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Cognitive biases reveal underlying affective state by indicating optimism and pessimism. This methodology may permit assessment of positive welfare in animals that have few validated positive welfare indicators, such as bears. Our goal was to validate a judgment bias test for assessing optimism in brown bears, using a touchscreen. After training the bears on a conditional discrimination, we compared responses to an ambiguous stimulus in a 2 × 2 nested design involving four experimental conditions representing presence or absence of a behind-the-scenes tour and presence or absence of a keeper training session with food reinforcement. We recorded bears' behavior during the conditions as a measure of convergent validity. Testing revealed the possibility of pessimism in the absence of reinforcement in one bear. More frustration behaviors were also observed during the no food reinforcement conditions. This is the first experimental demonstration of brown bears using a touchscreen and one of only three reports in which bears have been reported to perform a conditional discrimination. This method of assessing underlying affective state shows promise for the future.

Choice-based design allows animals in human care opportunities to move at will between multiple interconnected spaces. Some evidence suggests providing environmental choice confers benefits to animals, but there is a dearth of research in this area with large carnivores. To understand the effects of this design strategy on large felids, behavioral and space use data were collected on three Amur tigers housed in a new habitat at Cleveland Metroparks Zoo. Data were collected in two conditions: 1) restricted to a single habitat, and 2) access to two habitats. With choice, tigers were less frequently inactive ( p = 0.003), and locomoted more frequently ( p = 0.009). They also showed different preferences in space use with choice, and a strong preference for overhead runways between habitats ( E*  = 0.83, E*  = 0.78). These results add to what is known about environmental impacts on zoo animal behavior and suggest this design and management strategy may be effective in conferring positive welfare benefits to tigers and other large carnivores.

Human African Trypanosomiasis (HAT) is a vector-borne disease caused by kinetoplastid parasites of the Trypanosoma genus. The disease proceeds in two stages, with a hemolymphatic blood stage and a meningo-encephalic brain stage. In the latter stage, the parasite causes irreversible damage to the brain leading to sleep cycle disruption and is fatal if untreated. An orally bioavailable treatment is highly desirable. In this study, we present a brain-penetrant, parasite-selective 20S proteasome inhibitor that was rapidly optimized from an HTS singleton hit to drug candidate compound 7 that showed cure in a stage II mouse efficacy model. Here, we describe hit expansion and lead optimization campaign guided by cryo-electron microscopy and an in silico model to predict the brain-to-plasma partition coefficient K p as an important parameter to prioritize compounds for synthesis. The model combined with in vitro and in vivo experiments allowed us to advance compounds with favorable unbound brain-to-plasma ratios ( K p,uu ) to cure a CNS disease such as HAT.

Matschie's tree kangaroo (Dendrolagus matschiei) is an endangered arboreal marsupial native to Papua New Guinea. Detailed field studies of its behavior and ecology are scarce due largely to its occupation of remote cloud forests and cryptic nature. Although this species has been in human care since the 1950s, much of its biology is still unknown. The current ex situ population is not sustainable due to health and reproductive problems, believed to stem largely from issues with diet and obesity. To better assess potential discrepancies between energy requirements and energy intake, we sought to 1) quantify total energy expenditure (TEE) of two zoo-housed Matschie's tree kangaroos (body mass = 9.0-9.7 kg) on a diet composed largely of leafy browse; 2) quantify food and macronutrient intake, apparent dry matter macronutrient digestibility, and metabolizable energy (ME) intake over a 14-month period; and 3) test for seasonal changes in ME intake due to seasonal differences in the varieties of leafy browse offered. Using the doubly labeled water method, we determined TEE for the female (288 kcal day -1) and male (411 kcal day -1). Resulting mean TEE was well below the expected value for marsupials and macropods (i.e., ~60% of the expected value based on body mass). The mean calculated ME intakes for the female and male were 307 kcal day-1 and 454 kcal day-1, respectively. There were significant seasonal differences in ME intakes, driven by reduced intake in the autumn. These results demonstrate that Matschie's tree kangaroos can be maintained at healthy body weights and conditions on fiber-rich and browse-heavy diets. Our findings contribute important insights into tree kangaroo energetics and physiology and can be applied to help reformulate the diet of Matschie's tree kangaroos at captive facilities to improve population health and sustainability., Competing Interests: The authors have declared that no competing interests exist.

