Myanmar is one of the most biodiverse countries from a species perspective in Southeast Asia, yet there is minimal published data on zoonotic viruses in small mammals. From July 2017 to August 2018, wildlife sampling was conducted at human-animal interfaces at sites in the Yangon Region and Kayin State. To investigate virus diversity of commensal rodents and shrew, rectal swabs were collected from mice (Mus sp., N = 3), rats (Rattus norvegicus, N = 80; Rattus rattus, N = 6), and Southeast Asian shrews (Crocidura fuliginosa, N = 8). RNA was extracted from rectal swabs, made into cDNA, and subjected to metagenomic next-generation sequencing followed by phylogenetic analysis for virus identification and taxonomic placement. The study provides the first detection of Wencheng shrew virus (WESV) in Myanmar and the first report in C. fuliginosa. A novel member of the genus Cardiovirus was also detected in R. norvegicus and clustered with Cardiovirus theileri sequences previously identified in wild rats from China. Further characterization of viruses circulating in small mammals will help inform public health officials of potential zoonotic risks in a region with virus surveillance gaps and ongoing land use change which may be increasing the risk of zoonotic disease emergence.

Over the past century, lion (Panthera leo) populations across Africa have experienced rapid and severe declines. Despite this, East Africa is considered a modern day lion stronghold. Here, we use whole genome sequencing of both recent and historical lion populations, primarily collected during the Smithsonian Roosevelt East African (1909 to 1911) and Rainey (1911 to 1912) Expeditions, to investigate changes in population structure, connectivity, and diversity over the last ~100 years in East Africa. We find a clear signal of population fragmentation when comparing historical (1896 to 1946) and recent (1990 to present) lion populations. Our analyses reveal genetic distinctions between remaining lion populations in Kenya and Tanzania and document loss of genetic diversity over time including reduced heterozygosity and the accumulation of runs of homozygosity. We detect a severe bottleneck in both Kenya and Tanzania approximately 25 generations ago, coinciding with a severe rinderpest outbreak in the region that is known to have decimated bovid and ultimately carnivore populations in the area. Nevertheless, modern lions in East Africa still exhibit overall high levels of diversity and low levels of inbreeding. Our results provide direct evidence of the effects of increasing habitat fragmentation and the significance of temporal data for contextualizing current patterns of population connectivity and diversity. ### Competing Interest Statement The authors have declared no competing interest.

Estimates of de novo mutation rates are essential for phylogenetic and demographic analyses, but their inference has previously been impeded by high error rates in sequence data and uncertainty in the fossil record. Here, we directly estimate de novo germline mutation rates for all extant members of Panthera, as well as the closely related outgroup Neofelis nebulosa, using pedigrees. We use a previously validated pipeline (RatesTools) to calculate mutation rate for each species and subsequently explore the impacts of the novel rates on historic effective population size estimates in each of these charismatic felids of conservation concern. Importantly, we find that the choice of reference genome, the data type and coverage, and the individual all impact estimates of the mutation rate. Despite these stochastic effects, we inferred that base pair mutation rates for all species fell between 0.5 and 1.4e-08 per generation per base pair (mean 0.81e-08 +/- 0.35-08 across Pantherinae). Our results provide a cautionary view on inter-species mutation rate comparisons, given the error associated with the reference genome choice and sequencing depth of coverage of the individuals.

Avipoxvirus is an avian pathogen that likely contributed to the declines and extinctions of endemic Hawaiian birds since its 19th century introduction. We surveyed 719 DNA libraries, including 639 representing 440 Hawaiian bird specimens, for evidence of Avipoxvirus infection. We reconstructed a 5.2× Avipoxvirus genome from an 1898 Hawaii 'amakihi (Chlorodrepanis virens) specimen. Its sequence matched an extant Hawaiian Avipoxvirus strain, supporting the strain's persistence in Hawaii over the last century. We identified the earliest molecularly verified case of Avipoxvirus in the Hawaiian Islands in an 1887 'alalā (Corvus hawaiiensis) specimen and reconstructed a partial Avipoxvirus genome from this specimen. Both specimens' Avipoxvirus strains were most closely related to canarypox virus, suggesting that introduced passerines may be the source of Avipoxvirus in Hawaiian endemic land birds. These findings clarify the origins and evolution of Avipoxvirus in Hawaii and provide evidence for the broader role of pathogens in driving biodiversity loss.

