Large Clade Research Articles

DNA barcoding of sea urchin species in the bird’s head seascape Papua-Indonesia

The DNA barcoding of COI gene has been used for molecular identification of the numerous individuals of sea urchin present in Bird’s Head Seascape (BHS) Papua in Indonesia. We extracted, sequenced, and analysed DNA of a total of 29 individuals collected from Raja Ampat, Rumberpon, Fakfak and Manokwari. The fragment gene of cytochrome oxidase subunit 1, were amplified with polymerase chain reaction by using coif and coitri primers. Total 570 base pairs nucleotide from each sample were analysed with MEGA X. The results of sample sequence homology with the GenBank and BOLD System databases show that some samples can be identified at the species level, and some samples can only be identified at the genus level. The phylogenetic tree results show that all samples are separated into two large clades.

Molecular identification of sharks and rays species from Aceh waters, Indonesia

Shark is a cartilaginous fish that is widely hunted because it has high economic value. The waters of Aceh are directly adjacent to the Indian Ocean and the Malacca Strait making it a preferred habitat for pelagic fish, especially sharks. Information on shark species landed in the waters west and east of Aceh is very limited due to difficulties in identification and commonly used local names. This study aimed to determine the types of sharks landed in Banda Aceh, Meulaboh, Langsa, Southwest Aceh and Simeulu and to find out specifically what species of sharks are most often landed in Aceh waters. Primary data gathering consisted of four stages, namely sample collection, identification using identification books, molecular identification cytochrome oxidase subunit I (COI) and phylogenetic analysis. The results of the molecular analysis of 46 tissue samples from five locations identified 13 species of sharks, namely Carcharhinhus sorrah, Carcharhinhus amboinensis, Triaenodon obesus, Isurus oxyrinchus, Sphyrna zygaena, Sphyrna lewini, Loxodon macrorhinus, Hemipristis elongaria, Stagostoma fasciatum, Nebrius ferrugineus, Chilloscyllium punctatum, Isurus oxyrinchus, Alopias pelagicus, Alopias supercillosus and 1 species of rays, namely Rhynchobatus australiae. Phylogenetic tree reconstruction using the Neighbor Joining method of 610 basepairs consisting of two large clades separates the species Alopias pelagicus and Isurus oxyrichus with Carcharhinus sorrah, Sphyrna lewini, Loxodon macrorhinus and Rhyncobatus australiae with boostrap values of 87% and 64%. The haplotype diversity shown ranged from 0.667-0.889 while the nucleotide diversity ranged from 0.001-0.097. These values indicates high diversity because of the variance in the number of species found.Keywords:BarcodeCOISharks catchAlopias supercillosusSphyrna lewini

New species of Abyssocladia and two new cladorhizid genera (Porifera, Cladorhizidae) from New Zealand and Australia.

Seamounts on subantarctic New Zealand's Macquarie Ridge, including parts of Australia's Exclusive Economic Zone surrounding Macquarie Island, have been demonstrated to be a rich source of new species of carnivorous sponges (Demospongiae Sollas, Poecilosclerida Topsent, Cladorhizidae Dendy). Four new species of Abyssocladia Lévi, 1964, are described from Macquarie Ridge seamounts and at other disparate locations: Abyssocladia lanceola sp. nov. from Seamounts 7, 8, and 9 (Australia EEZ), Seamount 10 (International Waters), and the South Tasman Rise; Abyssocladia rowdeni sp. nov., first collected from diffuse hydrothermal vent sites at Brothers Seamount on the Southern Kermadec Ridge and recorded here from the non-venting seamounts on Chatham Rise to the east of the South Island of New Zealand; Abyssocladia tumulorum sp. nov., found exclusively on the Chatham Rise; and Abyssocladia sonnae sp. nov. from Monowai Seamount on the Tonga-Kermadec Ridge in International Waters, also found, surprisingly, on Macquarie Ridge's Seamount 8 (Australia EEZ). Patriciacladia gen. nov. has been established for a new species of Cladorhizidae discovered on Macquarie Ridge and Chatham Rise. Patriciacladia enigmatica gen. et sp. nov. is highly unusual in that it possesses palmate isochelae not typically found in Cladorhizidae and has a long branch in phylogenetic analysis of the family, supporting the establishment of a new genus and species for Abyssocladia n. sp. B (QM G339872, was NIWA 41033): 28S rDNA: LN870583, COI: LN870445, Macquarie Ridge) in Hestetun et al. (2016a: table 1; 2017: fig. 15). The discovery of two new species, again from the Macquarie Ridge and other New Zealand locations, expands support for the establishment of a new genus, Australocladia gen. nov., which contains several additional species nested as a monophyletic clade within the large, heterogenous, and paraphyletic Abyssocladia clade in molecular phylogenetic analyses. Australocladia sphaerichela gen. et sp. nov. and Au. alopecura gen. et sp. nov. both possess spherical abyssochelae, funnel-shaped expansions which may contain spermatophores on the body, substrongyles in the attachment base, and a generally southern hemisphere distribution.

