Changes in parrot diversity after human arrival to the Caribbean.

Humans did not arrive on most of the world's islands until relatively recently, making islands favorable places for disentangling the timing and magnitude of natural and anthropogenic impacts on species diversity and distributions. Here, we focus on Amazona parrots in the Caribbean, which have close relationships with humans (e.g., as pets as well as sources of meat and colorful feathers). Caribbean parrots also have substantial fossil and archaeological records that span the Holocene. We leverage this exemplary record to showcase how combining ancient and modern DNA, along with radiometric dating, can shed light on diversification and extinction dynamics and answer long-standing questions about the magnitude of human impacts in the region. Our results reveal a striking loss of parrot diversity, much of which took place during human occupation of the islands. The most widespread species, the Cuban Parrot, exhibits interisland divergences throughout the Pleistocene. Within this radiation, we identified an extinct, genetically distinct lineage that survived on the Turks and Caicos until Indigenous human settlement of the islands. We also found that the narrowly distributed Hispaniolan Parrot had a natural range that once included The Bahamas; it thus became "endemic" to Hispaniola during the late Holocene. The Hispaniolan Parrot also likely was introduced by Indigenous people to Grand Turk and Montserrat, two islands where it is now also extirpated. Our research demonstrates that genetic information spanning paleontological, archaeological, and modern contexts is essential to understand the role of humans in altering the diversity and distribution of biota.

Bird populations and species lost to Late Quaternary environmental change and human impact in the Bahamas.

Comparing distributional information derived from fossils with the modern distribution of species, we summarize the changing bird communities of the Bahamian Archipelago across deep ecological time. While our entire dataset consists of 7,600+ identified fossils from 32 sites on 15 islands (recording 137 species of resident and migratory birds), we focus on the landbirds from four islands with the best fossil records, three from the Late Pleistocene (∼25 to 10 ka [1,000 y ago]) and one from the Holocene (∼10 to 0 ka). The Late Pleistocene sites feature 51 resident species that have lost one or more Bahamian populations; 29 of these species do not occur in any of the younger Holocene sites (or in the Bahamas today). Of these 29 species, 17 have their closest affinities to species now or formerly living in Cuba and/or North America. A set of 27 species of landbirds, most of them extant somewhere today, was more widespread in the Bahamas in the prehistoric Holocene (∼10 to 0.5 ka) than they are today; 16 of these 27 species were recorded as Pleistocene fossils as well. No single site adequately captures the entire landbird fauna of the combined focal islands. Information from all sites is required to assess changes in Bahamian biodiversity (including endemism) since the Late Pleistocene. The Bahamian islands are smaller, flatter, lower, and more biotically depauperate than the Greater Antilles, resulting in more vulnerable bird communities.

Ancient DNA from the extinct Haitian cave-rail (Nesotrochis steganinos) suggests a biogeographic connection between the Caribbean and Old World.

Worldwide decline in biodiversity during the Holocene has impeded a comprehensive understanding of pre-human biodiversity and biogeography. This is especially true on islands, because many recently extinct island taxa were morphologically unique, complicating assessment of their evolutionary relationships using morphology alone. The Caribbean remains an avian hotspot but was more diverse before human arrival in the Holocene. Among the recently extinct lineages is the enigmatic genus Nesotrochis, comprising three flightless species. Based on morphology, Nesotrochis has been considered an aberrant rail (Rallidae) or related to flufftails (Sarothruridae). We recovered a nearly complete mitochondrial genome of Nesotrochis steganinos from fossils, discovering that it is not a rallid but instead is sister to Sarothruridae, volant birds now restricted to Africa and New Guinea, and the recently extinct, flightless Aptornithidae of New Zealand. This result suggests a widespread or highly dispersive most recent common ancestor of the group. Prior to human settlement, the Caribbean avifauna had a far more cosmopolitan origin than is evident from extant species.

Scaffold-hopping identifies furano[2,3-d]pyrimidine amides as potent Notum inhibitors.

The carboxylesterase Notum is a key negative regulator of the Wnt signaling pathway by mediating the depalmitoleoylation of Wnt proteins. Our objective was to discover potent small molecule inhibitors of Notum suitable for exploring the regulation of Wnt signaling in the central nervous system. Scaffold-hopping from thienopyrimidine acids 1 and 2, supported by X-ray structure determination, identified 3-methylimidazolin-4-one amides 20-24 as potent inhibitors of Notum with activity across three orthogonal assay formats (biochemical, extra-cellular, occupancy). A preferred example 24 demonstrated good stability in mouse microsomes and plasma, and cell permeability in the MDCK-MDR1 assay albeit with modest P-gp mediated efflux. Pharmacokinetic studies with 24 were performed in vivo in mouse with single oral administration of 24 showing good plasma exposure and reasonable CNS penetration. We propose that 24 is a new chemical tool suitable for cellular studies to explore the fundamental biology of Notum.

