Aquatic Lineages Research Articles

Secondary contact erodes Pleistocene diversification in a wide-ranging freshwater mussel (Quadrula).

The isolated river drainages of eastern North America serve as a natural laboratory to investigate the roles of allopatry and secondary contact in the evolutionary trajectories of recently diverged lineages. Drainage divides facilitate allopatric speciation, but due to their sensitivity to climatic and geomorphological changes, neighboring rivers frequently coalesce, creating recurrent opportunities of isolation and contact throughout the history of aquatic lineages. The freshwater mussel Quadrula quadrula is widely distributed across isolated rivers of eastern North America and possesses high phenotypic and molecular variation across its range. We integrate sequence data from three genomes, including female- and male-inherited mitochondrial markers and thousands of nuclear encoded SNPs with morphology and geography to illuminate the group's divergence history. Across contemporary isolated rivers, we found continuums of molecular and morphological variation, following a pattern of isolation by distance. In contact zones, hybridization was frequent with no apparent fitness consequences, as advanced hybrids were common. Accordingly, we recognize Q. quadrula as a single cohesive species with subspecific variation (Q. quadrula rumphiana). Demographic modeling and divergence dating supported a divergence history characterized by allopatric vicariance followed by secondary contact, likely driven by river rearrangements and Pleistocene glacial cycles. Despite clinal range-wide variation and hybridization in contact zones, the process-based species delimitation tool delimitR, which considers demographic scenarios like secondary contact, supported the delimitation of the maximum number of species tested. As such, when interpreting species delimitation results, we suggest careful consideration of spatial sampling and subsequent geographic patterns of biological variation, particularly for wide-ranging taxa.

Origin and evolution of the blue light receptor cryptochromes (CRY1/2) in aquatic angiosperms.

Cryptochromes (CRYs), which are responsible for sensing blue light in plants, play a critical role in regulating blue light signals and circadian rhythms. However, their functions extend beyond light detection, as they also aid plants in adapting to stress and potentially other regulatory mechanisms. Aquatic angiosperms, which independently evolved from various angiosperm lineages, have developed specific adaptations to unique light qualities and environmental stressors found in aquatic habitats compared to terrestrial ones. It was hypothesized that the sequences and regulatory networks of angiosperm CRY1/2 underwent adaptive evolution in different aquatic angiosperm lineages. To test this hypothesis, we compiled comprehensive datasets consisting of 55 green plant genomes (including 37 angiosperm genomes), 80 angiosperm transcriptomes, and 4 angiosperm expression networks. Through comparative analysis, we found that CRY1 originated from a common ancestor of seed plants, whereas CRY2 originated from a common ancestor of land plants. In angiosperms, the CRY1/2 sequences of aquatic lineages exhibited positive selection, and the conserved valine-proline (VP) motif of CRY2 showed a convergent loss in two aquatic species. Co-expressed genes associated with blue light receptors (CRY) showed adaptations to aquatic environments, specifically in relation to flooding and osmotic stress. These discoveries shed light on the adaptive evolution of CRY1/2, encompassing their origins, sequences, and regulatory networks. Furthermore, these results provide valuable insights for investigating the uncharacterized functions and regulatory pathways of CRY and offer potential targets for enhancing growth and adaptation in agricultural plants.

Metabarcoding reveals ecologically distinct fungal assemblages in river and groundwater along an Austrian alpine to lowland gradient.

Biodiversity, the source of origin, and ecological roles of fungi in groundwater are to this day a largely neglected field in fungal and freshwater ecology. We used DNA-based Illumina high-throughput sequence analysis of both fungal gene markers 5.8S and internal transcribed spacers region 2 (ITS2), improving taxonomic classification. This study focused on the groundwater and river mycobiome along an altitudinal and longitudinal transect of a pre-alpine valley in Austria in two seasons. Using Bayesian network modeling approaches, we identified patterns in fungal community assemblages that were mostly shaped by differences in landscape (climatic, topological, geological), and environmental conditions. While river fungi were comparatively more diverse, unique fungal assemblages could be recovered from groundwater, including typical aquatic lineages such as Rozellomycota and Olpidiomycota. The most specious assemblages in groundwater were not linked to the input of organic material from the surface, and as such, seems to be sustained by characteristic groundwater conditions. Based on what is known from closely related fungi, our results suggest that the present fungal communities potentially contribute to mineral weathering, carbon cycling and denitrification in groundwater. Furthermore, we were able to observe the effects of varying land cover due to agricultural practices on fungal biodiversity in groundwater ecosystems. This study contributes to improving our understanding of fungi in the subsurface aquatic biogeosphere.

