Occurrence of male-specific and somatic coliphages and relationship with rainfall in privately-owned wells from peri­urban and rural households.

Privately-owned drinking water wells serving fewer than 25 people (private wells) are prevalent and understudied across most of the US. Private wells primarily serve rural households located outside of municipal drinking water and sewerage service coverage areas. These wells are not regulated by United States Environmental Protection Agency (EPA) under the Safe Drinking Water Act, are not regularly monitored by any public agency or utility, and generally do not undergo disinfection treatment. Coliphages are a group of viruses that infect coliform bacteria and are useful viral surrogates for fecal contamination in water systems in much the same way that fecal indicator bacteria (FIB), such as E. coli and to a lesser extent total coliforms, are used to quantify fecal contamination. Coliphages are approved by the EPA for regulatory monitoring in groundwater wells in the USA, but are not routinely used for this purpose. The present study characterizes the occurrence of male-specific and somatic coliphages, along with FIB, in private wells (n = 122) across two different counties in North Carolina. While occurrences of E. coli were rare and frequency of total coliform was generally low (~20%), male-specific and somatic coliphages were detectable in 66% and 54% of samples, respectively. Concentrations of male-specific coliphages were higher than somatics at each county and on a monthly basis. Rainfall appears to be partly influencing higher coliphage concentrations in December, January and February. This research underscores the need for increased surveillance in private wells and consideration of using coliphages in order to better characterize occurrence of fecal contamination at the time of sampling, especially during rainier months.

Occurrence of Lead and Other Toxic Metals Derived from Drinking-Water Systems in Three West African Countries.

BACKGROUND: Exposure to toxic metals (TMs) such as lead can cause lifelong neurodevelopmental impairment and other adverse outcomes. TMs enter drinking water from human activity, geogenic contamination, and corrosion of water system components. Several studies report TM contamination in piped systems and private wells in high-income countries (HICs). However, few robust studies report on TM contamination in low- and middle-income countries (LMICs). OBJECTIVES: We characterized the occurrence and investigated sources of TM contamination in 261 rural water systems in three West African LMICs to inform prevention and management. METHODS: Water samples were collected from 261 community water systems (handpumps and public taps) across rural Ghana, Mali, and Niger. Scrapings were collected from accessible components of a subset of these systems using a drill with acid-washed diamond-tipped bits. Samples were analyzed by inductively coupled plasma (ICP) mass spectrometry or ICP optical emission spectroscopy. RESULTS: Of the TMs studied, lead most frequently occurred at levels of concern in sampled water system components and water samples. Lead mass fractions exceeded International Plumbing Code (IPC) recommended limits (0.25% wt/wt) for components in 82% (107/130) of systems tested; brass components proved most problematic, with 72% (26/36) exceeding IPC limits. Presence of a brass component in a water system increased expected lead concentrations in drinking-water samples by 3.8 times. Overall, lead exceeded World Health Organization (WHO) guideline values in 9% (24/261) of drinking-water samples across countries; these results are broadly comparable to results observed in many HICs. Results did not vary significantly by geography or system type. DISCUSSION: Ensuring use of lead-free (<0.25%) components in new water systems and progressively remediating existing systems could reduce drinking-water lead exposures and improve health outcomes for millions. However, reflexive decommissioning of existing systems may deprive users of sufficient water for health or drive them to riskier sources. Because supply chains for many water system components are global, TM monitoring, prevention, and management may be warranted in other LMICs beyond the study area as well. https://doi.org/10.1289/EHP7804.

WaSH CQI: Applying continuous quality improvement methods to water service delivery in four districts of rural northern Ghana.

