Palaeognaths reveal evolutionary ancestry of the avian Major Histocompatibility Complex class II.

The multigene family of the Major Histocompatibility Complex (MHC) codes for the key antigen-presenting molecules of the vertebrate immune system. In birds, duplicated MHC class II (MHC-II) genes are highly homogenized by concerted evolution and, thus, identification of their orthologous relationships across long evolutionary timescales remains challenging. Relatively low evolutionary rate of avian MHC class IIA genes has been expected to provide a promising avenue to allow such inferences, but availability of MHC-IIA sequences in non-model bird species has been limited until recently. Here, taking advantage from accumulating genomic resources, we identified and analysed MHC-IIA sequences from the most basal lineage of extant birds (Palaeognathae). Conserved region of the MHC-IIA membrane-proximal domain was used to search for orthologous relationships between palaeognath birds and non-avian reptiles. First, analyses of palaeognath sequences revealed the presence of a separate MHC-IIA gene lineage (DAA3) in kiwis, which did not cluster with previously described avian MHC-IIA lineages (DAA1 and DAA2). Next, phylogenetic reconstruction showed that kiwi DAA3 sequences form a single well supported cluster with turtle MHC-IIA. High similarity of these sequences most likely reflects their remarkable evolutionary conservation and retention of ancient orthologous relationships, which can be traced back to basal archosauromorphs ca. 250 million years ago. Our analyses offer novel insights into macroevolutionary history of the MHC and reinforce the view that rapid accumulation of high-quality genome assemblies across divergent non-model species can substantially advance our understanding of gene evolution.

Spatio-temporal distribution and international context of bovine viral diarrhoea virus genetic diversity in France.

Bovine viral diarrhoea (BVD) is one of the most economically damaging livestock enzootic diseases in the world. BVD aetiological agents are three pestiviruses (BVDV-1, -2 and HoBi-like pestivirus), which exhibit high genetic diversity and complex transmission cycles. This considerably hampers the management of the disease, which is why eradication plans have been implemented in several countries. In France, a national plan has been in place since 2019. Our understanding of its impact on the distribution of BVDV genotypes is limited by the availability of French genetic data. Here, we conducted a molecular epidemiology study to refine our knowledge of BVDV genetic diversity in France, characterise its international relationships, and analyse national spatio-temporal genotypic distribution. We collated 1037 BVDV-positive samples throughout France between 2011 and 2023, with a greater sampling effort in two major cattle production areas. We developed a high-throughput sequencing protocol which we used to complete the 5'UTR genotyping of this collection. We show that two main BVDV-1 genotypes, 1e and 1b, account for 88% of genotyped sequences. We also identified seven other BVDV-1 genotypes occurring at low frequencies and three BVDV-2 samples (genotype 2c). Phylogenetic analyses indicate different worldwide distribution patterns between the two main BVDV-1 genotypes. Their relative frequencies present no major changes in France since the 1990s and few variations at the national scale. We also found some degree of local spatial structuring in western France. Overall, our results demonstrate the potential of large-scale sequence-based surveillance to monitor changes in the epidemiological situation of enzootic diseases.

Genus-targeted markers for the taxonomic identification and monitoring of coagulase-positive and coagulase-negative Staphylococcus species.

The Staphylococcus genus comprises multiple pathogenic and opportunistic species that represent a risk to public health. Epidemiological studies require accurate taxonomic classification of isolates with enough resolution to distinguish clonal complexes. Unfortunately, 16 S rRNA molecular analysis and phenotypic characterization cannot distinguish all species and do not offer enough resolution to assess intraspecific diversity. Other approaches, such as Multilocus Sequence Tagging, provide higher resolution; however, they have been developed for Staphylococcus aureus and a few other species. Here, we developed a set of genus-targeted primers using five orthologous genes (pta, tuf, tpi, groEs, and sarA) to identify all Staphylococcus species within the genus. The primers were initially evaluated using 20 strains from the Collection of Microorganisms of Interest in Animal Health from AGROSAVIA (CMISA), and their amplified sequences were compared to a set of 33 Staphylococcus species. This allowed the taxonomic identification of the strains even on close species and the establishment of intraspecies diversity. To enhance the scope and cost-effectiveness of the proposed strategy, we customized the primer sets for an Illumina paired-end amplicon protocol, enabling gene multiplexing. We assessed five genes across 177 strains, generating 880 paired-end libraries from the CMISA. This approach significantly reduced sequencing costs, as all libraries can be efficiently sequenced in a single MiSeq run at a fraction (one-fourth or less) of the cost associated with Sanger sequencing. In summary, this method can be used for precise identification and diversity analysis of Staphylococcus species, offering an advancement over traditional techniques in both resolution and cost-effectiveness.

