Assessment of animal impacts on bacterial water quality in a South Carolina, USA tidal creek system.

Fecal pollution may adversely impact water quality in coastal ecosystems. The goal of this study was to determine whether cattle were a source of fecal pollution in a South Carolina watershed. Surface water samples were collected in June 2002 and February through March 2003 in closed shellfish harvesting waters of Toogoodoo Creek in Charleston County, SC. Fecal coliform concentrations in 70 % of the water samples taken for this study exceeded shellfish harvesting water standards. Ribotyping was performed in order to identify animal sources contributing to elevated fecal coliform levels. Escherichia coli isolates (n = 253) from surface water samples were ribotyped and compared to a ribotype library developed from known sources of fecal material. Ribotypes from water samples that matched library ribotypes with 90 % maximum similarity or better were assigned to that source. Less than half of the unknown isolates (38 %) matched with library isolates. About half (53 %) of the matched ribotypes were assigned to cattle isolates and 43 % to raccoon. Ribotyping almost exclusively identified animal sources. While these results indicate that runoff from cattle farms was a likely source of fecal pollution in the watershed, wildlife also contributed. Given the small size of the library, ribotyping was moderately useful for determining the impact of adjacent cattle farms on Toogoodoo Creek. Increasing the number and diversity of the wildlife sources from the area would likely increase the usefulness of the method.

F+ RNA coliphage typing for microbial source tracking in surface waters.

AIMS: The utility of coliphages to detect and track faecal pollution was evaluated using South Carolina surface waters that exceeded State faecal coliform standards. METHODS AND RESULTS: Coliphages were isolated from 117 surface water samples by single agar layer (SAL) and enrichment presence/absence (EP/A) methods. Confirmed F+ RNA coliphages were typed for microbial source tracking using a library-independent approach. Concentrations of somatic coliphages using 37 and 44.5 degrees C incubation temperatures were found to be significantly different and the higher temperature may be more specific for faecal contamination. The EP/A technique detected coliphages infecting Escherichia coli Famp in 38 (66%) of the 58 surface water samples negative for F+ coliphages by the SAL method. However, coliphages isolated by EP/A were found to be less representative of coliphage diversity within a sample. Among the 2939 coliphage isolates tested from surface water and known source samples, 813 (28%) were found to be F+ RNA. The majority (94%) of surface water F+ RNA coliphage isolates typed as group I. Group II and/or III viruses were identified from 14 surface water stations, the majority of which were downstream of wastewater discharges. These sites were likely contaminated by human-source faecal pollution. CONCLUSIONS: The results suggest that faecal contamination in surface waters can be detected and source identifications aided by coliphage analyses. SIGNIFICANCE AND IMPACT OF THE STUDY: This study supports the premise that coliphage typing can provide useful, but not absolute, information to distinguish human from animal sources of faecal pollution. Furthermore, the comparison of coliphage isolation methods detailed in this study should provide valuable information to those wishing to incorporate coliphage detection into water quality assessments.

Risk of leukemia in children treated with human growth hormone: review and reanalysis.

BACKGROUND: Data have suggested that any increased incidence of leukemia in growth-hormone (GH)-treated patients was limited to those with known risk factors for leukemia. However, previous studies may have overestimated the numbers of patient-years of risk by not excluding data from "positive-risk-factor" patients. This risk was reanalyzed by using data on children in the National Cooperative Growth Study (NCGS), with correction for this possible confounding factor. METHODS: The risk of leukemia in GH-treated patients without known risk factors was determined by using patient-years of GH therapy and patient-years since first exposure to GH therapy and the values obtained were compared with values from the Surveillance, Epidemiology, and End Results program of the National Cancer Institute. RESULTS: Three cases of leukemia in patients without known risk factors were found in the NCGS database; 3.42 cases would be expected in the 119,846 patient-years in the analysis using time since GH exposure. Two of these cases of leukemia occurred during GH therapy (67,773 patient-years); 2.13 cases would be expected. CONCLUSION: Excluding data on patients with known risk factors for leukemia provides a more accurate estimate of the risks in GH-treated patients. The incidence of leukemia in these patients is comparable to that in the general population of age-matched children.

