Revision of Trigastrotheca Cameron (Hymenoptera, Braconidae, Braconinae) with descriptions of 13 new species.

The Old World braconine wasp genus Trigastrotheca Cameron is revised. The genus is recorded from the island of Madagascar for the first time based on two new species, T.christianhenrichi Quicke & Butcher, sp. nov. and T.formosa Quicke & Friedman, sp. nov. Trigastrothecagriffini Quicke, sp. nov. is described from Australia; T.aethiopica Quicke & Friedman, sp. nov. is described from Ethiopia; T.braeti Quicke & Butcher, sp. nov. is described from Congo; T.simba van Noort, sp. nov. is described from Tanzania; T.freidbergi Quicke & Friedman, sp. nov., T.carinata Ranjith, sp. nov., T.flava Ranjith, sp. nov. and T.similidentata Ranjith, sp. nov. are described from India; T.khaoyaiensis Quicke & Butcher, sp. nov., T.naniensis Quicke & Butcher, sp. nov., and T.sublobata Quicke & Butcher, sp. nov. are described from Thailand. Trigastrothecatridentata is recorded from Thailand for the first time. A putative female of T.sureeratae is described for the first time. Acroceriliatricolor Quicke & Ingram, 1993 is transferred into Trigastrotheca, as T.acroceropsis nom. nov. A key is provided for the identification of species.

SACCHARIS v2: Streamlining Prediction of Carbohydrate-Active Enzyme Specificities Within Large Datasets.

Carbohydrates are chemically and structurally diverse, composed of a wide array of monosaccharides, stereochemical linkages, substituent groups, and intermolecular associations with other biological molecules. A large repertoire of carbohydrate-active enzymes (CAZymes) and enzymatic activities are required to form, dismantle, and metabolize these complex molecules. The software SACCHARIS (Sequence Analysis and Clustering of CarboHydrate Active enzymes for Rapid Informed prediction of Specificity) provides a rapid, easy-to-use pipeline for the prediction of potential CAZyme function in new datasets. We have updated SACCHARIS to (i) simplify its installation by re-writing in Python and packaging for Conda; (ii) enhance its usability through a new (optional) interactive GUI; and (iii) enable semi-automated annotation of phylogenetic tree output via a new R package or the commonly-used webserver iTOL. Significantly, SACCHARIS v2 has been developed with high-throughput omics in mind, with pipeline automation geared toward complex (meta)genome and (meta)transcriptome datasets to reveal the total CAZyme content ("CAZome") of an organism or community. Here, we outline the development and use of SACCHARIS v2 to discover and annotate CAZymes and provide insight into complex carbohydrate metabolisms in individual organisms and communities.

Paragoniastreavariabilis Kishi, Nomura & Fukami, sp. nov. (Cnidaria, Anthozoa, Scleractinia), a new coral species previously considered as a variant of Paragoniastreadeformis, from Japan and northern Taiwan.

A new zooxanthellate scleractinian coral, Paragoniastreavariabilis Kishi, Nomura & Fukami, sp. nov. (Scleractinia, Merulinidae), is described from non-coral reef regions of Japan and northern Taiwan. This new species was previously recognized as a morphological variant of Paragoniastreadeformis (Veron, 1990) and can be morphologically distinguished from that species by lacking groove-and-tube structures on corallite wall joints, and by having larger calices, numerous septa, and up to three corallites in one valley. The new species also formed an independent clade from its congeners, P.australensis (Milne Edwards & Haime, 1857), P.deformis and P.russelli (Wells, 1954), in the molecular phylogeny based on the mitochondrial intergenic region and nuclear ribosomal internal transcribed spacers.

Phylogenomics and biogeography of the small carpenter bees (Apidae: Xylocopinae: Ceratina).