Shining new light on naupliar eyes: A novel molecular phylogeny for Pleuromamma (Family: Metridinidae) and the characterization of luciferase and opsin expression.

Pleuromamma (Giesbrecht, 1898) is a cosmopolitan genus of metridinid copepods, with species that perform remarkable diel vertical migrations (DVM) and emit a bioluminescent secretion when disturbed that varies both spectrally and kinetically. Copepod bioluminescence is autogenic and uses luciferase enzymes that catalyze a luciferin, coelenterazine, to produce light. Pleuromamma possess naupliar eyes, relatively simple photosensitive structures used for many visually-guided behaviors. Yet the fundamental molecular unit for vision, the opsin protein, has not been previously described for the family. The light producers and detectors are important to study because DVM is a behavior that mediates significant active elemental fluxes between the upper ocean and midwaters across vast stretches of oceanic habitat, and DVM is guided by visual behaviors, with animals tracking an isolume. Here we provide the first fully resolved molecular phylogeny for Pleuromamma (Family: Metridinidae) and describe the luciferase and opsin gene diversity and expression using de novo assembled transcriptomes. We successfully sequenced and assembled transcriptomes for 10 of 11 described species of Pleuromamma as well as two other metridinid species: Metridia longa and Gaussia princeps. In all species, we obtained coding sequences of one putative rhabdomeric middle wavelength sensitive visual opsin gene, as well as several non-visual opsins - a c-type pteropsin and a tetra-opsin type peropsin. Furthermore, Pleuromamma express luciferases from each of two main evolutionary clades (Luc1 and Luc2), and a single paralog (Luc2a) dominates expression throughout the group. The variation in luciferase number, sequence, and expression among species could lead to different spectral and kinetic properties of bioluminescence and aid in congener recognition.

The dynamic history of prokaryotic phyla: discovery, diversity and division.

Here, I review the dynamic history of prokaryotic phyla. Following leads set by Darwin, Haeckel and Woese, the concept of phylum has evolved from a group sharing common phenotypes to a set of organisms sharing a common ancestry, with modern taxonomy based on phylogenetic classifications drawn from macromolecular sequences. Phyla came as surprising latecomers to the formalities of prokaryotic nomenclature in 2021. Since then names have been validly published for 46 prokaryotic phyla, replacing some established names with neologisms, prompting criticism and debate within the scientific community. Molecular barcoding enabled phylogenetic analysis of microbial ecosystems without cultivation, leading to the identification of candidate divisions (or phyla) from diverse environments. The introduction of metagenome-assembled genomes marked a significant advance in identifying and classifying uncultured microbial phyla. The lumper-splitter dichotomy has led to disagreements, with experts cautioning against the pressure to create a profusion of new phyla and prominent databases adopting a conservative stance. The Candidatus designation has been widely used to provide provisional status to uncultured prokaryotic taxa, with phyla named under this convention now clearly surpassing those with validly published names. The Genome Taxonomy Database (GTDB) has offered a stable, standardized prokaryotic taxonomy with normalized taxonomic ranks, which has led to both lumping and splitting of pre-existing phyla. The GTDB framework introduced unwieldy alphanumeric placeholder labels, prompting recent publication of over 100 user-friendly Latinate names for unnamed prokaryotic phyla. Most candidate phyla remain 'known unknowns', with limited knowledge of their genomic diversity, ecological roles, or environments. Whether phyla still reflect significant evolutionary and ecological partitions across prokaryotic life remains an area of active debate. However, phyla remain of practical importance for microbiome analyses, particularly in clinical research. Despite potential diminishing returns in discovery of biodiversity, prokaryotic phyla offer extensive research opportunities for microbiologists for the foreseeable future.

