Gene Segments Research Articles

Reassortment of newly emergent clade 2.3.4.4b A(H5N1) highly pathogenic avian influenza A viruses in Bangladesh

ABSTRACT Avian influenza active surveillance was conducted in Bangladesh from January 2022 to November 2023 in live-poultry markets (LPMs) and Tanguar Haor wetlands. The predominant viruses circulating in LPMs were low pathogenic avian influenza (LPAI) A(H9N2) and clade 2.3.2.1a highly pathogenic avian influenza (HPAI) A(H5N1) viruses. Non-H9N2 LPAIs were found at Tanguar Haor and at a lower prevalence in LPMs. Starting from June 2023, we detected novel genotypes of clade 2.3.4.4b A(H5N1) viruses from ducks in LPMs. The HA, NA, and M genes of these viruses are related to those of 2020 European clade 2.3.4.4b H5N1 viruses such as A/Eurasian Wigeon/Netherlands/1/2020 (Netherlands/1). However, analyses of the other five gene segments’ sequences identified three distinct genotypes (BD-G2, BD-G3, and BD-G4). BD-G2 viruses were closely related to the clade 2.3.4.4b H5N1 viruses that have been detected in Japan and nearby regions since November 2022. BD-G3 viruses were reassortants, with gene segments from other Eurasian LPAI viruses. BD-G4 viruses were similar to BD-G2 viruses, but their NS gene was accrued from contemporary Bangladeshi clade 2.3.2.1a A(H5N1) viruses. The ability of any of the clade 2.3.4.4b viruses to displace the long-entrenched 2.3.2.1a A(H5N1) viruses in Bangladesh is unknown.

Distinct T-cell receptor (TCR) gene segment usage and MHC-restriction between foetal and adult thymus.

Here, we sequenced rearranged TCRβ and TCRα chain sequences in CD4+CD8+ double positive (DP), CD4+CD8- single positive (SP4) and CD4-CD8+ (SP8) thymocyte populations from the foetus and young adult mouse. We found that life-stage had a greater impact on TCRβ and TCRα gene segment usage than cell-type. Foetal repertoires showed bias towards 3'TRAV and 5'TRAJ rearrangements in all populations, whereas adult repertoires used more 5'TRAV gene segments, suggesting that progressive TCRα rearrangements occur less frequently in foetal DP cells. When we synchronised young adult DP thymocyte differentiation by hydrocortisone treatment the new recovering DP thymocyte population showed more foetal-like 3'TRAV and 5'TRAJ gene segment usage. In foetus we identified less influence of MHC-restriction on α-chain and β-chain combinatorial VxJ usage and CDR1xCDR2 (V region) usage in SP compared to adult, indicating weaker impact of MHC-restriction on the foetal TCR repertoire. The foetal TCRβ repertoire was less diverse, less evenly distributed, with fewer non-template insertions, and all foetal populations contained more clonotypic expansions than adult. The differences between the foetal and adult thymus TCR repertoires are consistent with the foetal thymus producing αβT-cells with properties and functions that are distinct from adult T-cells: their repertoire is less governed by MHC-restriction, with preference for particular gene segment usage, less diverse with more clonotypic expansions, and more closely encoded by genomic sequence.

Influenza A(H5N1) Virus Clade 2.3.2.1a in Traveler Returning to Australia from India, 2024.

We report highly pathogenic avian influenza A(H5N1) virus clade 2.3.2.1a in a child traveler returning to Australia from India. The virus was a previously unreported reassortant consisting of clade 2.3.2.1a, 2.3.4.4b, and wild bird low pathogenicity avian influenza gene segments. These findings highlight surveillance gaps in South Asia.

Genome‑wide identifcation and expression analysis of growth-regulating factors under drought in Brassica juncea

Growth-regulating factors (GRFs) are plant-specific transcription factors involved in the regulation of plant growth, development, and abiotic stress processes. However, the function of Brassica juncea (L.) Czern & Coss GRFs remain largely unknown. In this study, 34 BjGRF genes were identified in B. juncea. BjGRF members of the same subfamily were found to share a similar motif composition and gene structure. In total, 663 cis-acting elements were found in the promoter regions of BjGRF genes, which were related to light response, hormone response, environmental stresses, and plant growth/development. Additionally, 48 pairs of segmental duplication genes were identified during gene duplication events, and no tandemly duplicated genes were identified. qRT-PCR analysis showed that the 34 BjGRF genes were primarily expressed in the roots, followed by the leaves. Furthermore, the 10 BjGRF genes were screened in response to drought stress, and the expression patterns of the genes were relatively consistent, with a maximum expression level at 3 or 24 h, and the selected BjGRF03 and BjGRF32 might be involved in drought stress. This preliminary study clarifies the response of the BjGRF gene family to drought stress and provides ideas for further analyses of the biological functions of BjGRF genes.