Continuous, safely managed water is critical to health and development, but rural service delivery faces complex challenges in low- and middle-income countries (LMICs). We report the first application of continuous quality improvement (CQI) methods to improve the microbial quality of household water for consumption (HWC) and the functionality of water sources in four rural districts of northern Ghana. We further report on the impacts of interventions developed through these methods. A local CQI team was formed and trained in CQI methods. Baseline data were collected and analyzed to identify determinants of service delivery problems and microbial safety. The CQI team randomized communities, developed an improvement package, iteratively piloted it in intervention communities, and used uptake survey data to refine the package. The final improvement package comprised safe water storage containers, refresher training for community WaSH committees and replacement of missing maintenance tools. This package significantly reduced contamination of HWC (p<0.01), and significant reduction in contamination persisted two years after implementation. Repair times in both intervention and control arms decreased relative to baseline (p<0.05), but differences between intervention and control arms were not significant at endline. Further work is needed to build on the gains in household water quality observed in this work, sustain and scale these improvements, and explore applications of CQI to other aspects of water supply and sanitation.

Indinavir Increases Midazolam N-Glucuronidation in Humans: Identification of an Alternate CYP3A Inhibitor Using an In Vitro to In Vivo Approach.

Midazolam is a widely used index substrate for assessing effects of xenobiotics on CYP3A activity. A previous study involving human hepatocytes showed the primary route of midazolam metabolism, 1'-hydroxylation, shifted to N-glucuronidation in the presence of the CYP3A inhibitor ketoconazole, which may lead to an overprediction of the magnitude of a xenobiotic-midazolam interaction. Because ketoconazole is no longer recommended as a clinical CYP3A inhibitor, indinavir was selected as an alternate CYP3A inhibitor to evaluate the contribution of the N-glucuronidation pathway to midazolam metabolism. The effects of indinavir on midazolam 1'-hydroxylation and N-glucuronidation were first characterized in human-derived in vitro systems. Compared with vehicle, indinavir (10 µM) inhibited midazolam 1'-hydroxylation by recombinant CYP3A4, human liver microsomes, and high-CYP3A activity cryopreserved human hepatocytes by ≥70%; the IC50 obtained with hepatocytes (2.7 µM) was within reported human unbound indinavir Cmax (≤5 µM). Midazolam N-glucuronidation in hepatocytes increased in the presence of indinavir in both a concentration-dependent (1-33 µM) and time-dependent (0-4 hours) manner (by up to 2.5-fold), prompting assessment in human volunteers (n = 8). As predicted by these in vitro data, indinavir was a strong inhibitor of the 1'-hydroxylation pathway, decreasing the 1'-hydroxymidazolam/midazolam area under the plasma concentration versus time curve (AUC)0-12h ratio by 80%. Although not statistically significant, the midazolam N-glucuronide/midazolam AUC0-12h ratio increased by 40%, suggesting a shift to the N-glucuronidation pathway. The amount of midazolam N-glucuronide recovered in urine increased 4-fold but remained <10% of the oral midazolam dose (2.5 mg). A powered clinical study would clarify whether N-glucuronidation should be considered when assessing the magnitude of a xenobiotic-midazolam interaction.

A bioassay-based protocol for chemical neutralization of human faecal wastes treated by physico-chemical disinfection processes: A case study on benzalkonium chloride.

In situ physico-chemical disinfection of high risk faecal waste is both effective and widely used as a sanitation management strategy for infection prevention and control. Systematic tests where the performance of alternative physico-chemical disinfection methods is systematically compared and optimized must be based on reliable protocols. These protocol are currently not adequately addressing the neutralization related issues: the neutralization of the tested disinfectant after specified conditions of concentration and contact time (CT) is necessary to prevent continued disinfection after the intended contact time; moreover such neutralization is often necessary in practice and on a large scale to prevent adverse health and ecological impacts from remaining disinfectant after the target CT is achieved. Few studies adequately assess the extent of neutralization of the chemical disinfectant and are intended to optimize on-site disinfection practices for waste matrices posing high microbial risks. Hence, there is a need for effective and reproducible neutralization protocols in chemical disinfection trials and practice. Furthermore, for most of chemical disinfectants used in healthcare settings there is no practical methodology to reliably and conveniently measure the residual disinfectant concentration after its neutralization and also determine the optimum concentration of the neutralizer. Because some neutralizing compounds can themselves be toxic to the test microorganisms, it is necessary to optimize neutralization procedures in disinfection experiments for the development of infection control practices using accepted positive control microbes. In the presented work, a stepwise bioassay-based protocol using representative faecal indicator microbes is described for optimizing chemical disinfection and subsequent disinfectant neutralization of any infectious faecal waste matrix. The example described is for the quaternary ammonium compound benzalkonium chloride and its recommended chemical neutralizer in a high strength human faecal waste matrix.

