Surveillance of RNase P, PMMoV, and CrAssphage in wastewater as indicators of human fecal concentration across urban sewer neighborhoods, Kentucky.

Wastewater surveillance has been widely used as a supplemental method to track the community infection levels of severe acute respiratory syndrome coronavirus 2. A gap exists in standardized reporting for fecal indicator concentrations, which can be used to calibrate the primary outcome concentrations from wastewater monitoring for use in epidemiological models. To address this, measurements of fecal indicator concentration among wastewater samples collected from sewers and treatment centers in four counties of Kentucky (N = 650) were examined. Results from the untransformed wastewater data over 4 months of sampling indicated that the fecal indicator concentration of human ribonuclease P (RNase P) ranged from 5.1 × 101 to 1.15 × 106 copies/ml, pepper mild mottle virus (PMMoV) ranged from 7.23 × 103 to 3.53 × 107 copies/ml, and cross-assembly phage (CrAssphage) ranged from 9.69 × 103 to 1.85 × 108 copies/ml. The results showed both regional and temporal variability. If fecal indicators are used as normalization factors, knowing the daily sewer system flow of the sample location may matter more than rainfall. RNase P, while it may be suitable as an internal amplification and sample adequacy control, has less utility than PMMoV and CrAssphage as a fecal indicator in wastewater samples when working at different sizes of catchment area. The choice of fecal indicator will impact the results of surveillance studies using this indicator to represent fecal load. Our results contribute broadly to an applicable standard normalization factor and assist in interpreting wastewater data in epidemiological modeling and monitoring.

Wastewater Sample Site Selection to Estimate Geographically Resolved Community Prevalence of COVID-19: A Sampling Protocol Perspective.

Wastewater monitoring for virus infections within communities can complement conventional clinical surveillance. Currently, most SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) clinical testing is voluntary and inconsistently available, except for a few occupational and educational settings, and therefore likely underrepresents actual population prevalence. Randomized testing on a regular basis to estimate accurate population-level infection rates is prohibitively costly and is hampered by a range of limitations and barriers associated with participation in clinical research. In comparison, community-level fecal monitoring can be performed through wastewater surveillance to effectively surveil communities. However, epidemiologically defined protocols for wastewater sample site selection are lacking. Herein, we describe methods for developing a geographically resolved population-level wastewater sampling approach in Jefferson County, Kentucky, and present preliminary results. Utilizing this site selection protocol, samples (n = 237) were collected from 17 wastewater catchment areas, September 8 to October 30, 2020 from one to four times per week in each area and compared to concurrent clinical data aggregated to wastewater catchment areas and county level. SARS-CoV-2 RNA was consistently present in wastewater during the studied period, and varied by area. Data obtained using the site selection protocol showed variation in geographically resolved wastewater SARS-CoV-2 RNA concentration compared to clinical rates. These findings highlight the importance of neighborhood-equivalent spatial scales and provide a promising approach for viral epidemic surveillance, thus better guiding spatially targeted public health mitigation strategies.

A rapid assessment of wastewater for genomic surveillance of SARS-CoV-2 variants at sewershed scale in Louisville, KY.

In this communication, we report on the genomic surveillance of SARS-CoV-2 using wastewater samples in Jefferson County, KY. In February 2021, we analyzed seven wastewater samples for SARS-CoV-2 genomic surveillance. Variants observed in smaller catchment areas, such as neighborhood manhole locations, were not necessarily consistent when compared to associated variant results in downstream treatment plants, suggesting catchment size or population could impact the ability to detect diversity.

Increased progesterone receptor concentrations in bladder lesions of estrogen-treated Syrian hamsters.

