Molecular Phylogenetics and Evolution

This investigation aimed at compiling all phylogenetic lineages within and around genus Cyanoboletus. The evolutionary inference obtained from the nuclear ribosomal genes internal transcribed spacer region (ITS) suggests that part of the species currently classified in Cyanoboletus belong in lineages separate from the genus, thus suggesting a narrower boundary that includes only the species that develop a strong staining reaction to touch and to air exposure of the context. The separate lineages are the monotypic Cupreoboletus genus and a few species that do not develop such reaction, which are part of a clade together with genera Cacaoporus and Acyanoboletus, thus broadening the concept of Cacaoporus to encompass all of them. The emerging 3C perspective of Cupreoboletus, Cacaoporus and Cyanoboletus offers a remarkably consistent morphological diagnosis, overcoming the problems of a too broad concept for Cyanoboletus. This work reveals that Boletus neotropicus, B. novae-zelandiae and B. sensibilis belong respectively in Cyanoboletus, Cacaoporus and Lanmaoa, and by studying multigene alignment concatenates it identifies lineages that probably represent undescribed species: at least four in Cacaoporus and at least five in Cyanoboletus. Diagnostic tables and dichotomic keys are presented by geographic region. The present work also includes a study of the phylogenetic position of Neoboletus flavosanguineus, a species once classified in Cyanoboletus. The complexity of assigning species epithets in some lineages is addressed, namely for the boundaries between Cacaoporus instabilis and Ca. fagaceophilus as well as the diversity under the names Cyanoboletus sinopulverulentus and Cy. pulverulentus. The overall picture of evolutionary lineages sets a framework for the choice of reference data that can provide, in future phylogenetic studies that involve the 3C, a balanced and efficient coverage. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=197 SRC="FIGDIR/small/724631v1_ufig1.gif" ALT="Figure 1"> View larger version (23K): org.highwire.dtl.DTLVardef@7f618corg.highwire.dtl.DTLVardef@dd6a14org.highwire.dtl.DTLVardef@5f7399org.highwire.dtl.DTLVardef@9e7443_HPS_FORMAT_FIGEXP M_FIG C_FIG

Forest trees pose numerous potential challenges to phylogenomic inference. Their large effective population sizes and relatively long generation times lead to deep allele coalescence and consequently incomplete lineage sorting (ILS), which biases inferences of divergence times toward older ages and introduces gene tree discordance. Deep phylogenetic divergences, reaching back into the Paleocene, introduce reference-mapping biases. Introgression--the movement of genes between lineages--may result in different phylogenies being inferred depending on which individuals are included in analysis, even if the plurality of the genome favors the divergence history unaffected by introgression. These factors influence phylogenetic inference across the Tree of Life but are particularly prevalent in forest trees. Oaks (Quercus) are notable for all three influences. In addition, our knowledge of the oak phylogeny is currently based strongly on restriction site associated DNA sequencing (RADseq) datasets published over the past decade, which may introduce additional sources of uncertainty. In this chapter, we analyze a 322-species RADseq dataset and genome resequencing data from across the genus to address sources of uncertainty in our understanding of the global oak phylogeny, which we hope will serve as a model for other research groups working on comparable woody plant groups.

The order Atheriniformes (silversides, rainbowfishes, and blue-eyes) is a globally distributed group of fishes with frequent evolutionary transitions between marine and freshwater ecosystems. However, understanding the tempo and mode of these transitions has been hampered by poor phylogenetic resolution and limited taxonomic sampling, particularly within the suborder Atherinoidei. We generated a phylogenomic dataset of 1,100 exon loci for 150 species to resolve interfamilial relationships and reconstruct the groups biogeographic history. We were also able to incorporate a large number of existing GenBank sequences, producing a phylogeny with 265 species sampled for at least some genetic data (67% of known species diversity). While the New World suborder Atherinopsidae is well-resolved, we found that the family Atherinidae is polyphyletic across all analyses. We propose a revised classification that restricts Atherinidae to the genus Atherina and recognizes Atherinomoridae and Craterocephalidae as separate families. Our biogeographic inferences using explicit geographic areas suggests more frequent marine-to-freshwater transitions than previously inferred with simplified binary (marine vs. freshwater) coding, and uncover habitat transitions where marine ancestors may have gone extinct. These results highlight how explicit geographic modeling can uncover marine ancestry erased by extinction, providing a robust phylogenetic framework for future evolutionary studies of Atheriniformes.

