Neotropics as a Cradle for Adaptive Radiations.

Neotropical ecosystems are renowned for numerous examples of adaptive radiation in both plants and animals resulting in high levels of biodiversity and endemism. However, we still lack a comprehensive review of the abiotic and biotic factors that contribute to these adaptive radiations. To fill this gap, we delve into the geological history of the region, including the role of tectonic events such as the Andean uplift, the formation of the Isthmus of Panama, and the emergence of the Guiana and Brazilian Shields. We also explore the role of ecological opportunities created by the emergence of new habitats, as well as the role of key innovations, such as novel feeding strategies or reproductive mechanisms. We discuss different examples of adaptive radiation, including classic ones like Darwin's finches and Anolis lizards, and more recent ones like bromeliads and lupines. Finally, we propose new examples of adaptive radiations mediated by ecological interactions in their geological context. By doing so, we provide insights into the complex interplay of factors that contributed to the remarkable diversity of life in the Neotropics and highlight the importance of this region in understanding the origins of biodiversity.

Does urbanisation lead to parallel demographic shifts across the world in a cosmopolitan plant?

Urbanisation is occurring globally, leading to dramatic environmental changes that are altering the ecology and evolution of species. In particular, the expansion of human infrastructure and the loss and fragmentation of natural habitats in cities is predicted to increase genetic drift and reduce gene flow by reducing the size and connectivity of populations. Alternatively, the 'urban facilitation model' suggests that some species will have greater gene flow into and within cities leading to higher diversity and lower differentiation in urban populations. These alternative hypotheses have not been contrasted across multiple cities. Here, we used the genomic data from the GLobal Urban Evolution project (GLUE), to study the effects of urbanisation on non-adaptive evolutionary processes of white clover (Trifolium repens) at a global scale. We found that white clover populations presented high genetic diversity and no evidence of reduced Ne linked to urbanisation. On the contrary, we found that urban populations were less likely to experience a recent decrease in effective population size than rural ones. In addition, we found little genetic structure among populations both globally and between urban and rural populations, which showed extensive gene flow between habitats. Interestingly, white clover displayed overall higher gene flow within urban areas than within rural habitats. Our study provides the largest comprehensive test of the demographic effects of urbanisation. Our results contrast with the common perception that heavily altered and fragmented urban environments will reduce the effective population size and genetic diversity of populations and contribute to their isolation.

Whole-genome analysis reveals the diversification of Galapagos rail (Aves: Rallidae) and confirms the success of goat eradication programs.

Similar to other insular birds around the world, the Galapagos rail (Laterallus spilonota Gould, 1841) exhibits reduced flight capacity following its colonization of the archipelago ~1.2 mya. Despite their short evolutionary history, rails have colonized seven different islands spanning the entire width of the archipelago. Galapagos rails were once common on islands with sufficiently high altitudes to support shrubs in humid habitats. After humans introduced goats, this habitat was severely reduced due to overgrazing. Habitat loss devastated some rail populations, with less than 50 individuals surviving, rendering the genetic diversity of Galapagos rail a pressing conservation concern. Additionally, one enigma is the reappearance of rails on the island of Pinta after they were considered extirpated. Our approach was to investigate the evolutionary history and geographic distribution of Galapagos rails as well as examine the genome-wide effects of historical population bottlenecks using 39 whole genomes across different island populations. We recovered an early divergence of rail ancestors leading to the isolated populations on Pinta and a second clade comprising the rest of the islands, historically forming a single landmass. Subsequently, the separation of the landmass ~900 kya may have led to the isolation of the Isabela population with more panmictic populations found on Santa Cruz and Santiago islands. We found that rails genomes contain long runs of homozygosity (>2Mb) that could be related to the introduction of goats. Finally, our findings show that the modern eradication of goats was critical to avoiding episodes of inbreeding in most populations.

DNA-metabarcoding supports trophic flexibility and reveals new prey species for the Galapagos sea lion.