Mirusviruses link herpesviruses to giant viruses

DNA viruses have a major influence on the ecology and evolution of cellular organisms1–4, but their overall diversity and evolutionary trajectories remain elusive5. Here we carried out a phylogeny-guided genome-resolved metagenomic survey of the sunlit oceans and discovered plankton-infecting relatives of herpesviruses that form a putative new phylum dubbed Mirusviricota. The virion morphogenesis module of this large monophyletic clade is typical of viruses from the realm Duplodnaviria6, with multiple components strongly indicating a common ancestry with animal-infecting Herpesvirales. Yet, a substantial fraction of mirusvirus genes, including hallmark transcription machinery genes missing in herpesviruses, are closely related homologues of giant eukaryotic DNA viruses from another viral realm, Varidnaviria. These remarkable chimaeric attributes connecting Mirusviricota to herpesviruses and giant eukaryotic viruses are supported by more than 100 environmental mirusvirus genomes, including a near-complete contiguous genome of 432 kilobases. Moreover, mirusviruses are among the most abundant and active eukaryotic viruses characterized in the sunlit oceans, encoding a diverse array of functions used during the infection of microbial eukaryotes from pole to pole. The prevalence, functional activity, diversification and atypical chimaeric attributes of mirusviruses point to a lasting role of Mirusviricota in the ecology of marine ecosystems and in the evolution of eukaryotic DNA viruses.

Tracing the spatiotemporal phylodynamics of Japanese encephalitis virus genotype I throughout Asia and the western Pacific.

Japanese encephalitis virus (JEV; Flaviridae: Flavivirus) causes Japanese encephalitis (JE), which is the most important arboviral disease in Asia and the western Pacific. Among the five JEV genotypes (GI-V), GI has dominated traditional epidemic regions in the past 20 years. We investigated the transmission dynamics of JEV GI through genetic analyses. We generated 18 JEV GI near full length sequences by using multiple sequencing approaches from mosquitoes collected in natural settings or from viral isolates obtained through cell culture. We performed phylogenetic and molecular clock analyses to reconstruct the evolutionary history by integrating our data with 113 publicly available JEV GI sequences. We identified two subtypes of JEV GI (GIa and GIb), with a rate of 5.94 × 10-4 substitutions per site per year (s/s/y). At present, GIa still circulates within a limited region, exhibited no significant growth, the newest strain was discovered in China (Yunnan) in 2017, whereas most JEV strains circulating belong to the GIb clade. During the past 30 years, two large GIb clades have triggered epidemics in eastern Asia: one epidemic occurred in 1992 [95% highest posterior density (HPD) = 1989-1995] and the causative strain circulates mainly in southern China (Yunnan, Shanghai, Guangdong, and Taiwan) (Clade 1); the other epidemic occurred in 1997 (95% HPD = 1994-1999) and the causative strain has increased in circulation in northern and southern China during the past 5 years (Clade 2). An emerging variant of Clade 2 contains two new amino acid markers (NS2a-151V, NS4b-20K) that emerged around 2005; this variant has demonstrated exponential growth in northern China. JEV GI stain circulating in Asia have shifted during the past 30 years, spatiotemporal differences were observed among JEV GI subclade. GIa is still circulating within a limited range, exhibite no significant growth. Two large GIb clades have triggered epidemics in eastern Asia, all JEV sequences identified in northern China during the past 5 years were of the new emerging variant of G1b-clade 2.

