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Invasive Burmese pythons (Python bivittatus) in the Everglades of Florida, United States, have drastically reduced populations of mammals, yet populations of some rodents appear unaffected by the invasion. To understand this pattern, we radio-tagged cotton rats (Sigmodon hispidus) in areas of high and low python occurrence densities (hereafter occurrence) and quantified the effects of python occurrence, seasonality, and sex on their survival and cause-specific mortality. Cotton rat survival was not influenced by difference in python occurrence (hazard ratio = 1.32, 95% CI = 0.77–2.26, P = 0.30). However, cotton rats were at greater risk from mortalities caused by mammals in areas of low python occurrence. In areas with elevated python occurrence, we attributed most cotton rat mortalities to birds of prey (48.6%) and reptiles (non-python = 24.3%, python = 16.2%). Where python occurrence was relatively low, we attributed cotton rat mortalities to native reptilian (28.6%), avian (35.7%), and mammalian predators (35.7%) with no python-related deaths. In total, pythons were responsible for 11.8% of all cotton rat mortalities. Finding no difference in the survival of cotton rats, despite differences in the causative agents of mortality, suggests that predation pressure from an invasive predator was compensatory for cotton rat population dynamics. This type of compensatory mortality is common for small mammals and helps explain why mammal communities in python-invaded portions of the Greater Everglades Ecosystem are increasingly dominated by cotton rats and other rodents.

Convergent adaptation to the same environment by multiple lineages frequently involves rapid evolutionary change at the same genes, implicating these genes as important for environmental adaptation. Such adaptive molecular changes may yield either change or loss of protein function; loss of function can eliminate newly deleterious proteins or reduce energy necessary for protein production. We previously found a striking case of recurrent pseudogenization of the Paraoxonase 1 (Pon1) gene among aquatic mammal lineages—Pon1 became a pseudogene with genetic lesions, such as stop codons and frameshifts, at least four times independently in aquatic and semiaquatic mammals. Here, we assess the landscape and pace of pseudogenization by studying Pon1 sequences, expression levels, and enzymatic activity across four aquatic and semiaquatic mammal lineages: pinnipeds, cetaceans, otters, and beavers. We observe in beavers and pinnipeds an unexpected reduction in expression of Pon3, a paralog with similar expression patterns but different substrate preferences. Ultimately, in all lineages with aquatic/semiaquatic members, we find that preceding any coding-level pseudogenization events in Pon1, there is a drastic decrease in expression, followed by relaxed selection, thus allowing accumulation of disrupting mutations. The recurrent loss of Pon1 function in aquatic/semiaquatic lineages is consistent with a benefit to Pon1 functional loss in aquatic environments. Accordingly, we examine diving and dietary traits across pinniped species as potential driving forces of Pon1 functional loss. We find that loss is best associated with diving activity and likely results from changes in selective pressures associated with hypoxia and hypoxia-induced inflammation.

Invasive alien species (IAS) are a rising threat to biodiversity, national security, and regional economies, with impacts in the hundreds of billions of U.S. dollars annually. Proactive or predictive approaches guided by scientific knowledge are essential to keeping pace with growing impacts of invasions under climate change. Although the rapid development of diverse technologies and approaches has produced tools with the potential to greatly accelerate invasion research and management, innovation has far outpaced implementation and coordination. Technological and methodological syntheses are urgently needed to close the growing implementation gap and facilitate interdisciplinary collaboration and synergy among evolving disciplines. A broad review is necessary to demonstrate the utility and relevance of work in diverse fields to generate actionable science for the ongoing invasion crisis. Here, we review such advances in relevant fields including remote sensing, epidemiology, big data analytics, environmental DNA (eDNA) sampling, genomics, and others, and present a generalized framework for distilling existing and emerging data into products for proactive IAS research and management. This integrated workflow provides a pathway for scientists and practitioners in diverse disciplines to contribute to applied invasion biology in a coordinated, synergistic, and scalable manner.