Target enrichment methods have provided unprecedented advances in phylogenomics. Targeting hundreds of conserved regions has proven to be a good tradeoff between cost and efficiency, while being useful for museomics and diversified non-model clades. Unfortunately, current methods used for identifying such regions involve high degrees of conservation within targeted elements, usually pushing researchers to rely on flanking data with little guarantee for homology. With a growing number of high quality genomes available throughout the Tree of Life emerges new opportunities to improve marker selection. In this study, we introduce GABBI, a new method for designing target capture probes by taking advantage of genome alignments, avoiding the selection of a single reference genome that can cause notable biases. We compare GABBI-derived markers to the most commonly used probe design method, PHYLUCE, at two taxonomic scales, the weevil superfamily Curculionoidea and the tribe Pachyrhynchini. At both taxonomic scales, results show that our new method allows identifying more variable loci that prove to be more phylogenetically resolutive than the PHYLUCE-derived ones. Doing so, we provide the first probe set specifically designed for weevils, targeting a wide set of 4,255 shared homologous regions, encouraging future research on systematics and macroevolution of one of the most diverse and economically important groups of insects. By providing GABBI as an automated and open-access pipeline, we hope to open new probe design opportunities to other taxonomic groups that face similar phylogenetic obstacles.

Phylogenetic network inference methods are increasingly used to detect hybridization and gene flow from genomic data, but their robustness to common sources of model violation remains poorly characterized. We conducted a simulation study to evaluate the effects of hidden paralogy and substitution rate variation on two widely used network inference methods: find_graphs from ADMIXTOOLS 2 and SNaQ. Using an eight-taxon species tree calibrated from an empirical reptile phylogeny, we simulated data under various levels of hidden paralogy (from none to strong) and three levels of rate variation (none, gene-specific, and lineage-specific). We found that hidden paralogy had limited impact on network inference under the conditions examined: both network methods correctly favored a tree without reticulation, and ASTRAL recovered the correct species tree every time. In contrast, lineage-specific rates severely biased find_graphs, inflating worst f-statistic residuals well beyond the standard acceptance threshold. SNaQ correctly selected a tree model almost always across all conditions, though its network with h = 1 reticulation displayed the true species tree with a lower probability under lineage-specific rates. We also show that the standard worst residuals threshold of 3 for find_graphs produces inflated type I error even without rate variation, and we recommend empirical calibration of this threshold within each study system.

The Dinaric karst of Croatia encompasses a network of over 10,000 caves and represents one of the worlds most important subterranean biodiversity hotspots. It is inhabited by remarkably diverse and often endemic species, including planarian flatworms, which are among the rarest macroinvertebrates encountered in cave habitats. Although the presence of cave planarians has long been known, no integrative research on this group has been conducted to date, and the evolutionary relationships between these animals and their surface water counterparts are currently unresolved. To address these gaps, we combined field sampling, phylogenetic analysis based on COI and 18S genes, and phenotypic characterization. Our results show that cave planariids in Croatia belong to at least three genera and are more widespread and diverse across both Croatia, and the broader Dinaric karst, than previously assumed. We increased the number of cave records in the Dinaric karst from 26 to 37 and documented cf. Atrioplanaria and Phagocata in Croatian caves for the first time. Phylogenetic reconstructions suggest numerous independent cave colonization events, including multiple instances within the genera Crenobia and cf. Atrioplanaria. Variation in pigmentation and eye reduction, both within and between populations, further reveal heterogeneous evolutionary trajectories of cave-associated phenotypes. The biogeographical patterns and high genetic diversity we report here point to a complex evolutionary history of planariids in the Dinarides. Our newly generated molecular phylogenies and systematic documentation of trait variability establish Planariidae as a valuable model for studying mechanisms underlying convergent evolution of pigment loss and eye reduction in cave environments.