Genome-Wide Identification of β-1,3-Glucanase Genes in Hevea brasiliensis

β-1,3-glucanase is one of the pathogenesis-related proteins well-known for their antifungal properties which can be abundantly found in Hevea brasiliensis. Utilization of β-1,3-glucanase in the genetic improvement of H. brasiliensis is very important as the high susceptibility to various fungal infections has challenged the current natural rubber industry. A few nucleotide sequences for β-1,3-glucanase have been reported and their role in biotic stress management has been demonstrated. Being a multigene family, it is necessary to identify and characterize more isoforms of β-1,3-glucanase to select the most suitable isoform to be utilized in genetic improvement. In the current study, we conducted a genome-wide identification of β-1,3-glucanases in H. brasiliensis, their classification based on the functional domains and phylogenetic analysis, using different bioinformatics tools. All publicly available nucleotide sequences were collected and curated by eliminating sequences that lack glycoside hydrolase family 17 (GH 17) domain as well as the partial and closely identical sequences and obtained 14 full-length sequences. The sequences were categorized into 4 distinct classes (I-IV) based on their functional domains and C-terminal extension. Class III and IV which lack the carbohydrate-binding C-terminal X8 domain are the largest classes identified with 5 β-1,3-glucanase each while 4 β-1,3-glucanase contain a variable C-terminal X8 domain. Phylogenetic analysis showed the clustering of β-1,3-glucanases into six major clades (I-VI) based on the domains. Clades I and II were identified as the largest clades with 4 β-1,3-glucanase in each. Several paralogous clusters have been observed for H. brasiliensis indicating the gene family expansion within the species or in the immediate ancestors with possible species-specific function. Further functional characterization is necessary to select the suitable gene to be utilized in genetic improvement and the present study provides a platform for it.

Reassessment of generic boundaries in Neotropical Chrysophylloideae (Sapotaceae): Eleven reinstated genera and narrowed circumscriptions of Chrysophyllum and Pouteria

AbstractClassifications of the pantropical plant family Sapotaceae based solely on morphology have historically recognized between 125 and 53 genera. Phylogenetic analyses using molecular data have repeatedly demonstrated that broad concepts of two large genera belonging to subfamily Chrysophylloideae, Chrysophyllum and Pouteria, are untenable and their narrowed delimitations have restricted them to the Neotropics. A recent phylogenetic study proposed further amendments by resurrecting the genera Achrouteria, Cornuella, Lucuma, Martiusella, Nemaluma, Prieurella and Ragala, and questioned the status of three generally accepted genera, Chromolucuma, Pradosia and Sarcaulus. We test this suggested classification using expanded sampling that comprises 122 terminals, including material of 29 of the 34 name‐bringing species for generic names historically regarded as synonyms of Chrysophyllum and Pouteria. We used sequence data from ribosomal nrDNA (ETS, ITS), the nuclear gene RPB2, two cpDNA spacers (petN‐psbM, trnH‐psbA), and indel information to estimate phylogenetic relationships in a Bayesian framework using BEAST. All sequences were newly realigned to test reproducibility, and 26 morphological characters were mapped on the resulting tree. Our analyses recovered three African genera embedded within a large Neotropical clade of Chrysophylloideae. We found strong support for the reinstatement of the seven genera listed above as well as for four other genera, viz. Chloroluma, Englerella, Labatia, and Peteniodendron. This subsequently leads to further amendments of Chrysophyllum and Pouteria, which are now limited to include 25–30 and 7 species, respectively. However, one clade that includes many name‐bringing lineages largely corresponds to Pouteria s.l., a group that needs further phylogenetic research to unravel relationships and generic limits. The hypothesis that Chrysophyllum cuneifolium had an inter‐continental hybrid origin involving genomes from Africa and South America is rejected because it is shown to have been based on erroneous results obtained from a contaminated DNA aliquot. A total of 73 genera are currently recognised in Sapotaceae, 21 of which are Neotropical members of Chrysophylloideae. We update the nomenclature and synonymy of 75 species, make 36 new combinations, and designate lectotypes for 31 names.

Phylotranscriptomics of Swertiinae (Gentianaceae) reveals that key floral traits are not phylogenetically correlated.