Small carpenter bees in the genus Ceratina are behaviourally diverse, species-rich, and cosmopolitan, with over 370 species and a range including all continents except Antarctica. Here, we present the first comprehensive phylogeny of the genus based on ultraconserved element (UCE) phylogenomic data, covering a total of 185 ingroup specimens representing 22 of the 25 current subgenera. Our results support most recognized subgenera as natural groups, but we also highlight several groups in need of taxonomic revision - particularly the nominate subgenus Ceratina sensu stricto - and several clades that likely need to be described as new subgenera. In addition to phylogeny, we explore the evolutionary history of Ceratina through divergence time estimation and biogeographic reconstruction. Our findings suggest that Ceratinini split from its sister tribe Allodapini about 72 million years ago. The common ancestor of Ceratina emerged in the Afrotropical realm approximately 42 million years ago, near the Middle Eocene Climatic Optimum. Multiple subsequent dispersal events led to the present cosmopolitan distribution of Ceratina, with the majority of transitions occurring between the Afrotropics, Indomalaya, and the Palearctic. Additional movements also led to the arrival of Ceratina in Madagascar, Australasia, and a single colonization of the Americas. Dispersal events were asymmetrical overall, with temperate regions primarily acting as destinations for migrations from tropical source regions.

Sequencing and analysis of the complete mitochondrial genome of Amrasca biguttula biguttula Ishida, 1913 (Hemiptera: Cicadellidae: Typhlocybinae: Empoascini).

The complete mitogenome of the cotton leafhopper, Amrasca biguttula biguttula Ishida, 1913, was sequenced and annotated. The mitogenome is 14,474 bp long and contains 37 genes, including 13 protein-coding genes (PCGs), two ribosomal RNA (rRNA) genes, and 22 transfer RNA (tRNA) genes, as well as a control region. The nucleotide composition of the mitogenome is as follows: A, 39.17%; T, 39.3%; C, 11.13%; and G, 10.39%. The total length of the 13 PCGs is 10,496 bp, which encodes 3503 amino acids. All PCGs start with the ATG codon, except for ATA, ATC, GTG, and ATT. Most of the PCGs stop with TGA, and the remaining with CCT, GAA, GGT, TCA, CCA, CTA, TTA, AAA, ATT, or ATA. The phylogenetic tree shows that A. biguttula biguttula belongs to Empoascini of the subfamily Typhlocybinae, but is different from other species within the subfamily.

Proterodiplostomid no longer: Molecular phylogeny reveals the true position of Proalarioides (Digenea: Diplostomoidea).

Proalarioides Yamaguti, 1933 (Digenea Carus, 1863: Diplostomoidea Poirier, 1886) is a small genus of proterodiplostomids parasitic in the intestines of snakes in Asia. Only two species are considered valid: Proalarioides serpentis Yamaguti, 1933 and Proalarioides tropidonotis Vidyarthi, 1937. Unlike other proterodiplostomids, Proalarioides spp. possess pseudosuckers and lack the paraprostate, otherwise extremely characteristic of the Proterodiplostomidae Dubois, 1936. In the present study, we describe the morphology of progenetic metacercariae of a Proalarioides sp. from bicolored frog, Clinotarsus curtipes (Jerdon), collected in India and provide the first DNA sequences from any member of the genus. These specimens differ from previously described metacercariae and adults of P. serpentis and P. tropidonotis in several ways, including body and organ sizes, sucker ratios, and distribution of vitellarium. The newly generated partial large ribosomal subunit (28S) rRNA gene sequence was used to test the phylogenetic position of the genus among other major lineages of diplostomoideans. Our 28S phylogeny clearly demonstrated Proalarioides sp. to be well-separated from other members of the Proterodiplostomidae. Based on morphological and molecular evidence, we transfer Proalarioides out of the Proterodiplostomidae into the Diplostomidae Poirier, 1886.

Phylogenetic analysis of Ellipsomyxa species (Myxosporea) and description of Ellipsomyxa gordeyi n. sp. from the gall bladder of mullets (Mugiliformes: Mugilidae) in Nha Trang Bay of the East Sea, Vietnam.