Understanding how global biodiversity patterns arise is a central theme of biogeography, with contemporary theory recognising the roles of both dispersal and vicariance. Genera that are broadly distributed can provide important systems for disentangling the relative influence of these processes across evolutionary timescales. However, many lesser-studied groups, particularly those in the tropics, lack a densely sampled phylogeny which hinders robust inference of their evolutionary and biogeographic history. This study investigates the global diversification and systematics of the putative pantropical moth genus Parasa Moore (Lepidoptera: Limacodidae), with the aim of assessing the relative importance of dispersal and vicariance in shaping its distribution. Medium-coverage whole genome sequencing of specimens predominantly from museum collections were used to generate a globally sampled time-calibrated phylogeny of Parasa and associated genera (the Parasa-complex). Ancestral range estimation analyses were employed to infer geographical origins and possible dispersal times between bioregions. The Parasa-complex originated in Africa in the late Oligocene ([~]24 Ma) and, through a series of long-distance dispersal events during the early-mid Miocene, expanded into Asia ([~]23 Ma) and the Americas ([~]21 Ma). Across all regions, dispersal was the dominant process shaping present-day distributions, with a limited role of vicariance in some subregions. Phylogenetic analyses further demonstrated that Parasa is not monophyletic, with multiple independent lineages contributing to its apparent pantropical distribution. These findings highlight a central role of long-distance dispersal in generating certain global distributions. The results support a dynamic model of range evolution involving rapid Miocene dispersal and subsequent regional diversification. In addition, the non-monophyly of Parasa requires substantial taxonomic revision, underscoring the importance of robust phylogenetic frameworks for interpreting global biodiversity patterns.

Aimto characterize the evolution of climatic niches during the diversification of the Phyllotis darwini species group, in order to assess the extent to which divergences involved in radiation were associated with patterns of conservatism or divergence of climatic niches, and whether the differentiation found among climatic niches correlated with species phylogenetic relationships. Locationsouth-central Andes, surrounding lowlands, and Patagonia, South America. Methodsspecies climatic niches were characterized by sampling contemporaneous precipitation and temperature conditions across occurrence locations and entire distributional ranges. Climatic niches were analyzed and modeled using multivariate statistics (PCA, PERMANOVA), a maximum entropy-based algorithm, and novel methods developed to explore levels of differentiation (niche overlap test) and divergence (niche divergence test) between realized and fundamental niches. Comparative phylogenetic methods were applied using a time-calibrated phylogeny and integrating climate niche data to estimate ancestral environmental niches within geographic and environmental spaces. Resultscomparisons revealed low levels of climatic niche overlap, both among species realized niches and among their fundamental niches, suggesting high levels of niche differentiation during the diversification of Phyllotis species. Quantifications of niche overlap further showed that observed differences among species lay primarily in the multidimensional nature of climatic niches, as unidimensional quantifications exhibited higher levels of overlap. Evolved differences among species climatic niches were better fitted to a Brownian motion model of evolution, but lacked phylogenetic signal and showed no significant association with species phylogenetic distances. Main conclusionslow levels of differentiation between ancestral climatic niches suggest that the early radiation of species in the Phyllotis darwini species group was promoted by geographic isolation, whereas the more recent diversification of extant species was accompanied by climatic niche differentiation, possibly involving local adaptation to regional ecoclimatic changes associated with Quaternary glacial cycles. The spatial separation of sister species, the complete divergence of their climatic niches, and the lack of phylogenetic signal in niche differences suggest a scenario of diversification in which divergences were prompted by the spatial isolation, but also by the divergent selection exerted by regional climatic differences.