Genome-Wide Characterization of Solanum tuberosum CCoAOMT Gene Family and Identification of StCCoAOMT Genes Involved in Anthocyanin Biosynthesis.

The caffeoyl-CoA-O methyltransferase (CCoAOMT) family plays essential roles in the methylation of various secondary metabolites, including anthocyanins. Despite the wide identification of the CCoAOMT family in plants, the characterization and function of CCoAOMT protein members in Solanum tuberosum remain poorly understood. In this study, a total of 12 StCCoAOMT members were identified in the genome of S. tuberosum using the Blastp and HMM search and were unevenly located on eight chromosomes. Collinearity analysis revealed that four tandem duplicated gene pairs and two segmental duplicated gene pairs existed in the S. tuberosum genome, demonstrating that duplication events play a key role in the expansion of the CCoAOMT family. All StCCoAOMTs were clustered into group I and group II based on phylogenetic analysis, which was further verified by the conserved motifs and gene structures analysis. The cis-acting elements analysis illustrated that StCCoAOMTs might be important for photosynthesis, hormone responses, and abiotic stress. Expression analysis demonstrated that StCCoAOMT genes have diverse transcript levels in various tissues and that StCCoAOMT10 was significantly expressed in purple potatoes with abundant anthocyanin content according to RNA-seq data and qRT-PCR assays. In addition, the subcellular localization assay validated that the StCCoAOMT10 protein was mainly localized in the cytoplasm and nucleus. These results will be of great importance for a better understanding of the features of CCoAOMT family members, especially of the candidate genes involved in the methylation of anthocyanins in S. tuberosum, and also for improving the nutritional quality of S. tuberosum.

2018-2019 human seasonal H3N2 influenza A virus spillovers into swine with demonstrated virus transmission in pigs were not sustained in the pig population.

Human seasonal H3 clade 3C3a influenza A viruses (IAV) were detected four times in U.S. pigs from commercial swine farms in Michigan, Illinois, and Virginia in 2019. To evaluate the relative risk of this spillover to the pig population, whole genome sequencing and phylogenetic characterization were conducted, and the results revealed that all eight viral gene segments were closely related to 2018-2019 H3N2 human seasonal IAV. Next, a series of in vitro viral kinetics, receptor binding, and antigenic characterization studies were performed using a representative A/swine/Virginia/A02478738/2018(H3N2) (SW/VA/19) isolate. Viral replication kinetic studies of SW/VA/19 demonstrated less efficient replication curves than all 10 swine H3N2 viruses tested but higher than three human H3N2 strains. Serial passaging experiments of SW/VA/19 in swine cells did not increase virus replication, but changes at HA amino acid positions 9 and 159 occurred. In swine transmission studies, wild-type SW/VA/19 was shed in nasal secretions and transmitted to all indirect contact pigs, whereas the human seasonal strain A/Switzerland/9715293/2013(H3N2) from the same 3C3a clade failed to transmit. SW/VA/19 induced minimal macroscopic and microscopic lung lesions. Collectively, these findings demonstrate that these human seasonal H3N2 3C3a-like viruses did not require reassortment with endemic swine IAV gene segments for virus shedding and transmission in pigs. Limited detections in the U.S. pig population in the subsequent period of time suggest a yet-unknown restriction factor likely limiting the spread of these viruses in the U.S. pig population.IMPORTANCEInterspecies human-to-swine IAV transmission occurs globally and contributes to increased IAV diversity in pig populations. We present data that a swine isolate from a 2018-2019 human-to-swine transmission event was shed for multiple days in challenged and contact pigs. By characterizing this introduction through bioinformatic, molecular, and animal experimental approaches, these findings better inform animal health practices and vaccine decision-making. Since wholly human seasonal H3N2 viruses in the United States were not previously identified as being transmissible in pigs (i.e., reverse zoonosis), these findings reveal that the interspecies barriers for transmission to pigs may not require significant changes to all human seasonal H3N2, although additional changes may be required for sustained transmission in swine populations.

Respiratory virus infections and adenovirus characteristics during acute exacerbation of chronic obstructive pulmonary disease.

