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Marsupials give birth to immunologically naïve young after a relatively short gestation period compared to eutherians. Consequently, the joey significantly relies on maternal protection, which is the focus of the present review. The milk and the pouch environment are essential contributors to maternal protection for the healthy development of joeys. In this review, we discuss bioactive components found in the marsupial pouch and milk that form cornerstones of maternal protection. These bioactive components include immune cells, immunoglobulins, the s100 family of calcium-binding proteins, lysozymes, whey proteins, antimicrobial peptides and other immune proteins. Furthermore, we investigated the possibility of the presence of plurifunctional components in milk and pouches that are potentially bioactive. These compounds include caseins, vitamins and minerals, oligosaccharides, lipids, and microRNAs. Where applicable, this review addresses variability in bioactive components during different phases of lactation, designed to fulfil the immunological needs of the growing pouch young. Yet there are numerous additional research opportunities to pursue, including uncovering novel bioactive components, investigating their mode of action, dynamics, stability, and ability to penetrate the gut epithelium ensuring systemic actions.

Amphibians represent a diverse group of tetrapods, marked by deep divergence times between their three systematic orders and families. Studying amphibian biology through the genomics lens increases our understanding of the features of this animal class and that of other terrestrial vertebrates. The need for amphibian genomic resources is more urgent than ever due to the increasing threats to this group. Amphibians are one of the most imperiled taxonomic groups, with approximately 41% of species threatened with extinction due to habitat loss, changes in land use patterns, disease, climate change, and their synergistic effects. Amphibian genomic resources have provided a better understanding of ontogenetic diversity, tissue regeneration, diverse life history and reproductive modes, anti-predator strategies, and resilience and adaptive responses. They also serve as essential models for studying broad genomic traits, such as evolutionary genome expansions and contractions, as they exhibit the widest range of genome sizes among all animal taxa and possess multiple mechanisms of genetic sex determination. Despite these features, genome sequencing of amphibians has significantly lagged behind that of other vertebrates, primarily due to the challenges of assembling their large, repeat-rich genomes and the relative lack of societal support. The emergence of long-read sequencing technologies, combined with advanced molecular and computational techniques that improve scaffolding and reduce computational workloads, is now making it possible to address some of these challenges. To promote and accelerate the production and use of amphibian genomics research through international coordination and collaboration, we launched the Amphibian Genomics Consortium (AGC, https://mvs.unimelb.edu.au/amphibian-genomics-consortium ) in early 2023. This burgeoning community already has more than 282 members from 41 countries. The AGC aims to leverage the diverse capabilities of its members to advance genomic resources for amphibians and bridge the implementation gap between biologists, bioinformaticians, and conservation practitioners. Here we evaluate the state of the field of amphibian genomics, highlight previous studies, present challenges to overcome, and call on the research and conservation communities to unite as part of the AGC to enable amphibian genomics research to "leap" to the next level., (© 2024. The Author(s).)

The swift parrot ( Lathamus discolor ) is a Critically Endangered migratory parrot that breeds in Tasmania and winters on the Australian mainland. Here we provide a reference genome assembly for the swift parrot. We sequence PacBio HiFi reads to create a high-quality reference assembly and identify a complete mitochondrial sequence. We also generate a reference transcriptome from five organs to inform genome annotation. The genome was 1.24 Gb in length and consisted of 847 contigs with a contig N50 of 18.97 Gb and L50 of 20 contigs. This study provides an annotated reference assembly and transcriptomic resources for the swift parrot to assist in future conservation genomic research., Competing Interests: No competing interests were disclosed., (Copyright: © 2024 Silver LW et al.)