Carrying water may be a major contributor to disability from musculoskeletal disorders in low income countries: a cross-sectional survey in South Africa, Ghana and Vietnam.

BACKGROUND: The Sustainable Development Goals include commitments to end poverty, and promote education for all, gender equality, the availability of water and decent work for all. An important constraint is the fact that each day, many millions of women and children, and much less frequently men, carry their household's water home from off-plot sources. The burden of fetching water exacerbates gender inequality by keeping women out of education and paid employment. Despite speculation about the potential health impacts of fetching water, there is very little empirical evidence. We report the first large study of the health impacts of carrying water on women and children. METHODS: A cross-sectional survey was conducted in South Africa, Ghana and Vietnam during 2012. It investigated water carrying methods and health status. Because areas of self-reported pain were correlated we undertook factor analysis of sites of reported pain, to interpret patterns of pain reporting. Regression analysis using Generalised Estimating Equations (GEE) investigated water carrying as a risk factor for general health and self-reported pain. RESULTS: People who previously carried water had increased relative risk of reporting pain in the hands (risk ratio RR 3.62, 95% confidence interval CI 1.34 to 9.75) and upper back (RR 2.27, 95% CI 1.17 to 4.40), as did people who currently carry water (RR hand pain 3.11, 95% CI 1.34 to 7.23; RR upper back pain 2.16, 95% CI 1.25 to 3.73). The factor analysis results indicate that factor 1, 'axial compression', which is correlated with pain in the head and upper back, chest/ribs, hands, feet and abdomen/stomach, is associated with currently (0.30, 95% CI 0.17 to 0.43) or previously (0.21, 95% CI 0.01 to 0.42) carrying water. Factor 2, 'soft tissue strain', which is correlated with pain in the neck, shoulders/arms, lower back and hips/pelvis or legs, is marginally negatively associated with currently (-0.18, 95% CI -0.32 to -0.04) carrying water. The factor 'axial compression' was more strongly associated with carrying water containers on the head. CONCLUSIONS: Participants who reported a history of current or past water carrying more frequently reported pain in locations most likely to be associated with sustained spinal axial compression in the cervical region. Given the fact that cervical spinal conditions are globally one of the more common causes of disability, our findings suggest that water carrying, especially by head loading is a major contributing factor in musculoskeletal disease burden in low income countries. Our findings support the proposed indicator for monitoring SDG6.1: "Percentage of population using safely managed drinking water services at home."

Rapid Detection of Escherichia coli in Water Using Sample Concentration and Optimized Enzymatic Hydrolysis of Chromogenic Substrates.

Methods for rapid detection of fecal indicator bacteria in water are important to ensure that water is safe for drinking, bathing, recreation, fishing and shellfish harvesting. In this study, we tested experimental conditions for bacterial hydrolysis of two promising enzymatic substrates, 5-Bromo-4-chloro-3-indolyl ß-D-glucuronide (X-Gluc) and Resorufin ß-D-glucuronide (REG), and optimized parameters such as temperature and pH to determine conditions for rapid reactions. We then innovated a membrane filter-based approach to facilitate more rapid enzyme-based detection of Escherichia coli in water based on the combination of an initial concentration step and optimized test conditions. For this approach, a water sample (10‒100 mL) is filtered through a 0.45-µm pore size filter with a diameter of 4 or 13 mm. After filtration, a newly designed rapid detection broth is added containing the enzymatic inducer Methyl-beta-D-Glucuronide sodium (MetGlu) and the substrate REG or X-Gluc. After a few (1‒7) hours of incubation at 35 °C, the filter shows pink color (for REG-containing broth) or green color (for X-Gluc containing broth) if E. coli is present. The study provides insights and approaches towards developing a simple, fast, and low-cost method to detect fecal indicator bacteria in water.