Utility of the National Cooperative Growth Study database for safety reporting.

The National Cooperative Growth Study (NCGS) maintains the largest database in the world, collecting information on patients with growth disorders treated with growth hormone (GH). More than 24,000 children have been monitored during its first decade (1985 through 1995). The database provides unique opportunities to learn about effectiveness of GH therapy in a real-world context. Its size also makes it possible to investigate rare adverse reactions, which is not reasonable in a randomized controlled trial (RCT). The frequency of adverse experiences reported in an observational study like the NCGS is lower than in an RCT, because the investigators in an observational study typically do not report events that they believe are clearly unrelated to the study drug. Nevertheless the NCGS has greatly facilitated collecting adverse-event data; approximately 75% of all adverse-event reports for GH received by the manufacturer are through NCGS data-collection forms. The NCGS has thus amassed a repository of GH safety data that is unparalleled. Furthermore, in contrast to an RCT, the NCGS database reflects real-world experience with long-term GH therapy in North America. Although the advantages of an observational study such as the NCGS must be recognized, such a study does differ from an RCT in important ways. For example, because the NCGS protocol allows customized patient treatment and individualization of GH therapy, it may be difficult to use the database to address questions (e. g., estimation of dose-response relationships, true incidence of adverse events) that require RCT designs.

Brain tumor recurrence in children treated with growth hormone: the National Cooperative Growth Study experience.

As of October 1993 the National Cooperative Growth Study included 1262 children with brain tumor who were treated with growth hormone. The type of brain tumor was specified in 947 (75%) of these children. The most common types were glioma, medulloblastoma, and craniopharyngioma, accounting for 91.3% of all those for which type was specified. Brain tumor recurred in 83 (6.6%) of the 1262 children over a total of 6115 patient-years at risk. The frequencies of tumor recurrence in children with low-grade glioma (18.1%), medulloblastoma (7.2%), and craniopharyngioma (6.4%) are lower than those in published reports of tumor recurrence in the general pediatric population with the same types of tumors. The analysis cannot conclusively show that no increased risk of tumor recurrence exists, however, because of the potential incompleteness of data reporting in the National Cooperative Growth Study. Nevertheless the findings are reassuring that children with the more common types of brain tumor who are treated with growth hormone do not seem to be at excessive risk for tumor recurrence.

Respirometric analysis of the biodegradation of organic contaminants in soil and water.

Client-funded bench-scale investigations concerning the likelihood of successfully applying biological remediation to hazardous wastes must be cost-effective, and they usually need only determine if biodegradation is likely to occur on site. To assess the potential for stimulating biodegradation, biochemical oxygen demand (BOD) was used to continuously monitor bacterial respiration during growth on mixed organic wastes from contaminated water and soil. Continuously collected oxygen-consumption data provided information on the overall metabolic activity of the resident bacterial population and permitted direct observation of the cessation of microbial respiratory activity and, thus, the termination of aerobic degradation. The correlation of biological oxygen utilization with biodegradation was confirmed using independent analytical methods. Continuous, long-term BOD analysis was applied to bench-scale studies to assess the biodegradation of mixed organic wastes from contaminated sites and industrial waste effluents. This information was used to make an initial determination regarding the need to further explore bioremediation as a potential remedial-action technology using on-site, pilot-scale testing.

Effect of oxygen on photoautotrophic and heterotrophic growth of Chlamydomonas reinhardtii in an anoxic atmosphere.

Photoautotrophic growth of Chlamydomonas reinhardtii was shown to be independent of the presence of atmospheric oxygen. Under constant light and photoautotrophic conditions, C. reinhardtii grew equally well in either air or 367 PPM CO2-in-He. During 12-h light-dark cycles, the cells in air grew substantially faster than those grown in CO2-in-He, indicating a significant role for O2 in dark metabolism. Although cells grown under CO2-in-He were not supplied any exogenous O2, photosynthetic water splitting resulted in the liberation of O2. The effect of photoevolved O2 on the growth of C. reinhardtii was examined (1) by measuring the amount of O2 consumed by photosynthesizing algae, (2) by growing the algae heterotrophically on acetate in the dark and supplied with O2 generated by photoautotrophically grown cells, and (3) by determining the minimum level of O2 needed to stimulate CO2 evolution from cells suspended in minimal medium supplemented with acetate. The results from these investigations indicated that exogenous O2 was not required for photoautotrophic growth by C. reinhardtii and that this alga grew in an anoxic environment if supplied with CO2 and light.