The present study aimed to investigate the diet of African manatees in Cameroon to better inform conservation decisions within protected areas. A large knowledge gap on diet and seasonal changes in forage availability limits the ability to develop informed local management plans for the African manatee in much of its range. This research took place in the Sanaga River Watershed, which includes two protected areas in the Littoral Region of Cameroon: the Douala‐Edea National Park and the Lake Ossa Wildlife Reserve. We analyzed 113 manatee fecal samples and surveyed shoreline emergent and submerged vegetation within the Sanaga River Watershed. We used microhistological analyses to determine the relative contribution of each plant species to African manatee diets and compared across locations and across seasons (wet vs. dry season). We found that the shoreline vegetation is diverse with over 160 plant species, unevenly distributed across space and season, and dominated by emergent vegetation mostly represented by the antelope grass (Echinochloa pyramidalis). We recorded a total of 36 plant species from fecal samples with a spatial and temporal distribution mostly reflecting that of the corresponding shoreline vegetation. African manatees appear to be primarily opportunistically feeding on available vegetation across the seasons and habitat. This work documents the current, but changing, state of plant availability in the Sanaga River Watershed and reports the African manatee diet in Cameroon for the first time. This information can play a critical role in successfully managing the species and these protected areas. If we wish to protect the African manatee and the aquatic ecosystems within the Sanaga River Watershed, we must understand how forage availability changes over time, especially as its waters become nutrient enriched, eutrophic, and exposed to invasive species of plants in a changing world., Due to a large knowledge gap on diet and seasonal changes in forage availability, this study aims to investigate the diet of threatened African manatees to better inform conservation decisions within protected areas of Cameroon. We analyzed 113 manatee fecal samples and surveyed shoreline emergent and submerged vegetation within the Sanaga River watershed and found that African manatees appear to be primarily opportunistically feeding on available vegetation across the seasons and habitat. If we wish to protect the African manatee and the aquatic ecosystems within the Sanaga River watershed, we must understand how forage availability changes over time, especially as its waters become nutrient enriched, eutrophic, and exposed to invasive species of plants in a changing world.

Genetic diversity among and within populations of all species is necessary for people and nature to survive and thrive in a changing world. Over the past three years, commitments for conserving genetic diversity have become more ambitious and specific under the Convention on Biological Diversity's (CBD) draft post-2020 global biodiversity framework (GBF). This Perspective article comments on how goals and targets of the GBF have evolved, the improvements that are still needed, lessons learned from this process, and connections between goals and targets and the actions and reporting that will be needed to maintain, protect, manage and monitor genetic diversity. It is possible and necessary that the GBF strives to maintain genetic diversity within and among populations of all species, to restore genetic connectivity, and to develop national genetic conservation strategies, and to report on these using proposed, feasible indicators. ispartof: CONSERVATION GENETICS vol:24 issue:2 pages:181-191 ispartof: location:Netherlands status: published

Non-invasively collected faecal samples are an alternative source of DNA to tissues, that may be used in genetic studies of wildlife when direct sampling of animals is difficult. Although several faecal DNA extraction methods exist, their efficacy varies between species. Previous attempts to amplify mitochondrial DNA (mtDNA) markers from faeces of wild dugongs have met with limited success and nuclear markers (microsatellites) have been unsuccessful. This study aimed to develop a new tool for sampling both mtDNA and nuclear DNA (nDNA) from dugong faeces by modifying approaches used in studies of other large herbivores. First, amplification success of genetic markers from dugong faeces was compared between an established QIAamp and a newly developed DNA extraction method. Faecal DNA extracted using a new ‘High Volume-CTAB-PCI’ (HV-CTAB-PCI) method was found to achieve comparable amplification results to extraction of dugong skin DNA. As most prevailing practices advocate sampling from the outer surface of a stool to maximise capture of sloughed intestinal cells, this study compared amplification success of mtDNA between the outer and inner layers of faeces, but no difference in amplification was found. Assessment of the impacts of faecal age or degradation on extraction, however, demonstrated that fresher faeces with shorter duration of environmental (seawater) exposure amplified mtDNA and nDNA better than eroded scats. Using the HV-CTAB-PCI method, nDNA was successfully amplified for the first time from dugong faeces. This novel DNA extraction protocol offers a new tool that will facilitate genetic studies of dugongs and other large and cryptic marine herbivores in remote locations.

