Evaluation of interference to conventional and real-time PCR for detection and quantification of fungi in dust

Advances in polymerase chain reaction (PCR) have permitted accurate, rapid and quantitative identification of microorganisms in pure cultures regardless of viability or culturability. In this study, a simple sample processing method was investigated for rapid identification and quantification of fungal spores from dust samples using both conventional and real-time PCR. The proposed method was evaluated for susceptibility to interference from environmental dust samples. Stachybotrys chartarum and Aspergillus fumigatus were used as test organisms. The sensitivity of detection in pure culture was 0.1 spore DNA equivalents per PCR reaction corresponding to 20 spores ml(-1) in the sample. However, 1 spore DNA equivalent per PCR reaction corresponding to 200 spores ml(-1) in the sample was the lowest amount of spores tested without interference in dust samples spiked with spores of either fungal species. The extent of inhibition was calculated using conventional and real-time PCR reactions containing fungal spores, specific primers, specific probes (for real-time PCR) and various amounts of dust. The results indicate that the extent of inhibition by dust on PCR varies with the type and amount of dust, and number of spores. No interference in the analysis of spiked samples was detected from 0.2 mg ml(-1) of four real-life dust samples at p-value >0.05 using 2 x 10(4) spores for conventional PCR and 2 x 10(5) spores for real-time PCR. However, samples containing >0.2 mg ml(-1) real-life dust compromised the PCR assay. These results suggest the potential usefulness of a simple sample processing method in conjunction with PCR for monitoring the fungal content of aerosols collected from indoor environments.     

Land User Participation in Developing a Computerised Decision Support System for Combating Desertification

Combating desertification in natural rangelands has recently become a priority in large parts of southern Africa. Rangeland managers, farmers, scientists, conservationists and land users have been applying a variety of restoration technologies to address this problem. Bush encroachment, as part of the desertification process, involves the natural replacement of the herbaceous plant cover by undesirable problem woody species. The active and passive restoration technologies that are applied, are mainly based on indigenous knowledge and include the chemical, mechanical or manual reclamation of unproductive rangelands, as well as the combating of woody and alien species encroachment. Indigenous practices and knowledge play a major role in the effectiveness and success rate of these technologies. This project faces the challenge of bringing together both local and scientific knowledge in a single user-friendly, computerised Decision Support System (DSS) which is directly accessible by land users to support them in the process of decision making, concerning the combating of desertification. Case studies from central and northern Namibia were used to combine qualitative and quantitative data to develop this Decision Support System. The DSS currently consists of two databases and an expert system, which evaluates the results of land users’ management practices, and provides easily accessible information and advice for participants in the system, based on the incorporated data. The DSS is also linked to national and international web sites and databases to offer a wider range of information on technologies concerning agricultural and conservation practices.     

Assessment of Metal Contamination in Doñana National Park (Spain) using Crayfish (Procamburus Clarkii)

Water quality assessment in the Aznalcollar area was attempted using multivariate methods based on heavy metal concentrations in red swamp crayfish (Procamburus clarkii). Trace levels of four heavy metals, copper (Cu), zinc (Zn), cadmium (Cd) and lead (Pb), were detected in crayfish from eleven different stations. Principal component analysis (PCA) highlighted a gradient of contamination between the sampling stations. Cluster analysis (CA) distinguished three groups of stations. Discriminant analysis also differentiated three groups. The group centroids of the first discriminant function were used to devise an index that varies according to the source of the crayfish. These standardized values are proposed for use as a water quality index. The ability of this index to successfully predict environmental quality was proved with random samples.     

Total versus inhalable sampler comparison study for the determination of asphalt fume exposures within the road paving industry

Exposure to asphalt fumes has a threshold limit value (TLV of 0.5 mg m(-3) (benzene extractable inhalable particulate) as recommended by the American Conference of Governmental Industrial Hygienists (ACGIH). This reflects a recent change (2000) whereby two variables are different from the previous recommendation. First is a 10-fold reduction in quantity from 5 mg m(-3) to 0.5 mg m(-3). Secondly, the new TLV specifies the "inhalable" fraction as compared to what is presumed to be total particulate. To assess the impact of these changes, this study compares the differences between measurements of paving asphalt fume exposure in the field using an "inhalable" instrument versus the historically used 'total' sampler. Particle size is also examined to assist in the understanding of the aerodynamic collection differences as related to asphalt fumes and confounders. Results show that when exposures are limited to asphalt fumes, a 1:1 relationship exists between samplers, showing no statistically significant differences in benzene soluble matter (BSM). This means that for the asphalt fume ACGIH TLV, the 'total' 37-mm sampler is an equivalent method to the "inhalable" method, referred to as IOM (Institute of Occupational Medicine), and should be acceptable for use against the TLV. However, the study found that when confounders (dust or old asphalt millings) are present in the workplace, there can be significant differences between the two samplers' reported exposure. The ratio of IOM/Total was 1.37 for milling asphalt sites, 1.41 for asphalt paving over granular base, and 1.02 for asphalt over asphalt pavements.     

