Comparison of two pollen counting methods of slides from a hirst type volumetric trap

Two of the most frequently used methods of pollen counting on slides from Hirst type traps are evaluated in this paper: the transverse traverse method and the longitudinal traverse method. The study was carried out during June–July 1996 and 1997 on slides from a trap at Worcester, UK. Three pollen types were selected for this purpose: Poaceae, Urticaceae and Quercus. The statistical results show that the daily concentrations followed similar trends (p < 0.01, R-values between 0.78–0.96) with both methods during the two years, although the counts were slightly higher using the longitudinal traverses method. Significant differences were observed, however, when the distribution of the concentrations during 24 hour sampling periods was considered. For more detailed analysis, the daily counts obtained with both methods were correlated with the total number of pollen grains for the taxon over the whole slide, in two different situations: high and low concentrations of pollen in the atmosphere. In the case of high concentrations, the counts for all three taxa with both methods are significantly correlated with the total pollen count. In the samples with low concentrations, the Poaceae and Urticaceae counts with both methods are significantly correlated with the total counts, but none of Quercus counts are. Consideration of the results indicates that both methods give a reasonable approximation to the count derived from the slide as a whole. More studies need be done to explore the comparability of counting methods in order to work towards a Universal Methodology in Aeropalynology. This revised version was published online in June 2006 with corrections to the Cover Date.     

Lipase kinetics at the triacylglycerol-water interface using surface tension measurements

Two methods, the so-called "oil drop" and "Teflon plunger" methods, were designed to monitor lipase hydrolysis of natural long-chain triacylglycerols through the variation with time of the oil-water interfacial tension. The first part of this work is devoted to the development of these two techniques using pure, well-characterized porcine pancreatic lipase. They gave linear responses with enzyme concentrations ranging from 1 x 10(-3) to 30 units x ml-1. We then applied them to a study of the optimal pH conditions for human gastric lipase which were found to range around 5, as previously observed. In the presence of variable concentrations of sodium taurodeoxycholate, these two methods also showed that human gastric lipase is active in the 8-13 dyn cm-1 range of interfacial tension. It is concluded that these two methods, based upon variations with time of the oil-water interfacial tension, constitute reliable, sensitive and convenient means of investigating lipase kinetics.     

Comparison between two methods of assaying relative microbial activity in marine environments.

Two methods for determining relative microbial activity in the marine environment were compared. In one method, a single concentration of a labeled substrate was used to calculate rates of substrate utilization; in the other, multiple concentrations of the same substrate (heterotrophic activity method) were used to calculate maximum potential substrate utilization rates. These studies were made on 232 seawater and 79 sediment samples taken from a variety of marine environments. The highest correlations between these two methods were seen in the sediment samples tested. The lowest correlation coerfficient seen in the sediment samples was 0.90, and the highest was 0.98. In seawater samples (six studies), the lowest correlation coefficient was 0.77 and the highest was 0.95. The correlation between these two methods was also substrate concentration dependent. Higher correlation coefficients were observed when higher substrate concentrations were used. Under certain conditions, these two methods appear to be comparable for estimating relative levels of microbial activity in the marine environment.     

Comparison between two methods of assaying relative microbial activity in marine environments.

Two methods for determining relative microbial activity in the marine environment were compared. In one method, a single concentration of a labeled substrate was used to calculate rates of substrate utilization; in the other, multiple concentrations of the same substrate (heterotrophic activity method) were used to calculate maximum potential substrate utilization rates. These studies were made on 232 seawater and 79 sediment samples taken from a variety of marine environments. The highest correlations between these two methods were seen in the sediment samples tested. The lowest correlation coerfficient seen in the sediment samples was 0.90, and the highest was 0.98. In seawater samples (six studies), the lowest correlation coefficient was 0.77 and the highest was 0.95. The correlation between these two methods was also substrate concentration dependent. Higher correlation coefficients were observed when higher substrate concentrations were used. Under certain conditions, these two methods appear to be comparable for estimating relative levels of microbial activity in the marine environment.     

