Insights into RAG Evolution from the Identification of "Missing Link" Family A RAGL Transposons.

A series of "molecular domestication" events are thought to have converted an invertebrate RAG-like (RAGL) transposase into the RAG1-RAG2 (RAG) recombinase, a critical enzyme for adaptive immunity in jawed vertebrates. The timing and order of these events are not well understood, in part because of a dearth of information regarding the invertebrate RAGL-A transposon family. In contrast to the abundant and divergent RAGL-B transposon family, RAGL-A most closely resembles RAG and is represented by a single orphan RAG1-like (RAG1L) gene in the genome of the hemichordate Ptychodera flava (PflRAG1L-A). Here, we provide evidence for the existence of complete RAGL-A transposons in the genomes of P. flava and several echinoderms. The predicted RAG1L-A and RAG2L-A proteins encoded by these transposons intermingle sequence features of jawed vertebrate RAG and RAGL-B transposases, leading to a prediction of DNA binding, catalytic, and transposition activities that are a hybrid of RAG and RAGL-B. Similarly, the terminal inverted repeats (TIRs) of the RAGL-A transposons combine features of both RAGL-B transposon TIRs and RAG recombination signal sequences. Unlike all previously described RAG2L proteins, RAG2L-A proteins contain an acidic hinge region, which we demonstrate is capable of efficiently inhibiting RAG-mediated transposition. Our findings provide evidence for a critical intermediate in RAG evolution and argue that certain adaptations thought to be specific to jawed vertebrates (e.g. the RAG2 acidic hinge) actually arose in invertebrates, thereby focusing attention on other adaptations as the pivotal steps in the completion of RAG domestication in jawed vertebrates.

Sequencing and characterization of human bocavirus genomes from patients diagnosed in Southern France between 2017 and 2022.

The diversity and evolution of the genomes of human bocavirus (HBoV), which causes respiratory diseases, have been scarcely studied. Here, we aimed to obtain and characterize HBoV genomes from patients's nasopharyngeal samples collected between 2017 and 2022 period (5 years and 7 months). Next-generation sequencing (NGS) used Illumina technology after having implemented using GEMI an in-house multiplex PCR amplification strategy. Genomes were assembled and analyzed with CLC Genomics, Mafft, BioEdit, MeV, Nextclade, MEGA, and iTol. A total of 213 genomes were obtained. Phylogeny classified them all as of Bocavirus 1 (HBoV1) species. Five HBoV1 genotypic clusters determined by hierarchical clustering analysis of 27 variable genome positions were scattered over the study period although with differences in yearly prevalence. A total of 167 amino acid substitutions were detected. Besides, coinfection was observed for 52% of the samples, rhinoviruses then adenoviruses (HAdVs) being the most common viruses. Principal component analysis showed that HBoV1 genotypic cluster α tended to be correlated with HAdV co-infection. Subsequent HAdV typing for HBoV1-positive samples and negative controls demonstrated that HAdVC species predominated but HAdVB was that significantly HBoV1-associated. Overall, we described here the first HBoV1 genomes sequenced for France. HBoV1 and HAdVB association deserves further investigation.

Insights into RAG evolution from the identification of "missing link" family A RAGL transposons.

A series of "molecular domestication" events are thought to have converted an invertebrate RAG-like (RAGL) transposase into the RAG1-RAG2 (RAG) recombinase, a critical enzyme for adaptive immunity in jawed vertebrates. The timing and order of these events is not well understood, in part because of a dearth of information regarding the invertebrate RAGL-A transposon family. In contrast to the abundant and divergent RAGL-B transposon family, RAGL-A most closely resembles RAG and is represented by a single orphan RAG1-like (RAG1L) gene in the genome of the hemichordate Ptychodera flava (PflRAG1L-A). Here, we provide evidence for the existence of complete RAGL-A transposons in the genomes of P. flava and several echinoderms. The predicted RAG1L-A and RAG2L-A proteins encoded by these transposons intermingle sequence features of jawed vertebrate RAG and RAGL-B transposases, leading to a prediction of DNA binding, catalytic, and transposition activities that are a hybrid of RAG and RAGL-B. Similarly, the terminal inverted repeats (TIRs) of the RAGL-A transposons combine features of both RAGL-B transposon TIRs and RAG recombination signal sequences. Unlike all previously described RAG2L proteins, PflRAG2L-A and echinoderm RAG2L-A contain an acidic hinge region, which we demonstrate is capable of efficiently inhibiting RAG-mediated transposition. Our findings provide evidence for a critical intermediate in RAG evolution and argue that certain adaptations thought to be specific to jawed vertebrates (e.g., the RAG2 acidic hinge) actually arose in invertebrates, thereby focusing attention on other adaptations as the pivotal steps in the completion of RAG domestication in jawed vertebrates.