First Report and Molecular Variability of Belonolaimus longicaudatus Associated with Turfgrass in Maryland.

Turfgrass is a crop used extensively in athletic fields and golf courses in Maryland. A soil sample collected in July 2023 from an athletic field in Baltimore County, Maryland, part of a turfgrass nematode survey, contained Belonolaimus longicaudatus. In the southeastern United States, B. longicaudatus is an economically important pathogen of warm season turfgrass. The density was four individuals/100 cm3 of soil, and no visual symptoms were observed in the bermudagrass field. Morphological features and morphometrics of males and females were consistent with B. longicaudatus and placed the Maryland population in a subclade that was geographically represented by populations from north and west Florida, Texas, and South Carolina. Sequencing of the internal transcribed spacer region ITS1 and ITS2 and 28S large ribosomal subunit D2-23 expansion region confirmed the species' identity. Phylogenetic trees and parsimony network analysis placed the Maryland isolate in a large grouping of B. longicaudatus populations including those from Alabama, Delaware, Florida, Indiana, Mississippi, South Carolina, and Texas. To our knowledge, this is the first report of B. longicaudatus in Maryland.

Molecular phylogenetics uncovers two new species in the genus Phyllobates (Anura, Dendrobatidae): the terrible frog gets two new sisters.

True poison-dart frogs (Phyllobates, Dendrobatidae) evolved the ability to secrete batrachotoxins, the most powerful alkaloids known to date. The genus comprises five species whose systematics, at first glance, appeared clear. The most derived clade would include two Colombian species (P.terribilis and P.bicolor) with the highest toxicity, the largest body size, and predominantly yellow body colouration. The other three species (P.aurotaenia, P.vittatus, and P.lugubris) are less toxic on average, have smaller size, and are predominantly black with bright dorsolateral stripes. Recent research has revealed the existence of two major lineages among the three Colombian species. The northern lineage appears to result from a complex evolutionary history, including perhaps introgression among yellow and black taxa. The southern lineage instead revealed the existence of new clades closely related to P.terribilis, black and yellow, that arguably deserve their recognition as new species. Here, available evidence is combined to support the erection of southern populations of P.aurotaenia as a new highly toxic species, sister to P.terribilis, and much closer to it than to any other yellow or black-bodied species, Phyllobatessamperi sp. nov. Their common ancestor is sister to an additional yellow species, which we also describe here as Phyllobatesbezosi sp. nov. Both new species can be externally diagnosed using colouration. Our previous and current analyses also suggest the existence of additional taxa and corroborate multiple transitions in colouration across these hypertoxic taxa.

ResumenLas verdaderas ranas de dardo venenoso (género Phyllobates) evolucionaron la capacidad de secretar batracotoxinas, los alcaloides más poderosos que se conocen hasta hoy. El género comprende cinco especies, cuya taxonomía parecía clara a primera vista. El clado más derivado incluye dos especies colombianas (P.terribilis and P.bicolor) con la mayor toxicidad, mayor tamaño y coloración predominantemente amarilla. Las otras tres especies (P.aurotaenia, P.vittatus and P.lugubris) son en general menos tóxicas, menores en tamaño y predominantemente negras con conspicuas manchas dorsolaterales.Una reciente investigación confirmó la existencia dos linajes mayores entre las tres especies colombianas. El del norte parece ser el producto de una historia evolutiva compleja, incluyendo quizás introgresión entre especies amarillas y negras. El del sur reveló en cambio la existencia de nuevos clados filogenéticamente cercanos a P.terribilis, uno negro y dos amarillos, que requieren reconocimiento formal como nuevas especies. Esta investigación combina la evidencia disponible para apoyar el reconocimiento de las poblaciones del sur de P.aurotaenia como una nueva especie negra, hermana de P.terribilis, y mucho más cercana a ella que a las otras especies negras o amarillas, Phyllobatessamperi sp. nov. Su ancestro común es a su vez el grupo hermano de una especie adicional, amarilla, que también describimos como Phyllobatesbezosi sp. nov. Las dos nuevas especies pueden ser externamente reconocidas por la coloración. Nuestros análisis previos y actuales también sugieren la existencia de otras especies no descritas y corroboran la existencia de múltiples transiciones de coloración en la evolución de las especies hipertóxicas.