Chronic obstructive pulmonary disease (COPD) is a common respiratory disease globally, characterized by obstructive ventilatory disorder under pulmonary function tests. Recent years have witnessed a yearly increase in the prevalence of COPD. To investigate the impact of respiratory virus infections on patients with acute exacerbation of chronic obstructive pulmonary disease (AECOPD), and to perform sequencing typing and mutation analysis of viruses with high detection rate. A total of 1523 inpatients with AECOPD admitted to our hospital from April 1,2020 to March 30,2022 were collected and divided into two groups: the infected group (n= 532) and the non-infected group (n= 991). The related indexes between the two groups were collected and compared (including clinical characteristics and laboratory tests that blood cell count, PCT, CRP, adenovirus, respiratory syncytial virus, rhinovirus, influenza A virus, influenza B virus, etc.). In the infected group, the proportion of patients with palpitations (49.44% VS 8.07%, P< 0.001), lipid metabolism abnormalities (18.42% VS 39.96%, P< 0.001), heart failure (39.85% VS 29.87%, P< 0.001), disease duration (17.48 ± 7.47 VS 12.45 ± 11.43 d, P< 0.001), and poor prognosis (69.55% VS 17.15%, P< 0.001) were higher than those in the non-infected group; Adenovirus (ADV) accounted for 75.94% (404/532) of all infected viruses. 31 virus strains could be categorized into 16 ADV-C1, one ADV-C5, two ADV-B3, three ADV-B7, two ADV-D17, two ADV-D19, and five ADV-D27, which were similar to the serotypes reported in severe pneumonia. Furthermore, three strains of C1 adenovirus were found to be highly homologous to the original strain AF534906 by sequencing, and the phylogenetic trees of the three main structural genes were all on the same branch as the original strain. Base mutations and amino acid variants were found in each structural gene segment. In clinical data, it's found that patients with mutations are worse than those without mutations. Respiratory viruses are common in patients with poor prognosis of AECOPD, especially adenovirus, respiratory syncytial virus. Respiratory virus infections will lead to the deterioration of patients with AECOPD, accompanied by longer treatment cycles and poor prognosis.

E-cadherin Single Nucleotide Variants Are Associated with Increasing Susceptibility to Periodontitis.

To investigate the association of E-cadherin single nucleotide polymorphisms (SNPs) with periodontitis and the potential of these SNPs for identifying susceptibility to periodontitis. Periodontal clinical parameters were recorded followed by collecting venous blood for DNA extraction. Polymerase chain reaction was used to amplify target segments of the E-cadherin gene. Determination of the genotype and allele frequencies was performed using Sanger sequencing. All statistical analyses were performed using GraphPad Prism (version 9) using a statistically significant difference of p < 0.05. A total of 207 participants were recruited into two groups of healthy controls (n = 105) and cases diagnosed with periodontitis stage 2 or 3, grade B or C (n = 102). Analyses indicated that the genotypes and alleles of rs3743674 and rs5030625 E-cadherin SNPs were significantly associated with periodontitis. Results from a binary regression model suggested that the presence of these SNPs may indicate susceptibility to periodontitis and increase the rate of progression. Linkage disequilibrium analysis indicated that E-cadherin variants rs3743674 and rs5030625, and rs10272115 and rs16260 were correlated in a nonrandom manner (r 2 = 0.638 and 0.495, respectively). E-cadherin gene variants, rs3743674 and rs5030625, were associated with the periodontitis phenotype. These biomarkers may identify individuals susceptible to periodontitis and the rate of disease progression.

Mutations in the main antigenic sites of VP7 and VP8* from G3P[8] rotavirus a strains circulating in Brazil may impact immune evasion to rotavirus vaccination.

In the post-rotavirus (RVA) vaccination era, uncommon and zoonotic strains have emerged as causative agents of acute gastroenteritis in humans, including the equine-like G3P[8] strains. First identified in 2013, this strain has quickly spread worldwide, reaching the position of the most prevalent genotype in many countries, including Brazil. Here, we report full genotype characterization and phylogenetic analysis of two equine-like G3P[8] strains detected in Goiás, a state in the Cerrado biome of the Brazilian Midwestern region, during the year of 2019. The strains were detected in different socioeconomic and demographic contexts: GO-MR from an asymptomatic adult living in a rural traditional community and GO-H5 from a symptomatic child from the state capital, with access to safe drinking water and essential sanitation services. These strains also displayed different backbone constellations considering the NSP2 gene segment (G3-P [8]-I2-R2-C2-M2-A2-N2-T2-E2-H2 for GO-MR and G3-P[8]-I2-R2-C2-M2-A2-N1-T2-E2-H2 for GO-H5). Furthermore, significant mutations in the main epitope sites of the VP7 and VP8* proteins of the detected strains, and other Brazilian G3P[8] viruses, were found with the comparison to RV1 and RV5 vaccine proteins, indicating a potential ability of these viruses to evade vaccine protection, which may contribute to their prevalence both nationally and globally. In summary, this study corroborates the genetic diversity of equine-like G3P[8] DS-1-like strains circulating worldwide, highlights the epidemiological importance of adults as reservoirs of RVA and shows the substantial differences between these emerging strains and the currently used anti-RVA vaccines, which may partially explain their predominance due to potential evasion of vaccine-induced protection.

