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.     

Numerical Analyses of Soft Bottom Macroinvertebrates to Diagnose the Pollution in Tropical Coastal Waters

Soft bottom benthic organisms especially polychaetes are known to adapt as r or k selected strategies to different gradients of pollution. This will result in changes of benthic community structure from that of normal structure. There are a number of techniques to assess the impact of pollution on benthic community structure. Hence, to test this hypotheses some of the univariate and multivariate techniques were applied to soft bottom macro-invertebrates data of coastal waters of Mangalore, central west coast of India, a hot-spot area. Univariate techniques such as Pearson Rosenberg Model (PRM), abundance biomass comparison curve (ABC), geometrical class distribution, dominance-diversity curve, benthic community structure indices and multivariate techniques such as cluster classification, multidimensional scaling (MDS) and principal component analysis (PCA) were used to discriminate and diagnose the disturbance among the sites. Effectiveness and applicability of some of the above techniques are highlighted and discussed with the present set of data for environmental impact assessment studies.     

Pool of Dust Particles over the Asian Continent: Balloon-borne Optical Particle Counter and Ground-based Lidar Measurements at Dunhuang, China

Measurements of aerosols were made in 2001 and 2002 at Dunhuang (40 degrees 00'N, 94 degrees 30'E), China to understand the nature of atmospheric particles over the desert areas in the Asian continent. Balloon-borne measurements with an optical particle counter suggested that particle size and concentration had noticeable peaks in super micron size range not only in the boundary mixing layer but also in the free troposphere. Super-micron particle concentration largely decreased in the mid tropopause (from 5 to 10 km; above sea level, a.s.l.). Lidar measurements made during August 2002 at Dunhuang suggested the possibility that mixing of dust particles occurred from near the ground to about 6 km even under calm weather conditions, and a large depolarization ratio of particulate matter was found in the aerosol layer. The top of the aerosol layer was found at heights of nearly 6 km (a.s.l.). It is strongly suggested that nonspherical dust particles (Kosa particles) frequently diffused in the free atmosphere over the Taklamakan desert through small-scale turbulences and are possible sources of dust particles of weak Kosa events that have been identified in the free troposphere not only in spring but also in summer over Japanese archipelago. Electron microscopic experiments of the particles collected in the free troposphere confirmed that coarse and nonspherical particles observed by the mineral particle were major components of coarse mode (diameter larger than 1 microm) below about 5 km over Dunhuang, China.     

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.     

Utility of dynamics as indicators of stress in population models

The relationships between the dynamics of environmentally and chemically stressed populations and indicators of the effects of the stressor are explored in a model framework. The physiologically structured population, represented by a system of McKendrick–von Foerster hyperbolic partial differential equations, includes the dynamics of numerous individuals distinguished by ecotype. Chemical uptake of nonpolar narcotics is modeled by first order kinetics. Classical methodologies, frequency analysis and phase space reconstruction, are explored in a search for indicators of magnitude of stress. When these techniques proved generally unsuccessful for the objective of indicator selection in our model setting, summary statistics, as related to bifurcation diagrams, were constructed and appear more useful as indicators. It is concluded that physiological structures generally lead to more feasible measurable indicators of magnitude of stress than do specifics of population dynamics.     

我国环境监测仪器向规范化方向发展刍议

从我国环境监测事业发展和建立社会主义市场经济体制的需要，论述了加速我国环境监测仪器规范化的必要和可行性，并阐述了具体实施意见。     

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.     

Genetic population structure of the recently introduced Asian clam,Potamocorbula amurensis,in San Francisco Bay

The genetic population structure of the recently introduced Asian clam, Potamocorbula amurensis, in San Francisco Bay was described using starch gel electrophoresis at eight presumptive loci. Specimens were taken from five environmentally distinct sites located throughout the bay. The population maintains a high degree of genetic variation, with a mean heterozygosity of 0.295, a mean polymorphism of 0.75, and an average of 3.70 alleles per locus. The population is genetically homogeneous, as evidenced from genetic distance values and F-statistics. However, heterogeneity of populations was indicated from a contingency chi-square test. Significant deviations from Hardy-Weinberg equilibrium and heterozygote deficiencies were found at the Lap-1 locus for all populations and at the Lap-2 locus for a single population. High levels of variability could represent a universal characteristic of invading species, the levels of variability in the source population(s), and/or the dynamics of the introduction. Lack of differentiation between subpopulations may be due to the immaturity of the San Francisco Bay population, the general purpose phenotype genetic strategy of the species, high rates of gene flow in the population, and/or the selective neutrality of the loci investigated.     

Growth and development of cellular slime mould Dictyostelium discoideum treated with DDT

The effects of dichlorodiphenyl trichloroethane (DDT) on the growth, phagocytic activity, ultrastructure and developmental stages of a well known species of the cellular slime mould, Dictyostelium discoideum were studied. DDT, at doses of 60 ppm and above, inhibited growth of the vegetative cells and this was also evident from the colony blots which were smaller in size. A dose-dependent inhibition occurred in the phagocytic activity and macromolecular syntheses of DDT-treated cells. The cytomorphology of growing Dictyostelium cells, as revealed under light and electron microscopes, was profoundly affected by DDT treatment. Further, a considerable delay occurred in the various morphogenetic events in the slime mould cells exposed to DDT.     

Induction of metamorphosis in larvae of the oyster Crassostrea gigas using neuroactive compounds

Chemical (neuroactive compounds at different concentrations and exposure times) and physical (water agitation, light) factors with potential effects on the metamorphosis of larvae of the oyster Crassostrea gigas (Thunberg) larvae have been studied. The neurotransmitters l-dihydroxyphenylalanine (DO), epinephrine (EP), norepinephrine (NE), and acetylcholine (AC) have been identified as very active inducers of metamorphosis, whilst serotonin (SE), dopamine (DA) and potassium (K) were less effective inducers. The -aminobutyric acid (GA) and ammonium (AM) were found ineffective at the concentrations tested. Exposure to 10^-4 M EP for 15 min was sufficient to promote >80% metamorphosis within 48 h, whereas NE required 2 h to exert comparable induction. Maximum induction by DO (>50%) was achieved after 2 h exposure to 10^-4 M. However, unlike EP and NE, DO was lethal at that concentration in the long term. Maximum induction by AC (30% metamorphosis) was achieved at a concentration of 10^-4 M. In contrast to other neurotransmitters, AC induced settlement behaviour, cementation and eventual metamorphosis, yielding postlarvae which were all attached to the substratum. EP and NE triggered the morphogenetic changes, by-passing settlement and leading to postlarvae not cemented to the substratum. DO induced mostly attached spat at low concentrations (10^-5 M) and unattached spat at high concentrations (10^-4 M), and a similar pattern was apparent for the weaker inducers SE and DA. Regarding physical factors, a highly reflectant surface significantly increased the percentage of attached spat obtained, compared to a dark bottom. No consistent effect of water current or light was detected on the production of unattached spat. The three different forms of induction are discussed in relation to different regulatory pathways of settlement and metamorphosis.