Spatio-temporal distribution of rhinovirus types in Kenya: a retrospective analysis, 2014.

The epidemiology and circulation patterns of various rhinovirus types within populations remains under-explored. We generated 803 VP4/VP2 gene sequences from rhinovirus-positive samples collected from acute respiratory illness (ARI) patients, including both in-patient and outpatient cases, between 1st January and 31st December 2014 from eleven surveillance sites across Kenya and used phylogenetics to characterise virus introductions and spread. RVs were detected throughout the year, with the highest detection rates observed from January to March and June to July. We detected a total of 114 of the 169 currently classified types. Our analysis revealed numerous virus introductions into Kenya characterized by local expansion and extinction, and extensive spatial mixing of types within the country due to the widespread transmission of the virus after an introduction. This work demonstrates that in a single year, the circulation of rhinovirus in Kenya was characterized by substantial genetic diversity, multiple introductions, and extensive geographical spread.

European Culex pipiens Populations Carry Different Strains of Wolbachia pipientis.

The mosquito Culex pipiens occurs in two ecotypes differing in their mating and overwintering behavior: pipiens mate in open environments and diapause, and molestus also mate in small spaces and is active throughout the year. Cx. pipiens carry Wolbachia endosymbionts of the wPip strain, but the frequency of infection differs between studied populations. Wolbachia infection affects the host reproductive success through cytoplasmic incompatibility. wPip Wolbachia is divided into five types, wPip I-V. The type of wPip carried varies among Cx. pipiens populations. In northern European locations different wPip types are found in the two ecotypes, whereas in southern locations, they often carry the same type, indicating differences in hybridization between ecotypes. In this study, Cx. pipiens specimens of both ecotypes were collected from Sweden and compared to specimens from Norway, England, Italy, and the Netherlands, as well as Cx. quinquefasciatus from Mali and Thailand. The abundance varied, but all specimens were infected by Wolbachia, while the tested specimens of other mosquito species were often uninfected. The wPip strains were determined through the sequence analysis of Wolbachia genes ank2 and pk1, showing that Cx. pipiens ecotypes in Scandinavia carry different wPip strains. The observed differences in wPip strains indicate that hybridization is not frequent and may contribute to barriers against hybridization of the ecotypes in Sweden and Norway.

Characterization of Skoliomonas gen. nov., a haloalkaliphilic anaerobe related to barthelonids (Metamonada).

Metamonads are a large and exclusively anaerobic group of protists. Additionally, they are one of the three clades proposed to ancestrally possess an "excavate" cell morphology, with a conspicuous ventral groove accompanied by a posterior flagellum with a vane. Here, we cultivate and characterize four anaerobic bacterivorous flagellates from hypersaline and alkaline soda lake environments, which represent a novel clade. Small subunit ribosomal RNA (SSU rRNA) gene phylogenies support recent phylogenomic analyses in placing them as the sister of barthelonids, a group that is itself sister to or deeply branching within Fornicata (Metamonada). The new isolates have a distinctive morphology: the hunchbacked cell body is traversed by a narrow ventral groove ending in a large opening to a conspicuous recurrent cytopharynx. The right margin of the groove is defined by a thin "lip." The posterior flagellum bears a wide ventral-facing vane. The narrow ventral groove and elongate cytopharynx are shared with barthelonids. We describe one isolate as Skoliomonas litria, gen. et sp. nov. Further investigation of their mitochondrial-related organelles (MROs) and detailed ultrastructural studies would be important to understanding the adaptation to anaerobic conditions in Metamonads-especially fornicates-as well as the evolution of the "excavate" cell architecture.