant at levels just above those for the lake for 3-5 cm into the sediment due to wave action or other turbulent mixing processes. There was then a sharp increase in concentration to values of approximately 8,000 mg Cl liter(-1), 370 mg SO4 liter(-1), and 120 mg Br liter(-1) at a depth of similar to 8.5 cm. Using an advection-diffusion model, the linear interstitial advection velocity (LIV) of the groundwater into the lake was calculated to vary between 140 and 275 cm yr(-1). The LN values from conventional seepage flux meters at the same site were 30 and 164 cm yr(-1). Differences between the LIV measurements of these two methods may be due to a number of possible factors, including groundwater flux heterogeneity.

Several Au–Ag gossans occur over massive sulphide deposits in the Ordovician Tetagouche Group near Bathurst, New Brunswick. The Murray Brook and Heath Steele B zone goethite gossans were about 45 and 15 m thick, respectively, prior to mining. They contain no minerals suitable for radiometric age dating. Geologically they must be younger than the Devonian Acadian orogeny and older than the last glaciation. Paleomagnetic methods were used to analyse specimens from 29 sites, mostly from ore on the pit walls. Host rocks and sulphide mineralization retain characteristic remanent magnetization directions in magnetite and pyrrhotite with a variety of directions that include possible Devonian overprints. Fertile and barren gossan specimens at 21 sites retain antiparallel normal and reversed A characteristic remanence components in goethite and (or) hematite. The A direction is D = 357.7°, I = 61.7 °(α95 = 4.5°, k = 31). Its pole of 134.2°E, 85.2°N (dp = 5.4°, dm = 7.0°) falls on the North American apparent polar wander path and circumscribes the Earth's present rotational axis, indicating that the gossans formed during the Pliocene–Pleistocene. Examination of the site locations in the pits, along with the remanence polarity of their goethite and hematite A components, suggests the gossans formed during Chrons 1 and 2 only, or in the past 2.3 ± 0.3 Ma.

A lakebed seepage meter for measuring groundwater seepage rates in the nonlittoral zones of lakes was designed and field tested. Ease of installation and operation, sampling integrity, and year-round stability in ice-bound lake environments were the main factors governing design. The design of the meter incorporates a lake-bottom seepage meter connected by a flexible conduit hose to a sampling station -2 m below lake surface. The operation and design of the meter includes methods for minimizing and monitoring meter settlement, protecting meter components from trawling fishermen and nibbling fish (seepage bags), visually monitoring flux levels in seepage bags to determine optimum time for sampling, and easy detection by a small boat-mounted sonar unit. Results from a seepage meter survey in a small lake situated in complex glacial stratigraphy show that the system is effective in measuring very low flux rates and mapping complex inflow and outflow groundwater regimes in lake environments. Detailed lake studies in glaciated terrains allow classification of lakes, with regard to characteristics and relative contributions of groundwater and surface water inputs, into the six main types shown in Fig. 1. Although subtypes can be created within this classification, Fig. 1 shows the complexity of relationships that can exist between lakes and surrounding groundwater regimes. In some cases, lakes cannot be classed as one type year-round; for example, Jaquet’s (1976) studies of the groundwater regime around Snake Lake (Wisconsin) show that this lake has a hydrology much like type d (Fig. 1) in late summer, fall, and winter, but in spring the flow characteristics are represented better by type b. The lake I use to evaluate a new seepage meter design (Alexander Lake) was, from initial analyses, thought to be type d or type b. Later detailed seepage meter analysis showed it to be type b. Figure 2 schematically depicts two extremes of limnological regimes and groundwater processes on a continuum of lakes in glaciated environments. Although hydrologists generally accept that most groundwater that enters a lake system does so through the littoral zone