Tropical ecosystems are challenging for pinnipeds due to fluctuating food availability. According to previous research, the Galapagos sea lion (GSL, Zalophus wollebaeki) adopts trophic flexibility to face such conditions. However, this hypothesis comes from studies using traditional methods (hard-parts analysis of scat and isotopic analysis from tissue). We studied the diet of five rookeries in the southeastern Galapagos bioregion (which harbors the highest GSL density), via DNA-metabarcoding of scat samples. The DNA-metabarcoding approach may identify consumed prey with a higher taxonomic resolution than isotopic analysis, while not depending on hard-parts remaining through digestion. Our study included five different rookeries to look for evidence of trophic flexibility at the bioregional level. We detected 98 prey OTUs (124 scats), mostly assigned to bony-fish taxa; we identified novel prey items, including a shark, rays, and several deep-sea fish. Our data supported the trophic flexibility of GSL throughout the studied bioregion since different individuals from the same rookery consumed prey coming from different habitats and trophic levels. Significant diet differentiations were found among rookeries, particularly between Punta Pitt and Santa Fe. Punta Pitt rookery, with a more pronounced bathymetry and lower productivity, was distinguished by a high trophic level and consumption of a high proportion of deep-sea prey; meanwhile, Santa Fe, located in more productive, shallow waters over the shelf, consumed a high proportion of epipelagic planktivorous fish. Geographic location and heterogeneous bathymetry of El Malecon, Española, and Floreana rookeries would allow the animals therein to access both, epipelagic prey over the shelf, and deep-sea prey out of the shelf; this would lead to a higher prey richness and diet variability there. These findings provide evidence of GSL adopting a trophic flexibility to tune their diets to different ecological contexts. This strategy would be crucial for this endangered species to overcome the challenges faced in a habitat with fluctuating foraging conditions.

Urban living can rescue Darwin's finches from the lethal effects of invasive vampire flies.

Human activity changes multiple factors in the environment, which can have positive or negative synergistic effects on organisms. However, few studies have explored the causal effects of multiple anthropogenic factors, such as urbanization and invasive species, on animals and the mechanisms that mediate these interactions. This study examines the influence of urbanization on the detrimental effect of invasive avian vampire flies (Philornis downsi) on endemic Darwin's finches in the Galápagos Islands. We experimentally manipulated nest fly abundance in urban and non-urban locations and then characterized nestling health, fledging success, diet, and gene expression patterns related to host defense. Fledging success of non-parasitized nestlings from urban (79%) and non-urban (75%) nests did not differ significantly. However, parasitized, non-urban nestlings lost more blood, and fewer nestlings survived (8%) compared to urban nestlings (50%). Stable isotopic values (δ15 N) from urban nestling feces were higher than those from non-urban nestlings, suggesting that urban nestlings are consuming more protein. δ15 N values correlated negatively with parasite abundance, which suggests that diet might influence host defenses (e.g., tolerance and resistance). Parasitized, urban nestlings differentially expressed genes within pathways associated with red blood cell production (tolerance) and pro-inflammatory response (innate immunological resistance), compared to parasitized, non-urban nestlings. In contrast, parasitized non-urban nestlings differentially expressed genes within pathways associated with immunoglobulin production (adaptive immunological resistance). Our results suggest that urban nestlings are investing more in pro-inflammatory responses to resist parasites but also recovering more blood cells to tolerate blood loss. Although non-urban nestlings are mounting an adaptive immune response, it is likely a last effort by the immune system rather than an effective defense against avian vampire flies since few nestlings survived.

Effect of urbanization and parasitism on the gut microbiota of Darwin's finch nestlings.

Host-associated microbiota can be affected by factors related to environmental change, such as urbanization and invasive species. For example, urban areas often affect food availability for animals, which can change their gut microbiota. Invasive parasites can also influence microbiota through competition or indirectly through a change in the host immune response. These interacting factors can have complex effects on host fitness, but few studies have disentangled the relationship between urbanization and parasitism on an organism's gut microbiota. To address this gap in knowledge, we investigated the effects of urbanization and parasitism by the invasive avian vampire fly (Philornis downsi) on the gut microbiota of nestling small ground finches (Geospiza fuliginosa) on San Cristóbal Island, Galápagos. We conducted a factorial study in which we experimentally manipulated parasite presence in an urban and nonurban area. Faeces were then collected from nestlings to characterize the gut microbiota (i.e. bacterial diversity and community composition). Although we did not find an interactive effect of urbanization and parasitism on the microbiota, we did find main effects of each variable. We found that urban nestlings had lower bacterial diversity and different relative abundances of taxa compared to nonurban nestlings, which could be mediated by introduction of the microbiota of the food items or changes in host physiology. Additionally, parasitized nestlings had lower bacterial richness than nonparasitized nestlings, which could be mediated by a change in the immune system. Overall, this study advances our understanding of the complex effects of anthropogenic stressors on the gut microbiota of birds.