Silk glands have been found in two groups of amphipods: the Corophiida and the Ampeliscidae. The silk glands in Ampeliscidae, however, have yet to be examined in detail. Here we report, for the first time, the morphology and distribution of pereopodal glands in the Ampeliscidae, in non-thread producing Synopiidae, and in the Paragammaropsidae. In the Ampeliscidae we found two gland types distributed throughout all pereopods which have the ability to create threads. Pereopods three and four have additional silk extrusion morphology at the tip of the dactylus in which silk is transformed into semi-cylindrical threads used for building domiciles. Synopiid outgroup species have one of the gland types but lack silk extrusion morphology. Using ancestral state reconstruction analysis, we find that glands in the Synopiidae are likely ancestral and hypothesize that silk glands in Ampeliscidae are derived from these ancestral glands. Silk-spinning pereopods in the Paragammaropsidae had similarities with both Corophiida and Ampeliscidae but had distinctions. Ampeliscidae silk-spinning systems bear surprising resemblance to the Corophiida which presents one to reconsider the taxonomic placement of Ampeliscidae and the origins of silk-spinning in amphipods. This is the first comprehensive study on the glandular systems of Ampeliscidae, Synopiidae, and Paragammaropsidae using advanced microscopy, providing pertinent morphological data to the study of arthropod silk gland evolution and complex traits.

For over four decades, the bivalve Anomalocardia flexuosa has been recorded in Hong Kong coastal waters. However, the known native distribution of this heavily exploited commercial species is restricted to the Atlantic coast of South America, raising questions about the biogeographical validity of the Hong Kong populations. By employing an integrative taxonomic approach combining morphological re-evaluations and molecular phylogenetic analysis of the COI gene, we confirm that the species in Shui Hau, Hong Kong, China, has been historically misidentified. The population belongs to Cryptonema producta (syn. Anomalocardia producta).

Computational workshops are common in evolutionary biology and are used to share discipline-specific tools and skills with researchers. Despite the perceived importance of these workshops, there is no common set of criteria for workshop success, and there are few peer-reviewed studies investigating the efficacy of workshops or assessing the value of particular instructional techniques in this context. Here, we focused on one key element of a successful workshop: its ability to increase participants motivation to use the methods and tools presented during the workshop. We analyzed the goals, perceptions, and future plans of research practitioners engaging in a workshop on phylogenetic methods of historical biogeography using pre- and post-workshop surveys. Overall, the workshop was successful at motivating participants, and survey responses provided insights into participants perceptions of different activities, including "participatory live coding". Apart from this case study, we aim to highlight the importance of developing a common set of workshop goals in collaboration with other workshop stakeholders and the need for specialized, validated tools for assessing the efficacy of computational workshops for researchers.

Sex ratio distorters (SRDs) are heritable elements that bias offspring sex ratios to enhance their transmission. In the terrestrial isopod Armadillidium vulgare, feminization of genetic males can occur through vertical transmission of the sex ratio distorter known as the f-element, as well as through infection by Wolbachia, a maternally inherited bacterial endosymbiont that can alter host reproduction. Previous studies have focused on the distribution of SRDs and their associations with mitochondrial haplotypes in native European populations, but these patterns are poorly understood in the United States. In this study, we sampled A. vulgare in 12 states, screening individuals for Wolbachia infection, the presence of the f-element, and mitochondrial haplotypes. We found that Wolbachia shows a heterogeneous distribution across populations and haplotypes, in contrast with stronger associations in Europe. The f-element occurred in lower overall frequencies but showed a strong association with mitochondrial haplotype VI. These results indicate that patterns associated with SRD differ from those observed in Europe and suggest that multiple introductions and population mixing have shaped these distributions.