Native Hawaiian forest birds are experiencing an unprecedented extinction crisis. In particular, the iconic Hawaiian honeycreeper radiation has declined to just 17 out of ∼60 species remaining, most threatened with extinction due to avian malaria. Here, we investigate the genomic signatures of these declines in three honeycreeper species: the critically endangered 'akikiki (Oreomystis bairdi) and 'akeke'e (Loxops caeruleirostris) and the extinct po'ouli (Melamprosops phaeosoma). Surprisingly, we find that Hawaiian honeycreepers, even the last known po'ouli individual, maintain high heterozygosity compared with other island birds, reflecting historically large population sizes. This high heterozygosity may contribute to an elevated impact of inbreeding depression, as evidenced by reduced survival and reproductive success among highly inbred 'akikiki. Demographic analysis revealed that recent precipitous declines in 'akikiki and 'akeke'e coincide with the spread of avian malaria in the late 20th century, consistent with malaria being the primary driver of population collapse. Using predictive population viability modeling, we explore potential recovery scenarios for 'akeke'e, which has recently declined below 100 individuals in the wild. Our models predict that, under current conditions, 'akeke'e is likely to go extinct in the near future. However, if mosquito control campaigns are effective at reducing malaria, recovery can still occur. These findings emphasize the urgency of ongoing mosquito control efforts, demonstrating hope for a species nearing the brink of extinction. More broadly, our study provides a detailed examination of genomic diversity, inbreeding depression, and extinction risk in a collapsing adaptive radiation, with implications for conservation of other endangered island species.

Introduced pathogens exert novel selection on hosts, and although many host species have experienced drastic population declines in the absence of adaptation, some hosts have adapted to highly virulent pathogens. For instance, mosquitoes and Plasmodium relictum introduced to the Hawaiian Islands have resulted in extinctions and catastrophic population declines due to avian malaria, particularly in the diverse clade of Hawaiian honeycreepers. However, some species, such as the Hawai'i 'amakihi (Chlorodrepanis virens), can survive infection. Immunity exists in low-elevation populations where mosquitoes are abundant, whereas high-elevation, unexposed populations of 'amakihi display greatly reduced immunity. To explore the basis of adaptation to P. relictum in low-elevation 'amakihi, we sequenced transcriptomes from 24 low-elevation and 15 high-elevation 'amakihi. We tested for differential gene expression between (i) infected and uninfected birds and (ii) low- and high-elevation birds. Infected birds showed significant differences in expression across many transcripts with diverse cellular functions involved in different pathways of immune response; eight of the top 13 transcripts blasted to genes previously implicated in immunity to malaria in 'amakihi, and 11 have been identified in other infectious disease systems. Thirteen transcripts showed a trend of higher expression in high-elevation birds. These transcripts blasted to genes involved in metabolism, blood coagulation, and immune response. Our results provide increasing support for a subset of genes involved in immunity to malaria in 'amakihi and hint at possible antagonistic interactions between response to pathogens and environmental characteristics associated with elevation. Further work clarifying the nature of these interactions could benefit conservation efforts of Hawaiian honeycreepers in upper elevation refugia that are increasingly subject to malaria exposure.