Evaluating Mobile Survey Tools (MSTs) for Field-Level Monitoring and Data Collection: Development of a Novel Evaluation Framework, and Application to MSTs for Rural Water and Sanitation Monitoring.

Information and communications technologies (ICTs) such as mobile survey tools (MSTs) can facilitate field-level data collection to drive improvements in national and international development programs. MSTs allow users to gather and transmit field data in real time, standardize data storage and management, automate routine analyses, and visualize data. Dozens of diverse MST options are available, and users may struggle to select suitable options. We developed a systematic MST Evaluation Framework (EF), based on International Organization for Standardization/International Electrotechnical Commission (ISO/IEC) software quality modeling standards, to objectively assess MSTs and assist program implementers in identifying suitable MST options. The EF is applicable to MSTs for a broad variety of applications. We also conducted an MST user survey to elucidate needs and priorities of current MST users. Finally, the EF was used to assess seven MSTs currently used for water and sanitation monitoring, as a validation exercise. The results suggest that the EF is a promising method for evaluating MSTs.

A Systematic Review and Meta-Analysis of Fecal Contamination and Inadequate Treatment of Packaged Water.

BACKGROUND: Packaged water products provide an increasingly important source of water for consumption. However, recent studies raise concerns over their safety. OBJECTIVES: To assess the microbial safety of packaged water, examine differences between regions, country incomes, packaged water types, and compare packaged water with other water sources. METHODS: We performed a systematic review and meta-analysis. Articles published in English, French, Portuguese, Spanish and Turkish, with no date restrictions were identified from online databases and two previous reviews. Studies published before April 2014 that assessed packaged water for the presence of Escherichia coli, thermotolerant or total coliforms were included provided they tested at least ten samples or brands. RESULTS: A total of 170 studies were included in the review. The majority of studies did not detect fecal indicator bacteria in packaged water (78/141). Compared to packaged water from upper-middle and high-income countries, packaged water from low and lower-middle-income countries was 4.6 (95% CI: 2.6-8.1) and 13.6 (95% CI: 6.9-26.7) times more likely to contain fecal indicator bacteria and total coliforms, respectively. Compared to all other packaged water types, water from small bottles was less likely to be contaminated with fecal indicator bacteria (OR = 0.32, 95%CI: 0.17-0.58) and total coliforms (OR = 0.10, 95%CI: 0.05, 0.22). Packaged water was less likely to contain fecal indicator bacteria (OR = 0.35, 95%CI: 0.20, 0.62) compared to other water sources used for consumption. CONCLUSIONS: Policymakers and regulators should recognize the potential benefits of packaged water in providing safer water for consumption at and away from home, especially for those who are otherwise unlikely to gain access to a reliable, safe water supply in the near future. To improve the quality of packaged water products they should be integrated into regulatory and monitoring frameworks.

Microbiological and Chemical Quality of Packaged Sachet Water and Household Stored Drinking Water in Freetown, Sierra Leone.

Packaged drinking water (PW) sold in bottles and plastic bags/sachets is widely consumed in low- and middle-income countries (LMICs), and many urban users in sub-Saharan Africa (SSA) rely on packaged sachet water (PSW) as their primary source of water for consumption. However, few rigorous studies have investigated PSW quality in SSA, and none have compared PSW to stored household water for consumption (HWC). A clearer understanding of PSW quality in the context of alternative sources is needed to inform policy and regulation. As elsewhere in SSA, PSW is widely consumed in Sierra Leone, but government oversight is nearly nonexistent. This study examined the microbiological and chemical quality of a representative sample of PSW products in Freetown, Sierra Leone at packaged water manufacturing facilities (PWMFs) and at points of sale (POSs). Samples of HWC were also analyzed for comparison. The study did not find evidence of serious chemical contamination among the parameters studied. However, 19% of 45 PSW products sampled at the PWMF contained detectable Escherichia coli (EC), although only two samples exceeded 10 CFU/100 mL. Concentrations of total coliforms (TC) in PSW (but not EC) increased along the supply chain. Samples of HWC from 60 households in Freetown were significantly more likely to contain EC and TC than PSW at the point of production (p<0.01), and had significantly higher concentrations of both bacterial indicators (p<0.01). These results highlight the need for additional PSW regulation and surveillance, while demonstrating the need to prioritize the safety of HWC. At present, PSW may be the least unsafe option for many households.