Establishing how lineages with similar traits are phylogenetically related remains critical for understanding the origin of biodiversity on Earth. Floral traits in plants are widely used to explore phylogenetic relationships and to delineate taxonomic groups. The subtribe Swertiinae (Gentianaceae) comprises more than 350 species with high floral diversity ranging from rotate to tubular corollas and possessing diverse nectaries. Here we performed phylogenetic analysis of 60 species from all 15 genera of the subtribe Swertiinae sensu Ho and Liu, representing the range of floral diversity, using data from the nuclear and plastid genomes. Extensive topological conflicts were present between the nuclear and plastome trees. Three of the 15 genera represented by multiple species are polyphyletic in both trees. Key floral traits including corolla type, absence or presence of lobe scales, nectary type, nectary position, and stigma type are randomly distributed in the nuclear and plastome trees without phylogenetic correlation. We also revealed the likely ancient hybrid origin of one large clade comprising 10 genera with diverse floral traits. These results highlight the complex evolutionary history of this subtribe. The phylogenies constructed here provide a basic framework for further exploring the ecological and genetic mechanisms underlying both species diversification and floral diversity.

Concurrent infections of Streptococcus iniae and Aeromonas veronii in farmed Giant snakehead (Channa micropeltes).

The giant snakehead, Channa micropeltes, is an increasingly important economic freshwater fish in Thailand and other regions of Asia. Presently, giant snakehead are cultured under intensive aquaculture conditions, leading to high stress and conditions favouring disease. In this study, we reported a disease outbreak in farmed giant snakehead with a cumulative mortality of 52.5%, continuing for 2 months. The affected fish exhibited signs of lethargy, anorexia and haemorrhage of the skin and eyes. Further bacterial isolations revealed two different types of colonies on tryptic soy agar: small white, punctate colonies of gram-positive cocci and cream-coloured, round and convex colonies of rod-shaped gram-negative bacteria. Additional biochemical and species-specific PCR analysis based on 16S rRNA confirmed the isolates as Streptococcus iniae and Aeromonas veronii. Multilocus sequence analysis (MLSA) placed the S.iniae isolate into a large clade of strains from clinically infected fish worldwide. Gross necropsy findings showed liver congestion, pericarditis and white nodules in the kidney and liver. Histologically, the affected fish showed focal to multifocal granulomas with inflammatory cell infiltration in kidney and liver, enlarged blood vessels with mild congestion within the meninges of the brain and severe necrotizing and suppurative pericarditis with myocardial infarction. Antibiotic susceptibility tests revealed that S.iniae was sensitive to amoxicillin, erythromycin, enrofloxacin, oxytetracycline, doxycycline and resistant to sulfamethoxazole-trimethoprim, while the A.veronii was susceptible to erythromycin, enrofloxacin, oxytetracycline, doxycycline, sulfamethoxazole-trimethoprim and resistant to amoxicillin. Conclusively, our findings highlighted the natural concurrent bacterial infections in cultured giant snakehead, which support the implementation of appropriate treatment and control strategies.

Complete chloroplast genome sequence of Solanum iopetalum, one of the tuber-bearing wild potato relatives

Solanum iopetalum belongs to the Solanaceae family and is one of the tuber-bearing wild Solanum species. In this study, chloroplast genome sequencing of the species, completed with Illumina sequencing technology, is presented. The length of the chloroplast genome is 155,625 bp with a GC content of 37.86%. It comprises a large single copy (LSC) region of 86,057 bp, a small single copy (SSC) region of 18,382 bp, and two inverted repeat regions (IRa and IRb) of 25,593 bp. Additionally, 158 functional genes in the genome are identified, including 105 protein-coding genes, 8 ribosomal RNAs, and 45 transfer RNAs. Phylogenetic analysis revealed that S. iopetalum is grouped into a large clade with other Solanum species, including cultivated potatoes (S. tuberosum) and is closely related to Mexican Solanum species (S. stoloniferum, S. verrucosum, S. hougasii, S. hjertingii, and S. demissum). This study provides useful genomic information for future breeding and evolutionary studies of S. iopetalum and other Solanum species.

Prevalence and Molecular Characterization of Bovine Parainfluenza Virus Type 3 in Cattle Herds in China.

