The effects of biostimulation and bioaugmentation on crude oil biodegradation in two adjacent terrestrial oil spills of different age, in a hyper-arid region.

This study deals with two adjacent terrestrial oil spills, with similar properties, located in a hyper-arid region in Israel, one from 1975 and the other from 2014. It tests the effect of biostimulation on crude oil degradation in both spills and whether biostimulated sediments from the 1975 spill can bioaugment crude oil degradation in the 2014 spill. Soil hydrophobicity, expressed as Water Drop Penetration Time (WDPT), and Gasoline Range Organics (GRO) and Diesel Range Organics (DRO) content in sediments were measured in one-month ex-situ experiments. No significant reduction in hydrophobicity and GRO + DRO content was observed in non-biostimulated controls. A combined treatment of mineral fertilization at t0 and maintaining 50% water saturation, significantly accelerated the decrease in hydrophobicity and GRO + DRO content in sediments of both spills. The addition of biostimulated sediments from the 1975 spill failed to accelerate the reduction of GRO + DRO content and hydrophobicity in the 2014 spill. Surprisingly, the GRO + DRO degradation rate in biostimulated sediments from the 2014 spill was 36% higher than in biostimulated sediments from the 1975 spill. Crude oil composition in both spills changes during its degradation and is characterized by an increase in the GRO fraction. To a certain range, WDPT was found to serve as a reliable indicator for oil content in the soil. We conclude that even in a hyper-arid region, oil bio-degradation capabilities develop in a relatively short time. Moreover, while biostimulation was effective in accelerating biodegradation, bioaugmentation with biostimulated sediments from a nearby older spill was found ineffective.

MLH1 mediates PARP-dependent cell death in response to the methylating agent N-methyl-N-nitrosourea.

BACKGROUND: Methylating agents such as N-methyl-N-nitrosourea (MNU) can cause cell cycle arrest and death either via caspase-dependent apoptosis or via a poly(ADP-ribose) polymerase (PARP)-dependent form of apoptosis. We wished to investigate the possible role of MLH1 in signalling cell death through PARP. METHODS: Fibroblasts are particularly dependent on a PARP-mediated cell death response to methylating agents. We used hTERT-immortalised normal human fibroblasts (WT) to generate isogenic MLH1-depleted cells, confirmed by quantitative PCR and western blotting. Drug resistance was measured by clonogenic and cell viability assays and effects on the cell cycle by cell sorting. Damage signalling was additionally investigated using immunostaining. RESULTS: MLH1-depleted cells were more resistant to MNU, as expected. Despite having an intact G(2)/M checkpoint, the WT cells did not initially undergo cell cycle arrest but instead triggered cell death directly by PARP overactivation and nuclear translocation of apoptosis-inducing factor (AIF). The MLH1-depleted cells showed defects in this pathway, with decreased staining for phosphorylated H2AX, altered PARP activity and reduced AIF translocation. Inhibitors of PARP, but not of caspases, blocked AIF translocation and greatly decreased short-term cell death in both WT and MLH1-depleted cells. This MLH1-dependent response to MNU was not blocked by inhibitors of ATM/ATR or p53. CONCLUSION: These novel data indicate an important role for MLH1 in signalling PARP-dependent cell death in response to the methylating agent MNU.

Variation in the extent of microsatellite instability in human cell lines with defects in different mismatch repair genes.

Mismatch repair deficiency results in the elevation of mutation rates in tumors, which is especially pronounced in simple repeat sequences (microsatellites). We have investigated the relationship between microsatellite mutagenesis and certain combinations of mutations in mismatch repair genes, using a frameshift reversion assay to determine the spontaneous mutation rates of a dinucleotide microsatellite in two cancer cell lines, HCT116, which has defects in hMLH1 and hMSH3, and HEC-1-A, which has defects in hPMS2 and hMSH6. We found a 10-fold difference in mutation rates between these two cell lines. In addition, a mutant hPMS2 allele, PMS134, which has been reported to have a dominant negative effect, was expressed in mismatch repair-proficient telomerase-immortalized hTERT-1604 fibroblasts and mutation rates were determined. Expression of PMS134 did not elevate mutation rates in hTERT-1604. Combined, these results suggest that mutations in different mismatch repair genes can lead to varying degrees of microsatellite instability. It is also likely that there is heterogeneity in the mutations that are acquired in the absence of mismatch repair, such that some mismatch repair-defective cancer cells also contain mutations in other genes coding for proteins involved in the maintenance of genetic stability.