Immunoglobulin genes are a central component of jawed-vertebrate adaptive immunity. A previous study showed that the blunt-snouted clingfish Gouania willdenowi lacks immunoglobulin genes and T-cell receptor gamma/delta loci, while retaining T-cell receptor alpha/beta genes, MHC genes, and RAG1 /RAG2. Here I extend that observation to the family Gobiesocidae using all seven chromosome-level Gobiesocidae genome assemblies currently available. Manual tblastn and synteny-guided searches found no convincing immunoglobulin heavy-chain or light-chain loci in G. willdenowi, Gouania pigra, Gobiesox punctulatus, Apletodon dentatus, Lepadogaster candolii, Lepadogaster purpurea, or Diplecogaster bimaculata. Thus, the absence of antibody genes is best interpreted as a root-level character of clingfishes. The latest seven-species screen of 40 additional immune-associated genes shifts the broader interpretation in the same direction: the B-cell/adaptive core genes CD79A, CD79B, CIITA, TNFRSF13B, and TNFSF13B lack strong tblastn support in all sampled Gobiesocidae, and 37 of the 40 tested targets show an all-zero binary pattern at the presence threshold. Only IL21R.1, TYROBP, and TNFRSF11A show strong hits in one or more species. I therefore interpret the principal immune-gene erosion as occurring at or near the Gobiesocidae root rather than as a recent Gouania-specific process, while keeping weak, paralog-sensitive, and patchy loci provisional. RAG2 comparisons show a shared Gobiesocidae PHD-domain C-to-S replacement in the zinc-binding motif, with apparently intact RAG2 coding sequence. A family-wide TRG/TRD screen did not recover TRGV V segments or accepted TRDC constant-region exons, but it did detect TRGC-like constant exons in several genomes. These TRGC-like sequences are probably not canonical TRG constant exons without further validation, so I treat the gamma/delta system as eroded or rearranged rather than as a complete root-level loss equivalent to the Ig loss. The RAG2 variant provides a plausible molecular context for antigen-receptor remodeling, but it is not evidence that RAG genes are pseudogenized, because TCR alpha/beta, MHC genes, and RAG1 /RAG2 are retained. Gobiesocidae are therefore best described as a vertebrate family with ancestral loss of canonical immunoglobulin genes and associated root-level erosion of B-cell and immune-related genes, not as a lineage lacking adaptive immunity in its entirety. HighlightsO_LISeven chromosome-level Gobiesocidae genomes lack convincing canonical IgH and IgL loci. C_LIO_LIThe strongest non-Ig losses map to the B-cell/adaptive core: CD79A, CD79B, CIITA, TNFRSF13B, and TNFSF13B. C_LIO_LITCR alpha/beta, MHC genes, and RAG1 /RAG2 are retained, so Gobiesocidae should not be described as lacking adaptive immunity in full. C_LIO_LIA shared Gobiesocidae RAG2 PHD-domain C-to-S variant provides candidate molecular context for antigen-receptor remodeling. C_LI

PremiseCentaurea melitensis (Asteraceae) is a problematic invader of grasslands globally, but little is known about its genetic makeup. Here we develop a reference genome to facilitate studies of its invasion history, genetic variation, and evolution. MethodsInbred offspring of a single individual of C. melitensis from its invasion of California, USA were used for flow cytometry to estimate genome size, and for genomic DNA extraction. DNA was sequenced with PacBio HiFi technology (yield = 85.7Gb). The genome was assembled with Hifiasm and annotated with BRAKER3. GENESPACE was used to compare gene order (synteny) with three other species within the subfamily Cichorioideae. ResultsWe estimated a mean genome size of 795.0 Mbp for C. melitensis, and our assembly totaled 696.6 Mbp in 48 contigs (N50 = 55.6 Mbp; BUSCO = 98%), with annotation of 25,157 protein-encoding genes. This included four telomere-to-telomere putative chromosomes, nine additional chromosome arms terminated by telomeric repeats, and a complete chloroplast genome. Synteny varied markedly across the genus and subfamily, suggesting a dynamic history of structural variation in the lineage of C. melitensis. DiscussionWe provide a highly complete and contiguous genome assembly to facilitate the further study of genomic variation in C. melitensis.