A Metamodeling Approach for Integrated Assessment of Air Quality Policies

A method is proposed to build integrated models (also called Metamodels) aimed at quantifying the economic efficiency of air quality policies. This Metamodeling approach is based on the coupling of two complementary models, that operate at different scales in space and time, and which represent the economic and the physical and chemical processes, respectively. The joint consideration of the physico-chemical and techno-economic structure of the pollution control problems permits a comprehensive evaluation of air pollution abatement strategies. The motivating pollution control problems include urban-regional air quality management through efficient energy and traffic control policies. A pilot study, exploiting data collected in the Geneva canton (Switzerland), is used to demonstrate the potential of the approach.     

monitoring bioaccumulation of contaminants in the belted kingfisher (ceryle alcyon)

The belted kingfisher (Ceryle alcyon), acommon piscivore in the eastern United States, hasbecome a common endpoint in ecological riskassessments (ERA) because of their high consumptionof potentially contaminated aquatic prey. Whilebioaccumulation data and biosurveys may be used tosupport conclusions of ERAs for kingfishers, thereare currently no published data on contaminantconcentrations in kingfishers. Additionally,methods available for collecting biological samples(e.g., feathers, eggs, food debris, etc.) fromkingfisher burrows can be detrimental to thereproductive success of the birds. We present amethod for obtaining samples from burrows during orfollowing the nesting season. The method wasapplied to kingfisher burrows on the Oak RidgeReservation (ORR) in eastern Tennessee. Feathers,eggshells, and nestlings were collected from burrows and analyzed. In addition, carcasses ofthree adult kingfishers found dead on the ORR wereanalyzed. Metals and radionuclides were accumulatedby both juvenile and adult birds. Body burdens ofcadmium, lead, and cesium-137 in adult birds were belowlevels associated with toxicity. Concentrations of selenium and mercury were observed at potentiallytoxic levels. Contaminants in eggshells andnestling feathers indicate exposure, however, thereis insufficient information to evaluate thetoxicological significance of this contamination.National Laboratory Oak Ridge National Laboratory is managed for the U.S. Department of Energy by Lockheed Martin Energy Research Corp. under contract DE-AC05-96OR22464     

Methods for constructing and balancing ecosystem flux charts: new techniques and software

We describe a general computer program which employs novel techniques to analyze carbon (energy) and nutrient flows in complex food webs. The program uses estimates of metabolic/physiological parameters and ecosystem fluxes to simultaneously calculate the most likely balanced flux charts for carbon and a nutrient. We give an example of how the model can explore the theoretical issues involved with nutrient limitation and recycling.     

Fractionation of butyltin species during sample extraction and preparation for analysis

Solvent extraction and evaporative concentration steps are often used in procedures for the measurement of butyltins in environmental samples. As part of a larger study utilizing radiolabeled butyltins, the loss and fractionation of butyltins during sample preparation was investigated. TBT, DBT, and MBT were extracted from acidified seawater by hexane with efficiencies of about 95–99, 50–60 and 11% respectively. In addition, losses of about 70% of DBT were found during evaporative concentration of hexane. A variety of sediment extraction procedures were tested and none were found to be highly efficient for total butyltin extraction.     

Sampling and analysis for radon-222 dissolved in ground water and surface water

Radon-222 is a naturally occurring radioactive gas in the uranium-238 decay series that has traditionally been called, simply, radon. The lung cancer risks associated with the inhalation of radon decay products have been well documented by epidemiological studies on populations of uranium miners.The realization that radon is a public health hazard has raised the need for sampling and analytical guidelines for field personnel. Several sampling and analytical methods are being used to document radon concentrations in ground water and surface water worldwide but no convenient, single set of guidelines is available. Three different sampling and analytical methods-bubbler, liquid scintillation, and field screening-are discussed in this paper. The bubbler and liquid scintillation methods have high accuracy and precision, and small analytical method detection limits of 0.2 and 10 pCi/l (picocuries per liter), respectively. The field screening method generally is used as a qualitative reconnaissance tool.     

Biodegradability: An assessment of commercial polymers according to the Canadian method for anaerobic conditions

The degradation of several biodegradable polymers was measured as a result of exposure to an anaerobic medium. The polymers investigated included materials based upon polylactic acid, polylactone, and poly(hydroxy butyrate/valerate) as well as those incorporating starch-based materials. The degradation was monitored by methane and carbon dioxide evolution. In addition, the physical and chemical changes were noted as a result of exposure. These measurements included changes in mass, dimension, and molecular weight. FTIR, UV-vis, proton, and¹³C NMR spectra were also recorded prior to and after exposure. The results clearly indicated that several biological and chemical degradation processes were occurring with the biodegradable polymers studied.Paper presented at the Bio/Environmentally Degradable Polymer Society—Second National Meeting, August 19–21, 1993, Chicago, Illinois.Issued as NRCC No. 37549.