The Eighth Survey of Staffing in Cardiology was conducted with an index date of 30 September 1992. The total number of cardiologists in England and Wales, defined as individuals trained in the specialty and spending at least 40% of their professional time working in it, was 358. Of these 11 were part time, defined as six sessions or less, giving a number in whole time (or near whole time) equivalents of 352.5. The number of individuals increased from 1991 by 18 (5.3%). There were 71 cardiologists in Scotland and Northern Ireland, making a total for the United Kingdom of 429 individuals (423.5 whole time equivalents), which is 7.3 per million population. A total of 44 Districts serving 8.8 million people have no resident cardiologist. There has been little improvement since the 1991 survey. An additional 34 Districts with populations greater than 250,000 have only one cardiologist: we have clear evidence of inadequate provision of care in most of these, a situation that is inevitable within the resources provided. A wider threat to the provision of a satisfactory level of cardiological care throughout the United Kingdom will follow from changes in the organisation of the National Health Service and in the new requirements for training of future cardiologists because these changes will make major new demands on consultant time which cannot be met within existing resources. A crisis will be averted only if a rapid and major expansion of the consultant grade can be achieved.

We demonstrate the hohlraum radiation temperature and symmetry required for ignition-scale inertial confinement fusion capsule implosions. Cryogenic gas-filled hohlraums with 2.2 mm-diameter capsules are heated with unprecedented laser energies of 1.2 MJ delivered by 192 ultraviolet laser beams on the National Ignition Facility. Laser backscatter measurements show that these hohlraums absorb 87% to 91% of the incident laser power resulting in peak radiation temperatures of T(RAD)=300 eV and a symmetric implosion to a 100 μm diameter hot core.

Glycoprotein D (gD) of equine herpesvirus 1 (EHV-1) was expressed at the surface of insect cells infected by a recombinant baculovirus. EHV-1 gD was detected as multiple forms (56, 52, and 48 kDa) from 18 to 96 h postinfection. Laboratory animals inoculated with the recombinant EHV-1 gD developed neutralizing antibody responses against both EHV-1 and EHV-4.

Genomics and bioinformatics are dynamic fields well-suited for capturing the imagination of undergraduates in both research laboratories and classrooms. Currently, raw nucleotide sequence is being provided, as part of several genomics research initiatives, for undergraduate research and teaching. These initiatives could be easily extended and much more effective if the source of the sequenced material and the subsequent focus of the data analysis were aligned with the research interests of individual faculty at undergraduate institutions. By judicious use of surplus capacity in existing nucleotide sequencing cores, raw sequence data could be generated to support ongoing research efforts involving undergraduates. This would allow these students to participate actively in discovery research, with a goal of making novel contributions to their field through original research while nurturing the next generation of talented research scientists.

The Sixth Biennial Survey of Staffing in Cardiology was conducted in July 1990. This report summarises the data that were collected, together with the results of a survey of facilities in cardiology made in 1989. The total number of cardiologists in the United Kingdom, defined as individuals trained in the specialty and spending at least 40% of their time working in it, is now 323. Six individuals work part time only, making 320 whole time posts. This number has increased over the two years from 1988 to 1990 by 32, of which 23 work only in the specialty and nine as general physicians with a major interest in cardiology. The rate of increase in numbers over the past decade has been reasonably consistent with an average of approximately 4.4% per year. Thirty one districts in England and Wales still have no cardiologist and 13 other districts have little provision with an average of three (visiting) sessions each per week. The population in these 44 districts is 8.3 million. Scotland also has an inadequate distribution of service in the specialty. If recommendations for cardiac surgery and angioplasty made in the Fourth Report of a Joint Cardiology Committee of the Royal College of Physicians of London and the Royal College of Surgeons of England are to be met, we calculate that we need 63 more cardiologists in our major centres. To provide one cardiologist in every district hospital and two for larger districts would require 94 more specialists, making a total shortfall of 157 individuals. We have no excess of senior registrars to provide for a major expansion at consultant level. Time spent within the senior registrar (or academic equivalent) grade has tended steadily to decline and very few now reach the end of their contracts. The need for more individuals to pass through the senior registrar grade will be met in part by a planned reduction in the training period to three years. This will be inadequate, however, because projected retirements show that the number of consultant vacancies will increase sharply from 1997. We believe that additional senior registrar posts must be created if a serious shortfall in service provision by consultants is to be avoided. The provision of non-invasive facilities in cardiology is reasonable. The need for additional equipment for invasive cardiology has not been assessed. The number of physiological measurement technicians varies considerably between regions and is generally inadequate.