A freshwater ecosystem once existed in the Rajasthan desert: evidence from a fossil of the aquatic herb Nelumbo

Nelumbonaceae encompasses aquatic plant lineages that represent major components in the modern tropical wetland ecosystems of India. The group is characterized by its distinctive reproductive receptacle incorporating in situ fruitlets and vegetative organs that are well-represented in the fossil record. Unusually, though, no Nelumbo reproductive fossils have yet been identified from the Cenozoic of India. Here, we report the first such fossil reproductive organ from the aquatic perennial herb Nelumbo, which was recovered in the lower Eocene Palana Formation at the Gurha lignite mine in Rajasthan, western India. The fossil preserves an aggregate fruit with 28 indehiscent, apocarpous and globose to oblong fruitlets embedded within an accrescent receptacle, and a single small central cavity or apical remains of a stigma in each fruitlet. Comparisons with extant and extinct Nelumbo species support attribution to Nelumbo sp. Our unexpected discovery of a fully aquatic plant in the Rajasthan Basin during the early Eocene indicates that a freshwater ecosystem existed there ca 55 ± 10 million years ago; this contrasts starkly with the arid climatic conditions predominant in Rajasthan today. Raman Patel† [ ramanpatel2142@gmail.com ], Department of Geology, Hemvati Nandan Bahuguna Garhwal University, Srinagar, 246174 Uttarakhand, India Ashif Ali† [ aashifli0011@gmail.com ], Palaeobotany, Palynology, and Plant Evolution Laboratory, Department of Botany, Sidho-Kanho-Birsha University, Ranchi Road, Purulia-723104, India Rajendra Singh Rana [ rajendra.rana1@gmail.com ], Department of Geology, Hemvati Nandan Bahuguna Garhwal University, Srinagar, 246174 Uttarakhand, India Mahasin Ali Khan* [ khan.mahasinali@gmail.com ], Palaeobotany, Palynology, and Plant Evolution Laboratory, Department of Botany, Sidho-Kanho-Birsha University, Ranchi Road, Purulia-723104, India.

A new giant Jurassic lacewing larva reveals a particular aquatic habit and its significance to the palaeoecology

AbstractNeuroptera, as a small relic group of Insecta undergoing a rapid species diversification during the Mesozoic Era, is known by diverse extinct endemic lineages preserved as impression fossils and in amber. The current understanding of Mesozoic neuropterans′ diversity has mainly focused on the adults, because the contemporaneous larvae have been fairly rare especially for the Jurassic lacewings. Herein, a new giant lacewing larva, Natator giganteus gen. et sp. nov., is described from the Middle Jurassic Daohugou Beds of China. The remarkable larva is characterized by its impressively large body size, distinctively elongated cervix, and presence of swimming hairs on legs, which provide direct evidence to reveal an aquatic habit for the Jurassic lacewing larva. The morphological analysis indicates this giant larva would have probably inhabited the benthic environments of Jurassic montane rivers and streams. In addition, its morphological specialization suggests that it might have adopted an ambush predation strategy to catch its prey. The finding enhances our knowledge of the species diversity and morphological plasticity for the Jurassic lacewing larvae, and reveals that the aquatic lineages of Neuroptera exhibited dramatically structural and ecological convergence across the evolutionary process.

Chromosome-scale genome assemblies of Himalopsyche anomala and Eubasilissa splendida (Insecta: Trichoptera)

Trichoptera is one of the most evolutionarily successful aquatic insect lineages and is highly valued value in adaptive evolution research. This study presents the chromosome-level genome assemblies of Himalopsyche anomala and Eubasilissa splendida achieved using PacBio, Illumina, and Hi-C sequencing. For H. anomala and E. splendida, assembly sizes were 663.43 and 859.28 Mb, with scaffold N50 lengths of 28.44 and 31.17 Mb, respectively. In H. anomala and E. splendida, we anchored 24 and 29 pseudochromosomes, and identified 11,469 and 10,554 protein-coding genes, respectively. The high-quality genomes of H. anomala and E. splendida provide critical genomic resources for understanding the evolution and ecology of Trichoptera and performing comparative genomics analyses.