The ichneumonid subfamily Eucerotinae has been thought to be almost absent from the tropics, with the only known Afrotropical species found in Madagascar. We report the subfamily to be present in the mainland Afrotropics, and describe a new species, Euceros species 1 from Uganda and Cameroon (name not yet shown in preprint). The subfamily had likely not been observed in the mainland Afrotropics before due to low abundances and insufficient sampling. More Eucerotinae likely remain to be discovered in tropical Africa and Asia, although tropical America may genuinely have few eucerotine species. Much more extensive sampling will be needed before it is possible to make confident estimates of how eucerotine diversity is distributed globally.

The genus Trichiurus is the most economically valuable fish in the family Trichiuridae, currently recognized to include 10 valid species. However, historically numerous morphologically similar congeners have been erroneously assigned as synonyms or subspecies of T. lepturus. In this study, we examined 16 hairtail specimens collected from the southern waters of Java Island, Indonesia. Integrated morphological and mitochondrial phylogenetic analyses (COX1 and 16S rRNA), compared against global Trichiurus sequences, revealed that these specimens form an independent lineage that diverged early from other congeners. Consequently, we describe this lineage as a previously undescribed cryptic species. Diagnostic characters include: first anal-fin spine below 36th-37th dorsal-fin rays; anus below 35th-36th dorsal-fin rays; anteriormost tip of supraoccipital well posterior to posterior distal margin of eye; anterior margin of the pectoral-fin spine non-serrated; fangs on both jaws with barb-like processes; upper jaw long, mean 16.6% (15.5-17.6%) of preanal length; snout short, 12.0% (10.9-13.1%) of preanal length; eye small, diameter 5.3% (4.3-5.7%) of preanal length; and absence of hyperostosis on dorsal cranium. We herein propose the name Trichiurus javaensis sp. nov., and provide a formal morphological description and diagnostic characterization of this species.

To preserve homeostasis in the face of continual chemical insult, animals evolved dedicated molecular systems that detect, detoxify, and eliminate foreign compounds. Collectively, these enzymes, transporters, and regulatory pathways constitute the chemical defensome. In cetaceans, the loss of two key nuclear receptors (NR1I2/PXR and NR1I3/CAR) suggests a profound rearrangement of the chemical defense systems. Therefore, we investigated the gene inventory of the chemical defensome in Cetacea and two other major marine mammal lineages (Pinnipedia and Sirenia), using their closest terrestrial relatives to understand the extent and patterns of chemical defensome remodelling. We demonstrate large-scale gene loss in chemical defensome genes of cetaceans, as well as smaller scale gene loss in the other two marine mammal lineages, indicating possible convergent evolution. Gene loss occurred predominantly in phase I and phase II biotransformation enzymes, including CYPs, FMOs, SULTs, and GSTs. Many of the lost genes in cetaceans are known to be regulated by PXR and/or CAR, while genes lost in multiple marine mammal lineages are often not regulated by these transcription factors. We hypothesize that the transition to aquatic environments, often accompanied by corresponding changes in feeding habits, led to convergent loss of chemical defensome genes, and loss of PXR and CAR in cetaceans accelerated these losses. These findings reveal systematic erosion of chemical defense capabilities across marine mammal lineages, suggesting that adaptation to marine life involves trade-offs in detoxification capacity that may have significant implications for these species responses to increasing chemical pollution in present-day ocean environments.