A novel method for collection and preservation of faeces for genetic studies

Quantitative and qualitative measurements of DNA were used to compare faecal sample storage in ethanol and silica with a novel method (two‐step) in which samples are soaked in ethanol and then desiccated with silica. Silica‐preserved samples had the lowest DNA concentrations. The two‐step method yielded significantly more DNA in high quality samples (average DNA concentrations > 100 pg/µL with all storage methods). However, for lower quality samples, the ethanol and two‐step methods performed similarly. The amounts and rates of sample degradation were not strongly affected by storage method and neither was the percentage of target DNA (< 1%) obtained from the samples.     

Comparison of gas chromatographic/mass spectrometric and microbiological assays for 5-fluorouracil in plasma

The concentrations of 5-fluorouracil in 99 plasma samples were determined by both microbiological and gas chromatographic/mass spectrometric assays. The values determined by the two methods were similar (correlation coefficient = 0.90). A regression of the natural logarithms of the concentrations (0.01-94 micrograms ml-1) determined by the two methods gave a line whose slope and intercept were not statistically different (p greater than 0.05) from 1 and 0 respectively. Thus, the microbiological assay has specificity over a sufficient concentration range to serve as a practical routine laboratory method for the determination of 5-fluorouracil.     

Aeroallergens: a comparative study of two monitoring methods

Olive and grass pollen grains are the major causes of hay fever in the Mediterranean region. A number of samplers and methods have been developed in recent years in order to obtain reliable data regarding airborne allergen concentrations. This paper reports on a detailed comparison of two samplers—Cyclone and ChemVol—and on the parameters that could influence their efficiency. Airborne concentrations of two key olive and grass allergens, Ole e 1 and Phl p 5, respectively, were monitored over two years with different weather patterns, 2012 and 2014. Allergenic particles were quantified by ELISA assay, and results were compared with pollen concentrations monitored using a Hirst-type volumetric spore trap over the same study periods. The influence of weather-related parameters on local airborne pollen and allergen concentrations was also analysed. Although a positive correlation was detected between results obtained using the two samplers during the pollen season, results for the cumulative annual Allergen Index varied considerably. The two samplers revealed a positive correlation between pollen concentrations and both minimum temperature during the warmer year (2012) and maximum temperature during the cooler year (2014); a negative significant correlation was observed in both cases with rainfall and relative humidity. In summary, although some differences were observed between the two samplers studied, both may be regarded as suitable for allergen detection.     

Effect of commercial processing on fumonisin concentrations of maize-based foods

Fumonisin-contaminated maize was used to study the effect of three cooking and food processing methods and residual contamination of the food product. Frying, autoclaving and extrusion were examined with naturally-contaminated maize meal, maize flour and sweet maize kernels, all at two fumonisin concentrations. High Pressure Liquid Chromatography determination of fumonisins B1 and B2 and hydrolized fumonisin B1 (AP1) were performed in unprocessed materials and at the end of the experimental runs. Reductions of fumonisins concentration in processed products were obtained for fried polenta and from one of the two runs of extruded maize batter. These reductions were consistent with previous studies of the thermal degradation of fumonisins. Autoclaving sweet maize kernels apparently resulted in reductions of fumonisin concentrations that were highly temperature dependent, but this needs further study. The unexpectedly large reduction in fumonisin concentrations in the extrusion processing of batter with high initial concentrations also needs further investigation. There was no evidence that AP1 was formed under any of the conditions tested.     

Extraction and cleaning methods to detect yessotoxins in contaminated mussels

Yessotoxin (YTX) and its analogues are a newly recognized group of toxins with increased presence in shellfish in recent years. They can be quantified by various functional assays due to their interaction with phosphodiesterases (PDEs). One of these assays detects the binding between the YTX and the fluorescently labeled PDE I using fluorescence polarization, a spectroscopic technique based on exciting a fluorescent molecule with plane-polarized light and measuring the polarization degree of the emitted light. The aim of this study was to develop a YTX extraction procedure from mussels that does not interfere with this detection method. YTX concentrations were measured in spiked mussel extracts obtained through use of different extraction methods and cleaning procedures. The percentages of toxin recovery in various steps of the processes were calculated using these concentrations. Six extraction methods and two cleaning steps were used and no matrix effects and high toxin recoveries were obtained in two cases. One case used acetone as extraction solvent followed by three dichloromethane partitions and the other case used methanol. The cleaning procedure includes a silica cartridge and a 10,000 NMWL filter. Finally these two extraction-cleaning-detection methods were applied to a naturally contaminated mussel sample and results showed that not only YTX but also homoYTX and hydroxyYTX can be quantified with a 85-90% recovery.     