The c protein (Ibc) of group B streptococci (GBS) is associated with at least four antigens (alpha, beta, gamma, delta). To assess the virulence potential of these antigens, 255 GBS isolates recovered from septic neonates, healthy neonates, and pregnant women were serotyped and surveyed for reactivity with sera to c protein and the four associated antigens. A radioimmunoassay using intact bacteria was used to detect the GBS antigens. In contrast to earlier reports, most (66%) of the type III strains expressed the c protein. Except for the gamma antigen, none of the other c protein-associated antigens showed an increased association with pathogenic strains independent of the polysaccharide antigens. The gamma antigen was expressed by 15 of 41 c protein-positive early-onset strains and by 4 of 38 c protein-positive late-onset strains (P = .007). This association was independent of the type-specific antigen, suggesting a potential role for the gamma antigen as a virulence factor in GBS strains causing early-onset sepsis.

Plasminogen receptors have been identified on the surface of a number of prokaryotic and eukaryotic cells. A receptor demonstrating high affinity for plasmin with minimal reactivity with the native zymogen Glu-plasminogen has been identified on the surface of certain group A streptococci. In this study the group A streptococcal plasmin receptor has been solubilized and purified to homogeneity. The isolated protein was an Mr approximately 41,000 molecule which retained its ability to bind plasmin following solubilization and affinity purification on a column of enzymatically inactivated human plasmin. The isolated plasmin receptor was compared functionally, antigenically, and physicochemically to the secreted plasminogen activator, streptokinase, produced by the same organism. The Mr approximately 41,000 surface plasmin receptor was shown to be functionally and antigenically distinct from the Mr approximately 48,000 streptokinase molecule produced by the same strain and lacked any plasminogen activator activity. The streptokinase molecule produced by this strain was shown to be closely related to the plasminogen activator protein secreted by other group A and C streptococci. This study represents the first report of the isolation of a plasmin receptor, either prokaryotic or eukaryotic, with functional activity.

This paper reports an assessment of the tolerance of STC’s bipolar, CMOS and ‘merged’ bipolar/CMOS processes to gamma total dose, dose rate and neutron fluence radiation effects. The objective of the assessment was to characterise process performance under worst case and near to worst case criteria when subjected to nuclear radiation up to the tactical level in accordance with BS 9000. Total dose irradiation was performed on the bipolar, CMOS and BiCMOS technologies with 9 Mrads (Si), 10-12 krads (Si) and 10-30 krads (Si) performance levels reported, respectively. Under dose rate conditions, transient upset, latch-up and photocurrent are reported with latch-up immunity reported for the bipolar 2 process and simultaneous latch-up and transient upset reported for both the bulk CMOS and BiCMOS processes. Neutron fluence testing was performed with the onset of failure for the CMOS reported at the 5 × 1014 neutrons/cm level and onset of failure for the bipolar and BiCMOS reported in the range 5 × 1014 to 5 × 1014 neutrons/cm2.

Eight commercial semiconductor grade epoxy compounds that are used to encapsulate 1C (integrated circuit) devices have been evaluated for their ability to minimize the development of thermal stresses which can cause failure during device temperature cycling. Thermal expansion, dynamic modulus and adhesion studies are used to describe the mechanical interaction between the plastic package and the silicon device it surrounds. A “figure of merit” is defined for the development of stress on the 1C device as it is cooled after the packaging process. The stress is shown to be proportional to the product of three terms: (αp-αs) Ep (Tanch-T) where αp and αs are the expansion coefficients for the plastic and silicon, respectively, Ep is the modulus of the epoxy and Tanch is the temperature at which the epoxy becomes anchored to the silicon device during transfer molding. In addition, the importance of good adhesion between the epoxy encapsulant and the silicon device to prevent package cracking has been demonstrated by finite element analysis and a novel adhesion test.