During studies of renal tumorigenesis induced by estrogen in Syrian hamsters, we have observed that about 15 to 20% of animals develop bladder lesions with an increased wet weight of tissue from 0.2 g to 0.4 to 1.7 g. Histological examination of the lesions showed a spectrum of changes from inflammatory reactions to squamous metaplasia and intense hyperplasia of the transitional epithelium. The concentration of progesterone-binding sites was increased in the bladders with lesions. No specific progesterone-binding sites could be detected in the cytosol of bladders from hamsters not treated with estrogen. The affinity constant for the progesterone-binding sites in cytosol from bladders with lesions was 10(9) M-1, the same as that reported for progesterone receptors in other target tissues for estrogen. The binding sites are specific for progesterone and are not competed for by 17 beta-estradiol, 5 alpha-dihydrotestosterone, or aldosterone.

Receptor characteristics of specific estrogen binding in the renal adenocarcinoma of the golden hamster.

Linear sucrose gradient analyses reveal that all estrogen-induced and -dependent primary renal tumor cytosols examined contain an 8 S and variable amounts of 4 S receptor in low ionic buffer concentrations. Similar results were obtained with extracts of primary metastases of these tumors. Sucrose gradients containing high salt (0.4 M KCl) convert the 8 S receptor in both the hamster renal tumor and uterus to a 4 to 5 S complex. Scatchard plot analysis reveals that the renal tumor cytosol estradiol-receptor complex has a Ka of 1.7 X 10(9) M-1 and 9.2 X 10(-10) M binding sites. Competition for the tritiated 17beta-estradiol binding sites in the renal tumor was similar to that in the uterus with respect to estrogenic compounds. Nonestrogenic steroids exhibited minimal competition at the same concentrations or higher. Substitution in the ring structure, particularly in position 3 of the phenolic A-ring, resulted in a considerable loss in the ability of such compounds to compete for these receptors. Aniestrogens were effective competitors for these estrogen receptors only at higher concentrations relative to the tritiated estradiol.

Pharmacodynamics and pharmacokinetics of higher-dose, double-bolus eptifibatide in percutaneous coronary intervention.

BACKGROUND: Pharmacodynamics of eptifibatide, a cyclic heptapeptide antagonist of platelet glycoprotein IIb/IIIa, are substantially altered by anticoagulants that chelate calcium, resulting in overestimation ex vivo of the in vivo effects of this agent. We conducted a dose-ranging study to characterize the pharmacodynamics and pharmacokinetics of eptifibatide under physiological conditions. METHODS AND RESULTS: Patients (n=39) undergoing elective percutaneous coronary intervention were randomly assigned to an eptifibatide bolus followed by an infusion (180-microgram/kg bolus followed by 2 microgram/kg per minute or 250-microgram/kg bolus followed by 3 microgram/kg per minute) for 18 to 24 hours. In a 2:1 ratio, these patients received either a second bolus of eptifibatide (90 microgram/kg or 125 microgram/kg for the initial 180-microgram/kg or 250-microgram/kg groups, respectively) or placebo 30 minutes after the initial bolus. Bleeding times, ex vivo platelet aggregation, receptor occupancy, and plasma eptifibatide levels at baseline and at 1, 2, 3, 4, 6, and 8 hours were evaluated. Platelet inhibition was dose dependent and >80% in all groups by steady state. The single-bolus regimens had a transient loss of inhibition at 1 hour, consistent with rapid distribution and drug elimination. Pharmacokinetic modeling suggested that optimal dosing of eptifibatide would be obtained with a 180-microgram/kg bolus and a 2-microgram/kg per minute infusion followed by a second 180-microgram/kg bolus 10 minutes later. CONCLUSIONS: A novel higher-dose, double-bolus regimen of eptifibatide in coronary intervention attains and maintains >90% inhibition of platelet aggregation in >90% of patients, providing the pharmacodynamic construct for the design of the Enhanced Suppression of the Platelet IIb/IIIa Receptor with Integrilin Therapy (ESPRIT) trial of adjunctive eptifibatide in coronary stent implantation.

Changes in estrogen receptor levels during deciduomata development in the pseudopregnant rat.