Microsatellite mutation rates are equivalent in normal and telomerase-immortalized human fibroblasts.

Telomerase-expressing human fibroblasts generally have the same properties as normal cells, except that they have an indefinite life span in culture. We have introduced a dinucleotide repeat sequence into the telomerase-expressing hTERT-1604 cell line and characterized the rates and types of frameshift mutations within this microsatellite. These data have been compared with those in diploid fibroblasts with a finite life span. The rates of mutation were found to be similar in the two cell types, indicating that DNA mismatch repair and other cellular processes responsible for maintenance of mutational stability are not disrupted by telomerase immortalization.

Exploring protein sequence space using knowledge-based potentials.

Knowledge-based potentials can be used to decide whether an amino acid sequence is likely to fold into a prescribed native protein structure. We use this idea to survey the sequence-structure relations in protein space. In particular, we test the following two propositions which were found to be important for efficient evolution: the sequences folding into a particular native fold form extensive neutral networks that percolate through sequence space. The neutral networks of any two native folds approach each other to within a few point mutations. Computer simulations using two very different potential functions, M. Sippl's PROSA pair potential and a neural network based potential, are used to verify these claims.

The geometry of shape space: application to influenza.

Shape space was proposed over 20 years ago as a conceptual formalism in which to represent antibody/antigen binding. It has since played a key role in computational immunology. Antigens and antibodies are considered to be points in an abstract "shape space", where coordinates of points in this space represent generalized physico-chemical properties associated with various (unspecified) physical properties related to binding, such as geometric shape, hydrophobicity, charge, etc. Distances in shape space between points representing antibodies and (the shape complement) of antigens are assumed to be related to their affinity, with small distances corresponding to high affinity. In this paper, we provide algorithms, related to metric and ordinal multidimensional scaling algorithms first developed in the mathematical psychology literature, which construct explicit, quantitative coordinates for points in shape space given experimental data such as hemagglutination inhibition assays, or other general affinity assays. Previously, such coordinates had been conceptual constructs and totally implicit. The dimension of shape space deduced from hemagglutination inhibition assays for influenza is low, approximately five dimensional. The deduction of the explicit geometry of shape space given experimental affinity data provides new ways to quantify the similarity of antibodies to antibodies, antigens to antigens, and the affinity of antigens to antibodies. This has potential utility in, e.g. strain selection decisions for annual influenza vaccines, among other applications. The analysis techniques presented here are not restricted to the analysis of antibody-antigen interactions and are generally applicable to affinity data resulting from binding assays.

Potential particulate impacts at the Grand Canyon from northwestern Mexico.

Project MOHAVE was a major air quality and visibility research program conducted from 1990 to 1999 to investigate the causes of visibility impairment in the Grand Canyon National Park region. At Meadview, a remote monitoring site just west of the Grand Canyon National Park, on September 1 and 2, 1992, the concentrations of sulfate (3.1 and 4.3 microg sulfate/m3) were the highest seen in 6 years of monitoring at this site. During this period, the concentrations of SO2 at Meadview were also abnormally high and approximately three times the sulfate concentrations, on a nmol/m3 basis. High concentrations of sulfate and SO2 extended south into southern Arizona and northwestern Mexico. Based on ambient atmospheric conditions, emissions from the Mohave Power Project (MPP) 110 km upwind of Meadview could not have been responsible for the majority of the regionally observed sulfur oxides. The geographical distribution of SO2 and sulfate, and available source information suggest that northwestern Mexico was a significant source of the unusually high observed sulfur oxides. A CMB model developed during Project MOHAVE was used to apportion sulfur oxides at Meadview and other sampling sites throughout the study region for August 31-September 2, 1992. The results indicate that the contribution of MPP to sulfate at Meadview was typical. However, the transport of SOx from northwestern Mexico was elevated throughout much of the region during this time period. This led to the large increase in sulfate concentrations at Meadview on September 1 and 2. These results indicate that emissions from Mexico can be a significant source of particulate material in the Grand Canyon.