The goal of the Bio-ICE Task Team, convened from July 2020 - January 2022, was to advance the integration of biological observations from local, regional, and federal sources using best practices to inform national needs and ultimately feed into GOOS data visualizations and reporting. Using the GOOS marine mammals and corals EOVs as examples, the Task Team sought to: 1. Reconcile U.S. federal collection of these variables with GOOS EOVs and the Group on Earth Observations’ EBVs, identifying synergies in spatial and temporal observing requirements and with existing observation infrastructure and delivery, including use of documented best practices and standards. 2. Identify and improve pathways for data flow for observations of these variables from the IOOS Regional Associations and federal sources into common data portals. This included a focus on identifying and implementing best practices for standardized data collection and delivery adhering to the FAIR and CARE data principles, as appropriate. The Task Team effort focused on corals and marine mammals to ensure the work of the task team was achievable within the time frame. These two EOVs were selected because of their importance to multiple stakeholders, as well as offering opportunities to tie-in to several critical U.S. priorities in ocean science. While the team chose this focus, it acknowledged that there are activities and communities working to advance the other biology and ecosystem essential variables in ways that might be similar to or synergistic with the task team, such as SCOR Working Group 158: Coordinated Global Research Assessment of Seagrass System (C-GRASS), global efforts to coordinate ocean sound monitoring and data, GOOS BioEco panel community workshops and ongoing panel activities, and other efforts worth tracking or considering for future engagement with IOOC.

Marine mammals are wide-ranging, relatively long-lived organisms that play a crucial role in maintaining healthy ocean ecosystems. Often referred to as ecosystem engineers and sentinel species in marine ecosystems, these charismatic megafauna feed at a variety of trophic levels, affecting food web dynamics and cycling of chemicals and nutrients in the water column as well as in benthic habitats, both nearshore and in the deep ocean. An understanding of their abundance and distribution is an essential starting point for evaluating their role in ocean ecosystems. Accordingly, marine mammals have been included among key variables to monitor in ocean observing systems, from core variablesfor the U.S. Integrated Ocean Observing System (IOOS) to an Essential Ocean Variable (EOV)for the Global Ocean Observing System (GOOS). They also contribute to several Essential Biodiversity Variables (EBVs)for the Group on Earth Observations Biodiversity Observation Network (GEO BON). Further, evaluation of the health of marine mammal populations will help deliver societal benefits by contributing to the UN Decade of Ocean Science for Sustainable Development; informing reporting activities such as the World Ocean Assessment; and supporting achievement of Sustainable Development Goal 14, the post-2020 framework for the Convention for Biological Diversity, and a new treaty for conservation and sustainable use of marine biodiversity beyond national jurisdiction. The goal of this Interagency Ocean Observation Committee task team was to advance the integration of existing biological observations from local, regional, and federal sources using best practices to inform national needs and ultimately feed seamlessly into the Global Ocean Observing System, as appropriate. To accomplish this goal the subgroup focused on marine mammals to: 1. Reconcile the IOOS core biological variables with GOOS EOVs and the GEO BON EBVs, identifying where there are clear synergies in terms of spatial and temporal observing requirements and existing observation infrastructure and delivery including best practices/standards. 2. Identify and improve pathways for data flow for observations of these variables from Federal sources, such as the stock assessments conducted by NMFS and FWS, into IOOS with a focus on identifying and implementing best practices surrounding standardized data collection and delivery adhering to the FAIR and CAREdata principles, as appropriate.

Genome-wide evaluations of genetic diversity and population structure are important for informing management and conservation of trailing-edge populations. North American moose (Alces alces) are declining along portions of the southern edge of their range due to disease, species interactions, and marginal habitat, all of which may be exacerbated by climate change. We employed a genotyping by sequencing (GBS) approach in an effort to collect baseline information on the genetic variation of moose inhabiting the species’ southern range periphery in the contiguous United States. We identified 1920 single nucleotide polymorphisms (SNPs) from 155 moose representing three subspecies from five states: A. a. americana (New Hampshire), A. a. andersoni (Minnesota), and A. a. shirasi (Idaho, Montana, and Wyoming). Molecular analyses supported three geographically isolated clusters, congruent with currently recognized subspecies. Additionally, while moderately low genetic diversity was observed, there was little evidence of inbreeding. Results also indicated > 20% shared ancestry proportions between A. a. shirasi samples from northern Montana and A. a. andersoni samples from Minnesota, indicating a putative hybrid zone warranting further investigation. GBS has proven to be a simple and effective method for genome-wide SNP discovery in moose and provides robust data for informing herd management and conservation priorities. With increasing disease, predation, and climate related pressure on range edge moose populations in the United States, the use of SNP data to identify gene flow between subspecies may prove a powerful tool for moose management and recovery, particularly if hybrid moose are more able to adapt.