Cheetahs have been the subject of reproductive study for over 35 years, yet steroid hormone activity remains poorly described after ovulation. Our objective was to examine and compare fecal progestagen (fPM), estrogen (fEM), and glucocorticoid (fGM) metabolite concentrations post-ovulation in pregnant and non-pregnant animals to better understand female physiology (1) during successful pregnancy, (2) surrounding frequent non-pregnant luteal phases, and (3) after artificial insemination (AI) to improve the low success rate. Secondarily, the authors also validated a urinary progestagen metabolite assay, allowing pregnancy detection with minimal sample collection. Fecal samples were collected from 12 females for ≥2 weeks prior to breeding/hormone injection (the PRE period) through 92 days post-breeding/injection. Samples were assessed for hormone concentrations using established enzyme immunoassays. Urine samples were collected for 13 weeks from 6 females after natural breeding or AI. There were no differences among groups in fGM, but in pregnant females, concentrations were higher (p < 0.01) in the last trimester than any other time. For pregnant females that gave birth to singletons, fGM was higher (p = 0.0205), but fEM tended to be lower (p = 0.0626) than those with multi-cub litters. Our results provide insight into the physiological events surrounding natural and artificially stimulated luteal activity in the cheetah.

With fewer than 7500 cheetahs remaining in the wild, ex situ cheetah populations serve as an insurance policy against extinction and a resource to study species' biology. This study aimed to identify the age of pubertal onset in ex situ female cheetahs using non-invasive faecal steroid hormone monitoring and body weights. Faecal samples from nine female cheetahs were collected two to three times weekly from 2 to 36months of age and body weights were recorded every 3months. Faecal oestrogen metabolites (FOM) and faecal glucocorticoid metabolites (FGM) were analysed using enzyme immunoassays and samples were categorised into 6-month intervals to compare endocrine characteristics. Faecal hormone and body weight data were analysed using generalised linear mixed models. Age was a significant predictor of mean and baseline FOM concentrations, number of FOM peaks, mean and maximum FOM peak concentrations and the number of cycles. Female cheetahs aged 24-30months exhibited a marked rise in mean FOM concentration and the number of FOM peaks and cycles increased with age until 24-30months. Females attained adult body weight by 21months of age. Mean and baseline FGM concentrations were highest at the 0-6 and 12-18months of age groups and did not follow the same FOM patterns. Based on body weight data, the FOM concentrations and peak patterning, females were considered pubertal from 24 to 30months of age. Characterisation of cheetah puberty has direct and significant implications for the improvement of management and reproductive success of cheetahs under human care. This information is particularly informative for identifying important windows of development, littermate dispersal and breeding introductions.

The North American cheetah population serves as a reservoir for the species, and acts as a research population to help understand the unique biology of the species. Little is known about the intrauterine physiology of the cheetah, including embryo differentiation, implantation, and the development of the placenta. After mating, cheetah females frequently experience (30-65% of matings) a non-pregnant luteal phase where progestogen metabolite levels match those found in pregnant females for the first ~55 days of gestation, but parturition does not occur. Immunoglobulin J chain (IgJ) is a molecule that is involved in the activation of the secretory immune response and has been found to be indicative of pregnancy in the cheetah using fecal monitoring. In this study, western blotting was employed to track IgJ abundance in pooled weekly fecal samples following natural breeding or exogenous stimulation to ovulate, and IgJ levels were compared between individuals undergoing a pregnant (n = 12) and non-pregnant (n = 19) luteal phase. It was revealed that IgJ abundance was increased in pregnant females compared to non-pregnant females at week 4 and week 8 post-breeding, indicating the potential modulation of maternal immunity in response to sensitive events such as implantation and the increased secretory activity of the placenta. IgJ levels also tended to be higher early after breeding in females that were bred naturally with intact males compared to exogenously stimulated females with no exposure to seminal plasma, potentially indicating a response to the act of intromission or the stress of breeding, or possibly demonstrating an immune response resulting in the promotion of maternal tolerance to seminal antigens present upon embryonic implantation. Monitoring fecal IgJ may be a potential method to determine gestational status in the cheetah and will aid future conservation efforts of the species., Competing Interests: The authors have declared that no competing interests exist.

