Evaluating putative ecological drivers of microcystin spatiotemporal dynamics using metabarcoding and environmental data.

Microcystin is a cyanobacterial hepatotoxin of global concern. Understanding the environmental factors that cause high concentrations of microcystin is crucial to the development of lake management strategies that minimize harmful exposures. While the literature is replete with studies linking cyanobacterial production of microcystin to changes in various nutrients, abiotic stressors, grazers, and competitors, no single biotic or abiotic factor has been shown to be reliably predictive of microcystin concentrations in complex ecosystems. We performed random forest regression analyses with 16S and 18S rRNA gene sequencing data and environmental data to determine which putative ecological drivers best explained spatiotemporal variation in total microcystin and several individual congeners in a eutrophic freshwater reservoir. Model performance was best for predicting concentrations of the congener MC-LR, with ca. 88% of spatiotemporal variance explained. Most of the variance was associated with changes in the relative abundance of the cyanobacterial genus Microcystis. Follow-up RF regression analyses revealed that factors that were the most important in predicting MC-LR were also the most important in predicting Microcystis population dynamics. We discuss how these results relate to prevailing ecological hypotheses regarding the function of microcystin.

Extraction and detection of arsenicals in seaweed via accelerated solvent extraction with ion chromatographic separation and ICP-MS detection.

An accelerated solvent extraction (ASE) device was evaluated as a semi-automated means of extracting arsenicals from ribbon kelp. The effect of the experimentally controllable ASE parameters (pressure, temperature, static time, and solvent composition) on the extraction efficiencies of arsenicals from seaweed was investigated. The extraction efficiencies for ribbon kelp (approximately 72.6%) using the ASE were fairly independent (< 7%) of pressure, static time and particle size after 3 ASE extraction cycles. The optimum extraction conditions for the ribbon kelp were obtained by using a 3 mL ASE cell, 30/70 (w/w) MeOH/H2O, 500 psi (1 psi = 7 KPa), ambient temperature, 1 min heat step, 1 min static step, 90% vol. flush, and a 120 s purge. Using these conditions, two other seaweed products produced extraction efficiencies of 25.6% and 50.5%. The inorganic species present in the extract represented 62.5% and 27.8% of the extracted arsenic. The speciation results indicated that both seaweed products contained 4 different arsenosugars, DMA (dimethylarsinic acid), and As(V). One seaweed product also contained As(III). Both of these seaweed products contained an arsenosugar whose molecular weight was determined to be 408 and its structure was tentatively identified using ion chromatography-electrospray ionization-mass spectrometry/mass spectrometry (IC-ESI-MS/MS).

An HPLC method with UV detection, pH control, and reductive ascorbic acid for cyanuric acid analysis in water.

Every year over 250 million pounds of cyanuric acid (CA) and chlorinated isocyanurates are produced industrially. These compounds are standard ingredients in formulations for household bleaches, industrial cleansers, dishwasher compounds, general sanitizers, and chlorine stabilizers. The method developed for CA using high-performance liquid chromatography (HPLC) with UV detection simplifies and optimizes certain parameters of previous methodologies by effective pH control of the eluent (95% phosphate buffer: 5% methanol, v/v) to the narrow pH range of 7.2-7.4. UV detection was set at the optimum wavelength of 213 nm where the cyanuric ion absorbs strongly. Analysis at the lower pH range of 6.8-7.1 proved inadequate due to CA keto-enol tautomerism, while at pHs of <6.8 there were substantial losses in analytical sensitivity. In contrast, pHs of >7.4 proved more sensitive but their use was rejected because of CA elution at the chromatographic void volume and due to chemical interferences. The complex equilibria of chlorinated isocyanurates and associated species were suppressed by using reductive ascorbic acid to restrict the products to CA. UV, HPLC-UV, and electrospray ionization mass spectrometry techniques were combined to monitor the reactive chlorinated isocyanurates and to support the use of ascorbic acid. The resulting method is reproducible and measures CA in the 0.5-125 mg/L linear concentration range with a method detection limit of 0.05 mg/L in water.

Analytical methods for water disinfection byproducts in foods and beverages.