The red lionfish Pterois volitans is a successful invasive predator across the western North Atlantic, Caribbean, and Gulf of Mexico. The southeast coast of Florida (USA) has been identified as the original introduction location, but genetic analyses including Florida lionfish have yet to investigate introduction scenarios. Here, we assessed the potential lionfish invasion pathways using 1795 sequences from previously published mitochondrial D-loop sequences (n = 1558) and new samples (n = 237) from 6 locations: The Bahamas, Florida Keys, northwest Florida, North Carolina, Panamá, and southeast Florida. None of the assessed Florida lionfish (n = 394) contained the H05-H09 D-loop haplotypes found in The Bahamas, North Carolina, and Bermuda (the Northern Region), indicating that Florida was not the source for these haplotypes. Assessing the mitochondrial population structure, the Florida east coast lionfish grouped with the Caribbean/Gulf of Mexico, as opposed to the Northern Region. To further explore connectivity and invasion pathways, 14 nuclear microsatellite loci were multiplexed on lionfish collected from 15 locations (n = 394). As found in other nuclear lionfish studies, the analyses identified a lack of population structure likely due to founding effects and/or inbreeding in aquaculture brood stocks. Together, the significant haplotype differences and H01-H04 haplotypes refute Florida as the sole source of red lionfish introduction. The results of this study support alternative invasion scenarios, in which Florida was colonized as a secondary introduction site or by individuals from the Northern Region. Understanding invasive species’ population boundaries and dispersal patterns informs local control efforts and management planning for future invasive species introductions.

The rapidly emerging field of macrogenetics focuses on analysing publicly accessible genetic datasets from thousands of species to explore large-scale patterns and predictors of intraspecific genetic variation. Facilitated by advances in evolutionary biology, technology, data infrastructure, statistics and open science, macrogenetics addresses core evolutionary hypotheses (such as disentangling environmental and life-history effects on genetic variation) with a global focus. Yet, there are important, often overlooked, limitations to this approach and best practices need to be considered and adopted if macrogenetics is to continue its exciting trajectory and reach its full potential in fields such as biodiversity monitoring and conservation. Here, we review the history of this rapidly growing field, highlight knowledge gaps and future directions, and provide guidelines for further research.