The presence of a cytoplasmic estradiol receptor with an affinity constant of 10(10)M-1 has been demonstrated in the differentiated fibroblasts comprising the deciduomata in pseudopregnant rats. The receptor had a sedimentation coefficient of 6S in sucrose density gradients containing a physiological concentration of salt, and estradiol binding was completely abolished by a hundred-fold excess of unlabeled estradiol. During the early stages of decidualization (days 2 and 3), the decidualized uterine horn or isolated deciduomal tissue contained a concentration of receptor comparable to that in non-gravid uteri from rats ovariectomized at estrus. By day 5 of decidualization, the concentration of estradiol binding in deciduomal tissue decreased to about one-half the concentration measurable at days 2 and 3 of decidualization despite continued tissue growth until day 7. By day 7 of decidualization, estradiol binding had decreased to about 20% of the concentration on days 2 and 3 of decidualization. Cytosol from untreated uterine horns of rats bearing deciduomata bound the same amount of estradiol through day 7 as that in uteri of rats ovariectomized at estrus. These observations are discussed in terms of steroid hormone involved in deciduomal growth and regression.

Changes in progesterone receptor levels during deciduomata development in the pseudopregnant rat.

A specific cytoplasmic progesterone receptor has been identified and quantified in the deciduomata of the pseudopregnant rat. The receptor had a sedimentation coefficient of 6-7S on sucrose density gradients and was inactivated by proteolytic enzymes, sulfhydryl blocking agents and elevated temperature. The equilibrium association constant for the binding of progesterone by the deciduomal receptor was determined to be approximately 10(9)M-1. The concentrations of progesterone receptor sites in cytosols prepared from deciduomata on day 3 and 5 of decidualization were 3.4 +/- 0.3 X 10(-10)M and 3.6 +/- 0.4 X 10(10)M, respectively, when normalized to a protein concentration of 1 mg/ml. These concentrations of progesterone receptor sites were similar to that measured in the uterine cytosol of estrous rats or in the contralateral untreated uterine horn of rats undergoing decidualization. Following day 5 of decidualization the concentrations of progesterone receptor sites decreased linearly so that by day 7 the concentration was approximately one-half that at days 3 and 5. The physiological significance of the progesterone receptor and the decrease in its concentration with time are discussed with regard to their influence on the decidualization reaction.

Biochemical comparison of estrogen receptors of the hamster hypothalamus and uterus.

Comparison of the soluble estradiol receptor proteins in the hypothalamus and uterus of the golden Syrian hamster revealed that both have sedimentation coefficients of 8S in a low ionic strength buffer and migrate similarly on analytical disc-gel electrophoresis in both 5% and 7% acrylamide gels. The competition of [3H]-17beta-estradiol binding by unlabeled estrogenic as well as nonestrogenic compounds was similar with the receptors present in the two tissues. The affinity constants for the hypothalamic and uterine receptors were 4.3 X 10-9M-1 and 1.6 X 10-10M-1, respectively; the number of binding sites when expressed on an equivalent protein basis (i.e. 1 mg/ml) was 0.18 X 10-10M and 3.1 X 10-10M, respectively. Identical amounts of soluble receptors with similar sedimentation characteristics were found in hypothalami from male and female animals gonadectomized 64 h prior to use. A titration of binding sites in the uteri of intact and ovariectomized animals revealed decreased binding in the supernatant preparation from intact animals, probably due to the presence of bound endogenous unlabeled hormone.

Genetic evidence for limited dispersal in the coastal California killifish, Fundulus parvipinnis.

The California killifish, Fundulus parvipinnis, is a marine species that lives in salt marshes, estuaries and wetlands along the California and Baja California coasts. In order to estimate levels of dispersal between different coastal habitats over its range, we have studied six populations using morphological and genetic markers. Lateral line scale and vertebrae counts showed significant differences between individuals collected north of Punta Eugenia and south of Punta Eugenia. Morphological differences across Punta Eugenia were accompanied by large genetic differences at the mitochondrial control region (5.8%). Gene flow was in general very reduced over the range of the species (pairwise average F(st)=0.70, Nm=0.30), with a strong break at Punta Eugenia (F(st)=0.95, Nm=0.03). Such limited interchanges between coastal habitats have important theoretical and conservation implications.