The fitness landscape of a community of Darwin's finches.

Divergent natural selection should lead to adaptive radiation-that is, the rapid evolution of phenotypic and ecological diversity originating from a single clade. The drivers of adaptive radiation have often been conceptualized through the concept of "adaptive landscapes," yet formal empirical estimates of adaptive landscapes for natural adaptive radiations have proven elusive. Here, we use a 17-year dataset of Darwin's ground finches (Geospiza spp.) at an intensively studied site on Santa Cruz (Galápagos) to estimate individual apparent lifespan in relation to beak traits. We use these estimates to model a multi-species fitness landscape, which we also convert to a formal adaptive landscape. We then assess the correspondence between estimated fitness peaks and observed phenotypes for each of five phenotypic modes (G. fuliginosa, G. fortis [small and large morphotypes], G. magnirostris, and G. scandens). The fitness and adaptive landscapes show 5 and 4 peaks, respectively, and, as expected, the adaptive landscape was smoother than the fitness landscape. Each of the five phenotypic modes appeared reasonably close to the corresponding fitness peak, yet interesting deviations were also documented and examined. By estimating adaptive landscapes in an ongoing adaptive radiation, our study demonstrates their utility as a quantitative tool for exploring and predicting adaptive radiation.

Genomic evidence for homoploid hybrid speciation in a marine mammal apex predator.

Hybridization is widespread and constitutes an important source of genetic variability and evolution. In animals, its role in generating novel and independent lineages (hybrid speciation) has been strongly debated, with only a few cases supported by genomic data. The South American fur seal (SAfs) Arctocephalus australis is a marine apex predator of Pacific and Atlantic waters, with a disjunct set of populations in Peru and Northern Chile [Peruvian fur seal (Pfs)] with controversial taxonomic status. We demonstrate, using complete genome and reduced representation sequencing, that the Pfs is a genetically distinct species with an admixed genome that originated from hybridization between the SAfs and the Galapagos fur seal (Arctocephalus galapagoensis) ~400,000 years ago. Our results strongly support the origin of Pfs by homoploid hybrid speciation over alternative introgression scenarios. This study highlights the role of hybridization in promoting species-level biodiversity in large vertebrates.

Molecular phylogeny of Psychodopygina (Diptera, Psychodidae) supporting morphological systematics of this group of vectors of New World tegumentary leishmaniasis.

New World sandflies are vectors of leishmaniasis, bartonellosis, and some arboviruses. A classification based on 88 morphological characters was proposed 27 years ago when the New World phlebotomines were organized into two tribes Hertigiini and Phlebotomini. The latter was structured into four subtribes (Brumptomyiina, Sergentomyiina, Lutzomyiina, and Psychodopygina) and 20 genera. The subtribe Psychodopygina, including most of the American vectors of tegumentary Leishmania comprises seven genera from which no molecular work has been produced to support this classification. Here, we carried out a molecular phylogeny based on combined sequences (1,334 bp) of two genes: partial 28S rDNA and mtDNA cytochrome b from 47 taxa belonging to the Psychodopygina. The Bayesian phylogenetic reconstruction agreed with the classification based on morphological characters, supporting the monophyly of the genera Psychodopygus and Psathyromyia, whereas Nyssomyia and Trichophoromyia seemed to be paraphyletic. The paraphylies of the two latter groups were exclusively caused by the doubtful position of the species Ny. richardwardi. Our molecular analysis provides additional support to adopt the morphologic classification of Psychodopygina.