Biodiversity underlies ecosystem resilience, ecosystem function, sustainable economies, and human well-being. Understanding how biodiversity sustains ecosystems under anthropogenic stressors and global environmental change will require new ways of deriving and applying biodiversity data. A major challenge is that biodiversity data and knowledge are scattered, biased, collected with numerous methods, and stored in inconsistent ways. The Group on Earth Observations Biodiversity Observation Network (GEO BON) has developed the Essential Biodiversity Variables (EBVs) as fundamental metrics to help aggregate, harmonize, and interpret biodiversity observation data from diverse sources. Mapping and analyzing EBVs can help to evaluate how aspects of biodiversity are distributed geographically and how they change over time. EBVs are also intended to serve as inputs and validation to forecast the status and trends of biodiversity, and to support policy and decision making. Here, we assess the feasibility of implementing Genetic Composition EBVs (Genetic EBVs), which are metrics of within-species genetic variation. We review and bring together numerous areas of the field of genetics and evaluate how each contributes to global and regional genetic biodiversity monitoring with respect to theory, sampling logistics, metadata, archiving, data aggregation, modeling, and technological advances. We propose four Genetic EBVs: (i) Genetic Diversity; (ii) Genetic Differentiation; (iii) Inbreeding; and (iv) Effective Population Size (Ne ). We rank Genetic EBVs according to their relevance, sensitivity to change, generalizability, scalability, feasibility and data availability. We outline the workflow for generating genetic data underlying the Genetic EBVs, and review advances and needs in archiving genetic composition data and metadata. We discuss how Genetic EBVs can be operationalized by visualizing EBVs in space and time across species and by forecasting Genetic EBVs beyond current observations using various modeling approaches. Our review then explores challenges of aggregation, standardization, and costs of operationalizing the Genetic EBVs, as well as future directions and opportunities to maximize their uptake globally in research and policy. The collection, annotation, and availability of genetic data has made major advances in the past decade, each of which contributes to the practical and standardized framework for large-scale genetic observation reporting. Rapid advances in DNA sequencing technology present new opportunities, but also challenges for operationalizing Genetic EBVs for biodiversity monitoring regionally and globally. With these advances, genetic composition monitoring is starting to be integrated into global conservation policy, which can help support the foundation of all biodiversity and species' long-term persistence in the face of environmental change. We conclude with a summary of concrete steps for researchers and policy makers for advancing operationalization of Genetic EBVs. The technical and analytical foundations of Genetic EBVs are well developed, and conservation practitioners should anticipate their increasing application as efforts emerge to scale up genetic biodiversity monitoring regionally and globally. ispartof: BIOLOGICAL REVIEWS vol:97 issue:4 pages:1511-1538 ispartof: location:England status: published

Invasive species surveillance programs can utilize environmental DNA sampling and analysis to provide information on the presence of invasive species. Wider utilization of eDNA techniques for invasive species surveillance may be warranted. This paper covers topics directed towards invasive species managers and eDNA practitioners working at the intersection of eDNA techniques and invasive species surveillance. It provides background information on the utility of eDNA for invasive species management and points to various examples of its use across federal and international programs. It provides information on 1) why an invasive species manager should consider using eDNA, 2) deciding if eDNA can help with the manager's surveillance needs, 3) important components to operational implementation, and 4) a high-level overview of the technical steps necessary for eDNA analysis. The goal of this paper is to assist invasive species managers in deciding if, when, and how to use eDNA for surveillance. If eDNA use is elected, the paper provides guidance on steps to ensure a clear understanding of the strengths and limitation of the methods and how results can be best utilized in the context of invasive species surveillance.

Background Environmental DNA (eDNA) analysis is increasingly being used to detect the presence and relative abundance of rare species, especially invasive or imperiled aquatic species. The rapid progress in the eDNA field has resulted in numerous studies impacting conservation and management actions. However, standardization of eDNA methods and reporting across the field is yet to be fully established, with one area being the calculation and interpretation of assay limit of detection (LOD) and limit of quantification (LOQ). Aims Here, we propose establishing consistent methods for determining and reporting of LOD and LOQ for single‐species quantitative PCR (qPCR) eDNA studies. Materials & Methods/ Results We utilize datasets from multiple cooperating laboratories to demonstrate both a discrete threshold approach and a curve‐fitting modeling approach for determining LODs and LOQs for eDNA qPCR assays. We also provide details of an R script developed and applied for the modeling method. Discussion/Conclusions Ultimately, standardization of how LOD and LOQ are determined, interpreted, and reported for eDNA assays will allow for more informed interpretation of assay results, more meaningful interlaboratory comparisons of experiments, and enhanced capacity for assessing the relative technical quality and performance of different eDNA qPCR assays.

A correction to this paper has been published: https://doi.org/10.1007/s10592-021-01376-9