In the current era, many terrestrial carnivore populations confront a multitude of threats and are rapidly shifting their ranges in response to human-induced modifications. Monitoring changes in genetic diversity and structure of such species in response to changing environmental conditions is important for understanding species' responses and designing effective conservation management strategies. In this study, we investigated the genetic status of the golden jackal, a widely distributed canid inhabiting human-dominated landscapes and exhibiting high dispersal capability. We collected and analyzed 141 jackal samples from across the distribution range in India, employing a combination of mitochondrial DNA (mtDNA) (cytochrome b and control region) and nuclear microsatellite (n = 25) markers to investigate patterns of genetic diversity, gene flow, demographic history and phylogeography. Our analyses showed substantial levels of genetic diversity within India surpassing levels observed in other global populations. Bayesian and non-Bayesian clustering analyses revealed low levels of genetic differentiation among sampled populations, except for the Southern Indian population. Demographic analysis using both mtDNA and microsatellites revealed that golden jackals in India have not experienced significant bottlenecks, while estimates of past effective population size suggested declines during the last 2,500 generations, which corresponds to 7,500 to 10,000 years before present. Global phylogeographic analysis highlighted the distinctiveness of Indian jackals compared with other populations from across the species' distribution, with the highest number of haplotypes observed in Indian populations and no shared haplotypes observed between Indian and Middle Eastern populations, or Indian and European populations. These findings are indicative of a long evolutionary history and bring new insights to inform targeted conservation management strategies for golden jackals, both locally and globally.

Owing in large part to Robert MacArthur’s classic research, wood warblers in the family Parulidae are textbook exemplars of species competition and niche partitioning. Conventional wisdom suggests that subtle differences in foraging behaviour are the principal means by which these nearly morphologically indistinguishable species are able to co-occur and avoid extinction. Yet, MacArthur’s study was in fact quite limited in scale, and he said little about the relevance of evolution to the study system. Here, we reassess MacArthur’s conclusions across an expanded set of syntopic warbler species in a forest in northern New York. We combine morphometrics, quantitative foraging data and faecal metabarcoding—a direct measure of warbler diet—to study competition and niche partitioning in an evolutionary framework. We find close relationships between morphology and foraging behaviour, but little connection between warbler ecomorphology and the 2237 invertebrate taxa detected in their diets. Instead, diet remains phylogenetically conserved—closely related warblers eat similar suites of invertebrates, regardless of where they forage. Finally, we present evidence that these species not only partition niche space in the present day but also that competition has shaped their behaviours over evolutionary time.

ABSTRACTThe animal gut microbiome can have a strong influence on the health, fitness, and behavior of its hosts. The composition of the gut microbial community can be influenced by factors such as diet, environment, and evolutionary history (phylosymbiosis). However, the relative influence of these factors is unknown in most bird species. Furthermore, phylosymbiosis studies have largely focused on clades that diverged tens of millions of years ago, and little is known about the degree of gut microbiome divergence in more recent species radiations. This study explores the drivers of microbiome variation across the unique and recent Hawaiian honeycreeper radiation (Fringillidae: Drepanidinae). Fecal samples were collected from 14 extant species spanning the main islands of the Hawaiian archipelago and were sequenced using three metabarcoding markers to characterize the gut microbiome, invertebrate diet, and plant diet of Hawaiian honeycreepers. We then used these metabarcoding data and the honeycreeper host phylogeny to evaluate their relative roles in shaping the gut microbiome. Microbiome variation across birds was highly individualized; however, source island had a small but significant effect on microbiome structure. The microbiomes did not recapitulate the host phylogenetic tree, indicating that evolutionary history does not strongly influence microbiome structure in the honeycreeper clade. These results expand our understanding of the roles of diet, geography, and phylogeny on avian microbiome structure, while also providing important ecological information about the diet and gut microbiota of wild Hawaiian honeycreepers.