Hydralazine dose-response curve analysis.

A multicenter, parallel, double-blind, 181-patient study compared the safety and antihypertensive efficacy of immediate-release (IR) and extended-release (ER) hydralazine. After 2 to 4 weeks on diuretic, patients were maintained on diuretic and randomized to a hydralazine treatment regimen: IR thrice daily, ER twice daily, or ER once daily. Daily doses of hydralazine were 75, 150, or 300 mg. Although designed as a titration study, important dose-response data were available for analysis with nonlinear mixed effect modeling (NONMEM). Sitting diastolic blood pressure (BP) was selected as the response variable. Several factors were tested for importance, including body weight, time (week) effects, concomitant beta-blocker (BB) therapy, acetylator class, and treatment regimen. All factors were important (p less than 0.05) except treatment regimen (p greater than 0.30). The maximum antihypertensive response (Emax) to hydralazine was 9.4 mm Hg. The daily dose that elicited 50% of the maximum response (D50) was 0.87 mg/kg for slow acetylators and 1.68 mg/kg for fast acetylators. BP fell 0.52 mm Hg per week independent of other effects, and concomitant BB therapy induced a drop of 6.6 mm Hg in addition to hydralazine, diuretic and week effects. NONMEM's use assisted in evaluations and provided information not obtainable through traditional means.

Application of NONMEM to routine bioavailability data.

Although NONMEM has been proposed as a modeling tool for sparse data sets, little work has described its application to pharmacokinetic data which is also amenable to typical evaluations. An analysis was performed with NONMEM using plasma concentration data obtained during the development of liquid and capsule extended-release (ER) pseudoephedrine products. A total of four studies (single dose and steady-state studies for both the liquid and capsule formulations) were evaluated, each with an immediate-release (IR) control, and consisting of 18 to 20 subjects. NONMEM analyses provided additional information which could not be obtained through traditional means. Specifically, NONMEM provided not only estimates of residual error from single dose and steady-state studies but also a stochastic measure of bioinequivalence and dose-dumping. It permitted hypothesis testing in the same process as pharmacokinetic parameter estimation, such as contrasting absorption rates from capsule and suspension ER products. A less biased estimate of absorption rate was obtainable for ER formulations by utilizing IR runs. Finally, these NONMEM runs confirmed that, even when data are plentiful and amenable to two-stage analyses, NONMEM provides estimates that may in fact be more meaningful and less susceptible to assay or residual variability. Fundamental differences between population and two-stage approaches are discussed.

The influence of assay variability on pharmacokinetic parameter estimation.

The impact of assay variability on pharmacokinetic modeling was investigated. Simulated replications (150) of three "individuals" resulted in 450 data sets. A one-compartment model with first-order absorption was simulated. Random assay errors of 10, 20, or 30% were introduced and the ratio of absorption rate (Ka) to elimination rate (Ke) constants was 2, 10, or 20. The analyst was blinded as to the rate constants chosen for the simulations. Parameter estimates from the sequential method (Ke estimated with log-linear regression followed by estimation of Ka) and nonlinear regression with various weighting schemes were compared. NONMEM was run on the 9 data sets as well. Assay error caused a sizable number of curves to have apparent multicompartmental distribution or complex absorption kinetic characteristics. Routinely tabulated parameters (maximum concentration, area under the curve, and, to a lesser extent, mean residence time) were consistently overestimated as assay error increased. When Ka/Ke = 2, all methods except NONMEM underestimated Ke, overestimated Ka, and overestimated apparent volume of distribution. These significant biases increased with the magnitude of assay error. With improper weighting, nonlinear regression significantly overestimated Ke when Ka/Ke = 20. In general, however, the sequential approach was most biased and least precise. Although no interindividual variability was included in the simulations, estimation error caused large standard deviations to be associated with derived parameters, which would be interpreted as interindividual error in a nonsimulation environment. NONMEM, however, acceptably estimated all parameters and variabilities. Routinely applied pharmacokinetic estimation methods do not consistently provide unbiased answers. In the specific case of extended-release drug formulations, there is clearly a possibility that certain estimation methods yield Ka and relative bioavailability estimates that would be imprecise and biased.