Mothers with disabilities: in their own voice.

OBJECTIVES: The purpose of this study was to gain an understanding of the personal experience of women with disabilities engaged in the occupation of mothering and their perceptions of their interpersonal environment, including interaction with family, professional caregivers, and the community. METHOD: In this qualitative study, in-depth interviews were conducted with a diverse sample (n = 8) of mothers with disabilities or chronic illnesses to uncover the nature of their mothering experience. Data were analyzed for themes with the constant comparison approach of grounded theory. RESULTS: The quality of the participants' experience varied with the degree of perceived similarity or dissimilarity with other mothers and acceptance of these differences. In addition, their perception of the supportive or nonsupportive nature of their interpersonal environment had an impact on their mothering experience. CONCLUSION: The results suggest that mothers with disabilities tend to value the importance of performing maternal occupations according to a perceived culturally common way, although self-acceptance of their own unique differences in performing these occupations facilitates maternal engagement as well.

Exploring spatial patterns of particulate sulfur and OMH from the Project MOHAVE summer intensive regional network using analyses of variance techniques and meteorological parameters as sort determinants.

Project MOHAVE (Measurements of Haze and Visual Effects) encompassed a 1-yr field study in the southwestern United States from September 1991 through August 1992. The congressionally mandated study was a joint partnership between the U.S. Environmental Protection Agency, Southern California Edison, and the National Park Service. A major objective of this study was to quantify the potential haze impacts on the nearby Grand Canyon National Park from the 1580 MW coal-fired MOHAVE Power Project (MPP). Any regional impacts from MPP were from secondary fine sulfate. In this paper, we explore the temporal and spatial patterns of particulate sulfur (Sp) and "organic mass by hydrogen" (OMH) during the summer intensive, conducted from mid-July through the end of August 1992. Using an innovative hierarchical pattern recognition classification scheme, we developed 6 groups of Sp and 8 groups of OMH temporally similar behaving patterns in the sampling region. From a regional understanding of synoptic meteorology, these Sp patterns were explainable. We observed two regional gradients. One gradient was a west-to-east decreasing gradient, most likely the result of major sources from urban southern California, including the San Joaquin Valley. The other decreasing gradient was from south-to-north, perhaps the result of emissions emanating from the large urban centers in northern Mexico. The patterns for OMH were not as regionally homogeneous as the patterns for Sp. A west-to-east decreasing gradient was observed for OMH, along with reduced values in the lower Colorado River Valley and some higher values in central and eastern Arizona. The west-to-east decreasing gradient suggests the presence of the Los Angeles urban plume, while the higher values in central and eastern Arizona may be due to biogenic emissions and increased seasonal fires.

Characterization of regional transport and dispersion using Project MOHAVE tracer data.

Perfluorocarbon tracers were released continuously from several surface locations and one power plant stack location during the winter (30 days) and summer (50 days) intensive studies as part of Project MOHAVE. Tracers were released in winter from the Mohave Power Plant (MPP) and Dangling Rope, UT, located on the shore of Lake Powell near Page, AZ; and in summer from MPP, the Tehachapi Pass between the Mojave Desert and the Central Valley in California, and El Centro, CA, on the California-Mexico border. At the Tehachapi tracer release site six-hour pulses of a separately identifiable perfluorocarbon tracer were released every four days in order to assess the time for the tracer to clear the monitoring network. Daily 24-hr integrated samples were collected at about 30 sites in four states. Limited tracer concentration data with higher time resolution is also available. Graphical displays and analyses identify several regional transport paths, including a convergence zone in the Mojave Desert, the importance of terrain channeling, especially in winter, and a relationship between 24-hr maximum influence function and distance that may prove useful as a scoping tool and to test regional scale air quality models. In winter, Dangling Rope tracer was routinely transported through the entire length of the Grand Canyon, while in summer, MPP tracer was routinely transported over most of Lake Mead.