Whole Genome Duplication Events Likely Contributed to the Aquatic Adaptive Evolution of Parkerioideae.

As the only aquatic lineage of Pteridaceae, Parkerioideae is distinct from many xeric-adapted species of the family and consists of the freshwater Ceratopteris species and the only mangrove ferns from the genus Acrostichum. Previous studies have shown that whole genome duplication (WGD) has occurred in Parkerioideae at least once and may have played a role in their adaptive evolution; however, more in-depth research regarding this is still required. In this study, comparative and evolutionary transcriptomics analyses were carried out to identify WGDs and explore their roles in the environmental adaptation of Parkerioideae. Three putative WGD events were identified within Parkerioideae, two of which were specific to Ceratopteris and Acrostichum, respectively. The functional enrichment analysis indicated that the lineage-specific WGD events have played a role in the adaptation of Parkerioideae to the low oxygen concentrations of aquatic habitats, as well as different aquatic environments of Ceratopteris and Acrostichum, such as the adaptation of Ceratopteris to reduced light levels and the adaptation of Acrostichum to high salinity. Positive selection analysis further provided evidence that the putative WGD events may have facilitated the adaptation of Parkerioideae to changes in habitat. Moreover, the gene family analysis indicated that the plasma membrane H+-ATPase (AHA), vacuolar H+-ATPase (VHA), and suppressor of K+ transport growth defect 1 (SKD1) may have been involved in the high salinity adaptation of Acrostichum. Our study provides new insights into the evolution and adaptations of Parkerioideae in different aquatic environments.

A Systematic Review of Insect Decline and Discovery: Trends, Drivers, and Conservation Strategies over the past Two Decades

Insects make up 75%–80% of all known animal species, with estimates ranging from 10 to 30 million, of which approximately 1.11–1.7 million are described. Despite varying estimates, 80% of insect species remain undiscovered. Unfortunately, insect populations have been declining globally in recent decades, which have significant implications for the overall health of ecosystems. Thus, this review article delves into the status of insects, exploring the causes behind their decline, highlighting discoveries made in recent years, and emphasizing the profound impact of this decline on ecosystems. Furthermore, it discusses the urgent need for conservation efforts and identifies promising areas for future insect research. To achieve these objectives, the researcher uses a systematic review of 156 distinct journals and websites. Insect decline is a worldwide issue caused by habitat loss, pesticide use, pollution, invading species, intensive agriculture, and climate change. It affects flying, ground, and aquatic lineages, with Lepidoptera, Hymenoptera, Coleoptera, Odonata, Plecoptera, Trichoptera, and Ephemeroptera being especially vulnerable. The decline of insects has significant impacts on ecosystems, including food chains, pollination, and nutrient cycling. Insect conservation strategies involve habitat restoration, sustainable land management, species‐specific conservation, and policy. Developing insect conservation laws, enforcing environmental laws, and promoting public education are also crucial for addressing insect declines and promoting biodiversity. Future research should focus on using insects for sustainable agriculture, understanding their behavior for conservation and technology, utilizing insect genetics for pest control, and improving insect classification for conservation.

Dollo meets Bergmann: morphological evolution in secondary aquatic mammals

Secondary transitions to aquatic environments are common among vertebrates, and aquatic lineages display several adaptations to this realm, some of which might make these transitions irreversible. At the same time, discussions about secondary transitions often focus only on the marine realm, comparing fully terrestrial with fully aquatic species. This, however, captures only a fraction of land-to-water transitions, and freshwater and semi-aquatic groups are often neglected in macroevolutionary studies. Here, we use phylogenetic comparative methods to unravel the evolution of different levels of aquatic adaptations across all extant mammals, testing if aquatic adaptations are irreversible and if they are related to relative body mass changes. We found irreversible adaptations consistent with Dollo's Law in lineages that rely strongly on aquatic environments, while weaker adaptations in semi-aquatic lineages, which still allow efficient terrestrial movement, are reversible. In lineages transitioning to aquatic realms, including semi-aquatic ones, we found a consistent trend towards an increased relative body mass and a significant association with a more carnivorous diet. We interpret these patterns as the result of thermoregulation constraints associated with the high thermal conductivity of water leading to body mass increase consistently with Bergmann's rule and to a prevalence of more nutritious diets.