[1] We report new Sr, S, and B isotopic data for evaporites (gypsum, anhydrite), carbonates, melt rocks, gneisses, and groundwaters recovered in and around the Lake St. Martin (LSM) impact structure, Interlake Region, Manitoba, Canada. The LSM meteorite impacted Devonian to Ordovician carbonates and sandstones of the eastern Williston Basin, resulting in partial melting of underlying Superior Province (∼2.5 Ga) gneisses of the Canadian Shield. Overlying the LSM melt rocks are red beds and evaporites (anhydrite/gypsum/glauberite) previously inferred to have been deposited during the Jurassic. The 87Sr/86Sr (lowest values cluster at 0.70836) and δ34SCDT (+23.7 ± 0.9‰) of the evaporites, combined with B isotope compositions of associated groundwaters (δ11BNBS951 = +25 to +28‰), are consistent with evaporite deposition within the impact structure near the edge of an ocean-connected salina. The establishment of a marine origin for the evaporites offers a method of age assignment using the secular variation of S and Sr isotopes in seawater. Comparison of Sr and S isotope results with the seawater curves precludes Jurassic deposition for the evaporites or correlation with Watrous and Amaranth formation evaporites, previously considered correlative with those at LSM. The lowest Sr and mean of S isotope values from the LSM evaporites are similar to seawater in the latest Devonian, consistent with conodonts recovered from carbonate breccia overlying melt rocks, and we suggest this as an alternative age of the evaporites. Data presented here preclude a Jurassic age for the evaporites and therefore for the impact event.

This document describes the development of a user-friendly and efficient groundwater management model of the Project Shoal Area (PSA and surrounding area that will allow the U.S. Department of Energy and State of Nevada personnel to evaluate the impact of proposed water-use scenarios. The management model consists of a simple hydrologic model within an interactive groundwater management framework. This framework is based on an object user interface that was developed by the U.S. Geological Survey and has been used by the Desert Research Institute researchers and others to couple disparate environmental resource models, manage temporal and spatial data, and evaluate model results for management decision making. This framework was modified and applied to the PSA and surrounding Fairview Basin. The utility of the management model was demonstrated through the application of hypothetical future scenarios including mineral mining, regional expansion of agriculture, and export of water to large urban areas outside the region. While the results from some of the scenarios indicated potential impacts to groundwater levels near the PSA and others did not, together they demonstrate the utility of the management tool for the evaluation of proposed changes in groundwater use in or near the PSA.

This paper illustrates a real environmental concern and draws attention to the fact that natural processes can mobilize thallium (Tl), a highly toxic metal, which may enter the food chain as a “hidden health killer” with severe health impacts on local human population. Natural processes may be exacerbated by human activities such as mining and farming, and may cause enrichment of Tl in the environment. In geochemically anomalous areas with concentrated levels of Tl in the surface environment (bedrocks, waters, soils, and crops), such as the Lanmuchang area in southwestern Guizhou Province, China, it is essential to establish base-level values and to pay heed to the geological context of “natural contamination,” as high concentrations of Tl in bedrocks/ores (6–35,000 mg/kg) can lead to enrichment of Tl in the aquatic system (0.005–1100 μg/l in groundwaters and 0.07–31 μg/l in surface waters) and soil layers (1.5–124 mg/kg). In sensitive areas such as the Yanshang area of southwestern Guizhou, elevated natural levels of Tl from bedrocks may also cause higher concentrations of Tl in the surface environment, and thus more attention must be paid to geoenvironmental management of human activities if socio-economic catastrophes are to be avoided. Due to high uptake of Tl by crops, Tl can be transferred from soils to crops and remarkably concentrated in food crops. Concentrations of 1–500 mg/kg Tl based on dry weight (DW) were determined in many food crops growing on Tl-contaminated arable soils from the Lanmuchang area. The daily intake of 1.9 mg of Tl from consumed food crops was estimated for the local adult inhabitant of Lanmuchang. Thus, Tl is regarded as a latent health hazard with potential risk of toxicity in humans within areas of “natural” contamination by Tl. Keywords: Thallium, Geoenvironment, Health hazard, Contamination, Chronic poisoning, Guizhou, China