A simple apparatus for screening absolute photosynthetic rates of single algal colonies in an anoxic atmosphere.

Photosynthetically generated O(2) was measured from single algal colonies in a He atmosphere, using an enhanced Hersch galvanic cell. The enhancement consisted of using ultrapure potassium hydroxide as the electrolyte and ultrapure lead as the anode. The galvanic cell was placed in a regulated helium-flow system containing a reaction cuvette with the colonies and an electrolysis cell for calibration. Colonies were individually irradiated using a He-Ne laser. Data collection and laser positioning for colony irradiation were microcomputer controlled. This assay system was capable of detecting O(2) production rates of 500 femtomoles per second with a signal to noise ratio of 2, a level of sensitivity that permitted the detection of photoevolved O(2) from single algal colonies. This capability provides, for the first time, an approach for quantitatively measuring the absolute rate of photosynthetic O(2) evolution from a single algal colony.

Pseudoephedrine absorption from controlled release formulations: absorption rate constant estimation methods.

Five methods of absorption rate (Ka) estimation were compared using data from a previously reported bioavailability study: Wagner-Nelson (WN), asymptotic WN (AWN), the Hendeles-Weinberger modification of the WN (HW), nonlinear regression performed on plasma concentration vs time data with fixed elimination rates (NL), and nonlinear regression performed on the cumulative sum of AUCs obtained during WN analysis (AUCNL). The maximum plasma concentration (Cmax) and the time at which Cmax occurred (Tmax) were predicted with each method's respective Ka's, and these were compared to observed values. The WN and HW were consistently biased in their predictions of Cmax and Tmax, providing slower Ka's than any of the other methods. AWN could only be used for the reference treatment. Both NL and AUCNL provided unbiased estimations, but AUCNL had a high level of imprecision. With this set of data, only NL was an acceptable method for accurately estimating Ka.

Establishment of control parameters for in situ, automated screening of sustained hydrogen photoproduction by individual algal colonies.

An apparatus was constructed which allowed automated screening of individual microalgal colonies for sustained ability to photoevolve H(2) during anaerobic photosynthesis. The main components of this apparatus were a microcomputer, a He-Ne laser mounted on a computer-controlled X-Y translation stage, a flow-through chamber which contained an agar plate of colonies, and a H(2) detector which interfaced with the microcomputer for data collection. The system was capable of detecting a minimum production rate of 1 nanomole of H(2) per hour per colony and provided an efficient means of screening relatively large numbers of algal colonies. Examination of the effect of the spacing of colonies on the agar plate, light intensity, stability of colonies within a screening period, colony age, chlorophyll content, and colony size on H(2) yield indicated that, under optimum conditions, yields from genetically uniform colonies varied by no more than a factor of 2 in their H(2)-producing ability. Therefore, colonies of algae whose H(2) yields lie outside this intrinsic twofold variability can be identified and selected as natural variants or mutants. A description of the construction and of the apparatus is presented, and the experimental results used to establish the control parameters for Chlamydomonas reinhardtii colonies are discussed.

Influence of a standard meal on the absorption of a controlled release pseudoephedrine suspension.

The influence of a standard meal on the extent and rate of absorption of pseudoephedrine from a liquid controlled release (CR) formulation (Pennkinetic System) was studied in 16 normal male volunteers. Equivalent single doses of an immediate release reference syrup (IR-Sudafed) and the CR suspension were each studied under both fasted and postprandial conditions. AUC results showed no significant influence of food or formulation, indicating that the CR formulation was absorbed to the same extent as the IR product under all conditions. CMAX and TMAX tabulations under fasted conditions indicated that the CR preparation peaked at a lower level at a later time. Food diminished IR CMAXs but the CR suspension's CMAXs and TMAXs were apparently unaffected by food. This study indicates that this CR pseudoephedrine suspension releases drug independently of food intake.