Bovine parainfluenza virus type 3 (BPIV3) is a common respiratory pathogen that causes respiratory illness in cattle and makes a major contribution to the bovine respiratory disease complex (BRDC); however, data on the prevalence and molecular features of BPIV3 are still scarce in China. To investigate the epidemiological characteristics of BPIV3 in China, between September 2020 and June 2022, 776 respiratory samples were received from 58 BRDC-affected farms located in 16 provinces and one municipality. Those were screened for BPIV3 using a reverse transcription insulated isothermal PCR (RT-iiPCR) assay. Meanwhile, the HN gene and complete genome sequence of strains from different provinces were amplified, sequenced, and analyzed. The tests showed that 18.17% (141/776) of samples tested were positive for BPIV3, which originated from 21 farms in 6 provinces. Moreover, 22 complete HN gene sequences and 9 nearly complete genome sequences were obtained from the positive samples. Phylogenetic analysis based on the HN gene and complete genome sequences revealed that the sequences were clustered in one large clade for all Chinese BPIV3 genotype C strains, while overseas strain sequences of BPIV3 genotype C clustered into other clades. Moving beyond the known complete genome sequences of BPIV3 in GenBank, a total of five unique amino acid mutations were found in N protein, F protein, and HN protein in Chinese BPIV3 genotype C strains. Taken together, this study reveals that BPIV3 genotype C strains, the dominant strains in China, have a broad geographical distribution and some unique genetic characteristics. These findings contribute to our understanding of the epidemiological characteristics and genetic evolution of BPIV3 in China.

Morphological and molecular characterization of Cosmocercoides amapari n. sp. (Nematoda: Cosmocercidae), parasitic in hylid frogs from the Brazilian Amazon.

Cosmocercoides Wilkie, 1930 are gastrointestinal parasites commonly found in amphibians and reptiles, with 4 species reported from the Neotropical region. In the present study, a new species of Cosmocercoides, namely Cosmocercoides amapari n. sp. is described using integrated approaches such as light and scanning microscopy, and DNA sequencing of the mitochondrial cytochrome c oxidase subunit 1 gene. The specimens were collected from the large intestine of 3 species of hylid frogs in Amapá, Brazil. The new species can be distinguished from its congeners by morphological traits, including the pattern of caudal papillae, absence of the gubernaculum, 2 poorly sclerotized spicules, presence of lateral alae and somatic papillae along the body. In addition, our molecular analyses and phylogenetic reconstructions strongly support the status of the new taxon, which clustered poorly with a large clade of Cosmocerca spp. Cosmocercoides amapari n. sp. is the 29th species of the genus, the 5th from the Neotropical region, the third reported in Brazil, the second described from the Amazon region and the first belonging to the Neotropical region with molecular data.

Farnesyl/geranylgeranyl diphosphate synthases regulate the biosynthesis of alarm pheromone in a unique manner in the vetch aphid Megoura viciae.

Farnesyl/geranylgeranyl diphosphate synthases (FPPS/GGPPS) as the short-chain prenyltransferases catalyse the formation of the acyclic precursors (E)-FPP and (E)-GGPP for isoprenoid biosynthesis. Here, we first cloned the cDNAs encoding FPPS and GGPPS in the vetch aphid Megoura viciae (designated as MvFPPS and MvGGPPS). They had an open reading frame of 1185 and 930 bp in length, encoding 395 and 309 amino acids, with a theoretical isoelectric point of 6.52 and 6.21, respectively. Sequence alignment and phylogenetic analysis showed that MvFPPS and MvGGPPS shared the conserved aspartate-rich motifs characterized by all prenyltransferases identified to date and were clustered with their homologues in two large clades. RNA interference (RNAi) combined with gas chromatography/mass spectrometry (GC-MS) analysis showed that both MvFPPS and MvGGPPS were involved in the biosynthesis of alarm pheromone. Spatiotemporal expression profiling showed that the expression of MvFPPS and MvGGPPS was significantly higher in embryos than in other tissues. RNAi and GC-MS performed specifically in embryos corroborated the function of MvFPPS and MvGGPPS. In vitro, enzymatic activity assay and product analysis demonstrated that MvFPPS could catalysed the formation of (E)-FPP using DMAPP or (E)-GPP as the allylic cosubstrates in the presence of IPP, while MvGGPPS could only use (E)-GPP or (E)-FPP as cosubstrates. Functional interaction analysis using RNAi revealed that MvGGPPS exerts unidirectional functional compensation for MvFPPS. Moreover, it can regulate the biosynthesis of alarm pheromone by imposing a negative feedback regulation on MvFPPS. Our study helps to understand the molecular regulatory mechanism of terpenoid biosynthesis in the aphid.

Mitochondrial phylogenomics reveals deep relationships of scarab beetles (Coleoptera, Scarabaeidae).