Two threatened new species of Podostemaceae belonging to the genus Inversodicraea, I. joulei and I. lebbiei, both from the Republic of Sierra Leone, are described and illustrated. A first record in Sierra Leone of the genus Lestestuella is also reported. Inversodicraea is the most species-rich genus of Podostemaceae in Africa and now comprises 38 species. Inversodicraea joulei is easily recognised because it has a persistent spine distally on the median rib of each fruit valve, and scattered, membranous scale-leaves with broadly rounded apices, while Inversodicraea lebbiei is distinct in having narrowly triangular robust scale-leaves which are inrolled, spreading distally, and completely covering the stem, arranged in five ranks. Inversodicraea joulei is known from a single location with three sites while I. lebbiei is known from two locations each with one site. Using the latest IUCN Red List guidance, Inversodicraea joulei is assessed as Critically Endangered and I. lebbiei is assessed as Endangered, due to threats from dam construction projects, agricultural practices and mining activities, resulting in high levels of siltation on rocks in the fast-flowing rivers where these species grow.

The diversity of body shapes is one of the most prominent features of phenotypic variation in mammals. Yet, mammalian body shapes are poorly quantified and the underlying components contributing to its diversity as well as its relationship to other components of the skeleton are rarely tested. Here, we use lagomorphs (hares, rabbits and pikas) as a model system to (1) investigate which components of the skeleton contributed the most to body shape diversity, (2) examine the relationships between body shape and relative limb lengths, and (3) test how body size, ecotype, burrowing behavior, and locomotor mode influenced variation in lagomorph body shape and appendicular morphology. We quantified the body shape and functional proxies of the appendicular skeleton in 40 lagomorph species from osteological specimens held at museum collections. Using phylogenetic comparative methods, we found the relative length of the ribs and elongation or shortening of the thoracic and lumbar regions contributed the most to body shape evolution across lagomorphs. Second, we found that only leporids (hares and rabbits) exhibited a significant relationship between limb length and body shape, where more elongate species exhibit relatively shorter forelimbs and hindlimbs. Lastly, we found that models incorporating body size were the best predictors of lagomorph body shape and the majority of the appendicular traits, whereas models incorporating burrowing behavior and locomotor mode were largely poor fits. Broadly, these results indicate that larger lagomorphs tend to exhibit more robust body shapes with longer, more gracile forelimbs, whereas smaller lagomorphs tend to exhibit more elongate body shapes with shorter, more robust forelimbs. Overall, this work contributes to the growing understanding of mammalian body shape evolution and demonstrates the importance of not omitting body size in ecomorphological analyses.

Plant receptor-like kinases (RLKs) are involved in diverse processes, ranging from growth and reproduction to interactions with microbes. Variation in the extracellular domains delineates several RLKs subfamilies, including the malectin-like domain leucine-rich repeat receptor-like kinases (MLD-LRR-RLKs). Symbiosis Receptor-like Kinase (SymRK) is the prototypical member of MLD-LRR-RLKs and is required for microbial accommodation in host roots during root endosymbiosis. Yet, comparative phylogenetic analysis of SymRK orthologs in the broader context of MLD-LRR-RLK subfamily evolution remains limited. In this study, we examined the inventory, phylogeny and clade-specific evolutionary and transcriptional characteristics of this receptor group. SymRK and its closest homologs are present in most land plant lineages and group into four major clades and six additional species-specific clades. These clades can be distinguished by their evolutionary characteristics as either conserved with reduced gene copy number changes (including SymRK) or expanded and diversified, as observed in clade IV. Clade IV dynamics are largely driven by tandem gene duplications, which often arise within gene clusters. We further analysed the evolutionary characteristics of MLD-LRR-RLKs at the population level in Arabidopsis thaliana accessions. We found that some genes are conserved across accessions and are therefore likely to be functionally important, whereas a subset of genes, often located within tandem clusters, are highly diverse and likely contribute to accession-specific adaptations. Finally, most MLD-LRR-RLKs in the A. thaliana Col-0 accession are expressed in roots and respond broadly to biotic stimuli at the transcriptional level. Notably, clustered genes frequently exhibited divergent expression profiles, suggesting transcriptional diversification. Together, we revealed two contrasting evolutionary characteristics among members of the MLD-LRR-RLK subfamily, potentially associated with their functions in plants.