Chronology of the differentiation of cotton (Gossypium hirsutum L.) fiber cells.

Cotton fibers are single elongated cells that develop from epidermal cells of the ovule. The chronology of fiber differentiation was investigated using cultured ovules. Epidermal cells differentiate into fiber cells approx. 3 d before anthesis. When ovules were cultured on a defined medium, fiber growth could be initiated on ovules any time between 2 d preanthesis and the time of anthesis by adding indole-3-acetic acid and gibberellic acid to the medium. In the absence of phytohormones, fibers did not grow, and when ovules between 2 d preanthesis and anthesis were cultured without hormones past the day of anthesis and hormones then added, most ovules failed to produce fibers. The results define the timing of fiber differentiation from epidermal cells, and also define a window of time when differentiated cells are capable of further development. During this window, fiber cells are latent awaiting appropriate stimulation which in the intact plant is apparently associated with anthesis.

Influence of a standard meal on the absorption of controlled-release pseudoephedrine capsules.

The influence of a standard meal on the extent and rate of absorption of pseudoephedrine from a controlled-release (CR) capsule formulation (Pennkinetic System) was studied in 16 normal male volunteers. Equivalent single doses of an immediate-release (IR) pseudoephedrine reference syrup and the CR capsules were each studied under both fasted and postprandial conditions. Pharmacokinetic analysis evaluated the fraction of drug absorbed over time as determined by the Wagner-Nelson method. The area under the drug concentration versus time curve (AUC) results were not influenced by food or formulation, indicating that the CR formulation was absorbed to the same extent as the IR syrup. The maximum plasma concentration (Cmax) and the time to maximum concentration (tmax) tabulations under fasted conditions indicated that the CR preparation peaked at a lower level and a later time than the IR syrup. Food minimally affected the Cmax of the IR formulation, but did not affect that of the CR formulation. Food delayed the tmax of both the IR and CR preparations by less than 1 h, but only the delay of the CR formulation was statistically significant. Neither delay was considered clinically meaningful. The results of this study strongly suggest that the pharmacokinetic profile of the CR pseudoephedrine capsules was minimally affected by the presence of food.

Patient-controlled analgesia in the terminally ill cancer patient.

Patient-controlled analgesia (PCA) is a relatively new therapeutic modality which has allowed postsurgical patients to safely and effectively self-administer doses of intravenous narcotics via a syringe pump and sequencing device. A pilot study was designed to evaluate PCA's safety and effectiveness in the terminally ill cancer patient. Eight patients whose chronic pain was not adequately controlled by oral narcotics were permitted to use PCA for a minimum of 48 hours. Respiratory rates, sedation rankings, and pain rankings indicated these patients achieved satisfactory analgesia with a minimum of sedation and experienced no respiratory depression. Three patients were switched to oral regimens using PCA dosing as a guide. Pain and sedation rankings were similar to those registered while exclusively on PCA. This self-dosing technique was judged to be safe, effective, and able to accommodate wide fluctuations in analgesic need when treating pain in the terminally ill cancer patient. The results obtained in these patients support further trials using PCA to individualize oral analgesic regimens.

Pharmacologic evaluation of loratadine (SCH 29851), chlorpheniramine and placebo.

The antihistaminic effect of loratadine (160 mg) was compared in twenty-four normal male volunteers to chlorpheniramine maleate (4 mg) and placebo in a double blinded 3-way cross-over study of latin square design. After receiving single oral doses of each medication, the wheal response to serial 0.1 ml intradermal histamine (2 micrograms) and saline (control) injections were recorded over a 24-h period. The calculated wheal areas were compared to base-line measurements. The results were analyzed by analysis of variance. Loratadine exhibited a more pronounced inhibition of histamine wheal formation than placebo or chlorpheniramine maleate (p less than 0.003). In contrast to chlorpheniramine maleate which had a duration of action of only 3 h, loratadine inhibited the response for the entire observation period between 1 and 24 h post-dose. Although sedation was observed less frequently with loratadine (Placebo, n = 2; chlorpheniramine, n = 3; and loratadine, n = 1), the relative incidence were not statistically significant.