Cheetahs are one of the most heavily studied felid species, with numerous publications on health, disease, and reproductive physiology produced over the last 30 years. Despite this relatively long history of research, there is a paucity of crucial biological data, such as pubertal onset, which has direct and significant applications to improved management of ex situ cheetah populations. This study aimed to determine age of pubertal onset in ex situ male cheetahs using non-invasive fecal steroid hormone monitoring and body weights. Fecal samples from 12 male cheetahs from four institutions were collected 2-3 times weekly from 1 to 42 months of age. Fecal androgen and glucocorticoid metabolites were analyzed using enzyme immunoassays previously validated for use with cheetah feces. Animal body weights were recorded monthly. Fecal hormone and body weight data were analyzed using generalized linear mixed models. Androgen concentrations exhibited an increase to levels similar to those observed in adult males by 18-24 months of age, and males attained adult body weights by 21 months of age. Based on these weight data and the initial increase in androgens toward adult concentrations, males were considered pubertal from 18 to 24 months of age. Glucocorticoid concentrations and amplitude of concentration over baseline were also increased during this period. Knowledge about the physiological changes associated with puberty is useful for management and improving reproductive success of cheetah populations under human care, particularly for determining timing of litter separation from dam, littermate dispersal and when to introduce potential breeding pairs., (Published by Elsevier Inc.)

Approximately 80% of cheetahs living in typical zoological collections never reproduce. In more than 60% of breedings, the female is confirmed to ovulate, but parturition fails to occur. It is unknown if these non-pregnant intervals of elevated progesterone (deemed luteal phases) are conception failures or a pregnancy terminating in embryonic/fetal loss. There have been recent advances in metabolic profiling and proteome analyses in many species with mass spectrometry used to identify 'biomarkers' and mechanisms indicative of specific physiological states (including pregnancy). Here, we hypothesized that protein expression in voided cheetah feces varied depending on pregnancy status. We: 1) identified the expansive protein profile present in fecal material of females; and 2) isolated proteins that may be candidates playing a role in early pregnancy establishment and diagnosis. Five hundred and seventy unique proteins were discovered among samples from pregnant (n = 8), non-pregnant, luteal phase (n = 5), and non-ovulatory control (n = 5) cheetahs. Four protein candidates were isolated that were significantly up-regulated and two were down-regulated in samples from pregnant compared to non-pregnant or control counterparts. One up-regulated candidate, immunoglobulin J chain (IGJ; an important component of the secretory immune system) was detected using a commercially available antibody via immunoblotting. Findings revealed that increased IGJ abundance could be used to detect pregnancy successfully in >80% of 23 assessed females within 4 weeks after mating. The discovery of a novel fecal pregnancy marker improves the ability to determine reproductive, especially gestational, status in cheetahs managed in an ex situ insurance and source population.