Small, fragmented, or isolated populations are at risk of population decline due to fitness costs associated with inbreeding and genetic drift. The King Island scrubtit Acanthornis magna greeniana is a critically endangered subspecies of the nominate Tasmanian scrubtit A. m. magna, with an estimated population of <100 individuals persisting in three patches of swamp forest. The Tasmanian scrubtit is widespread in wet forests on mainland Tasmania. We sequenced the scrubtit genome using PacBio HiFi and undertook a population genomic study of the King Island and Tasmanian scrubtits using a double-digest restriction site-associated DNA (ddRAD) dataset of 5,239 SNP loci. The genome was 1.48 Gb long, comprising 1,518 contigs with an N50 of 7.715 Mb. King Island scrubtits formed one of four overall genetic clusters, but separated into three distinct subpopulations when analyzed independently of the Tasmanian scrubtit. Pairwise FST values were greater among the King Island scrubtit subpopulations than among most Tasmanian scrubtit subpopulations. Genetic diversity was lower and inbreeding coefficients were higher in the King Island scrubtit than all except one of the Tasmanian scrubtit subpopulations. We observed crown baldness in 8/15 King Island scrubtits, but 0/55 Tasmanian scrubtits. Six loci were significantly associated with baldness, including one within the DOCK11 gene which is linked to early feather development. Contemporary gene flow between King Island scrubtit subpopulations is unlikely, with further field monitoring required to quantify the fitness consequences of its small population size, low genetic diversity, and high inbreeding. Evidence-based conservation actions can then be implemented before the taxon goes extinct., (© The American Genetic Association. 2024.)

The legume albumin-1 gene family, arising after nodulation, encodes linear a- and b-chain peptides for nutrient storage and defense. Intriguingly, in one prominent legume, Clitoria ternatea, the b-chains are replaced by domains producing ultra-stable cyclic peptides called cyclotides. The mechanism of this gene hijacking is until now unknown. Cyclotides require recruitment of ligase-type asparaginyl endopeptidases (AEPs) for maturation (cyclization), necessitating co-evolution of two gene families. Here we compare a chromosome-level C. ternatea genome with grain legumes to reveal an 8 to 40-fold expansion of the albumin-1 gene family, enabling the additional loci to undergo diversification. Iterative rounds of albumin-1 duplication and diversification create four albumin-1 enriched genomic islands encoding cyclotides, where they are physically grouped by similar pI and net charge values. We identify an ancestral hydrolytic AEP that exhibits neofunctionalization and multiple duplication events to yield two ligase-type AEPs. We propose cyclotides arise by convergence in C. ternatea where their presence enhances defense from biotic attack, thus increasing fitness compared to lineages with linear b-chains and ultimately driving the replacement of b-chains with cyclotides., (© 2024. The Author(s).)

Ninu (greater bilby, Macrotis lagotis) are desert-dwelling, culturally and ecologically important marsupials. In collaboration with Indigenous rangers and conservation managers, we generated the Ninu chromosome-level genome assembly (3.66 Gbp) and genome sequences for the extinct Yallara (lesser bilby, Macrotis leucura). We developed and tested a scat single-nucleotide polymorphism panel to inform current and future conservation actions, undertake ecological assessments and improve our understanding of Ninu genetic diversity in managed and wild populations. We also assessed the beneficial impact of translocations in the metapopulation (N = 363 Ninu). Resequenced genomes (temperate Ninu, 6; semi-arid Ninu, 6; and Yallara, 4) revealed two major population crashes during global cooling events for both species and differences in Ninu genes involved in anatomical and metabolic pathways. Despite their 45-year captive history, Ninu have fewer long runs of homozygosity than other larger mammals, which may be attributable to their boom-bust life history. Here we investigated the unique Ninu biology using 12 tissue transcriptomes revealing expression of all 115 conserved eutherian chorioallantoic placentation genes in the uterus, an XY 1 Y 2 sex chromosome system and olfactory receptor gene expansions. Together, we demonstrate the holistic value of genomics in improving key conservation actions, understanding unique biological traits and developing tools for Indigenous rangers to monitor remote wild populations., (© 2024. The Author(s).)