Environmental DNA (eDNA) detection methods can complement traditional biomonitoring to yield new ecological insights in aquatic systems. However, the conceptual and methodological frameworks for aquatic eDNA detection and interpretation were developed primarily in freshwater environments and have not been well established for estuaries and marine environments that are by nature dynamic, turbid, and hydrologically complex. Environmental context and species life history are critical for successful application of eDNA methods, and the challenges associated with eDNA detection in estuaries were the subject of a symposium held at the University of California Davis on January 29, 2020 (https:// marinescie nce.ucdavis. edu/engagement/past-events/edna). Here, we elaborate upon topics addressed in the symposium to evaluate eDNA methods in the context of monitoring and biodiversity studies in estuaries. We first provide a concise overview of eDNA science and methods, and then examine the San Francisco Estuary (SFE) as a case study to illustrate how eDNA detection can complement traditional monitoring programs and provide regional guidance on future potential eDNA applications. Additionally, we offer recommendations for enhancing communication between eDNA scientists and natural resource managers, which is essential for integrating eDNA methods into existing monitoring programs. Our intent is to create a resource that is accessible to those outside the field of eDNA, especially managers, without oversimplifying the challenges or advantages of these methods., Estuaries and coasts, 45, ISSN:1559-2731, ISSN:1559-2723

Millette et al. (Ecology Letters, 2020, 23:55-67) reported no consistent worldwide anthropogenic effects on animal genetic diversity using repurposed mitochondrial sequences. We describe limitations to this study, some of which are common to other macrogenetic studies, that may lead to misinterpretations and unintended consequences for conservation.

Reproductive strategies are an essential component of invasion ecology that influence invasion success and rates of population growth. Burmese Pythons (Python bivittatus) are large constrictor snakes that were introduced to the Greater Everglades Ecosystem of southern Florida, USA, from Asia. Since their introduction, these giant constrictors have spread throughout wetlands of southern Florida while increasing in abundance and causing declines in the native species upon which they prey. Multiple paternity in reproduction could facilitate invasion success by increasing the genetic diversity produced within each reproductive event. We used Diversity Arrays Technology genome-wide genotyping to assess multiple paternity in the progeny of wild Burmese Pythons in Florida. We analyzed >4,000 single nucleotide polymorphisms from 153 neonates belonging to 4 clutches collected in southwestern Florida. Complementary hierarchical and K-means clustering analyses of the genetic distances within clutches revealed that three clutches were each fertilized by two sires, with a fourth fertilized by a single sire. The proportions of offspring attributable to each sire within multiple paternity clutches ranged from nearly even to highly skewed. Analysis of multivariate dispersion showed significantly increased genetic variability in the multiple paternity clutches. These results improve our understanding of the reproductive strategy and invasion potential of a giant constrictor with significant ecological impacts.

With increased usage of long-read sequencing technologies to perform transcriptome analyses, there becomes a greater need to evaluate different methodologies including library preparation, sequencing platform, and computational analysis tools. Here, we report the study design of a community effort called the Long-read RNA-Seq Genome Annotation Assessment Project (LRGASP) Consortium, whose goals are characterizing the strengths and remaining challenges in using long-read approaches to identify and quantify the transcriptomes of both model and non-model organisms. The LRGASP organizers have generated cDNA and direct RNA datasets in human, mouse, and manatee samples using different protocols followed by sequencing on Illumina, Pacific Biosciences, and Oxford Nanopore Technologies platforms. Participants will use the provided data to submit predictions for three challenges: transcript isoform detection with a high-quality genome, transcript isoform quantification, and de novo transcript isoform identification. Evaluators from different institutions will determine which pipelines have the highest accuracy for a variety of metrics using benchmarks that include spike-in synthetic transcripts, simulated data, and a set of undisclosed, manually curated transcripts by GENCODE. We also describe plans for experimental validation of predictions that are platform-specific and computational tool-specific. We believe that a community effort to evaluate long-read RNA-seq methods will help move the field toward a better consensus on the best approaches to use for transcriptome analyses.