Previous in vitro studies with transgenic and gene-knockout mice have shown that lenses with elevated levels of glutathione peroxidase (GPX)-1 activity are able to resist the cytotoxic effect of H(2)O(2), compared with normal lenses and lenses from GPX-1-deficient animals. The purpose of this study was to investigate the functional role of this enzyme in antioxidant mechanisms of lens in vivo by comparing lens changes of gene-knockout mice with age-matched control animals.In vivo lens changes were monitored by slit lamp biomicroscopy, and enucleated lenses were examined under a stereomicroscope in gene-knockout animals and age-matched control animals ranging in age from 3 weeks to 18 months. Transmission (TEM) and confocal microscopy were performed on different regions of lenses after the mice were killed at various times.Slit lamp images showed an increase in nuclear light scattering (NLS) in gene-knockout mice compared with control animals. TEM revealed changes in the nucleus as early as 3 weeks of age by the appearance of waviness of fiber membranes. With increasing age, there was greater distortion of fiber membranes and distension of interfiber space at the apex of fiber cells compared with control mice. The changes in nuclear fiber membranes were even more dramatic, as observed by confocal microscopy, which was performed on thicker sections. In contrast to the changes in the lens nucleus, the morphology of the epithelium and superficial cortex remained unchanged in knockout animals during the same experimental period, consistent with slit lamp observations. Stereomicroscopy of ex vivo lenses demonstrated a significant increase in opacification in gene-knockout mice relative to control animals of the same age. This effect became evident in mice aged 5 to 9.9 months and persisted thereafter in older animals, resulting in mature cataracts after 15 months.The results demonstrate the critical role of GPX-1 in antioxidant defense mechanisms of the lens nucleus. The increased NLS appears to be associated with damage to fiber membranes in the nucleus, which is particularly susceptible to oxidative challenge because of the deficiency of GPX-1. It is suggested that the lens membrane changes in the knockout animals may be due to the formation of lipid peroxides, which serve as substrates for GPX-1. Cataract development in gene-knockout mice appeared to progress from focal opacities, apparent at an earlier age, to lamellar cataracts between 6 and 10 months, and finally to complete opacification in animals older than 15 months. This is the first reported phenotype in GPX-1-knockout mice.

This study compared the pathology and infection pattern of streptococcal pyrogenic exotoxin B-positive (SpeB(+)) and SpeB-negative (SpeB(-)) isogenic variants of an M1 isolate of Streptococcus pyogenes in a mouse skin air sac model. SpeB(+) strains resulted in severe local tissue damage that extended from the epidermis through the subcutaneous layers, whereas isogenic SpeB(-) variants had reduced gross pathology. At the histologic level, differences in necrosis and host responses to each variant were apparent. Injection of purified SpeB alone into a skin air sac failed to induce any significant tissue damage; however, coinjection of the enzyme with either the wild-type or the speB mutant resulted in increased and accelerated tissue necrosis. Surprisingly, coinjection of the enzyme with the spleen-recovered SpeB(-) variant failed to induce a lesion.

To test the hypothesis that crystallin proteins may actively participate in the differentiation of lens epithelial cells into fiber cells.Primary epithelial cells from adult bovine lenses were cultured at 37 degrees C until reaching 95-100% confluency in approximately 4-7 days. Using osmotic lysis of pinocytic vesicles, cells were then loaded with proteins labeled or unlabeled with the fluorescent marker Texas Red (TR). Fetal bovine proteins loaded into cells were lens water soluble fractions, HPLC-purified alpha-, beta-, and gamma-crystallin fractions, or bovine serum albumin (BSA). Cultures were then monitored for morphological changes over a 7 day period.Both TR-labeled and unlabeled water-soluble and a-crystallin fractions from bovine lenses resulted in morphological changes to epithelial cells during the first two h postloading. These changes included aggregation of epithelial cells into raised multilayered cell masses, as well as several cells losing attachment to the dish. The initial changes were subsequently followed by elongation of cells within the mass and an increase in size of the mass, so that by 4 days postloading the multilayered, multicellular structures could be visualized with the unaided eye. Differentiation was confirmed within these structures by expression of MIP 26, beta- and gamma-crystallin. These changes did not occur in cultures containing cells originally loaded with beta- or gamma-crystallin fractions, or with cells loaded with BSA.The results support the hypothesis that alpha-crystallins may actively participate in the differentiation of lens epithelial cells into fiber cells.