Origin, paleoecology, and extirpation of bluebirds and crossbills in the Bahamas across the last glacial-interglacial transition.

On low islands or island groups such as the Bahamas, surrounded by shallow oceans, Quaternary glacial-interglacial changes in climate and sea level had major effects on terrestrial plant and animal communities. We examine the paleoecology of two species of songbirds (Passeriformes) recorded as Late Pleistocene fossils on the Bahamian island of Abaco-the Eastern bluebird (Sialia sialis) and Hispaniolan crossbill (Loxia megaplaga). Each species lives today only outside of the Bahamian Archipelago, with S. sialis occurring in North and Central America and L. megaplaga endemic to Hispaniola. Unrecorded in the Holocene fossil record of Abaco, both of these species probably colonized Abaco during the last glacial interval but were eliminated when the island became much smaller, warmer, wetter, and more isolated during the last glacial-interglacial transition from ∼15 to 9 ka. Today's warming temperatures and rising sea levels, although not as great in magnitude as those that took place from ∼15 to 9 ka, are occurring rapidly and may contribute to considerable biotic change on islands by acting in synergy with direct human impacts.

Ancient DNA from a 2,500-year-old Caribbean fossil places an extinct bird (Caracara creightoni) in a phylogenetic context.

Since the late Pleistocene humans have caused the extinction of species across our planet. Placing these extinct species in the tree of life with genetic data is essential to understanding the ecological and evolutionary implications of these losses. While ancient DNA (aDNA) techniques have advanced rapidly in recent decades, aDNA from tropical species, especially birds, has been historically difficult to obtain, leaving a gap in our knowledge of the extinction processes that have influenced current distributions and biodiversity. Here we report the recovery of a nearly complete mitochondrial genome from a 2,500 year old (late Holocene) bone of an extinct species of bird, Caracara creightoni, recovered from the anoxic saltwater environment of a blue hole in the Bahamas. Our results suggest that this extinct species is sister (1.6% sequence divergence) to a clade containing the extant C. cheriway and C. plancus. Caracara creightoni shared a common ancestor with these extant species during the Pleistocene (1.2-0.4 MYA) and presumably survived on Cuba when the Bahamas was mostly underwater during Quaternary interglacial intervals (periods of high sea levels). Tropical blue holes have been collecting animals for thousands of years and will continue to improve our understanding of faunal extinctions and distributions. In particular, new aDNA techniques combined with radiocarbon dating from Holocene Bahamian fossils will allow us to place other extinct (species-level loss) and extirpated (population-level loss) vertebrate taxa in improved phylogenetic, evolutionary, biogeographic, and temporal contexts.

New approaches for the treatment of Alzheimer's disease.

Alzheimer's disease (AD) is the most prevalent chronic neurodegenerative disease. Current approved therapies are symptomatic treatments having some effect on cognitive function. Therapies that target ß-amyloid (Aß) have been the focus of efforts to develop a disease modification treatment for AD but these approaches have failed to show any clinical benefit so far. Beyond the 'Aß hypothesis', there are a number of newer approaches to treat AD with neuroinflammation emerging as a very active area of research based on risk gene analysis. This short review will summarize approved drug therapies, recent clinical trials and new approaches for the treatment of AD.

Predictable evolution toward flightlessness in volant island birds.

Birds are prolific colonists of islands, where they readily evolve distinct forms. Identifying predictable, directional patterns of evolutionary change in island birds, however, has proved challenging. The "island rule" predicts that island species evolve toward intermediate sizes, but its general applicability to birds is questionable. However, convergent evolution has clearly occurred in the island bird lineages that have undergone transitions to secondary flightlessness, a process involving drastic reduction of the flight muscles and enlargement of the hindlimbs. Here, we investigated whether volant island bird populations tend to change shape in a way that converges subtly on the flightless form. We found that island bird species have evolved smaller flight muscles than their continental relatives. Furthermore, in 366 populations of Caribbean and Pacific birds, smaller flight muscles and longer legs evolved in response to increasing insularity and, strikingly, the scarcity of avian and mammalian predators. On smaller islands with fewer predators, birds exhibited shifts in investment from forelimbs to hindlimbs that were qualitatively similar to anatomical rearrangements observed in flightless birds. These findings suggest that island bird populations tend to evolve on a trajectory toward flightlessness, even if most remain volant. This pattern was consistent across nine families and four orders that vary in lifestyle, foraging behavior, flight style, and body size. These predictable shifts in avian morphology may reduce the physical capacity for escape via flight and diminish the potential for small-island taxa to diversify via dispersal.