In this study, we newly sequenced the complete mitochondrial genomes (mitogenomes) of two phytophagous scarab beetles, and investigated the deep level relationships within Scarabaeidae combined with other published beetle mitogenome sequences. The complete mitogenomes of Dicronocephalus adamsi Pascoe (Cetoniinae) and Amphimallon sp. (Melolonthinae) are 15,563 bp and 17,433 bp in size, respectively. Both mitogenomes have the typical set of 37 genes (13 protein-coding genes, 22 transfer RNA genes, two ribosomal RNA genes) and an A+T-rich region, with the same gene arrangement found in the majority of beetles. The secondary structures for ribosomal RNA genes (rrnL and rrnS) were inferred by comparative analysis method. Results from phylogenetic analyses provide support for major lineages and current classification of Scarabaeidae. Amino acid data recovered Scarabaeidae as monophyletic. The Scarabaeidae was split into two clades. One clade contained the subfamilies Scarabaeinae and Aphodiinae. The other major clade contained the subfamilies Dynastinae, Rutelinae, Cetoniinae, Melolonthinae and Sericini. The monophyly of Scarabaeinae, Aphodiinae, Dynastinae, Cetoniinae and Sericini were strongly supported. The Scarabaeinae was the sister group of Aphodiinae. The Cetoniinae was sister to the Dynastinae + Rutelinae clade. The Melolonthinae was a non-monophyletic group. The removal of fast-evolving sites from nucleotide dataset using a pattern sorting method (OV-sorting) supported the family Scarabaeidae as a monophyletic group. At the tribe level, the Onthophagini was non-monophyletic with respect to Oniticellini. Ateuchini was sister to a large clade comprising the tribes Onthophagini, Oniticellini and Onitini. Eurysternini was a sister group of the Phanaeini + Ateuchini clade.

Testing the chilling- before drought-tolerance hypothesis in Pooideae grasses.

Temperate Pooideae are a large clade of economically important grasses distributed in some of the Earth's coldest and driest terrestrial environments. Previous studies have inferred that Pooideae diversified from their tropical ancestors in a cold montane habitat, suggesting that above-freezing cold (chilling) tolerance evolved early in the subfamily. By contrast, drought tolerance is hypothesized to have evolved multiple times independently in response to global aridification that occurred after the split of Pooideae tribes. To independently test predictions of the chilling-before-drought hypothesis in Pooideae, we assessed conservation of whole plant and gene expression traits in response to chilling vs. drought. We demonstrated that both trait responses are more similar across tribes in cold as compared to drought, suggesting that chilling responses evolved before, and drought responses after, tribe diversification. Moreover, we found significantly more overlap between drought and chilling responsive genes within a species than between drought responsive genes across species, providing evidence that chilling tolerance genes acted as precursors for the novel acquisition of increased drought tolerance multiple times independently, partially through the cooption of chilling responsive genes.

Multi-Locus Sequencing Reveals Putative Novel Anaplasmataceae Agents, 'Candidatus Ehrlichia dumleri' and Anaplasma sp., in Ring-Tailed Coatis (Carnivora: Nasua nasua) from Urban Forested Fragments at Midwestern Brazil.

The Anaplasmataceae family encompasses obligate intracellular α-proteobacteria of human and veterinary medicine importance. This study performed multi-locus sequencing to characterize Ehrlichia and Anaplasma in coati's blood samples in Midwestern Brazil. Twenty-five samples (25/165-15.1%) were positive in the screening PCR based on the dsb gene of Ehrlichia spp. and were characterized using 16S rRNA, sodB, groEL, and gltA genes and the 23S-5S intergenic space region (ITS). Phylogenetic analyses based on all six molecular markers positioned the sequences into a new clade, with a common origin of Ehrlichia ruminantium. Haplotype analyses of 16S RNA sequences revealed the presence of two distinct Ehrlichia genotypes. Six samples (6/165, 3.6%) were positive in the screening nPCR for the 16S rRNA gene of Anaplasma spp. and were submitted to an additional PCR targeting the ITS for molecular characterization. Phylogenetic analyses based on both 16S rRNA gene and ITS positioned the Anaplasma sp. detected in the present study in a large clade with other Anaplasma sp. previously detected in ticks and wild animals and in a clade with 'Candidatus Anaplasma brasiliensis', respectively. Based on distinct molecular markers, the present work described a putative novel Anaplasmataceae agent, namely 'Candidatus Ehrlichia dumleri', and Anaplasma sp. closely related to the previously described 'Candidatus Anaplasma brasiliensis'.