Environmental DNA (eDNA) detection of invasive species can be used to inform management decisions by delimiting occupied ranges and estimating detection probabilities. Environmental DNA is shed into the environment through skin cells and bodily fluids and can be detected in water samples collected from lakes, rivers, and swamps. In south Florida, USA, invasive Burmese pythons (Python bivittatus) occupy much of the Greater Everglades in mostly inaccessible habitat and are credited with causing severe declines in populations of native species. Detection of Burmese pythons by traditional methods, including trapping and visual searching, has been largely ineffective, making eDNA a superior method to identify invaded areas. We adapted a quantitative PCR eDNA assay for use in droplet digital PCR, a state-of-the-art method that improves the precision and accuracy of eDNA detection and quantification. From August 2014 to October 2016, locations in and around Arthur R. Marshall Loxahatchee National Wildlife Refuge in southeast Florida were surveyed for Burmese python eDNA. Positive eDNA detections were made in each of the five sampling events, assessing a total of 399 samples, with moderate-to-high occurrence (ψ = 58–91%) and moderate detection (p = 38–70%) probabilities, potentially reduced by a high presence of PCR-inhibiting compounds in the water. The high occurrence rates and geographic distribution of the positive samples within Loxahatchee suggests a steady release of python eDNA from resident Burmese pythons and reduces support for transport of eDNA by boats or flowing water from the north. The first confirmed sighting of a Burmese python in the Refuge occurred in September 2016 after eDNA testing had indicated the presence of pythons. While the established population boundary is thought to be south of Loxahatchee, the eDNA detections indicate a northern range limit at, or north of, the refuge on the eastern side of the Florida peninsula. Our study demonstrates the utility of eDNA for determining more accurate range limits and expansion information for Burmese pythons, and provides a means for assessing control efforts.

To inform management and conservation decisions, environmental DNA (eDNA) methods are used to detect genetic material shed into the water by imperiled and invasive species. Methodological enhancements are needed to reduce filter clogging, PCR inhibition, and false-negative detections when eDNA is at low concentrations. In the first of three simple experiments, we sought to ameliorate filter clogging from particulates and organic material through a scaled-up, multi-filter protocol. We combined four filters in a 5 mL Phenol-Chloroform-Isoamyl (PCI) procedure to allow for larger volumes of water (~1 L) to be filtered rapidly. Increasing the filtered water volume by four times resulted in 4.4X the yield of target DNA. Next, inhibition from organic material can reduce or block eDNA detections in PCR-based assays. To remove inhibitory compounds retained during eDNA isolation, we tested three methods to chemically strip inhibitors from eDNA molecules. The use of CTAB as a short-term (5–8 day) storage buffer, followed by a PCI isolation, resulted in the highest eDNA yields. Finally, as opposed to a linear relationship among increasing concentrations of filtered genomic eDNA, we observed a sharp change between the lower (70–280 ng) and higher (420–560 ng) amounts. This may be important for effectively precipitating eDNA during protocol testing.

International agreements such as the Convention on Biological Diversity (CBD) have committed to conserve, and sustainably and equitably use, biodiversity. The CBD is a vital instrument for global conservation because it guides 195 countries and the European Union in setting priorities and allocating resources, and requires regular reporting on progress. However, the CBD and similar policy agreements have often neglected genetic diversity. This is a critical gap because genetic diversity underlies adaptation to environmental change and ecosystem resilience. Here we aim to inform future policy, monitoring, and reporting efforts focused on limiting biodiversity loss by conducting the largest yet evaluation of how Parties to the CBD report on genetic diversity. A large, globally representative sample of 114 CBD National Reports was examined to assess reported actions, progress, values and indicators related to genetic diversity. Although the importance of genetic diversity is recognized by most Parties to the CBD, genetic diversity targets mainly addressed variation within crops and livestock (a small fraction of all species). Reported actions to conserve genetic diversity primarily concerned ex situ facilities and legislation, rather than monitoring and in situ intervention. The most commonly reported status indicators are not well correlated to maintaining genetic diversity. Lastly, few reports mentioned genetic monitoring using DNA data, indigenous use and knowledge of genetic diversity, or development of strategies to conserve genetic diversity. We make several recommendations for the post-2020 CBD Biodiversity Framework, and similar efforts such as IPBES, to improve awareness, assessment, and monitoring of genetic diversity, and facilitate consistent and complete reporting in the future.