An improved, scalable synthesis of Notum inhibitor LP-922056 using 1-chloro-1,2-benziodoxol-3-one as a superior electrophilic chlorinating agent.

Background: The carboxylesterase Notum has been shown to act as a key negative regulator of the Wnt signalling pathway by mediating the depalmitoleoylation of Wnt proteins. LP-922056 (1) is an orally active inhibitor of Notum. We are investigating the role of Notum in modulating Wnt signalling in the central nervous system and wished to establish if 1 would serve as a peripherally restricted control. An accessible and improved synthetic route would allow 1 to become more readily available as a chemical tool to explore the fundamental biology of Notum and build target validation to underpin new drug discovery programs. Results: An improved, scalable synthesis of 1 is reported. Key modifications include: (1) the introduction of the C7-cyclopropyl group was most effectively achieved with a Suzuki-Miyaura cross-coupling reaction with MIDA-boronate 11 (5 → 6), and (2) C6 chlorination was performed with 1-chloro-1,2-benziodoxol-3-one (12) (6 → 7) as a mild and selective electrophilic chlorination agent. This 7-step route from 16 has been reliably performed on large scale to produce multigram quantities of 1 in good efficiency and high purity. Pharmacokinetic studies in mouse showed CNS penetration of 1 is very low with a brain/plasma concentration ratio of just 0.01. A small library of amides 17 were prepared from acid 1 to explore if 1 could be modified to deliver a CNS penetrant tool by capping off the acid as an amide. Although significant Notum inhibition activity could be achieved, none of these amides demonstrated the required combination of metabolic stability along with cell permeability without evidence of P-gp mediated efflux. Conclusion: Mouse pharmacokinetic studies demonstrate that 1 is unsuitable for use in models of disease where brain penetration is an essential requirement of the compound but would be an ideal peripherally restricted control. These data will contribute to the understanding of drug levels of 1 to overlay with appropriate in vivo efficacy endpoints, i.e., the PK-PD relationship. The identification of a suitable analogue of 1 (or 17) which combines Notum inhibition with CNS penetration would be a valuable chemical probe for investigating the role of Notum in disease models.

Vertebrate community on an ice-age Caribbean island.

We report 95 vertebrate taxa (13 fishes, 11 reptiles, 63 birds, 8 mammals) from late Pleistocene bone deposits in Sawmill Sink, Abaco, The Bahamas. The >5,000 fossils were recovered by scuba divers on ledges at depths of 27-35 m below sea level. Of the 95 species, 39 (41%) no longer occur on Abaco (4 reptiles, 31 birds, 4 mammals). We estimate that 17 of the 39 losses (all of them birds) are linked to changes during the Pleistocene-Holocene Transition (PHT) (∼ 15-9 ka) in climate (becoming more warm and moist), habitat (expansion of broadleaf forest at the expense of pine woodland), sea level (rising from -80 m to nearly modern levels), and island area (receding from ∼ 17,000 km(2) to 1,214 km(2)). The remaining 22 losses likely are related to the presence of humans on Abaco for the past 1,000 y. Thus, the late Holocene arrival of people probably depleted more populations than the dramatic physical and biological changes associated with the PHT.

Tropical ancient DNA reveals relationships of the extinct Bahamian giant tortoise Chelonoidis alburyorum.

Ancient DNA of extinct species from the Pleistocene and Holocene has provided valuable evolutionary insights. However, these are largely restricted to mammals and high latitudes because DNA preservation in warm climates is typically poor. In the tropics and subtropics, non-avian reptiles constitute a significant part of the fauna and little is known about the genetics of the many extinct reptiles from tropical islands. We have reconstructed the near-complete mitochondrial genome of an extinct giant tortoise from the Bahamas (Chelonoidis alburyorum) using an approximately 1 000-year-old humerus from a water-filled sinkhole (blue hole) on Great Abaco Island. Phylogenetic and molecular clock analyses place this extinct species as closely related to Galápagos (C. niger complex) and Chaco tortoises (C. chilensis), and provide evidence for repeated overseas dispersal in this tortoise group. The ancestors of extant Chelonoidis species arrived in South America from Africa only after the opening of the Atlantic Ocean and dispersed from there to the Caribbean and the Galápagos Islands. Our results also suggest that the anoxic, thermally buffered environment of blue holes may enhance DNA preservation, and thus are opening a window for better understanding evolution and population history of extinct tropical species, which would likely still exist without human impact.