The determination of exposure to drinking water disinfection byproducts (DBPs) requires an understanding of how drinking water comes into contact with human through multiple pathways. In order to facilitate the investigation of human exposure to DBPs via foods and beverages, analytical method development efforts were initiated for haloacetonitriles, haloketones, chloropicrin, and the haloacetic acids (HAAs) in these matrices. The recoveries of the target analytes were investigated from composite foods and beverages. Individual foods and beverages used to investigate the general applicability of the developed methods were selected for testing based on their watercontent and frequency of consumption. The haloacetonitriles, the haloketones, and chloral hydrate were generally well recovered (70-130%), except for bromochloroacetonitrile (64%) and dibromoacetonitrile (55%), from foods spiked after homogenization and following extraction with methyl-t-butyl ether (MTBE); the addition of acetone was found to be necessary to improve recoveries from beverages. The process of homogenization resulted in decreased recoveries for the more volatile analytes despite the presence of dry ice. The HAAs were generally well recovered (70-130%), except for trichloroacetic acid (58%) and tribromoacetic acid (132%), from foods but low recoveries and emulsion formation were experienced with some beverages. With both groups of analytes, certain matrices were more problematic (as measured by volatility losses, emulsion formation) than others with regard to processing and analyte recovery.

Evaluation of solid phase microextraction for the analysis of hydrophilic compounds.

Two commercially available solid phase microextraction (SPME) fibers, polyacrylate and carboxen/polydimethylsiloxane (PDMS), were evaluated for their ability to extract hydrophilic compounds from drinking water. Conditions, such as desorption time, desorption temperature, sample temperature, sample stirring, methanol concentration in the sample, and ionic strength of the sample, were optimized for 12 hydrophilic compounds (e.g., amines and alcohols) with both fibers. Accuracy, precision, and method detection limits (MDLs) were determined for the target analytes with both fibers. In general, both fibers exhibited excellent accuracy and precision in the range of 91-110% and 1.0-13%, respectively. The carboxen/PDMS fiber extracted these hydrophilic compounds from water with 10 to 100 times lower MDLs (0.10 to 15 micrograms/l) than the polyacrylate fiber (1.5 to 80 micrograms/l). The MDLs of the carboxen/PDMS fiber demonstrate that SPME is a feasible approach for extracting hydrophilic compounds from drinking water.

Day-night difference in the preferred ambient temperature of human subjects.

To investigate the existence of a day-night difference in thermal preference, 32 college students (16 male, 16 female) were asked to indicate a threshold of discomfort in a climate-controlled environment. The experiment was performed at two distinct times of day that corresponded to the peaks and troughs of the subject's circadian rhythm of body temperature. Males, but not females, felt comfortable at a higher ambient temperature during the trough than during the peak of the body temperature rhythm. These data support the hypothesis (derived from animal studies) that behavioral thermoregulation opposes the circadian rhythm of body temperature.

Acute Effect of Topical (beta-Adrenergic Antagonists on Normal Perimacular Hemodynamics.

PURPOSE: beta-adrenergic antagonists are the most commonly prescribed antiglaucoma medications. The nonselective beta blocker timolol has been shown to either increase or to not affect retinal blood flow, despite the suggestion that the drug may cause vasoconstriction due to its blockade of beta-2 receptors. Other beta blockers with unique pharmacologies may produce different effects on blood flow. We studied the effects of betaxolol (beta-1 selective antagonist), carteolol (nonselective beta-antagonist with intrinsic sympathomimetic activity), and levobunolol (nonselective beta antagonist with intrinsic sympathomimetic activity), and levobunolol (nonselective beta antagonist with active polar metabolite) on perimacular hemodynamics. METHOD: On four separate occasions separated by at least 10 days, 16 normal subjects performed measurements of heart rate, blood pressure, IOP, and blue field entoptic simulation assessment of leukocyte velocity and density before and 2 h after instillation of one of the three drugs or an artificial tears placebo. RESULTS: IOP was significantly reduced by all three drugs as compared to placebo (p < 0.05, Bonferroni-corrected two-tailed paired t-test). No significant differences were found in heart rate, blood pressure, and perimacular leukocyte velocity or density in any of the treatment conditions as compared to placebo. CONCLUSIONS: Betaxolol, carteolol, and levobunolol do not appear to significantly alter perimacular hemodynamics 2 h after administration. The absence of any acute change in hemodynamics despite the drop in IOP suggests normal autoregulation maintained constant blood flow.