Salmonella enterica serovar Cerro displays a phylogenetic structure and genomic features consistent with virulence attenuation and adaptation to cattle.

Salmonella enterica subsp. enterica (S.) serovar Cerro is rarely isolated from human clinical cases of salmonellosis but represents the most common serovar isolated from cattle without clinical signs of illness in the United States. In this study, using a large, diverse set of 316 isolates, we utilized genomic methods to further elucidate the evolutionary history of S. Cerro and to identify genomic features associated with its apparent virulence attenuation in humans. Phylogenetic analyses showed that within this polyphyletic serovar, 98.4% of isolates (311/316) represent a monophyletic clade within section Typhi and the remaining 1.6% of isolates (5/316) form a monophyletic clade within subspecies enterica Clade A1. Of the section Typhi S. Cerro isolates, 93.2% of isolates (290/311) clustered into a large clonal clade comprised of predominantly sequence type (ST) 367 cattle and environmental isolates, while the remaining 6.8% of isolates (21/311), primarily from human clinical sources, clustered outside of this clonal clade. A tip-dated phylogeny of S. Cerro ST367 identified two major clades (I and II), one of which overwhelmingly consisted of cattle isolates that share a most recent common ancestor that existed circa 1975. Gene presence/absence and rarefaction curve analyses suggested that the pangenome of section Typhi S. Cerro is open, potentially reflecting the gain/loss of prophage; human isolates contained the most open pangenome, while cattle isolates had the least open pangenome. Hypothetically disrupted coding sequences (HDCs) displayed clade-specific losses of intact speC and sopA virulence genes within the large clonal S. Cerro clade, while loss of intact vgrG, araH, and vapC occurred in all section Typhi S. Cerro isolates. Further phenotypic analysis suggested that the presence of a premature stop codon in speC does not abolish ornithine decarboxylase activity in S. Cerro, likely due to the activity of the second ornithine decarboxylase encoded by speF, which remained intact in all isolates. Overall, our study identifies specific genomic features associated with S. Cerro's infrequent isolation from humans and its apparent adaptation to cattle, which has broader implications for informing our understanding of the evolutionary events facilitating host adaptation in Salmonella.

Molecular identification of a new species of Rhigonema (Nematoda: Rhigonematidae) and phylogenetic relationships within the infraorder Rhigonematomorpha

BackgroundThe infraorder Rhigonematomorpha comprises a group of obligate parasitic nematodes of millipedes (Arthropoda: Diplopoda). The current species identification of Rhigonematomorpha nematodes remains mainly based on morphological features, with molecular-based identification still in its infancy. Also, current knowledge of the phylogeny of Rhigonematomorpha is far from comprehensive.MethodsThe morphology of Rhigonematomorpha nematodes belonging to the genus Rhigonema, collected from the millipede Spirobolus bungii Brandt (Diplopoda: Spirobolida) in China, was studied in detail using light and scanning electron microscopy. Five different genetic markers, including the nuclear small ribosomal subunit (18S), internal transcribed spacer (ITS) and large ribosomal subunit (28S) regions and the mitochondrial cox1 and cox2 genes of these Rhigonematomorpha nematodes collected from China and Rhigonema naylae collected from Japan were sequenced and analyzed using Bayesian inference (BI) and Assemble Species by Automatic Partitioning (ASAP) methods. Phylogenetic analyses that included the most comprehensive taxa sampling of Rhigonematomorpha to date were also performed based on the 18S + 28S genes using maximum likelihood (ML) and BI methods.ResultsThe specimens of Rhigonema collected from S. bungii in China were identified as a new species, Rhigonema sinense n. sp. Striking variability in tail morphology was observed among individuals of R. sinense n. sp. ASAP analyses based on the 28S, ITS, cox1 and cox2 sequences supported the species partition of R. sinense n. sp. and R. naylae, but showed no evidence that the different morphotypes of R. sinense n. sp. represent distinct genetic lineages. BI analyses also indicated that R. sinense n. sp. represents a separated species from R. naylae based on the cox1 and cox2 genes, but showed that R. naylae nested in samples of R. sinense n. sp. based on the ITS and 28S data. Phylogenetic results showed that the representatives of Rhigonematomorpha formed two large clades. The monophyly of the families Carnoyidae and Ichthyocephalidae and the genus Rhigonema was rejected. The representatives of the family Ransomnematidae clustered together with the family Hethidae with strong support.ConclusionsA new species of Rhigonematomorpha, R. sinense n. sp. is described based on morphological and molecular evidence. ASAP analyses using 28S, ITS, cox1 and cox2 data indicate the striking variability in tail morphology of R. sinense n. sp. as intraspecific variation, and also suggest that partial 28S, ITS, cox1 and cox2 markers are effective for molecular identification of Rhigonematomorpha nematodes. The phylogenetic results support the traditional classification of Rhigonematomorpha into the two superfamilies Rhigonematoidea and Ransomnematoidea, and indicate that the families Carnoyidae and Ichthyocephalidae and the genus Rhigonema are non-monophyletic. The present phylogeny strongly supports resurrection of the family Brumptaemiliidae, and also indicates that the family Ransomnematidae is sister to the family Hethidae.Graphical