The coastal waters of Vietnam are home to a wide diversity of fishes, but the parasite diversity of these potential hosts is much less well characterized. To begin addressing this knowledge gap, we carried out surveys of myxozoan parasites in fishes collected from Nha Trang Bay in Vietnam's East Sea in 2018-2019. Mugilid fishes were collected in March-April 2018, January-February 2019, and November-December 2019, and examined for myxozoans. Myxospores consistent with those of the genus Ellipsomyxa were found in the gall bladder of four mullet species, and we thoroughly characterized those from Planiliza melinoptera. Myxospores were elliptoid and devoid of striation, with a distinct sinuous suture line. Polar capsules were pyriform and oriented toward the poles of the spore. Morphological features were compared to nominal species and this species from Vietnam was distinct. Phylogenetic analysis based on partial small subunit rDNA sequence revealed that broadly, Ellipsomyxa species split into three phylogenetic lineages, and although in some branches there are groupings by host family, habitat or locality, there are no clear phylogenetic patterns. The new species we encountered in P. melinoptera had a close sister relationship with Ellipsomyxa adlardi, with both species part of a larger subclade within the Ellipsomyxa lineage. Despite this phylogenetic similarity, these species were morphologically distinct, and partial large subunit DNA sequences were only 93% similar to each other. A combination of the morphological characteristics and molecular data suggest that this is an undescribed species and we propose the name Ellipsomyxa gordeyi n. sp.

Comparative Study of Two Himalayan Snow Trouts, Schizothorax esocinus and Schizothorax curvifrons Within the Schizothoracinae and Other Nearest Relatives of Cyprinidae, Inferred from Mitochondrial Sequences of Cytochrome b (Cyt-b) and Cytochrome Oxidase I (Co-I) Gene.

The Himalayan region encompasses varied aquatic ecosystems, characterized by the presence of diverse ichthyofauna, particularly represented by members of the Schizothorax genus, commonly referred to as snow trout. The primary objective of this work was to examine the molecular phylogeny of Schizothoracinae, specifically focusing on the two species, Schizothorax esocinus and Schizothorax curvifrons, which are known to inhabit the northern and north-eastern regions of the Himalayas. This investigation was conducted by analyzing the entire mitochondrial Cyt-b and Co-I gene sequences. The aligned Cyt-b and Co-I sequences for S. esocinus, S. curvifrons, and related members within the subfamily Schizothoracinae, spanned 1130 to 1141 and 1536 to 1551 base pairs, respectively. Using these gene, phylogenetic trees were created to compare Schizothoracinae species to other subfamilies of the family Cyprinidae (Barbinae, Alburninae, Leuciscinae, Xenocyprinae, Cyprininae, and Cultrinae). Genetic distances for Cyt-b and Co-I sequence at three hierarchical levels shows significant disparities in their average score. For Cyt-b, average p-distances for intraspecies, intragenus, and intrafamily were 2.13%, 4.1%, and 15.23%, respectively. Similarly, for Co-I, average p-distances were 1.19%, 3.6%, and 13.8% for intraspecies, intragenus, and intrafamily, respectively. Total number of haplotypes (h) based on Cyt-b and Co-I gene were 6 and 12 within the target Schizothorax spp. In the present study, the observed range of haplotype diversity (hd) for the Cyt-b gene varied from 0.00 to 0.847, with an average haplotype diversity of 0.847 ± 0.034. Similarly, for the Co-I gene, the observed haplotype diversity ranged from 0.00 to 0.931, with an average value of haplotype diversity estimated to be 0.931 ± 0.024. The results of the present study clearly shows that the representative species exhibited close affinities with members of Barbinae and Cyprininae, while other subfamilies formed distinct groups. The findings of the study also indicated that the Cyt-b and Co-I gene exhibits polymorphism and has the potential to serve as a marker for identifying genetic differentiation among populations based on ecological habitats. Mitochondrial Cyt-b and Co-I have been established as a universally accepted and validated genetic marker within a comprehensive bio-identification system at the species level.

Aspleniumguodanum (Aspleniaceae), a distinct new fern species from northern Guangdong, China, based on morphological data and molecular phylogeny.

A new spleenwort species, Aspleniumguodanum, was found and described from Danxia landform region in Guangdong, China. The new species has close resemblance to A.subcrenatum Ching ex S.H.Wu in morphology, but can be distinguished by having plants small, stipes and rachises not covered with fibrous scales, relatively fewer pairs of pinnae, pinnae short, pinna margin weakly biserrate, pinna apex acute and lower pinnae obviously reduced. Phylogenetic analyses, based on six plastid markers (atpB, rbcL, rps4 & rps4-trnS and trnL & trnL-F) of the new species and its relatives, support a close relationship between A.guodanum and A.subcrenatum. Only one population with no more than 50 individuals were found and, therefore, it is recommended to be classified as Critically Endangered (CR) following IUCN Red List Criteria.