Matrix-Encoding Gene Diversity of 624 Influenza A/H3N2 Genomes Does Not Show Association with Impaired Viral Detection by Commercialized qPCR Assays.

As for the case of SARS-CoV-2, genome sequencing of influenza viruses is of potential interest to raise and address virological issues. Recently, false-negativity of real-time reverse transcription-PCR (qPCR) assays that detect influenza A/H3N2 virus RNA were reported and associated with two mutations (A37T and C161T) in the Matrix-encoding (M1) gene located on viral segment 7. This triggered a national alert in France. The present study sought to assess the association between the presence of these mutations and potential false negative results of influenza A/H3N2 virus RNA detection by commercialized qPCR assays at the clinical virology laboratory of our university hospitals in southern France. This study focused on the genetic diversity in the M1 gene and segment 7 of 624 influenza A/H3N2 virus genomes obtained from respiratory samples having tested qPCR-positive with M1 gene-targeting assays in our clinical virology laboratory. A total of 585 among the 624 influenza A/H3N2 virus genomes (93.7%) were of clade 3C.2a1b.2a.2, and 39 (6.3%) were of clade 3C.2a1b.1a. M1 gene substitutions A37T and C161T were both present in 582 (93.3%) genomes, only of clade 3C.2a1b.2a.2. Substitution A37T was present in 621 (99.5%) genomes. Substitution C161T was present in 585 genomes (93.8%), all of clade 3C.2a1b.2a.2. Moreover, 21 other nucleotide positions were mutated in ≥90% of the genomes. The present study shows that A37T/C and C161T mutations, and other mutations in the M1 gene and segment 7, were widely present in influenza A/H3N2 virus genomes recovered from respiratory samples diagnosed qPCR-positive with commercialized assays.

SARS-CoV-2 Testing of Aircraft Wastewater Shows That Mandatory Tests and Vaccination Pass before Boarding Did Not Prevent Massive Importation of Omicron Variant into Europe.

BACKGROUND: Most new SARS-CoV-2 epidemics in France occurred following the importation from abroad of emerging viral variants. Currently, the risk of new variants being imported is controlled based on a negative screening test (PCR or antigenic) and proof of up-to-date vaccine status, such as the International Air Transport Association travel pass. METHODS: The wastewater from two planes arriving in Marseille (France) from Addis Ababa (Ethiopia) in December 2021 was tested by RT-PCR to detect SARS-CoV2 and screen for variants. These tests were carried out between landing and customs clearance and were then sequenced by MiSeq Illumina. Antigenic tests and sequencing by NovaSeq were carried out on respiratory samples collected from the 56 passengers on the second flight. RESULTS: SARS-CoV-2 RNA suspected of being from the Omicron BA.1 variant was detected in the aircraft's wastewater. SARS-CoV2 RNA was detected in 11 [20%) passengers and the Omicron BA.1 variant was identified. CONCLUSION: Our work shows the efficiency of aircraft wastewater testing to detect SARS-CoV-2 cases among travellers and to identify the viral genotype. It also highlights the low efficacy of the current control strategy for flights entering France from outside Europe, which combines a requirement to produce a vaccine pass and proof of a negative test before boarding.

Parallel evolution of amphioxus and vertebrate small-scale gene duplications.

BACKGROUND: Amphioxus are non-vertebrate chordates characterized by a slow morphological and molecular evolution. They share the basic chordate body-plan and genome organization with vertebrates but lack their 2R whole-genome duplications and their developmental complexity. For these reasons, amphioxus are frequently used as an outgroup to study vertebrate genome evolution and Evo-Devo. Aside from whole-genome duplications, genes continuously duplicate on a smaller scale. Small-scale duplicated genes can be found in both amphioxus and vertebrate genomes, while only the vertebrate genomes have duplicated genes product of their 2R whole-genome duplications. Here, we explore the history of small-scale gene duplications in the amphioxus lineage and compare it to small- and large-scale gene duplication history in vertebrates. RESULTS: We present a study of the European amphioxus (Branchiostoma lanceolatum) gene duplications thanks to a new, high-quality genome reference. We find that, despite its overall slow molecular evolution, the amphioxus lineage has had a history of small-scale duplications similar to the one observed in vertebrates. We find parallel gene duplication profiles between amphioxus and vertebrates and conserved functional constraints in gene duplication. Moreover, amphioxus gene duplicates show levels of expression and patterns of functional specialization similar to the ones observed in vertebrate duplicated genes. We also find strong conservation of gene synteny between two distant amphioxus species, B. lanceolatum and B. floridae, with two major chromosomal rearrangements. CONCLUSIONS: In contrast to their slower molecular and morphological evolution, amphioxus' small-scale gene duplication history resembles that of the vertebrate lineage both in quantitative and in functional terms.