Reduction of Paraoxonase Expression Followed by Inactivation across Independent Semiaquatic Mammals Suggests Stepwise Path to Pseudogenization.

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.

Highly Dynamic Gene Family Evolution Suggests Changing Roles for PON Genes Within Metazoa.

Change in gene family size has been shown to facilitate adaptation to different selective pressures. This includes gene duplication to increase dosage or diversification of enzymatic substrates and gene deletion due to relaxed selection. We recently found that the PON1 gene, an enzyme with arylesterase and lactonase activity, was lost repeatedly in different aquatic mammalian lineages, suggesting that the PON gene family is responsive to environmental change. We further investigated if these fluctuations in gene family size were restricted to mammals and approximately when this gene family was expanded within mammals. Using 112 metazoan protein models, we explored the evolutionary history of the PON family to characterize the dynamic evolution of this gene family. We found that there have been multiple, independent expansion events in tardigrades, cephalochordates, and echinoderms. In addition, there have been partial gene loss events in monotremes and sea cucumbers and what appears to be complete loss in arthropods, urochordates, platyhelminths, ctenophores, and placozoans. In addition, we show the mammalian expansion to three PON paralogs occurred in the ancestor of all mammals after the divergence of sauropsida but before the divergence of monotremes from therians. We also provide evidence of a novel PON expansion within the brushtail possum. In the face of repeated expansions and deletions in the context of changing environments, we suggest a range of selective pressures, including pathogen infection and mitigation of oxidative damage, are likely influencing the diversification of this dynamic gene family across metazoa.

New Caledonia's enigmatic terrestrial diving beetle Typhlodessus monteithi is a derived species of Paroster

AbstractThe classification of highly adapted species in well‐studied clades may be obscured by convergent character evolution. This is for example the case in aquatic lineages adapted to subterranean (and shallow subterranean) habitats, in which species usually possess reduced eyes and wings as well as translucent cuticles. In 1985, the terrestrial diving beetle genus and species Typhlodessus monteithi Brancucci (Coleoptera: Dytiscidae) was described from a shallow subterranean habitat in New Caledonia. Until now, the systematic classification of Typhlodessus remains unclear, as it could not be assigned to any of the known tribes of the subfamily Hydroporinae. Here, we reveal this species' phylogenetic position and evolutionary history. We obtained molecular data from an almost 30 years old museum specimen and performed phylogenetic analyses using complete mitochondrial genomes and nuclear markers, which resulted in the placement of Typhlodessus monteithi within the subtribe Sternopriscina, as a junior synonym of the genus Paroster. The genus Paroster contains epigean, stygobitic as well as two other terrestrial Australian species. Our finding extends the geographical distribution of Paroster to New Caledonia. Adaptation to terrestrial habitats in Paroster species may be a way to survive in hyper humid environments that however might lack suitable lentic habitats.

Insights into cryptic speciation of quillworts in China

Cryptic species are commonly misidentified because of high morphological similarities to other species. One group of plants that may harbor large numbers of cryptic species is the quillworts (Isoëtes spp.), an ancient aquatic plant lineage. Although over 350 species of Isoëtes have been reported globally, only ten species have been recorded in China. The aim of this study is to better understand Isoëtes species diversity in China. For this purpose, we systematically explored the phylogeny and evolution of Isoëtes using complete chloroplast genome (plastome) data, spore morphology, chromosome number, genetic structure, and haplotypes of almost all Chinese Isoëtes populations. We identified three ploidy levels of Isoëtes in China—diploid (2n = 22), tetraploid (2n = 44), and hexaploid (2n = 66). We also found four megaspore and microspore ornamentation types in diploids, six in tetraploids, and three in hexaploids. Phylogenetic analyses confirmed that I. hypsophila as the ancestral group of the genus and revealed that Isoëtes diploids, tetraploids, and hexaploids do not form monophyletic clades. Most individual species possess a single genetic structure; however, several samples have conflicting positions on the phylogenetic tree based on SNPs and the tree based on plastome data. All 36 samples shared 22 haplotypes. Divergence time analysis showed that I. hypsophila diverged in the early Eocene (∼48.05 Ma), and most other Isoëtes species diverged 3–20 Ma. Additionally, different species of Isoëtes were found to inhabit different water systems and environments along the Yangtze River. These findings provide new insights into the relationships among Isoëtes species in China, where highly similar morphologic populations may harbor many cryptic species.