Relating summer ambient particulate sulfur, sulfur dioxide, and light scattering to gaseous tracer emissions from the MOHAVE Power Project.

Project MOHAVE was initiated in 1992 to examine the role of emissions from the 1580 MW coal-fired MOHAVE Power Project (MPP) on haze at the Grand Canyon National Park (GCNP), located about 130 km north-north-east of the power plant. Statistical relationships were analyzed between summertime ambient concentrations of a gaseous perfluorocarbon tracer released from MPP and ambient SO2, particulate sulfur, and light scattering to evaluate whether MPP's emissions could be transported to the GCNP and then impact haze levels there. Spatial analyses indicated that particulate sulfur levels were strongly correlated across the monitoring network, regardless of whether the monitoring stations were upwind or downwind of MPP. This indicates that particulate sulfur levels in this region were influenced by distant regional emission sources. A significant particulate sulfur contribution from a point source such as MPP would result in a non-uniform pattern downwind. There was no suggestion of this in the data. Furthermore, correlations between the MPP tracer and ambient particulate sulfur and light scattering at locations in the park were virtually zero for averaging times ranging from 24 hr to 1 hr. Hour-by-hour MPP tracer levels and light scattering were individually examined, and still no positive correlations were detected. Finally, agreement between tracer and particulate sulfur did not improve as a function of meteorological regime, implying that, even during cloudy monsoon days when more rapid conversion of SO2 to particulate sulfur would be expected, there was no evidence for downwind particulate sulfur impacts. Despite the fact that MPP was a large source of SO2 and tracer, neither time series nor correlation analyses were able to detect any meaningful relationship between MPP's SO2 and tracer emission "signals" to particulate sulfur or light scattering.

Second generation chemical mass balance source apportionment of sulfur oxides and sulfate at the Grand Canyon during the Project MOHAVE summer intensive.

Receptor-based chemical mass balance (CMB) analysis techniques are designed to apportion species that are conserved during pollutant transport using conserved source profiles. The techniques will fail if non-conservative species (or profiles) are not properly accounted for in the CMB model. The straightforward application of the CMB model developed for Project MOHAVE using regional profiles resulted in a significant under-prediction of total sulfate oxides (SOx, SO2 plus fine particulate sulfate) for many samples at Meadview, AZ. In addition, for these samples the concentration of the inert tracer emitted from the MOHAVE Power Project (MPP), ocPDCH, was also under-predicted. A second-generation model has been developed which assumes that separation of particles and SO2 can occur in the MPP plume during nighttime stable plume conditions. This second-generation CMB model accounts for all SOx present at the various receptor sites. In addition, the concentrations of ocPDCH and the presence of other inert tracers of emission from regional sources are accurately predicted. The major source of SOx at Meadview was the MPP, but the major source of sulfate at this site was the Las Vegas urban area. At Hopi Point in the Grand Canyon, the Baja California region (Imperial Valley and northwestern Mexico) was the major source of both SOx and sulfate.

Evaluation of wind fields used in Grand Canyon Visibility Transport Commission analyses.