Evaluation of an indirect enzyme-linked immunosorbent assay for screening antibody against aflatoxins.

Enzyme-linked immunosorbent assay screening of antibody produced against aflatoxin was accomplished by a new and simple procedure. To demonstrate the new indirect ELISA technique used, antibody against aflatoxin M1 was produced in female BALB/CJ mice by immunization with an aflatoxin M1-bovine serum albumin conjugate. Instead of coating test-plate wells with purified antibody (direct ELISA) or synthesizing a second protein-aflatoxin conjugate (aflatoxin M1-poly-L-lysine) to coat test-plate wells, wells were coated with the readily available aflatoxin M1-bovine serum albumin and aflatoxin B1-bovine serum albumin. This method, applicable for any aflatoxin conjugated by the common cyclopentano-carboxymethoxyl-oxime technique, eliminates the more time-consuming and technically difficult portions of earlier direct and indirect ELISA. The new technique can be valuable in continued efforts toward development of new and improved immunoassays against aflatoxin metabolites.

Problem identification in apparently well neonates: implications for early discharge.

The frequency, time of identification, and type of problems of newborns in an urban indigent population were prospectively studied during their hospital stay to evaluate feasibility of early hospital discharge. Eight percent (563) of 7,021 term and near-term low-risk infants developed one or more predefined problems. Of those with problems, 42.1% received therapy and/or a higher level of care. Tachypnea, temperature instability, and cyanotic episodes were the most frequently treated problems. Nearly 69% of all problems were detected after the initial examination, and 31% developed problems after 24 hours of age; 5% were transferred to the NICU. Problems occurring after 24 hours of age emphasize the need for follow-up within days after hospital discharge in this population.

Computer-aided characterization and optimization of the Thumper compression waveform in closed-chest CPR.

Optimization of blood flow and blood pressure is critical for a positive outcome of cardiopulmonary resuscitation (CPR). The use of models can facilitate the analysis of different methods of performing CPR. Programming with SPICE software, the authors modified a hardware model electrically equivalent to the human cardiovascular system into a software model to analyze and predict the hemodynamic output parameters of different compression waveform inputs used in thoracic pump CPR. Thumper waveforms produced with the computer model predicted the same decaying frequency response as those found in other studies with humans and a recording manikin in which the generated pressures decreased with increased compression rates from 60 to 140 cycles/min. Using a square-wave input having a flat frequency response, the model predicted a constant diastolic pressure and a steady increase in systolic pressure from 94 to 109 mmHg while increasing compression rate from 60 to 140 cycles/min at a constant 50% duty cycle. Blood flows predicted with the use of a square-wave compression input were greater than the flows predicted with the use of a Thumper waveform, and increased with an increase in compression rate. The model also predicted increased mean systolic pressure when the duty cycle increased from 20% to 80% while diastolic pressure decreased slightly. Although mean systolic pressure increased with increased duty cycle, the model predicted a decrease in coronary flow at the higher duty cycles of all compression waveforms. Increases of 75% to 100% in systolic pressure and blood flow were produced when increases in chest compression force and the rapidity of the rise time, termed "spike-impulse" CPR, were simulated.

Sexual function and satisfaction in heterosexual couples when men are administered sildenafil citrate (Viagra) for erectile dysfunction: a multicentre, randomised, double-blind, placebo-controlled trial.

