Genomics-informed delineation of conservation units in a desert amphibian.

Delineating conservation units (CUs, e.g., evolutionarily significant units, ESUs, and management units, MUs) is critical to the recovery of declining species because CUs inform both listing status and management actions. Genomic data have strengths and limitations in informing CU delineation and related management questions in natural systems. We illustrate the value of using genomic data in combination with landscape, dispersal and occupancy data to inform CU delineation in Nevada populations of the Great Basin Distinct Population Segment of the Columbia spotted frog (Rana luteiventris). R. luteiventris occupies naturally fragmented aquatic habitats in this xeric region, but beaver removal, climate change and other factors have put many of these populations at high risk of extirpation without management intervention. We addressed three objectives: (i) assessing support for ESUs within Nevada; (ii) evaluating and revising, if warranted, the current delineation of MUs; and (iii) evaluating genetic diversity, effective population size, adaptive differentiation and functional connectivity to inform ongoing management actions. We found little support for ESUs within Nevada but did identify potential revisions to MUs based on unique landscape drivers of connectivity that distinguish these desert populations from those in the northern portion of the species range. Effective sizes were uniformly small, with low genetic diversity and weak signatures of adaptive differentiation. Our findings suggest that management actions, including translocations and genetic rescue, might be warranted. Our study illustrates how a carefully planned genetic study, designed to address priority management goals that include CU delineation, can provide multiple insights to inform conservation action.

Managing emerging mutagenicity risks: Late stage mutagenic impurity control within the atovaquone second generation synthesis.

The mutagenic-impurity control strategy for a second generation manufacturing route to the non-mutagenic antipneumocystic agent atovaquone (2-((1R,4R)-4-(4-chlorophenyl)cyclohexyl)-3-hydroxynaphthalene-1,4-dione) 1 is described. Preliminary assessment highlighted multiple materials of concern which were largely discharged either through returning a negative bacterial mutagenicity assay or through confidence that the impurity would be purged during the downstream processing from when it was first introduced. Additional genotoxicity testing highlighted two materials of concern where initial assessment suggested that testing for these impurities at trace levels within the drug substance would be required. Following a thorough review of process purging detail, spiking and purging experimentation, and an understanding of the process parameters to which they were exposed an ICH M7 Option 4 approach could be justified for their control. The development of two 1H NMR spectroscopy methods for measurement of these impurities is also described as well as a proposed summary table for describing the underlying rationale for ICH M7 control rationales to regulators. This manuscript demonstrates that process purging of potential mutagenic impurities can be realised even when they are introduced in the later stages of a process and highlights the importance of scientific understanding rather than relying on a stage-counting approach.

One science-driven approach for the regulatory implementation of alternative methods: A multi-sector perspective.

EU regulations call for the use of alternative methods to animal testing. During the last decade, an increasing number of alternative approaches have been formally adopted. In parallel, new 3Rs-relevant technologies and mechanistic approaches have increasingly contributed to hazard identification and risk assessment evolution. In this changing landscape, an EPAA meeting reviewed the challenges that different industry sectors face in the implementation of alternative methods following a science-driven approach. Although clear progress was acknowledged in animal testing reduction and refinement thanks to an integration of scientifically robust approaches, the following challenges were identified: i) further characterization of toxicity pathways; ii) development of assays covering current scientific gaps, iii) better characterization of links between in vitro readouts and outcome in the target species; iv) better definition of alternative method applicability domains, and v) appropriate implementation of the available approaches. For areas having regulatory adopted alternative methods (e.g., vaccine batch testing), harmonised acceptance across geographical regions was considered critical for broader application. Overall, the main constraints to the application of non-animal alternatives are the still existing gaps in scientific knowledge and technological limitations. The science-driven identification of most appropriate methods is key for furthering a multi-sectorial decrease in animal testing.

Establishing best practise in the application of expert review of mutagenicity under ICH M7.

The ICH M7 guidelines for the assessment and control of DNA reactive (mutagenic) impurities in pharmaceuticals allows for the consideration of in silico predictions in place of in vitro studies. This represents a significant advance in the acceptance of (Q)SAR models and has resulted from positive interactions between modellers, regulatory agencies and industry with a shared purpose of developing effective processes to minimise risk. This paper discusses key scientific principles that should be applied when evaluating in silico predictions with a focus on accuracy and scientific rigour that will support a consistent and practical route to regulatory submission.

Adult general psychiatric patients served in Nebraska's state hospitals: patient characteristics and needs.