The Grand Canyon Visibility Transport Commission (GCVTC) was established by the U.S. Congress to assess the potential impacts of projected growth on atmospheric visibility at Grand Canyon National Park and to make recommendations to the U.S. Environmental Protection Agency on what measures could be taken to avoid such adverse impacts. A critical input to the assessment tool used by the commission was three-dimensional model-derived wind fields used to transport the emissions. This paper describes the evaluation of the wind fields used at various stages in the assessment. Wind fields evaluated included those obtained from the Colorado State University Regional Atmospheric Modeling System (RAMS), the National Meteorological Center's Nested Grid Model (NGM), and the National Oceanic and Atmospheric Administration's Atmospheric Transport and Dispersion (ATAD) trajectory model. The model-derived wind fields were evaluated at multiple vertical levels at several locations in the southwestern United States by determining differences between model predicted winds and winds that were measured using radiosonde and radar wind profiler data. Model-derived winds were also evaluated by determining the percent of time that they were within acceptable differences from measured winds. All models had difficulties, generally meeting the acceptable criteria for less than 50% of the predictions. The RAMS model had a persistent bias toward southwesterly winds at the expense of other directions, especially failing to represent channeling by north-south mountain ranges in the lower levels. The NGM model exhibited a substantial bias in the summer months by extending northwesterly winds in the eastern Pacific Ocean well inland, in contrast to the observed southwesterlies at inland locations. The simpler ATAD trajectory model performed somewhat better than the other models, probably because of its use of more upper air sites. The results of the evaluation indicated that these wind fields could not be used to reliably predict source-receptor impacts on a particular day; thus, seasonally averaged impacts were used in the GCVTC assessment.

Assessment of the nested grid model estimates for driving regional visibility models in the southwestern United States.

The Nested Grid Model (NGM) is a primitive-equation meteorological model that is routinely exercised over North America for forecasting purposes by the National Meteorological Center. While prognostic meteorological models are being increasingly used to drive air quality models, their use in conducting annual simulations requires significant resources. NGM estimates of wind fields and other meteorological variables provide an attractive alternative since they are typically archived and readily available for an entire year. Preliminary evaluation of NGM winds during the summer of 1992 for application to the region surrounding the Grand Canyon National Park showed serious shortcomings. The NGM winds along the borders between California, Arizona and Mexico tend to be northwesterly with a speed of about 6 m/sec, while the observed flow is predominantly southerly at about 2-5 m/sec. The mesoscale effect of a thermal low pressure area over the highly heated Southern California and western Arizona deserts does not appear to be represented by the NGM because of its coarse resolution and the use of sparse observations in that region. Tracer simulations and statistical evaluation against special high resolution observations of winds in the southwest United States clearly demonstrate the northwest bias in NGM winds and its adverse effect on predictions of an air quality model. The "enhanced" NGM winds, in which selected wind observations are incorporated in the NGM winds using a diagnostic meteorological model provide additional confirmation on the primary cause of the northwest bias. This study has demonstrated that in situations where limited resources prevent the use of prognostic meteorological models, previously archived coarse resolution wind fields in which additional observations are incorporated to correct known biases provide an attractive option.

Hsp27 functions as a negative regulator of cytochrome c-dependent activation of procaspase-3.

The release of mitochondrial cytochrome c by genotoxic stress induces the formation of a cytosolic complex with Apaf-1 (mammalian CED4 homolog) and thereby the activation of procaspase-3 (cas-3) and procaspase-9 (cas-9). Here we demonstrate that heat-shock protein 27 (Hsp27) inhibits cytochrome c (cyt c)-dependent activation of cas-3. Hsp27 had no effect on cyt c release, Apaf-1 and cas-9 activation. By contrast, our results show that Hsp27 associates with cas-3, but not Apaf-1 or cas-9, and inhibits activation of cas-3 by cas-9-mediated proteolysis. Furthermore, the present results demonstrate that immunodepletion of Hsp27 depletes cas-3. Importantly, treatment of cells with DNA damaging agents dissociates the Hsp27/cas-3 complex and relieves inhibition of cas-3 activation. These findings define a novel function for Hsp27 and provide the first evidence that a heat shock protein represses cas-3 activation.

Serial evoked potential studies and MRI imaging in chronic progressive multiple sclerosis.