Phylogeny and biogeography of harmochirine jumping spiders (Araneae: Salticidae).

We use ultraconserved elements (UCE) and Sanger data to study the phylogeny, age, and biogeographical history of harmochirine jumping spiders, a group that includes the species-rich genus Habronattus, whose remarkable courtship has made it the focus of studies of behaviour, sexual selection, and diversification. We recovered 1947 UCE loci from 43 harmochirine taxa and 4 outgroups, yielding a core dataset of 193 UCEs with at least 50 % occupancy. Concatenated likelihood and ASTRAL analyses confirmed the separation of harmochirines into two major clades, here designated the infratribes Harmochirita and Pellenita. Most are African or Eurasian with the notable exception of a clade of pellenites containing Habronattus and Pellenattus of the Americas and Havaika and Hivanua of the Pacific Islands. Biogeographical analysis using the DEC model favours a dispersal of the clade's ancestor from Eurasia to the Americas, from which Havaika's ancestor dispersed to Hawaii and Hivanua's ancestor to the Marquesas Islands. Divergence time analysis on 32 loci with 85 % occupancy, calibrated by fossils and island age, dates the dispersal to the Americas at approximately 4 to 6 million years ago. The explosive radiation of Habronattus perhaps began only about 4 mya. The phylogeny clarifies both the evolution of sexual traits (e.g., the terminal apophyses was enlarged in Pellenes and not subsequently lost) and the taxonomy. Habronattus is confirmed as monophyletic. Pellenattus is raised to the status of genus, and 13 species moved into it as new combinations. Bianor stepposus Logunov, 1991 is transferred to Sibianor, and Pellenes bulawayoensis Wesolowska, 1999 is transferred to Neaetha. A molecular clock rate estimate for spider UCEs is presented and its utility to inform prior distributions is discussed.

Acquisition of green algal photobionts enables both chlorolichens and chloro-cyanolichens to activate photosynthesis at low humidity without liquid water.

Cyanobacteria require liquid water for photosynthesis, whereas green algae can photosynthesise with water vapour alone. We discovered that several Lobaria spp. which normally have cyanobacteria as the sole photobiont, in some regions of the trans-Himalayas also harboured green algae. We tested whether green algal acquisition was: limited to high elevations; obtained from neighbouring chloro-Lobaria species; enabled photosynthesis at low humidity. Lobaria spp. were collected from 2000 to 4000 m elevation. Spectrophotometry quantified green algal abundance by measuring chlorophyll b (absent in cyanobacteria). Thalli cross-sections visually confirmed green algal presence. We sequenced gene regions: Lobaria (ITS-EF-1α-RPB2), green algae (18S-RBC-L) and Nostoc (16S). Phylogenetic analysis determined myco-photobiont associations. We used a custom closed-circuit gas exchange system with an infrared gas analyser to measure CO2 exchange rates for desiccated specimens at 33%, 76%, 86% and 98% humidity. Cross-sections revealed that the photobiont layers in putative cyano-Lobaria contained both cyanobacteria and green algae, indicating that they should be considered chloro-cyanolichens. Chloro-Lobaria had no visible cephalodia nor cyanobacteria in the photobiont layer. Chloro-Lobaria and chloro-cyano-Lobaria had comparable levels of chlorophyll b. Chloro-Lobaria usually contained Symbiochloris. Chloro-cyano-Lobaria mainly associated with Parachloroidium and Nostoc; infrequently with Symbiochloris, Apatococcus, Chloroidium, Pseudochlorella, Trebouxia. Sequences from two green algal genera were obtained from within some thalli. Desiccated specimens of every Lobaria species could attain net photosynthesis with light exposure and 33% humidity. CO2 exchange dynamics over a five-day period differed between species. At all elevations, chloro-cyano-Lobaria spp. had abundant green algae in the photobiont layer, but green algal strains mostly differed to those of chloro-Lobaria spp. Both chloro-Lobaria and chloro-cyano-Lobaria were capable of conducting photosynthesis without liquid water. The data strongly suggest that they attained positive net photosynthesis.

Hivanua, a new genus of harmochirine jumping spiders from the Marquesas Islands (Araneae, Salticidae, Harmochirina).