Understanding handpump sustainability: Determinants of rural water source functionality in the Greater Afram Plains region of Ghana.

Safe drinking water is critical to human health and development. In rural sub-Saharan Africa, most improved water sources are boreholes with handpumps; studies suggest that up to one third of these handpumps are nonfunctional at any given time. This work presents findings from a secondary analysis of cross-sectional data from 1509 water sources in 570 communities in the rural Greater Afram Plains (GAP) region of Ghana; one of the largest studies of its kind. 79.4% of enumerated water sources were functional when visited; in multivariable regressions, functionality depended on source age, management, tariff collection, the number of other sources in the community, and the district. A Bayesian network (BN) model developed using the same data set found strong dependencies of functionality on implementer, pump type, management, and the availability of tools, with synergistic effects from management determinants on functionality, increasing the likelihood of a source being functional from a baseline of 72% to more than 97% with optimal management and available tools. We suggest that functionality may be a dynamic equilibrium between regular breakdowns and repairs, with management a key determinant of repair rate. Management variables may interact synergistically in ways better captured by BN analysis than by logistic regressions. These qualitative findings may prove generalizable beyond the study area, and may offer new approaches to understanding and increasing handpump functionality and safe water access.

Piperazinyl-oxadiazoles as selective sphingosine-1-phosphate receptor agonists.

The discovery of a new series of selective S1P1 agonists is described. This series of piperazinyl-oxadiazole derivatives was rapidly optimized starting from high-throughput screening hit 1 to afford potent and selective lead compound 10d. Further SAR studies showed that 10d was converted to the active phosphate metabolite 29 in vivo. Oral administration of compound 10d to rats was shown to induce lymphopenia at 3 mg/kg.

Global monitoring of water supply and sanitation: history, methods and future challenges.

International monitoring of drinking water and sanitation shapes awareness of countries' needs and informs policy, implementation and research efforts to extend and improve services. The Millennium Development Goals established global targets for drinking water and sanitation access; progress towards these targets, facilitated by international monitoring, has contributed to reducing the global disease burden and increasing quality of life. The experiences of the MDG period generated important lessons about the strengths and limitations of current approaches to defining and monitoring access to drinking water and sanitation. The methods by which the Joint Monitoring Programme (JMP) of WHO and UNICEF tracks access and progress are based on analysis of data from household surveys and linear regression modelling of these results over time. These methods provide nationally representative and internationally comparable insights into the drinking water and sanitation facilities used by populations worldwide, but also have substantial limitations: current methods do not address water quality, equity of access, or extra-household services. Improved statistical methods are needed to better model temporal trends. This article describes and critically reviews JMP methods in detail for the first time. It also explores the impact of, and future directions for, international monitoring of drinking water and sanitation.

Inactivation of Escherichia coli by polychromatic simulated sunlight: evidence for and implications of a fenton mechanism involving iron, hydrogen peroxide, and superoxide.

Sunlight inactivation of Escherichia coli has previously been shown to accelerate in the presence of oxygen, exogenously added hydrogen peroxide, and bioavailable forms of exogenously added iron. In this study, mutants unable to effectively scavenge hydrogen peroxide or superoxide were found to be more sensitive to polychromatic simulated sunlight (without UVB wavelengths) than wild-type cells, while wild-type cells grown under low-iron conditions were less sensitive than cells grown in the presence of abundant iron. Furthermore, prior exposure to simulated sunlight was found to sensitize cells to subsequent hydrogen peroxide exposure in the dark, but this effect was attenuated for cells grown with low iron. Mutants deficient in recombination DNA repair were sensitized to simulated sunlight (without UVB wavelengths), but growth in the presence of iron chelators reduced the degree of sensitization conferred by this mutation. These findings support the hypothesis that hydrogen peroxide, superoxide, and intracellular iron all participate in the photoinactivation of E. coli and further suggest that the inactivation rate of enteric bacteria in the environment may be strongly dependent on iron availability and growth conditions.