Florida manatees depend on freshwater environments as a source of drinking water and as warm-water refuges. These freshwater environments are in direct contact with human activities where glyphosate-based herbicides are being used. Glyphosate is the most used herbicide worldwide and it is intensively used in Florida as a sugarcane ripener and to control invasive aquatic plants. The objective of the present study was to determine the concentration of glyphosate and its breakdown product, aminomethylphosphonic acid (AMPA), in Florida manatee plasma and assess their exposure to manatees seeking a warm-water refuge in Crystal River (west central Florida), and in South Florida. We analyzed glyphosate’s and AMPA’s concentrations in Florida manatee plasma (n = 105) collected during 2009–2019 using HPLC-MS/MS. We sampled eight Florida water bodies between 2019 and 2020, three times a year: before, during and after the sugarcane harvest using grab samples and molecular imprinted passive Polar Organic Chemical Integrative Samplers (MIP-POCIS). Glyphosate was present in 55.8% of the sampled Florida manatees’ plasma. The concentration of glyphosate has significantly increased in Florida manatee samples from 2009 until 2019. Glyphosate and AMPA were ubiquitous in water bodies. The concentration of glyphosate and AMPA was higher in South Florida than in Crystal River, particularly before and during the sugarcane harvest when Florida manatees depend on warm water refuges. Based on our results, Florida manatees were chronically exposed to glyphosate and AMPA, during and beyond the glyphosate applications to sugarcane, possibly associated with multiple uses of glyphosate-based herbicides for other crops or to control aquatic weeds. This chronic exposure in Florida water bodies may have consequences for Florida manatees’ immune and renal systems which may further be compounded by other environmental exposures such as red tide or cold stress.

As an obligate salt marsh species, Seaside Sparrows (Ammospiza maritima) are vulnerable to numerous threats including climate change, coastal erosion, sea-level rise, and both natural and anthropogenic disasters. Of the 9 recognized subspecies, 2 are extinct and 1 is endangered. Previous genetic analyses of mitochondrial DNA (mtDNA) and microsatellite loci showed that current taxonomy does not accurately reflect underlying genetic diversity, with possible consequences for the distribution of conservation resources. To further inform Seaside Sparrow management, we comprehensively describe genetic structure among 24 range-wide sampling locations that include all extant subspecies. We inferred population structure from several thousand single-nucleotide polymorphisms collected from 272 individuals via restriction-site-associated DNA sequencing. Principal components, pairwise FST values, and clustering approaches suggest that Seaside Sparrows on the Atlantic and Gulf Coasts are distinct and consist of at least 5 genetic clusters: 1 in southern Texas, 1 ranging from Aransas County, Texas, to Mississippi; 1 in western Florida; and 2 or 3 genetic groups intermixed along a gradient on the Atlantic Coast. These genetic clusters are not consistent with current subspecies taxonomy and could be used as distinct population segments (DPSs) to inform the most efficient allocation of resources to Seaside Sparrow conservation. Our results regarding the endangered subspecies, A. m. mirabilis, from southern Florida are inconclusive due to low sample size, but indicate that it is distinct and may represent a sixth DPS. Based on our genetic results, we recommend additional song and morphometric analyses in western Florida and a closer study of the boundary between the breeding distributions of A. m. maritima and A. m. macgillivraii to ensure the proper identification of DPSs.

Cold stress syndrome (CSS) in the Florida manatee (Trichechus manatus latirostris) results in perturbations to many physiological pathways, often leading to further illness or death. In this study, we applied a non-targeted lipidomics approach with ultra-high performance liquid chromatography and high-resolution tandem mass spectrometry to characterize changes related to CSS in the lipidomic profiles of manatee plasma. Lipidomic analyses were conducted on healthy manatee (control) and cold-exposed manatee plasma samples with varying concentrations of Serum Amyloid A (SAA), an acute-phase protein that is associated with inflammatory disease. Control manatees (n = 10) were compared to all manatees exposed to cold temperatures (n = 17), and a subset of those manatees with SAA values > 120 μg/mL (n = 9). Increased SAA values were associated with higher levels of various acylcarnitine lipids, while several triacylglycerols and oxidized triacylglycerols were significantly lower in manatees with cold exposure. These identified lipids are critical molecules involved in the maintenance of energy homeostasis and could potentially be examined in conjunction with current physical parameters to characterize cold stress. The ability to detect such differences highlights the addition of lipidomics as a valuable tool in understanding cold stress and potentially other illnesses in manatees. Further investigation into the function of the altered lipids could greatly increase our understanding of lipid metabolism in physiologically stressed manatees as well as other marine mammals and inform future management recovery strategies.