Title: La phylogénie moléculaire des Psychodopygina (Diptera, Psychodidae) soutient la systématique morphologique de ce groupe de vecteurs de leishmaniose cutanée du Nouveau Monde. Abstract: Les phlébotomes du nouveau monde sont des vecteurs de leishmaniose, de bartonellose et de certains arbovirus. Une classification basée sur 88 caractères morphologiques a été proposée il y a 27 ans lorsque les phlébotomes du nouveau monde ont été organisés en deux tribus Hertigiini et Phlebotomini. Cette dernière était structurée en quatre sous-tribus (Brumptomyiina, Sergentomyiina, Lutzomyiina et Psychodopygina) et 20 genres. La sous-tribu des Psychodopygina, qui inclut la plupart des vecteurs américains de la leishmaniose cutanée, comprend sept genres mais aucun travail moléculaire n'a été produit pour soutenir cette classification. Dans cet article, nous avons réalisé une phylogénie moléculaire basée sur des séquences combinées (1334 pb) de deux gènes : ADNr 28S partiel et cytochrome b (ADNmt) chez 47 taxons appartenant aux Psychodopygina. La reconstruction phylogénique bayésienne est en accord avec la classification basée sur les caractères morphologiques, soutenant la monophylie des genres Psychodopygus et Psathyromyia, alors que Nyssomyia et Trichophoromyia semblent être paraphylétiques. Les paraphylies des deux derniers groupes sont exclusivement causées par la position douteuse de la seule espèce Ny. richardwardi. Notre analyse moléculaire fournit une raison supplémentaire pour adopter la classification morphologique des Psychodopygina.

Evolutionary genomics of oceanic island radiations.

A recurring feature of oceanic archipelagos is the presence of adaptive radiations that generate endemic, species-rich clades that can offer outstanding insight into the links between ecology and evolution. Recent developments in evolutionary genomics have contributed towards solving long-standing questions at this interface. Using a comprehensive literature search, we identify studies spanning 19 oceanic archipelagos and 110 putative adaptive radiations, but find that most of these radiations have not yet been investigated from an evolutionary genomics perspective. Our review reveals different gaps in knowledge related to the lack of implementation of genomic approaches, as well as undersampled taxonomic and geographic areas. Filling those gaps with the required data will help to deepen our understanding of adaptation, speciation, and other evolutionary processes.

Phenotypic divergence of traits that mediate antagonistic and mutualistic interactions between island and continental populations of the tropical plant, Tribulus cistoides (Zygophyllaceae).

Island systems have long served as a model for evolutionary processes due to their unique species interactions. Many studies of the evolution of species interactions on islands have focused on endemic taxa. Fewer studies have focused on how antagonistic and mutualistic interactions shape the phenotypic divergence of widespread nonendemic species living on islands. We used the widespread plant Tribulus cistoides (Zygophyllaceae) to study phenotypic divergence in traits that mediate antagonistic interactions with vertebrate granivores (birds) and mutualistic interactions with pollinators, including how this is explained by bioclimatic variables. We used both herbarium specimens and field-collected samples to compare phenotypic divergence between continental and island populations. Fruits from island populations were larger than on continents, but the presence of lower spines on mericarps was less frequent on islands. The presence of spines was largely explained by environmental variation among islands. Petal length was on average 9% smaller on island than continental populations, an effect that was especially accentuated on the Galápagos Islands. Our results show that Tribulus cistoides exhibits phenotypic divergence between island and continental habitats for antagonistic traits (seed defense) and mutualistic traits (floral traits). Furthermore, the evolution of phenotypic traits that mediate antagonistic and mutualistic interactions partially depended on the abiotic characteristics of specific islands. This study shows the potential of using a combination of herbarium and field samples for comparative studies on a globally distributed species to study phenotypic divergence on island habitats.

The terroir of the finch: How spatial and temporal variation shapes phenotypic traits in DARWIN'S finches.