Cheetah are induced ovulators, experiencing short, variable oestrogen waves year-round. Exogenous gonadotrophin administration induces ovulation, but success is variable and often improves if ovaries are quiescent. After affirming the presence of short-term oestrogenic waves, we examined the effect of the timing of administration of exogenous equine and human chorionic gonadotrophins (eCG-hCG) within the oestrogen concentration pattern on subsequent follicle development and oocyte and corpus luteum quality. We also investigated ovarian suppression using an oral progestin (Altrenogest, 7 days) and assessed whether Altrenogest moderated adrenal activity by reducing glucocorticoid metabolites. All cheetahs exhibited short (every ~7-10 days), sporadic, year-round increases in faecal oestradiol punctuated by unpredictable periods (4-10 weeks) of baseline oestradiol (anoestrous). Gonadotrophin (eCG-hCG) efficacy was not affected by oestradiol 'wave' pattern if administered ≥3 days after an oestrogen peak. Such cheetahs produced normative faecal progestagen patterns and higher numbers (P<0.06) of mature oocytes than females given gonadotrophins ≤2 days after an oestradiol peak. Altrenogest supplementation expanded the interval between oestradiol peaks to 12.9 days compared with 7.3 days without progestin pretreatment. Altrenogest-fed females excreted less (P<0.05) glucocorticoid metabolites than non-supplemented counterparts. Results show that Altrenogest is effective for suppressing follicular activity, may contribute to reduced glucocorticoid production and may result in more effective ovulation induction via gonadotrophin therapy.

Cheetahs in managed zoological collections do not reproduce efficiently, a problem that may be related to environmental/management stressors. In this study, we examined 17 adult female cheetahs to determine the influence of two environmental factors, (1) being housed on- or off-exhibit and (2) number of adult conspecifics (males and/or females) in nearby enclosures, on profiles and concentrations of ovarian and adrenal hormones. Secondarily, we assessed a subset of group-housed siblings (n=5 females in groups of 2 or 3) for effects of long-term cohabitation. All of the females demonstrated waves of estrogen excretion (indicative of ovarian activity) as well as occasional periods of no estrogen production (anestrus). Glucocorticoid and estrogen concentrations were correlated within an individual (r s =0.53; P<0.05), and overall there was a higher frequency of days with elevated glucocorticoid concentrations in association with elevated estrogen excretion. However, none of the management factors had an impact (P>0.05) on estrogen or glucocorticoid metabolite excretory patterns. Although we recently reported that public exposure can negatively affect sperm production, ovarian steroidogenesis in females was unaffected. There also was no evidence of hyper-adrenal activity. Thus, different methods of ex situ management appear to have minimal influence on ovarian function or stress susceptibility of female cheetahs., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Although the free-ranging cheetah is generally socially solitary, as many as 60% of males live in same-sex (usually sibling) coalitions. Under ex situ conditions, the cheetah experiences low reproductive success with only ~18% of males having ever produced young. Most male cheetahs (85%) are managed in captivity in coalitions, but with no data on the influence of social grouping on reproductive parameters. We examined the influence of singleton versus coalition management on various male cheetah physiological traits, including ejaculate quality and gonadal and adrenal hormone metabolite concentrations. We also assessed behaviour within coalitions for evidence of social hierarchy through initiation of interactions with group mates and relatedness to physiological traits. Ejaculate quality (including total motile and structurally normal spermatozoa per ejaculate) and androgen concentration profiles were higher (P<0.05) in coalition compared with singleton males. These results support the conclusion that testis function in the cheetah, specifically related to the development of normal, motile spermatozoa and androgen production, is influenced by management with same-sex conspecifics. The findings have implications for ex situ conservation breeding programs by suggesting that reproductive quality can be enhanced through group maintenance of cheetah males.