The genus Hivanuagen. nov. is established for the harmochirine jumping spiders of the Marquesas Islands, formerly placed in Habronattus F.O. Pickard-Cambridge, 1901 and Havaika Prószynski, 2002. The type species, Hivanuatekaosp. nov. is described, and five species described by Berland are re-illustrated and moved into the genus: Hivanuaflavipes (Berland, 1933), comb. nov., Hivanuanigrescens (Berland, 1933), comb. nov., Hivanuanigrolineata (Berland, 1933), comb. nov., Hivanuarufescens (Berland, 1934), comb. nov., and Hivanuatriangulifera (Berland, 1933), comb. nov. The female epigyne is much like that of Habronattus, Bianor Peckham & Peckham, 1896, and other harmochirines, with a centrally placed coupling pocket and two atria with crescent-shaped edges. The terminal apophysis of the male palp, which is variable throughout the pellenine subgroup of the Harmochirina, is absent in H.rufescens but present in H.tekaosp. nov., in which it is elbowed much as in Habronattus. These Pacific Island harmochirines, like the Havaika of Hawaii, appear to be largely foliage dwellers, unlike most of their continental relatives.

Four New Species and a New Combination of Boletaceae (Boletales) from Subtropical and Tropical China.

Previous studies have shown that boletes are abundant and diverse in China, especially in tropical and subtropical regions. In the present study, morphological, ecological, host relationship, and a four-locus (28S, tef1, rpb1, and rpb2) molecular phylogenetic analyses were used to study the family Boletaceae in subtropical and tropical China. Four new bluing species are described from three genera, viz. Boletellus verruculosus (Chinese name), Xerocomellus tenuis (Chinese name), Xer. brunneus (Chinese name), and Xerocomus zhangii (Chinese name). Moreover, the genus Nigroboletus is treated as a synonym of Xerocomellus, and a new combination, namely Xer. roseonigrescens (Chinese name), is proposed.

Coniocybe Ach. Revisited.

Calicioids form a research field that has encompassed ascomycetous fungi with stalked ascomata similar to those of the lichen genus Calicium. Early generic circumscriptions of calicioid lichens and fungi were mainly based on morphological and secondary chemistry information. After the introduction of molecular data, taxonomy in the group has been reconsidered. Here, based on a broad geographical sampling, Coniocybe Ach. was revised using molecular and morphological features. Three loci (ITS, LSU and rpb1) were compared to infer its phylogenetic position, and a total of 52 new sequences (14 ITS, 24 LSU and 14 rpb1) were produced. Apart from its type C. furfuracea, Coniocybe was revised and emended to also include C. brachypoda and C. confusa. In addition, a new species, Coniocybe eufuracea, was described, and a key to the species of Coniocybe was provided.

Revision of the Genus Rhagastis Rothschild & Jordan, 1903 (Lepidoptera: Sphingidae) from China, Based on Morphological and Phylogenetic Analyses.

Here, the taxonomy of the genus Rhagastis Rothschild & Jordan, 1903 (Lepidoptera, Sphingidae, Macroglossinae, Macroglossini) from China is revised based on differences in wing morphology, male and female genitalia, and the phylogenetic relationship of the DNA barcodes. Subspecies of Rhagastis albomarginatus (Rothschild, 1894) and R. castor (Walker, 1856) are treated as "good" species, namely Rhagastis dichroae Mell, 1922 stat. nov.; R. everetti Rothschild & Jordan, 1903 stat. nov.; R. aurifera (Butler, 1875) stat. rev.; R. chinensis Mell, 1922 stat. nov.; R. formosana Clark, 1925 stat. nov.; and R. jordani Oberthür, 1904 stat. rev. The distribution maps, biological notes, and ecological records of the genus Rhagastis Rothschild & Jordan, 1903 from China are given, and a species inventory of genus Rhagastis in the world is also included.

Distinct evolutionary lineages of Schistocephalus parasites infecting co-occurring sculpin and stickleback fishes in Alaska.