Determination of plasma catecholamines by high performance liquid chromatography with electrochemical detection: comparison with a radioenzymatic method.

High performance liquid chromatography with electrochemical detection has been compared to a radioenzymatic method for the determination of plasma catecholamines. With the use of an internal standard highly accurate determinations of plasma noradrenaline, adrenaline and dopamine were performed on 0.2–2 ml plasma with the chromatographic method. The radioenzymatic method required only 3 × 50 μl plasma. A comparison of noradrenaline and adrenaline concentrations measured by the two methods in a set of nine plasma samples showed an excellent agreement between the methods (r=0.993 and 0.994, respectively). Advantages and disadvantages with the two methods are discussed.     

An evaluation of two methods used for microscopic analysis of airborne fungal spore concentrations from the Burkard Spore Trap

The Burkard Volumetric Spore Trap is a common and efficient instrument used to collect outdoor air samples. In North America, two slide counting methods have been widely used by aerobiologists: the single longitudinal traverse method and the twelve transverse traverse method. The purpose of this study was to compare the two counting methods by assessing fungal spore concentrations of ascospores, basidiospores, smut teliospores, Cladosporium, Alternaria, Epicoccum, Curvularia, Drechslera, Pithomyces, other spores, and total spores at two metropolitan Tulsa, Oklahoma sites (Tulsa and Hectorville) during September 1996. Results showed that both methods were sensing parallel fluctuations in average daily spore concentration, although the twelve transverse traverse method usually resulted in higher concentrations. At the Tulsa site, the twelve transverse traverse method gave statistically higher concentrations than the single longitudinal traverse method except for Epicoccum, Pithomyces, smut teliospores, and other spores. At the Hectorville site, however, only Cladosporium and basidiospores showed that the twelve transverse traverse method was statistically higher than the single longitudinal traverse method. Comparison with concentrations obtained by counting the total slide surface of two slides indicated that neither method was equivalent to the total slide spore count, although the twelve transverse traverse method gave a lower absolute percent difference from the total slide surface concentration. While the twelve transverse traverse method gave slightly better approximations of the spore concentration, the increase in accuracy may not justify the extra effort required to analyze with this method.     

Mouse brain concentrations of 3-methoxytyramine and normetanephrine: a comparison of methods of sacrifice

The identification of sacrifice methods that produce reliable measures of baseline central nervous system neurotransmitter concentrations poses a challenge to analytical neurochemical investigation. In the present study, microwave irradiation (MWVI) was compared with in situ freezing, cervical dislocation, and simple decapitation, in an effort to examine their effects on whole mouse brain concentrations of 3-methoxytyramine (3MT) and normetanephrine (NMN), the O-methylated catecholamine metabolites believed to be sensitive indicators of release of CNS dopamine and norepinephrine, respectively. Both high-energy (6 kW, 0.3 s) and low-energy (2.5 kW, 1.5 s) MWVI produced the lowest mouse brain concentrations of 3MT and NMN when compared with other methods of sacrifice within experiments. In situ freezing resulted in values of 3MT and NMN that were slightly, yet significantly, higher than MWVI within experiments. The concentrations of 3MT and NMN obtained following either cervicle dislocation or simple decapitation were up to 9-fold greater than those produced by either of the two previous methods.     

A comparative study on three analytical methods for the determination of the neurotoxin BMAA in cyanobacteria