The dugong (Dugong dugon) is a marine mammal widely distributed throughout the Indo-Pacific and the Red Sea, with a Vulnerable conservation status, and little is known about many of the more peripheral populations, some of which are thought to be close to extinction. We present a de novo high-quality genome assembly for the dugong from an individual belonging to the well-monitored Moreton Bay population in Queensland, Australia. Our assembly uses long-read PacBio HiFi sequencing and Omni-C data following the Vertebrate Genome Project pipeline to reach chromosome-level contiguity (24 chromosome-level scaffolds; 3.16 Gbp) and high completeness (97.9% complete BUSCOs). We observed relatively high genome-wide heterozygosity, which likely reflects historical population abundance before the last interglacial period, approximately 125,000 yr ago. Demographic inference suggests that dugong populations began declining as sea levels fell after the last interglacial period, likely a result of population fragmentation and habitat loss due to the exposure of seagrass meadows. We find no evidence for ongoing recent inbreeding in this individual. However, runs of homozygosity indicate some past inbreeding. Our draft genome assembly will enable range-wide assessments of genetic diversity and adaptation, facilitate effective management of dugong populations, and allow comparative genomics analyses including with other sirenians, the oldest marine mammal lineage., (© The American Genetic Association. 2024.)

The number of genome-level resources for non-model species continues to rapidly expand. However, frog species remain underrepresented, with up to 90% of frog genera having no genomic or transcriptomic data. Here, we assemble the first genomic and transcriptomic resources for the recently described southern stuttering frog (Mixophyes australis). The southern stuttering frog is ground-dwelling, inhabiting naturally vegetated riverbanks in south-eastern Australia. Using PacBio HiFi long-read sequencing and Hi-C scaffolding, we generated a high-quality genome assembly, with a scaffold N50 of 369.3 Mb and 95.1% of the genome contained in twelve scaffolds. Using this assembly, we identified the mitochondrial genome, and assembled six tissue-specific transcriptomes. We also bioinformatically characterised novel sequences of two families of antimicrobial peptides (AMPs) in the southern stuttering frog, the cathelicidins and β-defensins. While traditional peptidomic approaches to peptide discovery have typically identified one or two AMPs in a frog species from skin secretions, our bioinformatic approach discovered 12 cathelicidins and two β-defensins that were expressed in a range of tissues. We investigated the novelty of the peptides and found diverse predicted activities. Our bioinformatic approach highlights the benefits of multi-omics resources in peptide discovery and contributes valuable genomic resources in an under-represented taxon., (© 2024. The Author(s).)

Objectives: Respiratory syncytial virus (RSV) is the leading cause of acute lower respiratory tract infection in young children worldwide. RSV is increasingly associated with severe respiratory disease in people aged >65 years. The heterogeneous landscape of RSV in Australia and New Zealand makes generalisation of results from global studies to local contexts difficult. Given the changing landscape of RSV, we aimed to examine the existing literature on the burden of RSV disease and identify evidence gaps in Australia and New Zealand., Study Design: Scoping review., Methods: We designed a scoping review protocol and searched the Web of Science and Scopus databases for eligible peer-reviewed publications. Data from eligible studies were charted and summarised in tabular and narrative form., Results: Of the 153 eligible publications identified, 123 investigated RSV disease in a hospital setting and six in primary care. Only six studies reported the economic burden of disease, all of which estimated direct healthcare costs associated with treatment and/or hospitalisation; no studies quantified the indirect costs or costs to families., Conclusions: In this scoping review, we describe the effect of RSV disease in several high-risk populations, including children and adults. An improved understanding of the RSV burden of disease, both in primary care settings and economically, within the local context will assist with the implementation of preventative strategies, including vaccination programmes. Future studies to determine the true burden of RSV-associated morbidity, mortality and economic burden across the entire patient journey and among different healthcare settings will help prioritise emerging RSV therapeutics., (Copyright © 2023 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

The Kroombit tinkerfrog ( Taudactylus pleione ) is a stream-dwelling amphibian of the Myobatrachidae family. It is listed as Critically Endangered and is at high risk of extinction due to chytridiomycosis. Here, we provide the first genome assembly of the evolutionarily distinct Taudactylus genus. We sequenced PacBio HiFi reads to assemble a high-quality long-read genome and identified the mitochondrial genome. We also generated a global transcriptome from a tadpole to improve gene annotation. The genome was 5.52 Gb in length and consisted of 4,196 contigs with a contig N50 of 8.853 Mb and an L50 of 153. This study provides the first genomic resources for the Kroombit tinkerfrog to assist in future phylogenetic, environmental DNA, conservation breeding, and disease susceptibility studies., Competing Interests: No competing interests were disclosed., (Copyright: © 2023 Farquharson KA et al.)