Ancient whale rhodopsin reconstructs dim-light vision over a major evolutionary transition: Implications for ancestral diving behavior

Cetaceans are fully aquatic mammals that descended from terrestrial ancestors, an iconic evolutionary transition characterized by adaptations for underwater foraging via breath-hold diving. Although the evolutionary history of this specialized behavior is challenging to reconstruct, coevolving sensory systems may offer valuable clues. The dim-light visual pigment, rhodopsin, which initiates phototransduction in the rod photoreceptors of the eye, has provided insight into the visual ecology of depth in several aquatic vertebrate lineages. Here, we use ancestral sequence reconstruction and protein resurrection experiments to quantify light-activation metrics in rhodopsin pigments from ancestors bracketing the cetacean terrestrial-to-aquatic transition. By comparing multiple reconstruction methods on a broadly sampled cetartiodactyl species tree, we generated highly robust ancestral sequence estimates. Our experimental results provide direct support for a blue-shift in spectral sensitivity along the branch separating cetaceans from terrestrial relatives. This blue-shift was 14 nm, resulting in a deep-sea signature (λmax = 486 nm) similar to many mesopelagic-dwelling fish. We also discovered that the decay rates of light-activated rhodopsin increased in ancestral cetaceans, which may indicate an accelerated dark adaptation response typical of deeper-diving mammals. Because slow decay rates are thought to help sequester cytotoxic photoproducts, this surprising result could reflect an ecological trade-off between rod photoprotection and dark adaptation. Taken together, these ancestral shifts in rhodopsin function suggest that some of the first fully aquatic cetaceans could dive into the mesopelagic zone (>200 m). Moreover, our reconstructions indicate that this behavior arose before the divergence of toothed and baleen whales.

Correlative adaptation between Rubisco and CO2-concentrating mechanisms in seagrasses.

Submerged angiosperms sustain some of the most productive and diverse ecosystems worldwide. However, their carbon acquisition and assimilation mechanisms remain poorly explored, missing an important step in the evolution of photosynthesis during the colonization of aquatic environments by angiosperms. Here we reveal a convergent kinetic adaptation of Rubisco in phylogenetically distant seagrass species that share catalytic efficiencies and CO2 and O2 affinities up to three times lower than those observed in phylogenetically closer angiosperms from terrestrial, freshwater and brackish-water habitats. This Rubisco kinetic convergence was found to correlate with the effectiveness of seagrass CO2-concentrating mechanisms (CCMs), which probably evolved in response to the constant CO2 limitation in marine environments. The observed Rubisco kinetic adaptation in seagrasses more closely resembles that seen in eukaryotic algae operating CCMs rather than that reported in terrestrial C4 plants. Our results thus demonstrate a general pattern of co-evolution between Rubisco function and biophysical CCM effectiveness that traverses distantly related aquatic lineages.

The unique rectus extraocular muscles of cetaceans: Homologies and possible functions.

