Mononuclear phagocytes (MNPs) encompass dendritic cells, monocytes, and macrophages (MoMac), which exhibit antimicrobial, homeostatic, and immunoregulatory functions. We integrated 178,651 MNPs from ...13 tissues across 41 datasets to generate a MNP single-cell RNA compendium (MNP-VERSE), a publicly available tool to map MNPs and define conserved gene signatures of MNP populations. Next, we generated a MoMac-focused compendium that revealed an array of specialized cell subsets widely distributed across multiple tissues. Specific pathological forms were expanded in cancer and inflammation. All neoplastic tissues contained conserved tumor-associated macrophage populations. In particular, we focused on IL4I1+CD274(PD-L1)+IDO1+ macrophages, which accumulated in the tumor periphery in a T cell-dependent manner via interferon-γ (IFN-γ) and CD40/CD40L-induced maturation from IFN-primed monocytes. IL4I1_Macs exhibited immunosuppressive characteristics through tryptophan degradation and promoted the entry of regulatory T cell into tumors. This integrated analysis provides a robust online-available platform for uniform annotation and dissection of specific macrophage functions in healthy and pathological states.
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•Cross-tissue integration of scRNA from monocytes and macrophages in health and disease•Conserved gene signatures of mononuclear phagocyte populations in human tissues•IL4I1+PD-L1+IDO1+ and TREM2+ TAM subsets accumulate in human tumors•IL4I1+PD-L1+IDO1+ TAM in the tumor periphery exhibit immunosuppressive characteristics
Mulder et al. integrate 178,651 human mononuclear phagocytes (MNPs) from 13 tissues across 41 datasets to generate a MNP single-cell RNA compendium (MNP-VERSE) that enables the definition of conserved gene signatures of MNP populations. This integrated approach provides a robust, online-available platform (https://gustaveroussy.github.io/FG-Lab/) for uniform annotation and dissection of specific macrophage functions in healthy and pathological states.
Brain macrophage populations include parenchymal microglia, border-associated macrophages, and recruited monocyte-derived cells; together, they control brain development and homeostasis but are also ...implicated in aging pathogenesis and neurodegeneration. The phenotypes, localization, and functions of each population in different contexts have yet to be resolved. We generated a murine brain myeloid scRNA-seq integration to systematically delineate brain macrophage populations. We show that the previously identified disease-associated microglia (DAM) population detected in murine Alzheimer’s disease models actually comprises two ontogenetically and functionally distinct cell lineages: embryonically derived triggering receptor expressed on myeloid cells 2 (TREM2)-dependent DAM expressing a neuroprotective signature and monocyte-derived TREM2-expressing disease inflammatory macrophages (DIMs) accumulating in the brain during aging. These two distinct populations appear to also be conserved in the human brain. Herein, we generate an ontogeny-resolved model of brain myeloid cell heterogeneity in development, homeostasis, and disease and identify cellular targets for the treatment of neurodegeneration.
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•M-Verse as a global cross-comparison of developing and adult murine brain macrophages•DAM correspond to a fetal-like reprogramming similar to Youth-Associated Microglia•DIMs appear during aging and increase in neurodegenerative diseases•DAM are embryonic derived, whereas DIMs are TREM2-independent monocyte derived
Through 6 scRNA-seq brain dataset integration, the authors generated a myeloid map called M-Verse to delineate macrophage population heterogeneity. M-Verse revealed two distinct macrophage populations expressing published disease-associated microglia (DAM) signature: embryonically derived TREM2-dependent DAM and monocyte-derived TREM2-independent disease inflammatory macrophages (DIMs).
Abstract Spatial omics data allow in-depth analysis of tissue architectures, opening new opportunities for biological discovery. In particular, imaging techniques offer single-cell resolutions, ...providing essential insights into cellular organizations and dynamics. Yet, the complexity of such data presents analytical challenges and demands substantial computing resources. Moreover, the proliferation of diverse spatial omics technologies, such as Xenium, MERSCOPE, CosMX in spatial-transcriptomics, and MACSima and PhenoCycler in multiplex imaging, hinders the generality of existing tools. We introduce Sopa ( https://github.com/gustaveroussy/sopa ), a technology-invariant, memory-efficient pipeline with a unified visualizer for all image-based spatial omics. Built upon the universal SpatialData framework, Sopa optimizes tasks like segmentation, transcript/channel aggregation, annotation, and geometric/spatial analysis. Its output includes user-friendly web reports and visualizer files, as well as comprehensive data files for in-depth analysis. Overall, Sopa represents a significant step toward unifying spatial data analysis, enabling a more comprehensive understanding of cellular interactions and tissue organization in biological systems.