Plasmodium parasites infect thousands of species and provide an exceptional system for studying hostpathogen dynamics, especially for multi-host pathogens. However, understanding these interactions requires an accurate assay of infection. Assessing Plasmodium infections using microscopy on blood smears often misses infections with low parasitemias (the fractions of cells infected), and biases in malaria prevalence estimates will differ among hosts that differ in mean parasitemias. We examined Plasmodium relictum infection and parasitemia using both microscopy of blood smears and quantitative polymerase chain reaction (qPCR) on 299 samples from multiple bird species in Hawai'i and fit models to predict parasitemias from qPCR cycle threshold (Ct) values. We used these models to quantify the extent to which microscopy underestimated infection prevalence and to more accurately estimate infection patterns for each species for a large historical study done by microscopy. We found that most qPCR-positive wild-caught birds in Hawaii had low parasitemias (Ct scores >= 35), which were rarely detected by microscopy. The fraction of infections missed by microscopy differed substantially among eight species due to differences in species' parasitemia levels. Infection prevalence was likely 4-5-fold higher than previous microscopy estimates for three introduced species, including Zosterops japonicus, Hawaii's most abundant forest bird, which had low average parasitemias. In contrast, prevalence was likely only 1.5-2.3-fold higher than previous estimates for Himatione sanguinea and Chlorodrepanis virens, two native species with high average parasitemias. Our results indicate that relative patterns of infection among species differ substantially from those observed in previous microscopy studies, and that differences depend on variation in parasitemias among species. Although microscopy of blood smears is useful for estimating the frequency of different Plasmodium stages and host attributes, more sensitive quantitative methods, including qPCR, are needed to accurately estimate and compare infection prevalence among host species. (c) 2023 Australian Society for Parasitology. Published by Elsevier Ltd. All rights reserved.

Island radiations, such as those of the Australo-Pacific, offer unique insight into diversification, extinction, and early speciation processes. Yet, their speciation and colonization histories are often obscured by conflicting genomic signals from incomplete lineage sorting (ILS) or hybridization. Here, we integrated mitogenomes and genome-wide SNPs to unravel the evolutionary history of one of the world's most geographically widespread island radiations. The Australo-Pacific reed warblers ( Acrocephalus luscinius complex) are a speciose lineage including five species that have become extinct since the 19th century and ten additional species of conservation concern. The radiation spans over 10,000 km across Australo-Papua, Micronesia and Polynesia, including the Mariana, Hawaii and Pitcairn Island archipelagos. Earlier mtDNA studies suggested a stepping-stone colonization process, resulting in archipelago-level secondary sympatry of divergent mtDNA lineages in the Mariana Islands and Marquesas. These studies hypothesized that morphologically similar species on neighboring islands arose from ecological convergence. Using DNA from historical museum specimens and modern genetic samples, we show that ILS and/or gene flow have shaped the radiation of Australo-Pacific reed warblers rather than secondary sympatry. The nuclear genome reconstructs a simpler biogeographic history than mtDNA, showing close relationships between species in the Mariana Islands and Marquesas despite their paraphyletic mtDNA lineages. Gene flow likely involved early and late colonizing waves of the radiation before the loss of ancestral dispersive ability. Our results highlight how collection genomics can elucidate evolutionary history and inform conservation efforts for threatened species.

Genetic and genomic data are increasingly used to aid conservation management of endangered species by providing insights into evolutionary histories, factors associated with extinction risks, and potential for future adaptation. For the 'Alalā, or Hawaiian crow (Corvus hawaiiensis), genetic concerns include negative correlations between inbreeding and hatching success. However, it is unclear if low genetic diversity and inbreeding depression are consequences of a historical population bottleneck, or if 'Alalā had historically low genetic diversity that predated human influence, perhaps as a result of earlier declines or founding events. In this study, we applied a hybridization-based sequence capture to generate a genome-wide single nucleotide polymorphism (SNP) dataset for comparing historical specimens collected in the 1890s, when 'Alalā were more numerous, to samples taken between 1973 and 1998, when 'Alalā population densities were near the lowest documented levels in the wild, prior to all individuals being collected for captive rearing. We found low genome-wide diversity in both sample groups, however, the modern sample group (1973 to 1998 cohort) exhibited relatively fewer polymorphic alleles, a lower proportion of polymorphic loci, and lower observed heterozygosity, consistent with a population decline and potential bottleneck effects. These results combined with a current low population size highlight the importance of continued efforts by conservation managers to mitigate inbreeding and maintain founder representation to preserve what genetic diversity remains.