Time-dependent inhibition and estimation of CYP3A clinical pharmacokinetic drug-drug interactions using plated human cell systems.

The current studies assessed the utility of freshly plated hepatocytes, cryopreserved plated hepatocytes, and cryopreserved plated HepaRG cells for the estimation of inactivation parameters k(inact) and K(I) for CYP3A. This was achieved using a subset of CYP3A time-dependent inhibitors (fluoxetine, verapamil, clarithromycin, troleandomycin, and mibefradil) representing a range of potencies. The estimated k(inact) and K(I) values for each time-dependent inhibitor were compared with those obtained using human liver microsomes and used to estimate the magnitude of clinical pharmacokinetic drug-drug interaction (DDI). The inactivation kinetic parameter, k(inact), was most consistent across systems tested for clarithromycin, verapamil, and troleandomycin, with a high k(inact) of 0.91 min(-1) observed for mibefradil in HepaRG cells. The apparent K(I) estimates derived from the various systems displayed a range of variability from 3-fold for clarithromycin (5.4-17.7 µM) to 6-fold for verapamil (1.9-12.6 µM). In general, the inactivation kinetic parameters derived from the cell systems tested fairly replicated what was observed in time-dependent inhibition studies using human liver microsomes. Despite some of the observed differences in inactivation kinetic parameters, the estimated DDIs derived from each of the tested systems generally agreed with the clinically reported DDI within approximately 2-fold. In addition, a plated cell approach offered the ability to conduct longer primary incubations (greater than 30 min), which afforded improved ability to identify the weak time-dependent inhibitor fluoxetine. Overall, results from these studies suggest that in vitro inactivation parameters generated from plated cell systems may be a practical approach for identifying time-dependent inhibitors and for estimating the magnitude of clinical DDIs.

Solar water disinfection (SODIS) of Escherichia coli, Enterococcus spp., and MS2 coliphage: effects of additives and alternative container materials.

The use of alternative container materials and added oxidants accelerated the inactivation of MS2 coliphage and Escherichia coli and Enterococcus spp. bacteria during solar water disinfection (SODIS) trials. Specifically, bottles made from polypropylene copolymer (PPCO), a partially UVB-transparent plastic, resulted in three-log inactivation of these organisms in approximately half the time required for disinfection in bottles made from PET, polycarbonate, or Tritan(®), which absorb most UVB light. Furthermore, the addition of 125 mg/L sodium percarbonate in combination with either citric acid or copper plus ascorbate tended to accelerate inactivation by factors of 1.4-19. Finally, it was observed that the inactivation of E. coli and enterococci derived from local wastewater was far slower than the inactivation of laboratory-cultured E. coli and Enterococcus spp., while the inactivation of MS2 was slowest of all. These results highlight the importance of UVB in SODIS under certain conditions, and also the greater sunlight resistance of some viruses and of bacteria of fecal origin, as compared to the laboratory-cultured bacteria commonly used to model their inactivation. Furthermore, this study illustrates promising new avenues for accelerating the inactivation of bacteria and viruses by solar disinfection.

Characterization of aldehyde oxidase enzyme activity in cryopreserved human hepatocytes.