Measurements of serial evoked potential latencies and plaque burden on MRI scans are often obtained during clinical studies of multiple sclerosis patients to provide additional information to the disability-based primary endpoints. The ideal laboratory-based marker of progression would be expected to significantly change over the time period of study. Serial visual (VEP) and brainstem auditory evoked potentials (BAEP) and MRI scans of 11 chronic progressive MS patients were obtained over a 1.5 year period in a clinical study. Over this period, there was no significant change in disability as measured by the Kurtzke EDSS, Ambulation Index or Neurological Rating Score. The VEP P100 significantly progressed over the period of study. However, the MRI T(2) plaque burden and BAEP I-V intrapeak latency did not significantly progress over the 1.5 years. We conclude that, in chronic progressive MS, serial visual evoked potential tests may complement standard disability-based endpoints to assess disease progression.

Relative stabilities of dinucleotide and tetranucleotide repeats in cultured mammalian cells.

The differences in rates of frameshift mutations between a dinucleotide repeat sequence [(CA)(17)] and a tetranucleotide repeat sequence [(GAAA)(17)] have been determined in immortalized, non-tumorigenic, mismatch repair-proficient mouse cells and in mismatch repair-defective human colorectal cancer cells. Clones with mutations were selected on the basis of restoration of activity of a bacterial neomycin resistance gene whose reading frame was disrupted by insertion of the microsatellite upstream of the translation initiation codon. This gene was introduced into the cells on a plasmid, which integrated into the genome of the host cells. Mutation rates of the tetra-nucleotide repeat were much lower than those of the dinucleotide repeat in both cell types. In addition, independent subclones of the colorectal cancer cell line were assayed by PCR for instability of endo-gen-ous tetranucleotide and dinucleotide repeat sequen-ces. In all cases, the mutation frequencies of the dinucleotide repeats were higher than those of the tetranucleotide repeats.

A double-blind, placebo-controlled trial of extracorporeal photopheresis in chronic progressive multiple sclerosis.

Extracorporeal photopheresis is a safe therapy for cutaneous T-cell lymphoma and may have efficacy in certain autoimmune disorders. We performed a randomized, double-blinded, placebo-controlled trial of monthly photopheresis therapy in 16 patients with clinically definite multiple sclerosis (MS). All patients had progressed during the preceding year with entry Expanded Disability Status Scale (EDSS) scores between 3.0 and 7.0. Patients received photopheresis or sham therapy for 1 year and were followed for an additional 6 to 12 months. Patients were clinically evaluated by three disability scales: (1) EDSS; (2) Ambulation index and (3) Scripp's quantitative neurologic assessment. No serious side effects occurred in either group. There were no differences between the photopheresis and sham therapy groups by the disability measures. Additionally, there were no differences in progression of MRI plaque burden or evoked potential latencies. In this limited study, photopheresis was found to be safe but did not significantly alter the course of chronic progressive MS.

Relative rates of insertion and deletion mutations in a microsatellite sequence in cultured cells.

A cell culture system has been used to determine the relative rates of insertions and deletions of integral numbers of dinucleotide repeats in a microsatellite sequence. A plasmid was constructed that contained 17 repeats of poly(dC-dA).poly(dG-dT) near the 5' end of a bacterial neomycin-resistance (neo) gene, such that the neo gene was translated in the (+1) reading frame. The plasmid was introduced into mismatch-repair-proficient and mismatch-repair-deficient mammalian cell lines. Rates of mutation to resistance to the neomycin analogue G418 were measured, and the nature of the mutations was determined. The mutations were all gains or losses of integral numbers of repeats, and mutations involving a single repeat greatly predominated over those involving multiple repeats. The data obtained from these studies were compared with results previously obtained with cells transfected with a similar plasmid in which the sequence of the oligonucleotide insert placed the neo gene in the (-1) reading frame. This experimental design made it possible to make direct comparisons between insertions and deletions of a single repeat unit. A significant excess of insertions over deletions was found in both repair-proficient and repair-deficient cell lines, although the few mutations involving more than two repeats were deletions.