Test/Retest reproducibility of color Doppler imaging assessment of blood flow velocity in orbital vessels.

PURPOSE: The purpose of this study was to evaluate the test-retest reproducibility of measures of blood flow velocities using color Doppler imaging (CDI) in orbital arteries. PATIENTS AND METHODS: Measures of peak systolic velocity, end-diastolic velocity, and resistive index were performed in a group of 15 normal tension glaucoma patients and in 15 healthy subjects using the Siemens Quantum 2000 CDI system with a 7.5 MHz linear probe. After each velocity measure in each vessel, the probe was removed and then replaced to repeat the measurement -2 min later. RESULTS: For all subjects, the coefficients of reliability for measures of peak systolic velocity were 12% in the ophthalmic artery, 25% in the central retinal artery, and 19% in a short posterior ciliary artery. For end diastolic velocity, the coefficients were 6%, 11%, and 25%, respectively. The coefficients for resistive index were 4%, 11%, and 38%. CONCLUSION: CDI produces highly reproducible measures in the ophthalmic artery. Measures in the central retinal artery are somewhat more variable but seem reasonably reproducible. Short posterior ciliary measurements were the most variable, suggesting that current methods for assessing these vessels may not be sufficiently reliable.

Color Doppler analysis of ocular vessel blood velocity in normal-tension glaucoma.

The pathogenesis of normal-tension glaucoma remains unknown. Because ocular vasospasm has been proposed as a possible mechanism, we investigated ocular vessel flow velocity in normal-tension glaucoma patients at rest and under treatment with a cerebral vasodilator. Ten normal-tension glaucoma patients and nine age- and gender-matched controls had flow velocity measured in three vessels (ophthalmic artery, central retinal artery, and temporal short posterior ciliary artery) by using color Doppler imaging, under baseline conditions and during carbon dioxide supplementation sufficient to increase end-tidal PCO2 by 15%. Peak systolic and end-diastolic velocities were measured, and the resistance index (peak systolic velocity minus end-diastolic velocity, divided by peak systolic velocity) was calculated. Compared with controls, these normal-tension glaucoma patients had significantly lower end-diastolic velocities (P = .002) and higher resistance indices (P = .007) in the ophthalmic artery at baseline. When PCO2 was increased, control subjects remained unchanged, whereas it increased end-diastolic velocity in patients (P = .003) and abolished the difference in resistance index between the two groups. Patients and control subjects differed little in their baseline or carbon dioxide response velocities or in resistance in the other two vessels. These results indicate that at baseline these normal-tension glaucoma patients may have increased vascular resistance distal to the ophthalmic artery, although this increased resistance cannot be specifically ascribed to the central retinal arterial or to temporal short posterior ciliary arterial vascular beds. The responsiveness of these patients to a cerebral vasodilator (increased PCO2) indicates further that the increased resistance distal to the ophthalmic artery may be the reversible result of vasospasm.

Regulation of retinal blood flow during blood gas perturbation.

In this review, we discuss the regulation of ocular blood flow, the role it may play in disease, and the physiologic methods used to assess it. Altered perfusion may play a role in several ocular diseases. In particular, derangements in ocular blood flow have been offered as possible factors in the development of glaucomatous optic neuropathy. In fact, recent studies have suggested that glaucomatous eyes may lack the degree of circulatory autoreg-ulation found in healthy eyes. The autoregulation of ocular blood flow is in response to tissue pressure, myogenic changes, and metabolic levels. To assess the eye's vascular autoregulation, physiologic perturbation of these mechanisms during hemodynamic assessment provides an autoregulatory "stress test." Along these lines, we have refined the technique of altering arterial levels of oxygen and carbon dioxide, thus provoking a metabolic autoregulatory response, during measures of blood velocities in ocular vessels. Using somewhat similar methods, others have documented changes in ocular hemodynamics by altering blood gas levels. Results of our own work suggest that retinal blood flow may be regulated to maintain constant oxygen delivery. Likewise, our studies using carbon dioxide suggest that retinal blood flow may be steeply dependant on Pco2, for these results to be placed in their proper context, future studies directed at better defining normal autoregulation are needed.