Until recently many historical museum specimens were largely inaccessible to genomic inquiry, but high‐throughput sequencing (HTS) approaches have allowed researchers to successfully sequence genomic ...DNA from dried and fluid‐preserved museum specimens. In addition to preserved specimens, many museums contain large series of allozyme supernatant samples, but the amenability of these samples to HTS has not yet been assessed. Here, we compared the performance of a target‐capture approach using alternative sources of genomic DNA from 10 specimens of spring salamanders (Plethodontidae: Gyrinophilus porphyriticus) collected between 1985 and 1990: allozyme supernatants, allozyme homogenate pellets and formalin‐fixed tissues. We designed capture probes based on double‐digest restriction‐site associated sequencing (RADseq) derived loci from frozen blood samples available for seven of the specimens and assessed the success and consistency of capture and RADseq approaches. This study design enabled direct comparisons of data quality and potential biases among the different data sets for phylogenomic and population genomic analyses. We found that in phylogenetic analyses, all enrichment types for a given specimen clustered together. In principal component space all capture‐based samples clustered together, but RADseq samples did not cluster with corresponding capture‐based samples. Single nucleotide polymorphism calls were on average 18.3% different between enrichment types for a given individual, but these discrepancies were primarily due to differences in heterozygous/homozygous single nucleotide polymorphism calls. We demonstrate that both allozyme supernatant and formalin‐fixed samples can be successfully used for population genomic analyses and we discuss ways to identify and reduce biases associated with combining capture and RADseq data.
A period of isolation in allopatry typically precedes local adaptation and subsequent divergence among lineages. Alternatively, locally adapted phenotypes may arise and persist in the face of gene ...flow, resulting in strong correlations between ecologically‐relevant phenotypic variation and corresponding environmental gradients. Quantifying genetic, ecological, and phenotypic divergence in such lineages can provide insights into the abiotic and biotic mechanisms that structure populations and drive the accumulation of phenotypic and taxonomic diversity. Low‐vagility organisms whose distributions span ephemeral geographic barriers present the ideal evolutionary context within which to address these questions. Here, we combine genetic (mtDNA and genome‐wide SNPs) and phenotypic data to investigate the divergence history of caecilians (Amphibia: Gymnophiona) endemic to the oceanic island of São Tomé in the Gulf of Guinea archipelago. Consistent with a previous mtDNA study, we find two phenotypically and genetically distinct lineages that occur along a north‐to‐south axis with extensive admixture in the centre of the island. Demographic modelling supports divergence in allopatry (~300 kya) followed by secondary contact (~95 kya). Consequently, in contrast to a morphological study that interpreted latitudinal phenotypic variation in these caecilians as a cline within a single widespread species, our analyses suggest a history of allopatric lineage divergence and subsequent hybridization that may have blurred species boundaries. We propose that late Pleistocene volcanic activity favoured allopatric divergence between these lineages with local adaptation to climate maintaining a stable hybrid zone in the centre of São Tomé Island. Our study joins a growing number of systems demonstrating lineage divergence on volcanic islands with stark environmental transitions across small geographic distances.