The term terroir is used in viticulture to emphasize how the biotic and abiotic characteristics of a local site influence grape physiology and thus the properties of wine. In ecology and evolution, such terroir (i.e., the effect of space or "site") is expected to play an important role in shaping phenotypic traits. Just how important is the pure spatial effect of terroir (e.g., differences between sites that persist across years) in comparison to temporal variation (e.g., differences between years that persist across sites), and the interaction between space and time (e.g., differences between sites change across years)? We answer this question by analyzing beak and body traits of 4388 medium ground finches (Geospiza fortis) collected across 10 years at three locations in Galápagos. Analyses of variance indicated that phenotypic variation was mostly explained by site for beak size (η 2 = 0.42) and body size (η 2 = 0.43), with a smaller contribution for beak shape (η 2 = 0.05) and body shape (η 2 = 0.12), but still higher compared to year and site-by-year effects. As such, the effect of terroir seems to be very strong in Darwin's finches, notwithstanding the oft-emphasized interannual variation. However, these results changed dramatically when we excluded data from Daphne Major, indicating that the strong effect of terroir was mostly driven by that particular population. These phenotypic results were largely paralleled in analyses of environmental variables (rainfall and vegetation indices) expected to shape terroir in this system. These findings affirm the evolutionary importance of terroir, while also revealing its dependence on other factors, such as geographical isolation.

Lack of genetic differentiation in yellowfin tuna has conservation implications in the Eastern Pacific Ocean.

Yellowfin tuna, Thunnus albacares, is an important global fishery and of particular importance in the Eastern Pacific Ocean (EPO). According to the 2019 Inter-American Tropical Tuna Commission (IATTC) assessment, yellowfin tuna within the EPO is a single stock, and is being managed as one stock. However, previous studies indicate site fidelity, or limited home ranges, of yellowfin tuna which suggests the potential for multiple yellowfin tuna stocks within the EPO, which was supported by a population genetic study using microsatellites. If numerous stocks are present, management at the wrong spatial scales could cause the loss of minor yellowfin tuna populations in the EPO. In this study we used double digestion RADseq to assess the genetic structure of yellowfin tuna in the EPO. A total of 164 yellowfin tuna from Cabo San Lucas, México, and the Galápagos Islands and Santa Elena, Ecuador, were analysed using 18,011 single nucleotide polymorphisms. Limited genetic differentiation (FST = 0.00058-0.00328) observed among the sampling locations (México, Ecuador, Peru, and within Ecuador) is consistent with presence of a single yellowfin tuna population within the EPO. Our findings are consistent with the IATTC assessment and provide further evidence of the need for transboundary cooperation for the successful management of this important fishery throughout the EPO.

Population Genetics and Phylogeography of Galapagos Fur Seals.

Pinnipeds found across islands provide an ideal opportunity to examine the evolutionary process of population subdivision affected by several mechanisms. Here, we report the genetic consequences of the geographic distribution of rookeries in Galapagos fur seals (GFS: Arctocephalus galapagoensis) in creating population structure. We show that rookeries across four islands (nine rookeries) are genetically structured into the following major groups: 1) a western cluster of individuals from Fernandina; 2) a central group from north and east Isabela, Santiago, and Pinta; and possibly, 3) a third cluster in the northeast from Pinta. Furthermore, asymmetric levels of gene flow obtained from eight microsatellites found migration from west Isabela to Fernandina islands (number of migrants Nm = 1), with imperceptible Nm in any other direction. Our findings suggest that the marked structuring of populations recovered in GFS is likely related to an interplay between long-term site fidelity and long-distance migration in both male and female individuals, probably influenced by varying degrees of marine productivity.

The genome sequence of the avian vampire fly (Philornis downsi), an invasive nest parasite of Darwin's finches in Galápagos.