Suppression of type 1 pilus assembly in uropathogenic Escherichia coli by chemical inhibition of subunit polymerization.

OBJECTIVES: To identify and to characterize small-molecule inhibitors that target the subunit polymerization of the type 1 pilus assembly in uropathogenic Escherichia coli (UPEC). METHODS: Using an SDS-PAGE-based assay, in silico pre-filtered small-molecule compounds were screened for specific inhibitory activity against the critical subunit polymerization step of the chaperone-usher pathway during pilus biogenesis. The biological activity of one of the compounds was validated in assays monitoring UPEC type 1 pilus biogenesis, type 1 pilus-dependent biofilm formation and adherence to human bladder epithelial cells. The time dependence of the in vivo inhibitory activity and the overall effect of the compound on UPEC growth were determined. RESULTS: N-(4-chloro-phenyl)-2-{5-[4-(pyrrolidine-1-sulfonyl)-phenyl]-[1,3,4]oxadiazol-2-yl sulfanyl}-acetamide (AL1) inhibited in vitro pilus subunit polymerization. In bacterial cultures, AL1 disrupted UPEC type 1 pilus biogenesis and pilus-dependent biofilm formation, and resulted in the reduction of bacterial adherence to human bladder epithelial cells, without affecting bacterial cell growth. Bacterial exposure to the inhibitor led to an almost instantaneous loss of type 1 pili. CONCLUSIONS: We have identified and characterized a small molecule that interferes with the assembly of type 1 pili. The molecule targets the polymerization step during the subunit incorporation cycle of the chaperone-usher pathway. Our discovery provides new insight into the design and development of novel anti-virulence therapies targeting key virulence factors of bacterial pathogens.

Investigating the reaction mechanism and organocatalytic synthesis of α,α'-dihydroxy ketones.

A biomimetic TK one-pot reaction using hydroxypyruvate and aldehydes to generate α,α'-dihydroxy ketones in water has recently been described. To investigate this tertiary-amine mediated reaction mechanism two approaches were used. Firstly, (13)C labelled lithium hydroxypyruvate was synthesised and used to establish where hydroxypyruvate is incorporated in the product. In separate experiments reaction intermediates were also successfully intercepted and structurally identified using ESI-MS with tandem mass spectrometry ESI-MS/MS. These studies indicated that two mechanisms appear to be operating, one involving the addition of the tertiary amine catalyst to hydroxypyruvate, the other an aldol-based mechanism. Since the first mechanism may enable facial stereodifferentiation in the addition of intermediates to the aldehyde, a preliminary study on the use of chiral catalysts was performed and the first asymmetric organocatalytic synthesis of α,α'-dihydroxy ketones in aqueous media achieved, in up to 50% ee, using a quinine ether catalyst.

Rational substrate and enzyme engineering of transketolase for aromatics.

The uses of 3-formylbenzoic acid and 4-formylbenzoic acid as molecular probes along with previous and new transketolase mutants revealed the factors governing the rate of reaction between transketolase and aromatic aldehydes. The novel α,α-dihydroxyketones were produced at 15 to 30-fold higher yields and up to 250-fold higher specific activities with D469T TK when compared to those obtained for benzaldehyde.

Virtual Screening Directly Identifies New Fragment-Sized Inhibitors of Carboxylesterase Notum with Nanomolar Activity.

Notum is a negative regulator of Wnt signaling acting through the hydrolysis of a palmitoleoylate ester, which is required for Wnt activity. Inhibitors of Notum could be of use in diseases where dysfunctional Notum activity is an underlying cause. A docking-based virtual screen (VS) of a large commercial library was used to shortlist 952 compounds for experimental validation as inhibitors of Notum. The VS was successful with 31 compounds having an IC50 < 500 nM. A critical selection process was then applied with two clusters and two singletons (1-4d) selected for hit validation. Optimization of 4d guided by structural biology identified potent inhibitors of Notum activity that restored Wnt/ß-catenin signaling in cell-based models. The [1,2,4]triazolo[4,3-b]pyradizin-3(2H)-one series 4 represent a new chemical class of Notum inhibitors and the first to be discovered by a VS campaign. These results demonstrate the value of VS with well-designed docking models based on X-ray structures.