The synthesis of densly functionized α-silyl-β-hydroxyl vinylsilanes via ruthenium-catalyzed multicomponent reaction (MCR) is reported herein. Exceptionally high regio- and diastereoselectivity was achieved by employing an unprecedented hydrosilylation of bifunctional silyl-propargyl boronates. The simple protocol, mild reaction conditions, and unique tolerability of this method make it a valuable tool for the synthesis of highly elaborated building blocks. The one-pot synthesis of stereodefined olefins, the generation of a valuable cyclohexene building block through a four-component MCR, and further functionalization in an abundance of diastereoselective reactions is disclosed herein., (© 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The synthesis of vinyl boronates and vinyl silanes was achieved by employing a Ru-catalyzed alkene-alkyne coupling reaction of allyl boronates or allyl silanes with various alkynes. The double bond geometry in the generated vinyl boronates can be remotely controlled by the juxtaposing boron- and silicon groups on the alkyne substrate. The synthetic utility of the coupling products has been demonstrated in a variety of synthetic transformations, including iterative cross-coupling reactions, and a Chan-Lam-type allyloxylation followed by a Claisen rearrangement. A sequential one-pot alkene-alkyne-coupling/allylation-sequence with an aldehyde to deliver a highly complex α-silyl-β-hydroxy olefin with a handle for further functionalization was also realized., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The collective cheetah (Acinonyx jubatus) population in zoological institutions has never been self-sustaining because of challenges in natural reproduction. A retrospective analysis of North American zoo-breeding records has revealed that >90% of litters produced since 2003 occurred in facilities 'off-display' from the public. We examined seminal, endocrine, and behavioral traits of 29 adult male cheetahs that were: 1) managed in public exhibit or off-display facilities; 2) maintained by different numbers of cheetah-specific care-givers; and 3) living adjacent to varying numbers of adult conspecifics. Cheetahs housed off-display produced more total motile sperm/ejaculate (P = 0.04) than on-exhibit males. This finding was mirrored in our laboratory's historical records where two-fold more total motile sperm (P < 0.01) were measured in ejaculates from individuals with no public exposure (n = 43) compared to on-exhibit (n = 116) counterparts. Males at institutions with ≤3 care-givers also produced more total motile sperm/ejaculate (P < 0.03) and spent more time behaviorally active (P < 0.01) than at facilities using >3 care-givers. Exposure to high numbers of conspecifics within the same institution did not impact (P > 0.05) seminal traits, and presence of the public, care-giver number, or animals/facility had no influence (P > 0.05) on androgen or glucocorticoid excretion or other behavioral metrics. Findings indicate that male cheetahs are sensitive to general public exposure and too many care-givers, resulting in compromised motile sperm output/ejaculate with mechanism of action unrelated to altered androgen or glucocorticoid excretion.

The development and application of a new generation of non-C2-symmetric ProPhenol ligands is reported herein. Rational design of the ProPhenol ligand paved the way to the first catalytic and asymmetric vinylation of N-Boc imines via hydrozirconation giving rise to valuable allylic amines in excellent yields and enantioselectivities. The utility of this method was demonstrated by developing the shortest reported asymmetric synthesis of the selective serotonine reuptake inhibitor (SSRI) (-)-dapoxetine.

Biologically interesting fused oligocyclic lactams have been prepared via an intramolecular redox-neutral cyclization process. By the proper choice of the substrates with a wide variety of tethered olefins, the less favored C-H bond can be activated and functionalized. This C-H activation proceeds under mild conditions, obviates the need for external oxidants, and displays a broad scope with respect to the substituents.

Multisubstituted isoquinoline and pyridine N-oxides have been prepared by Rh(III)-catalyzed cyclization of oximes and diazo compounds via aryl and vinylic C-H activation. This intermolecular annulation involving tandem C-H activation, cyclization, and condensation steps proceeds under mild conditions, obviates the need for oxidants, releases N2 and H2O as the byproducts, and displays a broad substituent scope.

The cooperative catalysis by palladium and triphenylborane effects the intramolecular oxycyanation of alkenes through the cleavage of O-CN bonds and the subsequent insertion of double bonds. The use of 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene (Xantphos) as a ligand for palladium is essential for allowing the transformation to proceed with high chemo- and regioselectivity. Variously substituted dihydrobenzofurans with both a tetra-substituted carbon and cyano functionality are accessed by the newly developed methodology., (© 2012 American Chemical Society)

Herein, we report on our findings of the Sonogashira-Hagihara reaction with 1-iodinated and 2-brominated glycals using several aromatic and aliphatic alkynes. This Pd-catalyzed cross-coupling reaction presents a facile access to alkynyl C-glycosides and sets the stage for a reductive/oxidative refunctionalization of the enyne moiety to regenerate either C-glycosidic structures or pyran derivatives with a substituent in position 2.