This study identified the characteristics and needs of adults discharged from state psychiatric hospitals. A retrospective analysis of data on patients discharged from adult psychiatric units of three state psychiatric hospitals in Nebraska 2005-2008 was conducted. Diagnoses were classified into six groups, and Axis III data from the state psychiatric hospitals provided information about medical comorbidity. Only 12% of admitted patients had private insurance or could pay for their own treatment. Almost all discharged patients (95%) had a diagnosis of serious mental illness, and substance abuse (68%) and personality disorder (68%) were common, as were significant general health problems. Fourteen percent of patients used emergency services five or more times during the study period. Greater efforts must be made to diagnose, treat, and monitor major somatic illnesses and to better understand the factors that contribute to readmission and emergency service use in this population.

Permitted daily exposure limits for noteworthy N-nitrosamines.

A genotoxic carcinogen, N-nitrosodimethylamine (NDMA), was detected as a synthesis impurity in some valsartan drugs in 2018, and other N-nitrosamines, such as N-nitrosodiethylamine (NDEA), were later detected in other sartan products. N-nitrosamines are pro-mutagens that can react with DNA following metabolism to produce DNA adducts, such as O6 -alkyl-guanine. The adducts can result in DNA replication miscoding errors leading to GC>AT mutations and increased risk of genomic instability and carcinogenesis. Both NDMA and NDEA are known rodent carcinogens in male and female rats. The DNA repair enzyme, methylguanine DNA-methyltransferase can restore DNA integrity via the removal of alkyl groups from guanine in an error-free fashion and this can result in nonlinear dose responses and a point of departure or "practical threshold" for mutation at low doses of exposure. Following International recommendations (ICHM7; ICHQ3C and ICHQ3D), we calculated permissible daily exposures (PDE) for NDMA and NDEA using published rodent cancer bioassay and in vivo mutagenicity data to determine benchmark dose values and define points of departure and adjusted with appropriate uncertainty factors (UFs). PDEs for NDMA were 6.2 and 0.6 µg/person/day for cancer and mutation, respectively, and for NDEA, 2.2 and 0.04 µg/person/day. Both PDEs are higher than the acceptable daily intake values (96 ng for NDMA and 26.5 ng for NDEA) calculated by regulatory authorities using simple linear extrapolation from carcinogenicity data. These PDE calculations using a bench-mark approach provide a more robust assessment of exposure limits compared with simple linear extrapolations and can better inform risk to patients exposed to the contaminated sartans.

Investigation of a Largescale Common Murre ( Uria aalge) Mortality Event in California, USA, in 2015.

From August through December 2015, beachcast bird survey programs reported increased deposition of Common Murres ( Uria aalge) on central and northern California beaches, but not on southern California beaches. Coastal wildlife rehabilitation centers received more than 1,000 live, stranded, and debilitated murres from Sonoma County to San Luis Obispo County during August-October. Approximately two-thirds of admitted birds were after-hatch-year birds in emaciated body condition and in various stages of molt, with extremely worn plumage. Necropsies were done on a sample ( n=35) of birds to determine the probable cause of death of beachcast carcasses. Most birds examined during necropsy were emaciated, with starvation the most likely cause of death. Birds were also tested for underlying infectious diseases at the US Geological Survey National Wildlife Health Center and harmful algal bloom toxins at the University of California at Santa Cruz and the National Oceanographic and Atmospheric Administration's Northwest Fisheries Science Center. Twenty-four out of 29 tested birds had detectable levels of domoic acid, and no indication of infectious disease was found. Emaciation is thought to be the cause of death for these birds, with a large warm water anomaly and harmful algal bloom playing a secondary detrimental role.

Entanglements of marine mammals and seabirds in central California and the north-west coast of the United States 2001-2005.

Entanglement records for seabirds and marine mammals were investigated for the period 2001-2005. The entanglement records were extracted from databases maintained by seven organizations operating along the west coast of the United States of America. Their programmes included beach monitoring surveys, rescue and rehabilitation and regional pinniped censuses. Records of 454 entanglements were documented in live animals and in carcasses for 31 bird species and nine marine mammal species. The most frequently entangled species were Common Murres, Western Gulls and California sea lions. The entanglement materials identified were primarily fishing related. Entanglements were recorded every year suggesting that although the incidence level differs annually, entanglement is a persistent problem. It is recommended that each programme records details in standardized categories to determine entanglement material sources. Numbers of entanglements observed during these surveys are likely to be a conservative view of the actual entanglement rate taking place at sea.