Structural Analysis and Development of Notum Fragment Screening Hits.

The Wnt signaling suppressor Notum is a promising target for osteoporosis, Alzheimer's disease, and colorectal cancers. To develop novel Notum inhibitors, we used an X-ray crystallographic fragment screen with the Diamond-SGC Poised Library (DSPL) and identified 59 fragment hits from the analysis of 768 data sets. Fifty-eight of the hits were found bound at the enzyme catalytic pocket with potencies ranging from 0.5 to >1000 µM. Analysis of the fragments' diverse binding modes, enzymatic inhibitory activities, and chemical properties led to the selection of six hits for optimization, and five of these resulted in improved Notum inhibitory potencies. One hit, 1-phenyl-1,2,3-triazole 7, and its related cluster members, have shown promising lead-like properties. These became the focus of our fragment development activities, resulting in compound 7d with IC50 0.0067 µM. The large number of Notum fragment structures and their initial optimization provided an important basis for further Notum inhibitor development.

Phylogenomic evidence for multiple losses of flight in ratite birds.

Ratites (ostriches, emus, rheas, cassowaries, and kiwis) are large, flightless birds that have long fascinated biologists. Their current distribution on isolated southern land masses is believed to reflect the breakup of the paleocontinent of Gondwana. The prevailing view is that ratites are monophyletic, with the flighted tinamous as their sister group, suggesting a single loss of flight in the common ancestry of ratites. However, phylogenetic analyses of 20 unlinked nuclear genes reveal a genome-wide signal that unequivocally places tinamous within ratites, making ratites polyphyletic and suggesting multiple losses of flight. Phenomena that can mislead phylogenetic analyses, including long branch attraction, base compositional bias, discordance between gene trees and species trees, and sequence alignment errors, have been eliminated as explanations for this result. The most plausible hypothesis requires at least three losses of flight and explains the many morphological and behavioral similarities among ratites by parallel or convergent evolution. Finally, this phylogeny demands fundamental reconsideration of proposals that relate ratite evolution to continental drift.

Ancient mitogenomics elucidates diversity of extinct West Indian tortoises.

We present 10 nearly complete mitochondrial genomes of the extinct tortoise Chelonoidis alburyorum from the Bahamas. While our samples represent morphologically distinct populations from six islands, their genetic divergences were shallow and resembled those among Galápagos tortoises. Our molecular clock estimates revealed that divergence among Bahamian tortoises began ~ 1.5 mya, whereas divergence among the Galápagos tortoises (C. niger complex) began ~ 2 mya. The inter-island divergences of tortoises from within the Bahamas and within the Galápagos Islands are much younger (0.09-0.59 mya, and 0.08-1.43 mya, respectively) than the genetic differentiation between any other congeneric pair of tortoise species. The shallow mitochondrial divergences of the two radiations on the Bahamas and the Galápagos Islands suggest that each archipelago sustained only one species of tortoise, and that the taxa currently regarded as distinct species in the Galápagos should be returned to subspecies status. The extinct tortoises from the Bahamas have two well-supported clades: the first includes one sample from Great Abaco and two from Crooked Island; the second clade includes tortoises from Great Abaco, Eleuthera, Crooked Island, Mayaguana, Middle Caicos, and Grand Turk. Tortoises belonging to both clades on Great Abaco and Crooked Island suggest late Holocene inter-island transport by prehistoric humans.

Structural Insights into Notum Covalent Inhibition.

The carboxylesterase Notum hydrolyzes a palmitoleate moiety from Wingless/Integrated(Wnt) ligands and deactivates Wnt signaling. Notum inhibitors can restore Wnt signaling which may be of therapeutic benefit for pathologies such as osteoporosis and Alzheimer's disease. We report the identification of a novel class of covalent Notum inhibitors, 4-(indolin-1-yl)-4-oxobutanoate esters. High-resolution crystal structures of the Notum inhibitor complexes reveal a common covalent adduct formed between the nucleophile serine-232 and hydrolyzed butyric esters. The covalent interaction in solution was confirmed by mass spectrometry analysis. Inhibitory potencies vary depending on the warheads used. Mechanistically, the resulting acyl-enzyme intermediate carbonyl atom is positioned at an unfavorable angle for the approach of the active site water, which, combined with strong hydrophobic interactions with the enzyme pocket residues, hinders the intermediate from being further processed and results in covalent inhibition. These insights into Notum catalytic inhibition may guide development of more potent Notum inhibitors.