Genomics can play important roles in biodiversity conservation, especially for Extinct-in-the-Wild species where genetic factors greatly influence risk of total extinction and probability of successful reintroductions. The Christmas Island blue-tailed skink (Cryptoblepharus egeriae) and Lister's gecko (Lepidodactylus listeri) are two endemic reptile species that went extinct in the wild shortly after the introduction of a predatory snake. After a decade of management, captive populations have expanded from 66 skinks and 43 geckos to several thousand individuals; however, little is known about patterns of genetic variation in these species. Here, we use PacBio HiFi long-read and Hi-C sequencing to generate highly contiguous reference genomes for both reptiles, including the XY chromosome pair in the skink. We then analyse patterns of genetic diversity to infer ancient demography and more recent histories of inbreeding. We observe high genome-wide heterozygosity in the skink (0.007 heterozygous sites per base-pair) and gecko (0.005), consistent with large historical population sizes. However, nearly 10% of the blue-tailed skink reference genome falls within long (>1 Mb) runs of homozygosity (ROH), resulting in homozygosity at all major histocompatibility complex (MHC) loci. In contrast, we detect a single ROH in Lister's gecko. We infer from the ROH lengths that related skinks may have established the captive populations. Despite a shared recent extinction in the wild, our results suggest important differences in these species' histories and implications for management. We show how reference genomes can contribute evolutionary and conservation insights, and we provide resources for future population-level and comparative genomic studies in reptiles., (© 2023 The Authors. Molecular Ecology Resources published by John Wiley & Sons Ltd.)

Conservation breeding programs aim to maintain 90% wild genetic diversity, but rarely assess functional diversity. Here, we compare both genome-wide and functional diversity (in over 500 genes) of Tasmanian devils ( Sarcophilus harrisii ) within the insurance metapopulation and across the species' range (64,519 km 2 ). Populations have declined by 80% since 1996 due to a contagious cancer, devil facial tumor disease (DFTD). However, predicted local extinctions have not occurred. Recent suggestions of selection for "resistance" alleles in the wild precipitated concerns that insurance population devils may be unsuitable for translocations. Using 830 wild samples collected at 31 locations between 2012 and 2021, and 553 insurance metapopulation devils, we show that the insurance metapopulation is representative of current wild genetic diversity. Allele frequencies at DFTD-associated loci were not substantially different between captive and wild devils. Methods presented here are valuable for others investigating evolutionary potential in threatened species, particularly ones under significant selective pressures., Competing Interests: The authors declare that they have no conflict of interest., (© 2022 The Author(s).)

Classic Mendelian inheritance is the bedrock of population genetics and underpins pedigree-based management of animal populations. However, assumptions of Mendelian inheritance might not be upheld in conservation breeding programmes if early viability selection occurs, even when efforts are made to equalise genetic contributions of breeders. To test this possibility, we investigated deviations from Mendelian proportions in a captive metapopulation of the endangered Tasmanian devil. This marsupial population is ideal for addressing evolutionary questions in conservation due to its large size, range of enclosure types (varying in environmental conditions), good genomic resources (which aid interpretation), and the species' biology. Devil mothers give birth to more offspring than they can nurse in the pouch, providing the potential for intense viability selection amongst embryos. We used data from 140 known sire-dam-offspring triads to isolate within-family selection from population-level mechanisms (such as mate choice or inbreeding), and compared observed offspring genotypes at 123 targeted SNPs to neutral (i.e., Mendelian) expectations. We found lower offspring heterozygosity than expected, and subtle patterns that varied across a gradient of management intensity from zoo-like enclosures to semi-wild environments for some loci. Meiotic drive or maternal-foetal incompatibilities are consistent with our results, although we cannot statistically confirm these mechanisms. We found some evidence that maternal genotype affects annual litter size, suggesting that family-level patterns are driven by differential offspring mortality before birth or during early development. Our results show that deviations from Mendelian inheritance can occur in conservation programmes, despite best-practice management to prevent selection., (© 2021 John Wiley & Sons Ltd.)