Studies were performed to determine if p53 mutations identified in rheumatoid arthritis (RA) synovial tissue are dominant negative.Site-directed mutagenesis was used to produce 2 RA-derived mutants: asparagine--serine at codon 239 (N239S) and arginine--stop at codon 213 R213*). HS68 dermal fibroblasts were transfected with either empty vector, wild-type p53 cDNA (wt), or the N239S or R213* mutant p53 cDNA clones. Interleukin-6 (IL-6) and bax gene expression were determined by Northern blot analysis. Bax transcription was determined using a bax promoter/reporter gene construct (bax-luc).Transfection of HS68 cells with wt increased bax mRNA levels. This process was blocked by cotransfection with either mutant. The mutant p53 genes also increased IL-6 gene expression. Low levels of bax promoter activity were detected in HS68 cells co-transfected with bax-luc and empty vector, N239S, or R213*, indicating that the RA mutants lacked transcriptional activity. Transfection with wt and bax-luc led to a 10-fold increase in luciferase expression. When the wt gene was cotransfected with either of the mutants, there was a dose-dependent inhibition of bax promoter activity.These data indicate that at least 2 of the p53 mutants identified in RA joint samples are dominant negative and suppress endogenous wild-type p53 function.

The p53 tumor suppressor gene is overexpressed in synovial tissue (ST) from patients with longstanding rheumatoid arthritis (RA), and may contain somatic mutations. The aim of this study was to determine p53 expression in ST from RA patients in different stages of the disease, compared with disease controls.ST biopsy specimens were obtained from the knee joints of 31 RA patients in varying disease phases, 8 patients with reactive arthritis (ReA), 10 patients with inflammatory osteoarthritis (OA), and 6 control patients (4 with meniscus pathology, 2 with vascular insufficiency). ST was also obtained from the clinically uninvolved knee joints of 9 RA patients. Expression of p53 was determined by immunohistology with DO1 monoclonal antibody (mAb) in all patients and by Western blot analysis with DO7 mAb in a subgroup of the patients.The p53 protein was detected by immunohistology in 10 of the 13 patients with early RA (duration6 months) and in 12 of the 14 patients with longstanding RA (duration5 years). The p53 protein was also demonstrated in clinically uninvolved knee joints. Western blots revealed immunoreactive p53 in ST extracts from all RA patients. Expression of p53 was about twice as high in ST from patients with longstanding RA as in early RA samples, but the difference did not reach statistical significance. Small amounts of p53 were also detected in ST from ReA and OA patients, although the expression in RA synovium was significantly higher. Immunohistologic analysis of normal ST gave negative results for p53.This study shows that p53 overexpression is specific for RA, compared with OA and ReA. This phenomenon is probably secondary to increased production of wild-type p53 protein in response to DNA damage and secondary to somatic mutations caused by the genotoxic local environment in inflamed ST. Of interest, p53 overexpression can also be found in the earliest stages of RA and in clinically uninvolved joints.

Malignant ovarian tumours of the fibrothecoma group are rare. The clinicopathological features of a case of ovarian malignant fibrothecoma in which there was metastatic disease in the small intestine and peritoneum at presentation are described. A number of differential diagnoses were considered but positive immunohistochemical staining of the resected ovarian and small intestinal neoplasms with anti-inhibin was of value in confirming a sex cord-stromal tumour and in excluding other lesions. The two tumours were also ultrastructurally identical. Classical malignant fibrothecomas are said to show four or more mitotic figures per 10 high power fields (HPF). Although the intestinal secondary was mitotically active, the primary ovarian tumour contained only one to two mitoses per 10 HPF, showing that formal mitotic counts are not an absolute indicator of malignant behaviour in this group of tumours.

We sought to assess the angiographic outcome, complication rates and clinical features of percutaneous transluminal coronary angioplasty (PTCA) after failed thrombolysis for acute myocardial infarction."Rescue angioplasty" refers to mechanical reopening of an occluded infarct-related artery (IRA) after failed intravenous thrombolysis. Although the procedure is commonly performed, data describing its technical and clinical outcome are sparse. Early reports suggested that rescue PTCA is less often successful and produces more complications than primary PTCA. Other reports have described beneficial effects of successful rescue PTCA but adverse outcomes when PTCA is unsuccessful.Using data from the Global Utilization of Streptokinase and Tissue Plasminogen Activator for Occluded Coronary Arteries (GUSTO-1) angiographic substudy, we compared clinical and angiographic outcomes of 198 patients selected for a rescue PTCA attempt with those of 266 patients with failed thrombolysis but managed conservatively and, for reference, with those of 1,058 patients with successful thrombolysis.Patients offered rescue PTCA had more impaired left ventricular function than those in whom closed vessels were managed conservatively. Rescue successfully opened 88.4% of closed arteries, with 68% attaining Thrombolysis in Myocardial Infarction (TIMI) grade 3 flow. The interventions did not increase catheterization laboratory or postprocedural complication rates. Multivariate analysis identified severe heart failure to be a determinant of a failed rescue attempt. Successful rescue PTCA resulted in superior left ventricular function and 30-day mortality outcomes, comparable to outcomes in patients with closed IRAs managed conservatively, but less favorable than in patients in whom thrombolytic therapy was initially successful. The mortality rate after a failed rescue attempt was 30.4%; however, five of the seven patients who died after failed rescue PTCA were in cardiogenic shock before the procedure.Rescue PTCA tends to be selected for patients with clinical predictors of a poor outcome. It is effective in restoring patency. Patients who die after a failed rescue attempt are often already in extremis before the angioplasty attempt.