Sculpins (coastrange and slimy) and sticklebacks (ninespine and threespine) are widely distributed fishes cohabiting 2 south-central Alaskan lakes (Aleknagik and Iliamna), and all these species are parasitized by cryptic diphyllobothriidean cestodes in the genus Schistocephalus. The goal of this investigation was to test for host-specific parasitic relationships between sculpins and sticklebacks based upon morphological traits (segment counts) and sequence variation across the NADH1 gene. A total of 446 plerocercoids was examined. Large, significant differences in mean segment counts were found between cestodes in sculpin (mean = 112; standard deviation [s.d.] = 15) and stickleback (mean = 86; s.d. = 9) hosts within and between lakes. Nucleotide sequence divergence between parasites from sculpin and stickleback hosts was 20.5%, and Bayesian phylogenetic analysis recovered 2 well-supported clades of cestodes reflecting intermediate host family (i.e. sculpin, Cottidae vs stickleback, Gasterosteidae). Our findings point to the presence of a distinct lineage of cryptic Schistocephalus in sculpins from Aleknagik and Iliamna lakes that warrants further investigation to determine appropriate evolutionary and taxonomic recognition.

A review of Nearctic Lathrobium (Coleoptera, Staphylinidae), with revision and descriptions of new flightless species from the mountains of the southeastern U.S.

Species of the genus Lathrobium Gravenhorst (Coleoptera: Staphylinidae: Paederinae) from North America north of Mexico are reviewed and 41 species are recognized. Morphology and mitochondrial COI sequence data were used to guide species designations in three flightless lineages endemic to the southern Appalachian Mountains, a biologically diverse region known for cryptic diversity. Using a combination of phylogeny, algorithm-based species delimitation analyses, and genitalic morphology, five new cryptic species are described and possible biogeographic scenarios for their speciation hypothesized: L.balsamense Haberski & Caterino, sp. nov., L.camplyacra Haberski & Caterino, sp. nov., L.islae Haberski & Caterino, sp. nov., L.lividum Haberski & Caterino, sp. nov., L.smokiense Haberski & Caterino, sp. nov. Five additional species are described: L.absconditum Haberski & Caterino, sp. nov., L.hardeni Haberski & Caterino, sp. nov., L.lapidum Haberski & Caterino, sp. nov., L.solum Haberski & Caterino, sp. nov., and L.thompsonorum Haberski & Caterino, sp. nov. Two species are transferred from Lathrobium to Pseudolathra Casey: Pseudolathraparcum (LeConte, 1880), comb. nov. and Pseudolathratexana (Casey, 1905), comb. nov. Twenty-six names are reduced to synonymy. Lectotypes are designated for 47 species. Larvae are described where known, and characters of possible diagnostic value are summarized. Species diagnoses, distributions, illustrations of male and female genitalia, and a key to Lathrobium species known from the Nearctic region (including several introduced species) are provided.

From Caves to the Savannah, the Mitogenome History of Modern Lions (Panthera leo) and Their Ancestors.

Lions (Panthera leo) play a crucial ecological role in shaping and maintaining fragile ecosystems within Africa. Conservation efforts should focus on genetic variability within wild populations when considering reintroduction attempts. We studied two groups of lions from two conservation sites located in Zambia and Zimbabwe to determine their genetic make-up, information that is usually unknown to the sites. In this study, we analysed 17 specimens for cytb and seven microsatellite markers to ascertain family relationships and genetic diversity previously obtained by observational studies. We then produced a standardised haplogroup phylogeny using all available entire mitogenomes, as well as calculating a revised molecular clock. The modern lion lineage diverged ~151 kya and was divided into two subspecies, both containing three distinct haplogroups. We confirm that Panthera leo persica is not a subspecies, but rather a haplogroup of the northern P.l. leo that exited Africa at least ~31 kya. The progenitor to all lions existed ~1.2 Mya, possibly in SE Africa, and later exited Africa and split into the two cave lion lineages ~175 kya. Species demography is correlated to major climactic events. We now have a detailed phylogeny of lion evolution and an idea of their conservation status given the threat of climate change.

Of sea, rivers and symbiosis: Diversity, systematics, biogeography and evolution of the deeply diverging florideophycean order Hildenbrandiales (Rhodophyta).