Loss of Avian Intromittent Organs as a Sperm Competition Strategy: A Race to Be Last.

AbstractThe loss of the intromittent organ (IO) in the majority of birds remains unexplained. Here, I propose that IO loss results from sperm competition in the context of the unique avian sperm storage system. The first stage of fertilization is the movement of sperm through the vagina from the site of ejaculation to the sperm storage tubules (SSTs) at the uterovaginal junction. In a second stage, sperm are released from the SSTs and move through the uterus to the site of fertilization in the infundibulum. Last-male fertilization precedence can occur if sperm that arrive later at the uterovaginal junction occupy uterus-side SSTs, which then have a head start in the race to the infundibulum with each ovulation. Under such "last-in, first-out" conditions, there is strong sperm competition to arrive later at the SSTs. Consequently, the optimal male strategy would be placement of the ejaculate at the cloacal opening to allow any other sperm in the vagina to reach the SSTs first. Cloacal placement is effectively achieved by loss of the IO. The evolution of altricial development in Neoaves, the largest clade that lacks IOs, created conditions that favor IO loss. Specifically, the smaller clutch sizes and hatching asynchrony of altricial birds increase the intensity of sperm competition for fertilization of early eggs in the laying sequence and thus the selective advantage of later arrival at the SSTs. The rarity of IO loss among all animals suggests that the complex mechanism of avian fertilization creates unique conditions for sperm competition.

Genomic Assembly of Clinical Candida glabrata (Nakaseomyces glabrata) Isolates Reveals within-Species Structural Plasticity and Association with In Vitro Antifungal Susceptibility.

The opportunistic human pathogen Candida glabrata has become an increasingly important threat to human health, with infections globally characterized by high mortality rates and multidrug resistance. To face this threat, more efficient diagnostic and therapeutic approaches are required, underpinning research to help define the intraspecies epidemiology, genetic variability, and therefore, diagnostic and therapeutic target stability. Previous comparative genetics studies conducted on limited numbers of strains only revealed partial resolution of chromosomal settings. In this study, by combining short- and long-read genome sequencing, phenotypic characterization, and comparative genomics over a large set of strains, we detected strict relationships between large chromosomal rearrangements and phylogenetic clades, genes subjected to different selective pressures, and new sets of genes associated with resistance to antifungals. Overall, these results not only provide a fundamental contribution to our knowledge of C. glabrata evolution and epidemiology but may also lay the foundations for the future development of tailored therapeutic approaches. IMPORTANCE The human pathogen Candida glabrata has become a global threat to human health, with infections characterized by high mortality and multidrug resistance. We have obtained nine fully assembled genomes from clinical isolates through a combination of short- and long-read sequencing approaches. The quality and completeness of such genomes and their subsequent comparison to the broadest set of genomes so far allowed us to pinpoint chromosomal rearrangements in several genomes and detect phylogenetic clades that were not associated with geographic location or isolation source. We identified a new set of genes associated with resistance to antifungals coding for adhesin or adhesin-like proteins, suggesting C. glabrata resists antifungals by forming aggregates or adhering to the host tissue. These results, which provide a fundamental contribution to our knowledge of C. glabrata evolution and epidemiology, may initiate the development of precision medicine interventions for patients with suspected or proven invasive fungal infections.