The cyanobacterial neurotoxin β-N-methylamino-L-alanine (BMAA) has been considered a serious health threat because of its putative role in multiple neurodegenerative diseases. First reports on BMAA concentrations in cyanobacteria were alarming: nearly all cyanobacteria were assumed to contain high BMAA concentrations, implying ubiquitous exposure. Recent studies however question this presence of high BMAA concentrations in cyanobacteria. To assess the real risk of BMAA to human health, this discrepancy must be resolved. We therefore tested whether the differences found could be caused by the analytical methods used in different studies. Eight cyanobacterial samples and two control samples were analyzed by three commonly used methods: HPLC-FLD analysis and LC-MS/MS analysis of both derivatized and underivatized samples. In line with published results, HPLC-FLD detected relatively high BMAA concentrations in some cyanobacterial samples, while both LC-MS/MS methods only detected BMAA in the positive control (cycad seed sarcotesta). Because we could eliminate the use of different samples and treatments as causal factors, we demonstrate that the observed differences were caused by the analytical methods. We conclude that HPLC-FLD overestimated BMAA concentrations in some cyanobacterial samples due to its low selectivity and propose that BMAA might be present in (some) cyanobacteria, but in the low μg/g or ng/g range instead of the high μg/g range as sometimes reported before. We therefore recommend to use only selective and sensitive analytical methods like LC-MS/MS for BMAA analysis. Although possibly present in low concentrations in cyanobacteria, BMAA can still form a health risk. Recent evidence on BMAA accumulation in aquatic food chains suggests human exposure through consumption of fish and shellfish which expectedly exceeds exposure through cyanobacteria.     

Concentration of prostaglandins E and F,fructose and glycerylphosphorylcholine in ram semen obtained by electro-ejaculation or artificial vagina and in vesicular fluid

The concentration of spermatozoa in electrically ejaculated ram semen was lower than in semen obtained by an artificial vagina. Glycerylphosphorylcholine concentrations were also lower in the electrically ejaculated semen and there was a high correlation between sperm and glycerylphosphorylcholine concentrations.Seminal fructose, prostaglandin E (PGE) and prostaglandin F (PGF) levels did not differ significantly between the two methods of collection but there was greater variability between rams when they were electrically ejaculated.The concentration of fructose in the vesicular secretion of rams was less variable and higher than in seminal plasma whereas PGE or PGF concentrations were not significantly different in the two fluids.     

Bioanalysis of N-acetyl-aspartyl-glutamate as a marker of glutamate carboxypeptidase II inhibition

We report the characterization of two methods for the analysis of N-acetyl-aspartyl-glutamate (NAAG) in biological fluids. In the first method, NAAG concentrations were calculated based on differences between glutamate concentrations before and after NAAG hydrolysis with exogenous glutamate carboxypeptidase II (GCP II) using high-performance liquid chromatography (HPLC) followed by fluorescence detection. In the second method, NAAG levels were quantified directly using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Analyses of NAAG levels in human cerebrospinal fluid samples using either method gave similar results within experimental error, confirming the validity of the two independent measurements. These methods will be useful in future clinical trials to assess drug-induced GCP II inhibition in biological matrices.     

Evaluation of culture methods to identify bovine feces with high concentrations of Escherichia coli O157

Our objective was to evaluate methods for identifying cattle with high concentrations of Escherichia coli O157 in their feces. In two experiments, feces were collected from cattle orally inoculated with nalidixic acid (Nal)-resistant E. coli O157, and direct plating of diluted feces on sorbitol MacConkey agar with cefixime and potassium tellurite (CT-SMAC) containing Nal was considered the gold standard (GS) method. In experiment 1, methods evaluated were preenrichment direct streak, immunomagnetic separation with most probable number (MPN), and postenrichment direct streak with MPN, all using CT-SMAC. The mean concentration of Nal-resistant E. coli O157 in samples (n = 59) by use of the GS was 3.6 log10 CFU/g. The preenrichment streak detected >3.0 log10 CFU/g samples with a 74.4% sensitivity and 68.8% specificity. Postenrichment direct streak-MPN and immunomagnetic separation-MPN concentrations were correlated significantly with GS concentrations (r = 0.53 and r = 0.39, respectively). In experiment 2 (480 samples), pre- and postenrichment direct streaking performed in triplicate and spiral plating on CT-SMAC were evaluated. For preenrichment streaks, sensitivity was 79.7% and specificity was 96.7% for detecting >3.0 log10 CFU/g when the criterion was positive cultures on at least two plates. For spiral plating at that concentration, sensitivity and specificity were 83.9% and 56.3%, respectively. Postenrichment streaking performed relatively poorly. Triplicate preenrichment streaks of 1:10-diluted feces on CT-SMAC may be useful for identifying cattle shedding high concentrations of E. coli O157. Estimates of sensitivity and specificity enable appropriate application of methods and interpretation of results and may enhance applied research, surveillance, and risk assessments.     