The Hildenbrandiales, a typically saxicolous red algal order, is an early diverging florideophycean group with global significance in marine and freshwater ecosystems across diverse temperature zones. To comprehensively elucidate the diversity, phylogeny, biogeography, and evolution of this order, we conducted a thorough re-examination employing molecular data derived from nearly 700 specimens. Employing a species delimitation method, we identified Evolutionary Species Units (ESUs) within the Hildenbrandiales aiming to enhance our understanding of species diversity and generate the first time-calibrated tree and ancestral area reconstruction for this order. Mitochondrial cox1 and chloroplast rbcL markers were used to infer species boundaries, and subsequent phylogenetic reconstructions involved concatenated sequences of cox1, rbcL, and 18S rDNA. Time calibration of the resulting phylogenetic tree used a fossil record from a Triassic purportedly freshwater Hildenbrandia species and three secondary time points from the literature. Our species delimitation analysis revealed an astounding 97 distinct ESUs, quintupling the known diversity within this order. Our time-calibration analysis placed the origin of Hildenbrandiales (crown age) in the Ediacaran period, with freshwater species emerging as a monophyletic group during the later Permian to early Triassic. Phylogenetic reconstructions identified seven major clades, experiencing early diversification during the Silurian to Carboniferous period. Two major evolutionary events-colonization of freshwater habitats and obligate systemic symbiosis with a marine fungus-marked this order, leading to significant morphological alterations without a commensurate increase in species diversification. Despite the remarkable newly discovered diversity, the extant taxon diversity appears relatively constrained when viewed against an evolutionary timeline spanning over 800 million years. This limitation may stem from restricted geographic sampling or the prevalence of asexual reproduction. However, species richness estimation and rarefaction analyses suggest a substantially larger diversity yet to be uncovered-potentially four times greater. These findings drastically reshape our understanding of the deeply diverging florideophycean order Hildenbrandiales species diversity, and contribute valuable insights into this order's evolutionary history and ecological adaptations. Supported by phylogenetic, ecological and morphological evidence, we established the genus Riverina gen. nov. to accommodate freshwater species of Hildenbrandiales, which form a monophyletic clade in our analyses. This marks the first step toward refining the taxonomy of the Hildenbrandiales, an order demanding thorough revisions, notably with the creation of several genera to address the polyphyletic status of Hildenbrandia. However, the limited diagnostic features pose a challenge, necessitating a fresh approach to defining genera. A potential solution lies in embracing a molecular systematic perspective, which can offer precise delineations of taxonomic boundaries.

Multi-gene phylogeny of the primary freshwater crab genus Ptychophallus Smalley, 1964 (Pseudothelphusidae: Ptychophallinae) from the Neotropical region.

The diversity of primary freshwater crabs of Central America is underestimated and poorly known, specially when considering both Atlantic and Pacific drainages distribution. Among the family Pseudothelphusidae Ortmann, 1893, the genus Ptychophallus Smalley, 1964 was recently revised using morphological data and encompassing 14 valid species, which are distributed exclusively in Costa Rica and Panama. Despite the informative scenario constructed with this previous study, some uncertainties remained such as those related to the populations distributed along different drainages and the lack of complete information regarding the phylogenetic relationships among the species that belong to this genus. Our phylogenetic reconstruction based on multigenes revealed four lineages. Ptychophallus costaricensis, P. colombianus, P. lavallensis, P. montanus, and P. tumimanus are well-established species, while P. paraxanthusi, P. exilipes and P. tristani were recovered as non-monophyletic in some relationships. Ptychophallus tristani and P. paraxanthusi were determined to be close relatives one with other, but with some representatives showing no clear correspondence and positioning in the molecular phylogeny, which raised doubts about the validity as separate species and/or the accuracy of identifications for some GenBank sequences. Ptychophallus tumimanus and P. montanus are morphologically very similar, but they are positioned in distinct clades. The genetic results confirmed a previous proposal that P. barbillaensis is a junior synonym of P. uncinatus, and P. bilobatus is closely related to P. uncinatus. The findings revealed the presence of one peculiar lineage with one species from Costa Rica compound by the resurrected P. campylus and another species from Costa Rica, originally identified as "P. cf. exilipes"; its morphology, however, did not match with any other analyzed species, which requires further analysis. Finally, possible misidentification in some sequences deposited in the GenBank were detected and should be reviewed. The reconstructed molecular phylogeny for the genus that inhabits both Atlantic and Pacific drainages of Central America provides a better understanding of the knowledge on the evolution of freshwater crabs and represents key information that will serve as baseline for further taxonomic studies on the cryptic biodiversity of this group.