Resumo
Um período de isolamento em alopatria geralmente precede adaptação local e divergência subsequente entre linhagens evolutivas. Alternativamente, fenótipos adaptados localmente podem surgir e persistir apesar de fluxo gênico, resultando em fortes correlações entre variação fenotípica ecologicamente relevante e os gradientes ambientais correspondentes. Quantificar divergência genética, ecológica e fenotípica em tais linhagens pode ajudar a clarificar os mecanismos abióticos e bióticos que estruturam as populações e levam ao acúmulo de diversidade fenotípica e taxonômica. Organismos de baixa vagilidade, cujas áreas de distribuição incluem barreiras geográficas efêmeras, representam um contexto evolutivo ideal para abordar essas questões. Neste estudo, combinamos dados genéticos (mtDNA e SNPs genômicos) e fenotípicos para investigar a história de divergência de cecílias endêmicas da ilha oceânica de São Tomé, no arquipélago do Golfo da Guiné. Consistentemente com um estudo anterior de mtDNA, encontramos duas linhagens fenotipicamente e geneticamente distintas que ocorrem ao longo de um eixo norte‐sul, com extensa mistura genética no centro da ilha. Modelagem demográfica suportou um cenário de divergência em alopatria (~ 300 mil anos atrás) seguida de contato secundário (~ 95 mil anos atrás). Ao contrário de um estudo morfológico que interpretou a variação fenotípica latitudinal nessas cecílias como uma clina dentro de uma única espécie amplamente difundida, nossas análises sugerem uma história de divergência de linhagens em alopatria e subsequente hibridização que pode ter confundido os limites das espécies. Propomos que atividade vulcânica durante o Pleistoceno tardio favoreceu divergência alopátrica entre essas linhagens, com adaptação local ao clima mantendo uma zona híbrida estável no centro da Ilha de São Tomé. Nosso estudo se une a um número crescente de sistemas que demonstram divergência entre linhagens em ilhas vulcânicas com transições ambientais marcantes ao longo de distâncias geográficas curtas.
The amphibian disease chytridiomycosis, caused by the fungus
Batrachochytrium dendrobatidis
(
Bd
), has contributed to the decline of Chiricahua leopard frogs (
Rana chiricahuensis
), a federally ...threatened species native to the Southwestern United States. We characterized immunogenetic variability in
R. chiricahuensis
by sequencing an expressed Major Histocompatibility Complex (MHC) class IIβ gene across 13 natural populations in Arizona, USA, as well as 283 individuals that were captive reared from two egg masses. We recovered a total of five class IIβ MHC alleles compared to 84 alleles previously characterized in eight natural populations of the Arizona congener
R. yavapaiensis
, demonstrating reduced MHC diversity in
R. chiricahuensis
. One allele was fixed in five populations but none of the
R. chiricahuensis
alleles were closely related to
R. yavapaiensis
allele Q, which is significantly associated with chytridiomycosis resistance in laboratory trials. Nine of 13
R. chiricahuensis
population localities were
Bd
positive, and bearing allele RachDRB*04 was the best genetic predictor of an individual being infected with
Bd
. A total of three class IIβ alleles were recovered from captive reared individuals, which were released to two natural population localities followed by recapture surveys to assess MHC-based survival over winter, the time when chytridiomycosis outbreaks are most severe. At one site, all released animals were fixed for a single allele and MHC-based survival could not be assessed. At the second site, fewer than half of the released but all of the recaptured individuals were homozygous for RachDRB*05, indicating that MHC genotype is important in determining
Bd
survival under natural field conditions. We conclude that the limited MHC variation in
R. chiricahuensis
is likely the consequence rather than the cause of natural selection favoring alleles that promote survival in the face of
Bd
. Our study highlights that preserving even low levels of functional genetic variation may be essential for population persistence, and that local disease adaptation may present as a reduction in genetic diversity. These finding also suggest that for populations that have declined due to a specific infectious pathogen, MHC-based genetically-informed reintroduction approaches may enhance species recovery efforts.
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•We identified five independent transitions to pueriparity in the genus Salamandra.•Transitions to pueriparity occurred at different evolutionary timescales.•We confirm pueriparity as ...the prevailing reproductive mode in S. s. bernardezi.•Introgression is common, but reproductive mode follows nuclear history.•Exome-capture and previously published RAD-data reveal similar evolutionary histories.
The ability to bear live offspring, viviparity, has evolved multiple times across the tree of life and is a remarkable adaptation with profound life-history and ecological implications. Within amphibians the ancestral reproductive mode is oviparity followed by a larval life stage, but viviparity has evolved independently in all three amphibian orders. Two types of viviparous reproduction can be distinguished in amphibians; larviparity and pueriparity. Larviparous amphibians deliver larvae into nearby ponds and streams, while pueriparous amphibians deliver fully developed juveniles and thus do not require waterbodies for reproduction. Among amphibians, the salamander genus Salamandra is remarkable as it exhibits both inter- and intraspecific variation in the occurrence of larviparity and pueriparity. While the evolutionary relationships among Salamandra lineages have been the focus of several recent studies, our understanding of how often and when transitions between modes occurred is still incomplete. Furthermore, in species with intraspecific variation, the reproductive mode of a given population can only be confirmed by direct observation of births and thus the prevalence of pueriparous populations is also incompletely documented. We used sequence capture to obtain 1,326 loci from 94 individuals from across the geographic range of the genus, focusing on potential reproductive mode transition zones. We also report additional direct observations of pueriparous births for 20 new locations and multiple lineages. We identify at least five independent transitions from the ancestral mode of larviparity to pueriparity among and within species, occurring at different evolutionary timescales ranging from the Pliocene to the Holocene. Four of these transitions occurred within species. Based on a distinct set of markers and analyses, we also confirm previous findings of introgression between species and the need for taxonomic revisions in the genus. We discuss the implications of our findings with respect to the evolution of this complex trait, and the potential of using five independent convergent transitions for further studies on the ecological context in which pueriparity evolves and the genetic architecture of this specialized reproductive mode.