The invasive avian vampire fly (Philornis downsi, Diptera: Muscidae) is considered one of the greatest threats to the endemic avifauna of the Galápagos Islands. The fly larvae parasitize nearly every passerine species, including Darwin's finches. Most P. downsi research to date has focused on the effects of the fly on avian host fitness and mitigation methods. A lag in research related to the genetics of this invasion demonstrates, in part, the need to develop full-scale genomic resources with which to address further questions within this system. In this study, an adult female P. downsi was sequenced to generate a high-quality genome assembly. We examined various features of the genome (e.g., coding regions and noncoding transposable elements) and carried out comparative genomics analysis against other dipteran genomes. We identified lists of gene families that are significantly expanding or contracting in P. downsi that are related to insecticide resistance, detoxification, and counter defense against host immune responses. The P. downsi genome assembly provides an important resource for studying the molecular basis of successful invasion in the Galápagos and the dynamics of its population across multiple islands. The findings of significantly changing gene families associated with insecticide resistance and immune responses highlight the need for further investigations into the role of different gene families in aiding the fly's successful invasion. Furthermore, this genomic resource provides a necessary tool to better inform future research studies and mitigation strategies aimed at minimizing the fly's impact on Galápagos birds.

Urbanization alters interactions between Darwin's finches and Tribulus cistoides on the Galápagos Islands.

Emerging evidence suggests that humans shape the ecology and evolution of species interactions. Islands are particularly susceptible to anthropogenic disturbance due to the fragility of their ecosystems; however, we know little about the susceptibility of species interactions to urbanization on islands. To address this gap, we studied how the earliest stages of urban development affect interactions between Darwin's finches and its key food resource, Tribulus cistoides, in three towns on the Galápagos Islands. We measured variation in mericarp predation rates, mericarp morphology, and finch community composition using population surveys, experimental manipulations, and finch observations conducted in habitats within and outside of each town. We found that both seed and mericarp removal rates were higher in towns than natural habitats. We also found that selection on mericarp size and defense differed between habitats in the survey and experimental populations and that towns supported smaller and less diverse finch communities than natural habitats. Together, our results suggest that even moderate levels of urbanization can alter ecological interactions between Darwin's finches and T. cistoides, leading to modified natural selection on T. cistoides populations. Our study demonstrates that trophic interactions on islands may be susceptible to the anthropogenic disturbance associated with urbanization. Despite containing the highest diversity in the world, studies of urbanization are lacking from the tropics. Our study identified signatures of urbanization on species interactions in a tropical island ecosystem and suggests that changes to the ecology of species interactions has the potential to alter evolution in urban environments.

International fisheries threaten globally endangered sharks in the Eastern Tropical Pacific Ocean: the case of the Fu Yuan Yu Leng 999 reefer vessel seized within the Galápagos Marine Reserve.

Shark fishing, driven by the fin trade, is the primary cause of global shark population declines. Here, we present a case study that exemplifies how industrial fisheries are likely depleting shark populations in the Eastern Tropical Pacific Ocean. In August 2017, the vessel Fu Yuan Yu Leng 999, of Chinese flag, was detained while crossing through the Galápagos Marine Reserve without authorization. This vessel contained 7639 sharks, representing one of the largest seizures recorded to date. Based on a sample of 929 individuals (12%), we found 12 shark species: 9 considered as Vulnerable or higher risk by the IUCN and 8 listed in CITES. Four species showed a higher proportion of immature than mature individuals, whereas size-distribution hints that at least some of the fishing ships associated with the operation may have been using purse-seine gear fishing equipment, which, for some species, goes against international conventions. Our data expose the magnitude of the threat that fishing industries and illegal trade represent to sharks in the Eastern Tropical Pacific Ocean.

Urban living influences the nesting success of Darwin's finches in the Galápagos Islands.

Urbanization is expanding worldwide with major consequences for organisms. Anthropogenic factors can reduce the fitness of animals but may have benefits, such as consistent human food availability. Understanding anthropogenic trade-offs is critical in environments with variable levels of natural food availability, such as the Galápagos Islands, an area of rapid urbanization. For example, during dry years, the reproductive success of bird species, such as Darwin's finches, is low because reduced precipitation impacts food availability. Urban areas provide supplemental human food to finches, which could improve their reproductive success during years with low natural food availability. However, urban finches might face trade-offs, such as the incorporation of anthropogenic debris (e.g., string, plastic) into their nests, which may increase mortality. In our study, we determined the effect of urbanization on the nesting success of small ground finches (Geospiza fuliginosa; a species of Darwin's finch) during a dry year on San Cristóbal Island. We quantified nest building, egg laying and hatching, and fledging in an urban and nonurban area and characterized the anthropogenic debris in nests. We also documented mortalities including nest trash-related deaths and whether anthropogenic materials directly led to entanglement- or ingestion-related nest mortalities. Overall, urban finches built more nests, laid more eggs, and produced more fledglings than nonurban finches. However, every nest in the urban area contained anthropogenic material, which resulted in 18% nestling mortality while nonurban nests had no anthropogenic debris. Our study showed that urban living has trade-offs: urban birds have overall higher nesting success during a dry year than nonurban birds, but urban birds can suffer mortality from anthropogenic-related nest-materials. These results suggest that despite potential costs, finches benefit overall from urban living and urbanization may buffer the effects of limited resource availability in the Galápagos Islands.