The chytrid fungus Batrachochytrium dendrobatidis (Bd) was discovered in 1998 as the cause of chytridiomycosis, an emerging infectious disease causing mass declines in amphibian populations worldwide. The rapid population declines of the 1970s-1990s were likely caused by the spread of a highly virulent lineage belonging to the Bd-GPL clade that was introduced to naïve susceptible populations. Multiple genetically distinct and regional lineages of Bd have since been isolated and sequenced, greatly expanding the known biological diversity within this fungal pathogen. To date, most Bd research has been restricted to the limited number of samples that could be isolated using culturing techniques, potentially causing a selection bias for strains that can grow on media and missing other unculturable or fastidious strains that are also present on amphibians. We thus attempted to characterize potentially non-culturable genetic lineages of Bd from distinct amphibian taxa using sequence capture technology on DNA extracted from host tissue and swabs. We focused our efforts on host taxa from two different regions that likely harbored distinct Bd clades: (1) wild-caught leopard frogs (Rana) from North America, and (2) a Japanese Giant Salamander (Andrias japonicus) at the Smithsonian Institution's National Zoological Park that exhibited signs of disease and tested positive for Bd using qPCR, but multiple attempts failed to isolate and culture the strain for physiological and genetic characterization. We successfully enriched for and sequenced thousands of fungal genes from both host clades, and Bd load was positively associated with number of recovered Bd sequences. Phylogenetic reconstruction placed all the Rana-derived strains in the Bd-GPL clade. In contrast, the A. japonicus strain fell within the Bd-Asia3 clade, expanding the range of this clade and generating additional genomic data to confirm its placement. The retrieved ITS locus matched public barcoding data from wild A. japonicus and Bd infections found on other amphibians in India and China, suggesting that this uncultured clade is widespread across Asia. Our study underscores the importance of recognizing and characterizing the hidden diversity of fastidious strains in order to reconstruct the spatiotemporal and evolutionary history of Bd. The success of the sequence capture approach highlights the utility of directly sequencing pathogen DNA from host tissue to characterize cryptic diversity that is missed by culture-reliant approaches.

Museum genomics provide an opportunity to investigate population demographics of extinct species, especially valuable when research prior to extinction was minimal. The Bachman's warbler (Vermivora bachmanii) is hypothesized to have gone extinct due to loss of its specialized habitat. However, little is known about other potential contributing factors such as natural rarity or changes to connectivity following habitat fragmentation. We examined mitochondrial DNA (mtDNA) and genome-wide SNPs using specimens collected from breeding and migration sites across the range of the Bachman's warbler. We found no signals of strong population structuring across the breeding range of Bachman's warblers in both mtDNA and genome-wide SNPs. Thus, long-term population isolation did not appear to be a significant contributor to the extinction of the Bachman's warbler. Instead, our findings support the theory that Bachman's warblers underwent a rapid decline likely driven by habitat destruction, which may have been exacerbated by the natural rarity, habitat specificity and low genetic diversity of the species.

Dynamic interactions between host, pathogen and host-associated microbiome dictate infection outcomes. Pathogens including Batrachochytrium dendrobatidis (Bd) threaten global biodiversity, but conservation efforts are hindered by limited understanding of amphibian host, Bd and microbiome interactions. We conducted a vaccination and infection experiment using Eastern hellbender salamanders (Cryptobranchus alleganiensis alleganiensis) challenged with Bd to observe infection, skin microbial communities and gene expression of host skin, pathogen and microbiome throughout the experiment. Most animals survived high Bd loads regardless of their vaccination status and vaccination did not affect pathogen load, but host gene expression differed based on vaccination. Oral vaccination (exposure to killed Bd) stimulated immune gene upregulation while topically and sham-vaccinated animals did not significantly upregulate immune genes. In early infection, topically vaccinated animals upregulated immune genes but orally and sham-vaccinated animals downregulated immune genes. Bd increased pathogenicity-associated gene expression in late infection when Bd loads were highest. The microbiome was altered by Bd, but there was no correlation between anti-Bd microbe abundance or richness and pathogen burden. Our observations suggest that hellbenders initially generate a vigorous immune response to Bd, which is ineffective at controlling disease and is subsequently modulated. Interactions with antifungal skin microbiota did not influence disease progression.