Each rectus extraocular muscle in cetaceans divides into two portions: a massive palpebral belly that inserts into the deep surface of the eyelids and a smaller scleral belly that inserts onto the eyeball. While the cetacean palpebral insertions have long been recognized, their homologies and functions remain unclear. To compare cetacean rectus EOM insertions with the global and orbital rectus EOM insertions of other mammals we dissected orbital contents of 20 odontocete species, 2 mysticete species and 18 non-cetacean species, both aquatic and terrestrial. Four cetacean species were also examined with magnetic resonance imaging (MRI). All four rectus muscles in cetaceans had well-developed palpebral bellies and insertions. Adjacent palpebral bellies showed varying degrees of fusion, from near independence to near complete fusion. Fusion was most complete towards palpebral insertions and less towards origins. A medial moiety of the superior rectus palpebral belly is likely the levator palpebrae superioris. Smaller but still robust scleral insertions were present on all recti, with the medial rectus (MR) being significantly more muscular than the others. All non-cetacean species examined had recti with distinct global and orbital insertions, the latter generally onto Tenon's capsule. Orbital insertions in pygmy hippopotamus and Florida manatee extended into the deep surfaces of the eyelids, hence qualifying as palpebral insertions. Our results suggest that rectus EOMs of mammals generally have both global and orbital insertions, and that palpebral bellies of cetaceans and other species are modified homologs of the orbital insertions. The presence of palpebral insertions in pygmy hippopotamus and absence in other cetartiodactyls suggests an intermediate condition between terrestrial cetartiodactyls and cetaceans. Palpebral insertions in Florida manatee and reports of their presence in some pinnipeds suggest parallel evolution in multiple aquatic lineages. Various functions of cetacean palpebral recti have been proposed, including eyelid dilators, protection during diving and thermogenesis for warming eye and brain. For further insight into their possible functions, we observed eye movements of captive bottlenose dolphins (Tursiops truncatus) at the U.S. National Aquarium. Our observations showed that in addition to rotation of the eyeball the entire surrounding palpebral region also moves during gaze changes. For example during upward gaze the globe not only rotates in supraduction but translates dorsally as well. It appears the rectus palpebral bellies are responsible for flexing the palpebral structures and thus also translating the globe, while the scleral insertions act directly for ocular rotation. Along with frequent non-conjugate eye movements, the oculomotor mechanics and repertoire of cetaceans are thus quite distinctive. Summarily, axial displacement within the orbit is a major 'eye movement' in cetaceans, with protrusion and retraction mediated by well-developed circular muscles and retractor bulbi respectively. Torsional eye movements driven by elaborate oblique EOMs are likewise significant. The roles of rectus EOMs for ocular rotation via their scleral insertions, especially the highly muscular MR, are for typical supra/infraductions and nasal/temporal ductions. The palpebral bellies accentuate these ductions by translating the globe and surrounding structures in the same direction.

A new species of mammalian trypanosome, Trypanosoma (Megatrypanum) bubalisi sp. nov., found in the freshwater leech Hirudinaria manillensis

Leeches have long been considered potential vectors for the aquatic lineage of trypanosomes, while bloodsucking insects are generally considered as the vectors for the terrestrial lineage of trypanosomes. The freshwater leech, Hirudinaria manillensis, is a widely distributed species in southern China and could potentially act as the vector for trypanosomes. Prior to this study, no trypanosomes had been reported from this leech. However, in this study, leeches were collected from three different places in Guangdong province, China, and a large number of flagellates were isolated and successfully cultured in vitro. Based on morphology, these flagellates looked like a typical trypanosome species. Analysis was carried out on the molecular sequences of the 18S rRNA gene and the glycosomal glyceraldehyde-3-phosphate dehydrogenase (gGAPDH) gene. To our surprise, these flagellates were identified as likely to be a mammalian trypanosome belonging to the clade containing Trypanosoma (Megatrypanum) theileri but they are significantly different from the typical TthI and TthII stocks. Analyses of blood composition indicated that the source of the blood meal in these leeches was from the water buffalo (Bubalus bubalis). To further test if this flagellate from the freshwater leech was indeed a mammalian trypanosome, we transferred the trypanosomes cultured at 27–37 °C and they were able to successfully adapt to this mammalian body temperature, providing further supporting evidence. Due to the significant genetic differences from other related trypanosomes in the subgenus Megatrypanum, we propose that this flagellate, isolated from H. manillensis, is a new species and have named it Trypanosoma bubalisi. Our results indicate that freshwater leeches may be a potential vector of this new mammalian trypanosome.