To visualize the structure and determine the continuity of lipid membranes in lens fiber cells (LFCs) from human aged normal and cataractous lenses.Thick sections from human nuclear cataracts and aged normal lenses were stained with the lipophilic probe DiI, and then analyzed by confocal microscopy. Staining patterns of membranes were observed in individual optical sections or three-dimensional projections of z-series taken in longitudinal section and cross-section of LFCs from different regions within the lens nucleus.DiI bound to and delineated the plasma membrane of LFCs from all regions of the lens nucleus. Three-dimensional projections of z-series from aged normal and cataractous lenses suggested that some of the stained lipid membranes were not continuous with LFC plasma membrane of cataractous lenses.The results obtained using these methods demonstrated that lipid membranes, discontinuous with the plasma membrane of LFCs, were indicative of a novel process occurring predominately in cataractous human lenses.

OBJECTIVES: To identify all patients with diabetes in a community using electronic record linkage of multiple data sources and to compare this method of case ascertainment with registers of diabetic patients derived from primary care. DESIGN: Electronic capture-recapture linkage of records included data on all patients attending hospital diabetes clinics, all encashed prescriptions for diabetes related drugs and monitoring equipment, all patients discharged from hospital, patients attending a mobile unit for eye screening, and results for glycated haemoglobin and plasma glucose concentrations from the regional biochemistry database. Diabetes registers from primary care were from a random sample of eight Tayside general practices. A detailed manual study of relevant records for the 35,144 patients registered with these eight general practices allowed for validation of the case ascertainment. SETTING: Tayside region of Scotland, population 391,274 on 1 January 1996. MAIN OUTCOME MEASURES: Prevalence of diabetes; population of patients identified by different data sources; sensitivity and positive predictive value of ascertainment methods. RESULTS: Electronic record linkage identified 7596 diabetic patients, giving a prevalence of known diabetes of 1.94% (0.21% insulin dependent diabetes, 1.73% non-insulin dependent): 63% of patients had attended hospital diabetes clinics, 68% had encashed diabetes related prescriptions, 72% had attended the mobile eye screening unit, and 48% had biochemical results diagnostic of diabetes. A further 701 patients had isolated hyperglycaemia (plasma glucose > 11.1 mmol/l) but were not considered diabetic by general practitioners. Validation against the eight general practices (636 diabetic patients) showed electronic linkage to have a sensitivity of 0.96 and a positive predictive value of 0.95 for ascertainment of known diabetes. General practice lists had a sensitivity of 0.91 and a positive predictive value of 0.98. CONCLUSIONS: Electronic record linkage was more sensitive than general practice registers in identifying diabetic subjects and identified an additional 0.18% of the population with a history of hyperglycaemia who might warrant screening for undiagnosed diabetes.