A flexible and robust method for the assembly of (1-->6)-linked C-glycosidic disaccharides is presented. The key reaction is a Pd-catalyzed coupling of 1-iodo- or 1-triflato-glycals with alkynyl glycosides. Reinstallation of the native hydroxyl group pattern is achieved after selective hydrogenation of the triple bond using Raney-nickel. Epoxidation with DMDO and reductive epoxide opening gives access to either the alpha- or the beta-derivative, depending on the hydride source.

The effects of perinatal exposure to progesterone (P) and estradiol (E) on sexual differentiation of behavior and morphology were examined by treating male and female gray short-tailed opossums on postnatal day 8 with progesterone alone (P), P plus estradiol (E) (PE), the P receptor antagonist mifepristone/RU486 (MIF), or corn oil control (C) and gonadectomizing them before puberty. When given female hormone replacement therapy in adulthood and tested with intact stimulus males, MIF animals showed less female-typical aggressive threat behavior than animals in other treatment groups. Stimulus males scent marked in more tests involving females than males and in more tests involving MIF animals than animals in other treatment groups. Body weight was lower in females than in males and was lower in MIF animals than in animals in other treatment groups, and P females failed to show female-typical genital locks after copulation. Sexual receptivity was similar in males and females and, while not decreased by any perinatal hormone treatment, was higher in PE males than in animals of either sex in any treatment group. These findings suggest that perinatal exposure to P is associated with the organization of feminine threat behavior and the defeminization of attractivity, body weight and genital anatomy in this marsupial. Reasons for these findings and for why female sexual receptivity is enhanced by perinatal exposure to exogenous E only in an endogenous masculine environment are discussed., (Copyright 2010 Elsevier Inc. All rights reserved.)

Herein we describe the synthesis of highly substituted chromans and isochromans using carbohydrates as starting materials. The key step of our synthetic approach is the annelation of the benzene moiety via a highly efficient Pd-catalyzed domino reaction. This powerful approach led to a small library of highly substituted chromans and isochromans by making use of a variety of different diynes and bromoglycals. We investigated several Pd-catalysts in order to improve the yields and to enlarge the scope of the domino reaction. Furthermore, we elucidated the mechanistic picture of the reaction with isotope-labelling experiments. Most probably the reaction proceeds via an oxidative addition followed by two carbopalladation steps and a final cyclization reaction.

Herein we describe the synthesis of highly substituted chromans and isochromans using carbohydrates as starting materials. Our approach makes use of a Pd-catalyzed domino reaction consisting of oxidative addition, followed by two carbopalladation steps and completed by a cyclization to annelate the benzene moiety. The versatility of this route has been demonstrated by a small library of highly substituted chromans and isochromans.

Female gray short-tailed opossums (Monodelphis domestica) lack an estrous cycle and are induced into estrus by exposure to a pheromone in male scent marks. Behavioral and physiological responses of females to the volatile and nonvolatile components of scent marks were examined in two experiments. Young females (n=9) were tested prior to and during their first estrus for behavioral responses to scent marks, collected on a 7-ml glass vial rubbed over the suprasternal gland of a mature male. The response to volatile components of the scent mark, recorded when marked and unmarked vials were covered with a perforated shield, was compared to the response to these vials when unshielded. Estrous females nuzzled the shields over marked vials (55.8+/-8.5 nuzzles/10 min) more than the shielded clean vial (10.9+/-2.4) (P<0.05); a similar response was observed in anestrous females. Nuzzling of unshielded, scent-marked vials was higher (P<0.05) during anestrus than in the same females when in estrus. The role of nonvolatile pheromones in reproductive activation was tested in adult females (n=11) exposed for up to 14 days to a shielded, marked vial or to an unshielded, marked vial in a crossover design. All females exposed to unshielded vials expressed estrus, and 10 copulated. Only 2 females expressed estrus (significantly fewer, P<0.05), when exposed to shielded marked vials, and neither copulated. These results demonstrate that females detect and respond behaviorally to both volatile and nonvolatile components of male suprasternal gland secretion, but the estrus-inducing pheromone in these secretions is nonvolatile.