Heme auxotrophy in abundant aquatic microbial lineages

Heme, a porphyrin ring complexed with iron, is a metalloprosthetic group of numerous proteins involved in diverse metabolic and respiratory processes across all domains of life, and is thus considered essential for respiring organisms. Several microbial groups are known to lack the de novo heme biosynthetic pathway and therefore require exogenous heme from the environment. These heme auxotroph groups are largely limited to pathogens, symbionts, or microorganisms living in nutrient-replete conditions, whereas the complete absence of heme biosynthesis is extremely rare in free-living organisms. Here, we show that the acI lineage, a predominant and ubiquitous free-living bacterial group in freshwater habitats, is auxotrophic for heme, based on the experimental or genomic evidence. We found that two recently cultivated acI isolates require exogenous heme for their growth. One of the cultured acI isolates also exhibited auxotrophy for riboflavin. According to whole-genome analyses, all (n = 20) isolated acI strains lacked essential enzymes necessary for heme biosynthesis, indicating that heme auxotrophy is a conserved trait in this lineage. Analyses of >24,000 representative genomes for species clusters of the Genome Taxonomy Database revealed that heme auxotrophy is widespread across abundant but not-yet-cultivated microbial groups, including Patescibacteria, Marinisomatota (SAR406), Actinomarinales (OM1), and Marine groups IIb and III of Euryarchaeota Our findings indicate that heme auxotrophy is a more common phenomenon than previously thought, and may lead to use of heme as a growth factor to increase the cultured microbial diversity.

Development of the Myzozoan Aquatic Parasite Perkinsus marinus as A Versatile Experimental Genetic Model Organism

The phylum Perkinsozoa is an aquatic parasite lineage that has devastating effects on commercial and natural mollusc populations, and also comprises parasites of algae, fish and amphibians. They are related to dinoflagellates and apicomplexans and thus offer excellent genetic models for both parasitological and evolutionary studies. Genetic transformation was previously achieved for Perkinsus spp. but with few tools for transgene expression and limited selection efficacy. We sought to expand the power of experimental genetic tools for Perkinsus using P. marinus as a model. We constructed a modular plasmid assembly system for expression of multiple genes simultaneously. We developed efficient selection systems for three drugs, puromycin, bleomycin and blasticidin, that are effective in as little as three weeks. We developed eleven new promoters of variable expression strength. Furthermore, we identified that genomic integration of transgenes is predominantly via non-homologous recombination but with transgene fragmentation including deletion of some elements. To counter these dynamic processes, we show that bi-cistronic transcripts using the viral 2A peptides can couple selection to the maintenance of the expression of a transgene of interest. Collectively, these new tools and insights provide great new capacity to genetically modify and study Perkinsus as an aquatic parasite and evolutionary model.

Integrative phylogenomics reveals a Permian origin of Adephaga beetles

AbstractBeetles are arguably the most diverse group of animals on Earth with over 400 000 described species. Yet the timing of main diversification events among these insects remains debated. The use of phylogenomic data generated using next‐generation sequencing recently resolved most recalcitrant phylogenetic relationships across Coleoptera. However, limited taxon sampling for some major clades still prevents the use of important fossil calibrations that could provide more accurate estimates of the timing of lineage diversification events among beetles. Here, we present a new fossil‐based dated framework with a focus on the suborder Adephaga. We rely on an integrative phylogenomic approach using a combination of genomic, ultraconserved element and RNAseq transcriptomic datasets, further revealing the prevalent exonic nature of ultraconserved elements in Coleoptera. We infer a robust phylogenomic tree under various optimality criteria and analytical conditions. Our preferred phylogenetic reconstruction is consistent with those of previous phylogenomic studies in recovering the paraphyly of ‘Hydradephaga’, Gyrinidae as sister to all other Adephaga, the monophyly of Geadephaga and Haliplidae as sister to a monophyletic Dytiscoidea. We further recover strong support for paraphyly of Aspidytidae and the placement of Hygrobiidae as part of a clade comprising Aspidytes, Ribera, Beutel, Balke & Vogler, Sinaspidytes Balke, Beutel & Ribera and Amphizoidae. Using 23 carefully chosen fossil calibrations across Coleoptera, we infer the origin of modern beetles ca. 317 Ma in the mid‐Carboniferous, the divergence between Archostemata and Adephaga ca. 296 Ma in the early Permian, and the crown of Adephaga in the end‐Permian ca. 255 Ma. Importantly, our analyses provide more precise estimates of divergence times for internal splits within Adephaga, including ground beetles, tiger beetles and aquatic adephagan lineages. These results represent a major step forward in our understanding of beetle diversification and will serve as a fundamental framework to unravel the evolutionary history of many clades within Adephaga.