DNA damage induces p53 tumor suppressor gene expression and protein production, which in turn facilitates DNA repair or apoptosis. Wild-type p53 protein has a short half-life, so it is rarely detected in non-neoplastic tissue. Because DNA fragmentation is abundant in the intimal lining in rheumatoid arthritis (RA) synovial tissue (ST) using in situ end-labeling (Firestein GS, Yeo M, Zvaifler NJ: Apoptosis in rheumatoid arthritis synovium. J Clin Invest 1995, 96:1631-1638), we assessed ST p53 expression. Immunohistochemical analysis of fixed RA synovium using antibody PAb 1801 showed prominent p53 staining in the cytoplasm and nuclei of intimal lining cells. Noninflammatory and osteoarthritis (OA) ST had significantly less p53 in the lining. These data were confirmed by Western blot analysis of ST extracts, with abundant p53 found in RA compared with OA. p53 expression in cultured fibroblast-like synoviocytes (FLS) was then examined. Flow cytometry on permeabilized cells showed that RA FLS constitutively express p53 protein. Western blots showed that RA FLS expressed significantly more p53 than either OA FLS or dermal fibroblasts. Immunohistochemistry of FLS cultured in chamber slides localized the p53 to the cytoplasm of most resting FLS, with nuclear staining in only 10.7 +/- 2.4%. Exposure to hydrogen peroxide for increased nuclear staining to 70.7 +/- 12.8% after 8 hours (P = 0.003). These data indicate that p53 is overexpressed in RA ST in the intimal lining, which is the primary site of DNA damage, and is constitutively expressed by FLS.

The M protein has been postulated to be a major group A streptococcal (GAS) virulence factor because of its contribution to the bacterial resistance to opsonophagocytosis. Direct evidence of this was only provided for GAS strains which expressed a single M protein. The majority of GAS express additional, structurally similar M-related proteins, Mrp and Enn, which have been described as IgG- and IgA-binding proteins. To determine the involvement of Mrp and M protein in phagocytosis resistance, the mrp and emm genes from serotypes M2, M4, and M49 as well as from M-untypeable strain 64/14 were insertionally inactivated. The mrp and emm mutants were subjected to direct bactericidal assays. As judged by numbers of surviving colony-forming units, all mutant strains with the exception of the mrp4 mutant exhibited reduced multiplication factors as compared to the isogenic wild-type strains. Subsequent analysis of phagocytosis by flow cytometry, measuring association of BCECF/AM-labelled bacteria and granulocytes, paralleled the results from direct bactericidal assays regardless of whether isolated granulocytes or whole blood were utilized. Resistant wild-type GAS strains bound to less than 24% of granulocytes, whereas phagocytosis-sensitive controls attached to more than 90% of the white blood cells. 40 to 60% of the granulocytes associated with the mrp and emm mutants within 1 h of co-incubation. Kinetic data suggested that attachment to granulocytes proceeds faster for emm mutants than for corresponding mrp mutants. By adding a dihydro-rhodamine123 stain and measuring fluorescence induced by oxidative burst, the experimental data suggested that bacteria bound to granulocytes were also engulfed and integrated into phagolysosomes. Thus, these data indicated that, if present, both mrp and emm gene products contribute to phagocytosis resistance by decreasing bacterial binding to granulocytes.

To measure the effect of methotrexate (MTX) treatment in rheumatoid arthritis (RA) on the expression of synovial collagenase, stromelysin, and tissue inhibitor of metalloproteinase 1 (TIMP-1) gene expression in a prospective study.Serial percutaneous synovial biopsies (pretreatment and after 3-4 months) were performed on the knees of 8 patients (7 with RA, 1 with seronegative arthritis) who were beginning oral MTX therapy. Synovial gene expression was determined by quantitative in situ hybridization using computer-assisted image analysis.After therapy, patients had decreased joint counts, morning stiffness, and erythrocyte sedimentation rates. Synovial inflammation in the biopsy tissues was slightly decreased after therapy. In situ hybridization on pretreatment and posttreatment frozen sections was performed to quantify synovial messenger RNA (mRNA) levels. Collagenase gene expression significantly decreased after MTX therapy (P = 0.006) even though cell density in the region was unchanged. TIMP-1 and stromelysin mRNA levels were not changed by MTX therapy. To study the mechanism of MTX action in vitro, MTX-treated and control fibroblast-like synoviocytes were stimulated with interleukin-1 beta (IL-1 beta). MTX did not alter collagenase or TIMP-1 mRNA levels after IL-1 exposure.MTX therapy decreases collagenase gene expression but not TIMP-1 or stromelysin gene expression in the synovium. This action is probably an indirect effect due to an alteration in the synovial cytokine milieu, rather than a direct effect on gene expression.