Sex identification in embryos and adults of Darwin's finches.

Darwin's finches are an iconic example of adaptive radiation and evolution under natural selection. Comparative genetic studies using embryos of Darwin's finches have shed light on the possible evolutionary processes underlying the speciation of this clade. Molecular identification of the sex of embryonic samples is important for such studies, where this information often cannot be inferred otherwise. We tested a fast and simple chicken embryo protocol to extract DNA from Darwin's finch embryos. In addition, we applied minor modifications to two of the previously reported PCR primer sets for CHD1, a gene used for sexing adult passerine birds. The sex of all 29 tested embryos of six species of Darwin's finches was determined successfully by PCR, using both primer sets. Next to embryos, hatchlings and fledglings are also impossible to distinguish visually. This extends to juveniles of sexually dimorphic species which are yet to moult in adult-like plumage and beak colouration. Furthermore, four species of Darwin's finches are monomorphic, males and females looking alike. Therefore, sex assessment in the field can be a source of error, especially with respect to juveniles and mature monomorphic birds outside of the mating season. We caught 567 juveniles and adults belonging to six species of Darwin's finches and only 44% had unambiguous sex-specific morphology. We sexed 363 birds by PCR: individuals sexed based on marginal sex specific morphological traits; and birds which were impossible to classify in the field. PCR revealed that for birds with marginal sex specific traits, sexing in the field produced a 13% error rate. This demonstrates that PCR based sexing can improve field studies on Darwin's finches, especially when individuals with unclear sex-related morphology are involved. The protocols used here provide an easy and reliable way to sex Darwin's finches throughout ontogeny, from embryos to adults.

Avian disease surveillance on the island of San Cristóbal, Galápagos.

Endemic island species face unprecedented threats, with many populations in decline or at risk of extinction. One important threat is the introduction of novel and potentially devastating diseases, made more pressing due to accelerating global connectivity, urban development, and climatic changes. In the Galápagos archipelago two important wildlife diseases: avian pox (Avipoxvirus spp.) and avian malaria (Plasmodium spp. and related Haemosporidia) challenge endemic species. San Cristóbal island has seen a paucity of disease surveillance in avian populations, despite the island's connectedness to the continent and the wider archipelago. To survey prevalence and better understand the dynamics of these two diseases on San Cristóbal, we captured 1205 birds of 11 species on the island between 2016 and 2020. Study sites included urban and rural lowland localities as well as rural highland sites in 2019. Of 995 blood samples screened for avian haemosporidia, none tested positive for infection. In contrast, evidence of past and active pox infection was observed in 97 birds and identified as strains Gal1 and Gal2. Active pox prevalence differed significantly with contemporary climatic conditions, being highest during El Niño events (~11% in 2016 and in 2019 versus <1% in the La Niña year of 2018). Pox prevalence was also higher at urban sites than rural (11% to 4%, in 2019) and prevalence varied between host species, ranging from 12% in medium ground finches (Geospiza fortis) to 4% in Yellow Warblers (Setophaga petechial aureola). In the most common infected species (Small Ground Finch: Geospiza fuliginosa), birds recovered from pox had significantly longer wings, which may suggest a selective cost to infection. These results illustrate the threat future climate changes and urbanization may present in influencing disease dynamics in the Galápagos, while also highlighting unknowns regarding species-specific susceptibilities to avian pox and the transmission dynamics facilitating outbreaks within these iconic species.