While the effects of barriers to dispersal such as population declines, habitat fragmentation, and geographic distance have been well-documented in terrestrial wildlife, factors impeding the dispersal of highly vagile taxa such as seabirds are less well understood. The roseate tern (Sterna dougallii) is a globally distributed seabird species, but populations tend to be both fragmented and small, and the species is declining across most of its range. We evaluated structuring of roseate tern populations in the Northwestern Atlantic, the Caribbean, and the Azores using both microsatellite markers and single-nucleotide polymorphisms generated through targeted sequencing of Ultra-conserved Elements. For both marker types, we found significant genetic differentiation among all 3 populations and evidence for moderate contemporary unidirectional gene flow from the Caribbean to the Azores, but not between other populations. Within the Caribbean population, we found high rates of unidirectional migration from the Virgin Islands to Florida, potentially indicative of movement from source population to sink or an artifact of dispersal among other unsampled populations in the Caribbean region. These observations have significance for species persistence in the Atlantic, as our results indicate that loss of genetic diversity within populations is unlikely to be buffered by inflow of new alleles from other breeding populations.

In response to a growing need to foster ethical behavior within scientific societies, the American Ornithological Society's (AOS) professional ethics committee conducted a survey of members in spring 2021 to identify the primary challenges and ethical conduct concerns. The survey indicated that the AOS has a strong culture of professional ethics and highlighted areas needing improvement. Participants identified discrimination and lack of inclusivity (44%), scientific fraud and abuse in data and publications (35%), and sexual harassment (31%) as the highest potential risks for unethical behavior in our organization. Moreover, approximately one-third of respondents (34%) had personally witnessed or experienced unethical behavior as an AOS member. A smaller proportion (16%) felt pressure to compromise their work standards in ornithology. These findings are likely representative of broader patterns that professional, scientific societies face as they seek to provide safe, welcoming, and thoughtful environments for researchers to share their work, gain valuable feedback, and develop collaborations. The survey results also create a framework for workshops, training opportunities, and disseminating information within the AOS and, ideally, with the broader, international community of ornithologists. Lay Summary center dot Scientific societies have an obligation to foster ethical behavior of their membership. center dot The American Ornithological Society (AOS) has conducted a survey to help identify the primary challenges and ethical conduct concerns that face the ornithological community. center dot We disseminate the major findings of the survey here and discuss implications and future steps the AOS to address member concerns. center dot Overall, AOS ranked admirably regarding the overall ethical culture of our professional society, but the survey also identified room for improvement.

The unprecedented rise in the number of new and emerging infectious diseases in the last quarter century poses direct threats to human and wildlife health. The introduction to the Hawaiian archipelago of Plasmodium relictum and the mosquito vector that transmits the parasite has led to dramatic losses in endemic Hawaiian forest bird species. Understanding how mechanisms of disease immunity to avian malaria may evolve is critical as climate change facilitates increased disease transmission to high elevation habitats where malaria transmission has historically been low and the majority of the remaining extant Hawaiian forest bird species now reside. Here, we compare the transcriptomic profiles of highly susceptible Hawai'i 'amakihi (Chlorodrepanis virens) experimentally infected with P. relictum to those of uninfected control birds from a naïve high elevation population. We examined changes in gene expression profiles at different stages of infection to provide an in-depth characterization of the molecular pathways contributing to survival or mortality in these birds. We show that the timing and magnitude of the innate and adaptive immune response differed substantially between individuals that survived and those that succumbed to infection, and likely contributed to the observed variation in survival. These results lay the foundation for developing gene-based conservation strategies for Hawaiian honeycreepers by identifying candidate genes and cellular pathways involved in the pathogen response that correlate with a bird's ability to recover from malaria infection.