Antifungal susceptibility testing of Trichosporon beigelii to imidazole compounds

Twenty-two strains of Trichosporon beigelii have been tested for susceptibility to imidazole compounds. Ten strains were isolated from untreated genital white piedra lesions and 12 were from the same patients following treatment failure with imidazole compounds. Agar dilution and disk elution methods were compared using two media: yeast nitrogen base and antibiotic assay medium 3 (Difco). Antifungal agents tested were econazole, miconazole, ketoconazole, clotrimazole, and amphotericin B in concentrations of 0.0625-32 micrograms/mL. The most consistent results occurred with antibiotic assay medium 3 and the agar dilution method giving minimal inhibitory concentrations between 0.0625 and 0.25 micrograms/mL. Using yeast nitrogen base agar, minimal inhibitory concentrations were higher ranging from 0.0625 to 2.0 micrograms/mL. End points of growth in the disk elution method were not clearly delineated and ranged from 0.0625 to 8.0 micrograms/mL. The distribution of minimal inhibitory concentrations obtained using different media and methods were compared by chi 2 analysis, and the medium was found to significantly change the minimal inhibitory concentrations. There was no difference in the susceptibility of strains of T. beigelii to imidazole compounds whether isolated before or after treatment. It was concluded that in vitro susceptibility of T. beigelii to imidazole compounds did not necessarily predict efficacy in vivo.     

TRANSIENT AMMONIUM UPTAKE IN THE MACROALGA ULVA LACTUCA (CHLOROPHYTA): NATURE,REGULATION, AND THE CONSEQUENCES FOR CHOICE OF MEASURING TECHNIQUE1

The nature of transient ammonium uptake by the macroalga Ulva lactuca L. was studied from the depletion of ammonium after single additions of ammonium to batch cultures. The experiments were carried out by the application of two different experimental setups: the “multiple flask” and the “perturbation” techniques. Uptake rate was nonlinear with time, and three distinct, succeeding phases of uptake were identified: 1) “surge” uptake, i.e. transiently enhanced uptake that lasted for a few hours only, followed by 2) “internally” controlled uptake, a relatively constant phase occurring at high substrate concentrations, and finally 3) the “externally” controlled uptake phase, which was substrate-dependent and occurred at low substrate concentrations. Surge uptake occurred over a broad range of substrate concentrations but was concentration-dependent and, so, equalled externally controlled uptake rates at substrate concentrations below 3–10 μM. The transient nature of ammonium uptake rate seemed related to rapid changes in small intracellular pools of inorganic nitrogen or amino acids rather than to changes in total N content of the algae. The transient nature of ammonium uptake has important implications for the measurement of uptake rates when either of the two standard methods, the multiple flask and the perturbation technique, are used, and I recommend that a combination of the two methods be used for future uptake experiments.     

超氧化物歧化酶两种邻苯三酚自氧化测定活力方法的比较

在同一反应条件下,以酵母SOD为原料,对2种邻苯三酚自氧化测定SOD的方法进行了比较,实验结果表明325 nm法测定的酶活力单位与420 nm法测定的酶活力单位的比值约为2.7,考察了不同浓度邻苯三酚对SOD活力的影响。     

AVOIDING THE CENTERING BIAS OR RANGE EFFECT WHEN DETERMINING AN OPTIMUM LEVEL OF SWEETNESS IN LEMONADE1

When two or more levels of an ingredient are presented for evaluation of an ideal level of that ingredient, the centering bias is introduced and the true optimum level is unlikely to be determined. Poulton (1979) and McBride (1982) have suggested methods for avoiding the centering bias, but their methods have never been tested. The purpose of this study was to check the validity of their methods. Three different ranges of sucrose concentrations were used to determine the optimum level of sucrose in lemonade. Twenty different judges were assigned to each of the three ranges and rated the samples on a just right scale. The unbiased just right sucrose concentrations, determined by the methods suggested by Poulton (1979) and McBride (1982), were 6.7% and 6.9%, respectively. A single stimulus presentation confirmed the results as being an unbiased optimum concentration.