Conservation breeding programs such as zoos play a major role in preventing extinction, but their sustainability may be impeded by neutral and adaptive population genetic change. These changes are difficult to detect for a single species or context, and impact global conservation efforts. We analyse pedigree data from 15 vertebrate species - over 30,000 individuals - to examine offspring survival over generations of captive breeding. Even accounting for inbreeding, we find that the impacts of increasing generations in captivity are highly variable across species, with some showing substantial increases or decreases in offspring survival over generations. We find further differences between dam and sire effects in first- versus multi-generational analysis. Crucially, our multispecies analysis reveals that responses to captivity could not be predicted from species' evolutionary (phylogenetic) relationships. Even under best-practice captive management, generational fitness changes that cannot be explained by known processes (such as inbreeding depression), are occurring.

As species extinction rates increase, genomics provides a powerful tool to support intensive management of threatened species. We use the Tasmanian devil (Sarcophilus harrisii) to demonstrate how conservation genomics can be implemented in threatened species management. We conducted whole genome sequencing (WGS) of 25 individuals from the captive breeding programme and reduced-representation sequencing (RRS) of 98 founders of the same programme. A subset of the WGS samples was also sequenced by RRS, allowing us to directly compare genome-wide heterozygosity with estimates from RRS data. We found good congruence in interindividual variation and gene-ontology classifications between the two data sets, indicating that our RRS data reflect the genome well. We also attempted genome-wide association studies with both data sets (regarding breeding success), but the genomic data suffered from small sample size, while the RRS data suffered from lack of precision, highlighting a key trade-off in the design of conservation genomic research. Nevertheless, we identified a number of candidate genes that may be associated with variation in breeding success. Individual heterozygosity, as measured by WGS or RRS, was not associated with breeding success in captivity but was negatively associated with litter sizes of breeding females in the RRS data set. Our findings enable conservation managers to have confidence in RRS data while understanding its limitations, and provide avenues for further investigation into which processes underlie variation in breeding success in captive Tasmanian devils. We caution, however, that deep functional insights using RRS may be impaired by a lack of precision, especially when marker density is low., (© 2020 John Wiley & Sons Ltd.)

Incorporating mate choice into conservation breeding programs can improve reproduction and the retention of natural behaviors. However, different types of genetic-based mate choice can have varied consequences for genetic diversity management. As a result, it is important to examine mechanisms of mate choice in captivity to assess its costs and benefits. Most research in this area has focused on experimental pairing trials; however, this resource-intensive approach is not always feasible in captive settings and can interfere with other management constraints. We used generalized linear mixed models and permutation approaches to investigate overall breeding success in group-housed Tasmanian devils at three nonmutually exclusive mate choice hypotheses: (a) advantage of heterozygous individuals, (b) advantage of dissimilar mates, and (c) optimum genetic distance, using both 1,948 genome-wide SNPs and 12 MHC-linked microsatellites. The managed devil insurance population is the largest such breeding program in Australia and is known to have high variance in reproductive success. We found that nongenetic factors such as age were the best predictors of breeding success in a competitive breeding scenario, with younger females and older males being more successful. We found no evidence of mate choice under the hypotheses tested. Mate choice varies among species and across environments, so we advocate for more studies in realistic captive management contexts as experimental or wild studies may not apply. Conservation managers must weigh up the need to wait for adequate sample sizes to detect mate choice with the risk that genetic changes may occur during this time in captivity. Our study shows that examining and integrating mate choice into the captive management of species housed in realistic, semi-natural group-based contexts may be more difficult than previously considered., Competing Interests: None declared., (© 2020 The Authors. Evolutionary Applications published by John Wiley & Sons Ltd.)