Effective conservation and management of pond‐breeding amphibians depends on the accurate estimation of population structure, demographic parameters, and the influence of landscape features on ...breeding‐site connectivity. Population‐level studies of pond‐breeding amphibians typically sample larval life stages because they are easily captured and can be sampled nondestructively. These studies often identify high levels of relatedness between individuals from the same pond, which can be exacerbated by sampling the larval stage. Yet, the effect of these related individuals on population genetic studies using genomic data is not yet fully understood. Here, we assess the effect of within‐pond relatedness on population and landscape genetic analyses by focusing on the barred tiger salamanders (Ambystoma mavortium) from the Nebraska Sandhills. Utilizing genome‐wide SNPs generated using a double‐digest RADseq approach, we conducted standard population and landscape genetic analyses using datasets with and without siblings. We found that reduced sample sizes influenced parameter estimates more than the inclusion of siblings, but that within‐pond relatedness led to the inference of spurious population structure when analyses depended on allele frequencies. Our landscape genetic analyses also supported different models across datasets depending on the spatial resolution analyzed. We recommend that future studies not only test for relatedness among larval samples but also remove siblings before conducting population or landscape genetic analyses. We also recommend alternative sampling strategies to reduce sampling siblings before sequencing takes place. Biases introduced by unknowingly including siblings can have significant implications for population and landscape genetic analyses, and in turn, for species conservation strategies and outcomes.
We use SNP data from the pond‐breeding amphibian Ambystoma mavortium generated using a ddRADseq approach to test the effect of within‐pond relatedness on population and landscape genetic analyses. We find that both the inclusion of siblings, but also reduced sample sizes following sibling removal, affected analyses. In particular, we find that analyses dependent on allele frequencies are especially susceptible to sibling‐induced biases and discuss the implications of these findings to future studies.
•Target enrichment can directly sequence host-associated fungal pathogens.•Fungal pathogen load is positively associated with enrichment success.•Resulting genetic data can identify phylogenetic ...placements of fastidious strains.•Salamander at US zoo was infected with Asian-origin Batrachochytrium strain.•Method can be applied to other unculturable fungal samples to identify cryptic strains.
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.
Due to their limited geographic distributions and specialized ecologies, cave species are often highly endemic and can be especially vulnerable to habitat degradation within and surrounding the cave ...systems they inhabit. We investigated the evolutionary history of the West Virginia Spring Salamander (
Gyrinophilus subterraneus)
, estimated the population trend from historic and current survey data, and assessed the current potential for water quality threats to the cave habitat. Our genomic data (mtDNA sequence and ddRADseq-derived SNPs) reveal two, distinct evolutionary lineages within General Davis Cave corresponding to
G. subterraneus
and its widely distributed sister species,
Gyrinophilus porphyriticus
, that are also differentiable based on morphological traits. Genomic models of evolutionary history strongly support asymmetric and continuous gene flow between the two lineages, and hybrid classification analyses identify only parental and first generation cross (F1) progeny. Collectively, these results point to a rare case of sympatric speciation occurring within the cave, leading to strong support for continuing to recognize
G. subterraneus
as a distinct and unique species. Due to its specialized habitat requirements, the complete distribution of
G. subterraneus
is unresolved, but using survey data in its type locality (and currently the only known occupied site), we find that the population within General Davis Cave has possibly declined over the last 45 years. Finally, our measures of cave and surface stream water quality did not reveal evidence of water quality impairment and provide important baselines for future monitoring. In addition, our unexpected finding of a hybrid zone and partial reproductive isolation between
G. subterraneus
and
G. porphyriticus
warrants further attention to better understand the evolutionary and conservation implications of occasional hybridization between the species.
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