OBJECTIVE: To investigate the effect of improvement in erectile dysfunction (ED) on sexual function and satisfaction measures in heterosexual couples in which the woman reports that sexual intercourse is unsatisfactory at least half of the time. DESIGN: Multicentre, double-blind, placebo-controlled study. SETTING: Outpatient medical clinics. POPULATION: Hundred and eighty men with ED and their female partners in whom sexual intercourse was satisfactory about half the time or less (score of < or =3 on the Female Partner of ED Subject Questionnaire question 3 [FePEDS Q3]). METHODS: Men were randomised to flexible-dose sildenafil (25, 50, and 100 mg) or placebo as needed for 12 weeks. MAIN OUTCOME MEASURES: Primary: FePEDS Q3 ('Over the past four weeks, when you had sexual intercourse, how often was it satisfactory for you?') scored as 0 (no sexual activity) and 1 (almost never or never) to 5 (almost always or always). Secondary, partners: Sexual Function Questionnaire, Female Sexual Function Index (FSFI), and ED Inventory of Treatment Satisfaction (EDITS) partner version (EDITS-Partner). Secondary, men: International Index of Erectile Function (IIEF), General Efficacy Questions, event log data, Self-Esteem And Relationship questionnaire, and EDITS. Secondary, partners and men: Dyadic Adjustment Scale. RESULTS: The intention-to-treat population included 85 sildenafil recipients (mean age 59 +/- 12 years) and 91 placebo recipients (mean age 57 +/- 11 years). Most partners (aged 20-79 years; mean, 54 years) were postmenopausal. Sildenafil compared with placebo couples had greater improvement in the primary outcome (FePEDS Q3 [P < 0.0001]) and in sexual function, intercourse success rates, and secondary sexual satisfaction measures (FSFI satisfaction domain [P < 0.0001] and IIEF satisfaction domains [P < 0.001]) and had higher treatment satisfaction (EDITS and EDITS-Partner; P < 0.0001). Several predictors of improvement were identified, and improvement in one member of the couple correlated positively with improvement in the other member. CONCLUSIONS: The interdependence of sexual function and sexual satisfaction measures between members of couples consisting of men with ED and sexually healthy women reporting infrequent satisfactory sexual intercourse underscores the importance of including partners in ED treatment discussions.

Clinical laboratory testing for the diagnosis of systemic disease associated with anterior uveitis.

In many cases, anterior uveitis is not an isolated ocular entity but the harbinger of systemic disease. The diagnosis of an underlying disease is confirmed through laboratory testing and physical examination. Rather than ordering a broad spectrum of unnecessary tests, the clinician must correlate the patient's history with symptoms and physical signs to tailor laboratory testing to the individual. Subsequent co-management of the patient with an internist or appropriate specialist ensures that total patient care, ocular and systemic, will be achieved. This discussion addresses when and what laboratory testing to order in anterior uveitis cases with specific attention to why each auxilliary test is appropriate.

Laser therapy for open-angle glaucoma.

Management of open-angle glaucoma is complex and consists of medications, laser therapy, and conventional surgery. Laser therapy is a vital component of management, with the most common procedure being a trabeculoplasty, which reduces intraocular pressure by increasing aqueous outflow. This review discusses the indications, contraindications, and specific techniques for laser trabeculoplasty. The use of holmium sclerostomy is also discussed. With recent advances in laser and fiberoptic technology, sclerostomy is proving to be a simpler procedure than conventional filtering surgery, while providing comparable effectiveness.

Traumatic anterior uveitis.

Traumatic anterior uveitis is a common result of ocular contusion. Patients require careful evaluation to determine the degree of injury and the risk of further complications. This discussion updates the practitioner on the pathophysiology, diagnosis, and treatment of anterior uveitis secondary to a nonpenetrating ocular injury.

1987 survey of military optometrists: activities, roles and attitudes.

A worldwide survey of active duty military optometrists (Air Force, Army and Navy) was conducted in the spring of 1987. The survey response rate was 79%. Statistically significant differences (alpha = 0.01) in their roles, activities and attitudes are presented and discussed. Significant interservice differences were found in: types of duties, profession/status of immediate supervisors, work assignments, frequency and types of patient encounters, awareness of therapeutic drug policies, access to specific clinical instruments, and frequency of routine clinical tests. Attitude and opinion tabulations suggest an overall satisfaction with professional experiences and interprofessional working relationships. However, there was a common perception that optometrists did not receive appropriate promotional consideration or career opportunities within this multidisciplinary setting. Implications of survey results for non-military multidisciplinary health care institutions are also discussed.