Background: Recent advances in genomics have greatly increased research opportunities for non-model species. For wildlife, a growing availability of reference genomes means that population genetics is no longer restricted to a small set of anonymous loci. When used in conjunction with a reference genome, reduced-representation sequencing (RRS) provides a cost-effective method for obtaining reliable diversity information for population genetics. Many software tools have been developed to process RRS data, though few studies of non-model species incorporate genome alignment in calling loci. A commonly-used RRS analysis pipeline, Stacks, has this capacity and so it is timely to compare its utility with existing software originally designed for alignment and analysis of whole genome sequencing data. Here we examine population genetic inferences from two species for which reference-aligned reduced-representation data have been collected. Our two study species are a threatened Australian marsupial (Tasmanian devil Sarcophilus harrisii; declining population) and an Arctic-circle migrant bird (pink-footed goose Anser brachyrhynchus; expanding population). Analyses of these data are compared using Stacks versus two widely-used genomics packages, SAMtools and GATK. We also introduce a custom R script to improve the reliability of single nucleotide polymorphism (SNP) calls in all pipelines and conduct population genetic inferences for non-model species with reference genomes., Results: Although we identified orders of magnitude fewer SNPs in our devil dataset than for goose, we found remarkable symmetry between the two species in our assessment of software performance. For both datasets, all three methods were able to delineate population structure, even with varying numbers of loci. For both species, population structure inferences were influenced by the percent of missing data., Conclusions: For studies of non-model species with a reference genome, we recommend combining Stacks output with further filtering (as included in our R pipeline) for population genetic studies, paying particular attention to potential impact of missing data thresholds. We recognise SAMtools as a viable alternative for researchers more familiar with this software. We caution against the use of GATK in studies with limited computational resources or time.

Successfully establishing captive breeding programs is a priority across diverse industries to address food security, demand for ethical laboratory research animals, and prevent extinction. Differences in reproductive success due to birth origin may threaten the long-term sustainability of captive breeding. Our meta-analysis examining 115 effect sizes from 44 species of invertebrates, fish, birds, and mammals shows that, overall, captive-born animals have a 42% decreased odds of reproductive success in captivity compared to their wild-born counterparts. The largest effects are seen in commercial aquaculture, relative to conservation or laboratory settings, and offspring survival and offspring quality were the most sensitive traits. Although a somewhat weaker trend, reproductive success in conservation and laboratory research breeding programs is also in a negative direction for captive-born animals. Our study provides the foundation for future investigation of non-genetic and genetic drivers of change in captivity, and reveals areas for the urgent improvement of captive breeding.

Captive breeding programs are an increasingly popular tool to augment the conservation of threatened wild populations. Many programs keep detailed pedigrees, which are used to prescribe breeding targets to meet demographic and genetic goals. Annual breeding targets are based on previous productivity, but do not account for changes in reproductive success that may occur over generations in captivity and which may impair the ability of a program to meet its goals. We utilize a large studbook from the Tasmanian devil (Sarcophilus harrisii) captive breeding program to investigate biological, genetic, and environmental factors that affect variation in reproductive success among individuals and over generations of captive breeding. Reproductive success declined with increasing generations in captivity: wild-born females had a 56.5% chance of producing a litter compared to a 2.8% chance for generation 5 captive-born females (N = 182) and when they did, wild-born females produced more offspring (3.1 joeys, 95% CI: 2.76-3.38, compared to 2.7 joeys, 95% CI: 2.55-2.90, in captive-born females [N = 105]). Reproductive success also declined as dam age at first breeding increased. Our results reveal a conflict with the widely cited conservation strategy to limit opportunity for selection by extending generation length through delaying reproduction, as captive breeding programs that delay female breeding with this goal in mind risk reduced productivity. Our data demonstrate the benefit of pedigree analysis to identify biological processes that reveal crucial trade-offs with conservation best-practice., (© The American Genetic Association 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)