Substrates of aldehyde oxidase (AO), for which human clinical pharmacokinetics are reported, were selected and evaluated in pooled mixed-gender cryopreserved human hepatocytes in an effort to quantitatively characterize AO activity. Estimated hepatic clearance (Cl(h)) for BIBX1382, carbazeran, O6-benzylguanine, zaleplon, and XK-469 using cryopreserved hepatocytes was 18, 17, 12, <4.3, and <4.3 ml · min⁻¹ · kg⁻¹, respectively. The observed metabolic clearance in cryopreserved hepatocytes was confirmed to be a result of AO-mediated metabolism via two approaches. Metabolite identification after incubations in the presence of H2¹8O confirmed that the predominant oxidative metabolite was generated by AO, as expected isotope patterns in mass spectra were observed after analysis by high-resolution mass spectrometry. Second, clearance values were efficiently attenuated upon coincubation with hydralazine, an inhibitor of AO. The low exposure after oral doses of BIBX1382 and carbazeran (∼5% F) would have been fairly well predicted using simple hepatic extraction (f(h)) values derived from cryopreserved hepatocytes. In addition, the estimated hepatic clearance value for O6-benzylguanine was within ∼80% of the observed total clearance in humans after intravenous administration (15 ml · min⁻¹ · kg⁻¹), indicating a reasonable level of quantitative activity from this in vitro system. However, a 3.5-fold underprediction of total clearance was observed for zaleplon, despite the 5-oxo metabolite being clearly observed. These data taken together suggest that the use of cryopreserved hepatocytes may be a practical approach for assessing AO-mediated metabolism in discovery and potentially useful for predicting hepatic clearance of AO substrates.

Simulated sunlight action spectra for inactivation of MS2 and PRD1 bacteriophages in clear water.

Action spectra for simulated sunlight were measured in clear water for two viruses: PRD1, a double-stranded DNA bacteriophage, and MS2, a single-stranded RNA bacteriophage. Viruses were diluted into phosphate buffered saline (20 mM PBS, pH 7.5) and exposed for 22 h to simulated sunlight either directly or through one of six glass filters with 50% cutoff wavelengths ranging from 280 to 350 nm. Virus survival was measured using the double agar layer plaque method. Both UVA (320-400 nm) and UVB (280-320 nm) light were found to contribute to PRD1 inactivation, while only UVB inactivated MS2. A computational model was developed for interpreting these action spectra with 3-nm resolution. Using these methods, we provide detailed estimates of the sensitivity of MS2 and PRD1 to photoinactivation from 285 to 345 nm. The resulting sensitivity coefficients can be combined with solar spectra to estimate inactivation rates in clear water under different sunlight conditions. This approach will be useful for modeling the inactivation of viruses and other microorganisms in sunlit natural and engineered systems.

Discovery of CP-690,550: a potent and selective Janus kinase (JAK) inhibitor for the treatment of autoimmune diseases and organ transplant rejection.

There is a critical need for safer and more convenient treatments for organ transplant rejection and autoimmune disorders such as rheumatoid arthritis. Janus tyrosine kinases (JAK1, JAK3) are expressed in lymphoid cells and are involved in the signaling of multiple cytokines important for various T cell functions. Blockade of the JAK1/JAK3-STAT pathway with a small molecule was anticipated to provide therapeutic immunosuppression/immunomodulation. The Pfizer compound library was screened against the catalytic domain of JAK3 resulting in the identification of a pyrrolopyrimidine-based series of inhibitors represented by CP-352,664 (2a). Synthetic analogues of 2a were screened against the JAK enzymes and evaluated in an IL-2 induced T cell blast proliferation assay. Select compounds were evaluated in rodent efficacy models of allograft rejection and destructive inflammatory arthritis. Optimization within this chemical series led to identification of CP-690,550 1, a potential first-in-class JAK inhibitor for treatment of autoimmune diseases and organ transplant rejection.

Recent advances in high throughput screening for ADME properties.

With the increase in the numbers of molecules synthesized in a typical drug discovery program, as well as the large amount of information utilized in the selection of a drug candidate, there is a need for a plethora of drug metabolism and pharmacokinetic (DMPK) information to be regularly generated in discovery. Over the past decade, many in vitro, and even in vivo, DMPK screens have been developed and routinely deployed to generate this information in support of drug discovery efforts. In the past few years, newer methods, or adaptations to methods, have been published, and this review attempts to summarize these advances. In particular, advances have been reported for experimental approaches to metabolic clearance, CYP inhibition, in vivo exposure, and distribution, as well as in silico determinations of absorption, distribution, metabolism, and excretion (ADME) properties. Bioanalytical approaches aimed at optimizing analyte method development, sample preparation, and analyte detection, have also been reported. Future advances will further improve the ability to make decisions on molecules earlier in drug discovery.