Unraveling the architecture of major histocompatibility complex class II haplotypes in rhesus macaques.

The regions in the genome that encode components of the immune system are often featured by polymorphism, copy number variation, and segmental duplications. There is a need to thoroughly characterize these complex regions to gain insight into the impact of genomic diversity on health and disease. Here we resolve the organization of complete major histocompatibility complex (MHC) class II regions in rhesus macaques by using a long-read sequencing strategy (Oxford Nanopore Technologies) in concert with adaptive sampling. In particular, the expansion and contraction of the primate DRB-region appear to be a dynamic process that involves the rearrangement of different cassettes of paralogous genes. These chromosomal recombination events are propagated by a conserved pseudogene, DRB6, which features the integration of two retroviral elements. In contrast, the DRA locus appears to be protected from rearrangements, which may be owing to the presence of an adjacently located truncated gene segment, DRB9 With our sequencing strategy, the annotation, evolutionary conservation, and potential function of pseudogenes can be reassessed, an aspect that was neglected by most genome studies in primates. Furthermore, our approach facilitates the characterization and refinement of an animal model essential to study human biology and disease.

Insights into Genetic and Antigenic Characteristics of Influenza A(H1N1)pdm09 Viruses Circulating in Sicily During the Surveillance Season 2023-2024: The Potential Effect on the Seasonal Vaccine Effectiveness.

After disruption in the influenza circulation due to the emergence of SARS-CoV-2, the intensity of seasonal outbreaks has returned to the pre-pandemic levels. This study aimed to evaluate the evolution and variability of whole-genome sequences of A(H1N1)pdm09, the predominant influenza virus in Sicily (Italy) during the season 2023-2024. The potential vaccine efficacy was calculated using the pepitope model based on amino acid changes in the dominant epitope of hemagglutinin. The HA gene sequences showed several amino acid substitutions, some of which were within the major antigenic sites. The phylogenetic analysis showed that Sicilian strains grouped into two main genetic clades (6B.1A.5a.2a.1 and 6B.1A.5a.2a) and several subclades. Notably, about 40% of sequences partially drifted from the WHO-recommended vaccine strain A/Victoria/4897/2022 for the Northern Hemisphere. These sequences mostly belonged to the subclades C.1.8 and C.1.9 and harboured the amino acid mutations responsible for the modest predicted vaccine efficacy (E = 38.12% of 53%, pepitope = 0) against these viruses. Amino acid substitutions in other gene segments were also found. Since influenza viruses are constantly evolving, genomic surveillance is crucial in monitoring their molecular evolution and the occurrence of genetic and antigenic changes, and, thus, their potential impact on vaccine efficacy.

Ultrasensitive allele inference from immune repertoire sequencing data with MiXCR.

Allelic variability in the adaptive immune receptor loci, which harbor the gene segments that encode B cell and T cell receptors (BCR/TCR), is of critical importance for immune responses to pathogens and vaccines. Adaptive immune receptor repertoire sequencing (AIRR-seq) has become widespread in immunology research making it the most readily available source of information about allelic diversity in immunoglobulin (IG) and T cell receptor (TR) loci. Here we present a novel algorithm for extra-sensitive and specific variable (V) and joining (J) gene allele inference, allowing reconstruction of individual high-quality gene segment libraries. The approach can be applied for inferring allelic variants from peripheral blood lymphocyte BCR and TCR repertoire sequencing data, including hypermutated isotype-switched BCR sequences, thus allowing high-throughput novel allele discovery from a wide variety of existing datasets. The developed algorithm is a part of the MiXCR software. We demonstrate the accuracy of this approach using AIRR-seq paired with long-read genomic sequencing data, comparing it to a widely used algorithm, TIgGER. We applied the algorithm to a large set of IG heavy chain (IGH) AIRR-seq data from 450 donors of ancestrally diverse population groups, and to the largest reported full-length TCR alpha and beta chain (TRA; TRB) AIRR-seq dataset, representing 134 individuals. This allowed us to assess the genetic diversity within the IGH, TRA and TRB loci in different populations and to establish a database of alleles of V and J genes inferred from AIRR-seq data and their population frequencies with free public access through an online database.

Strain-specific differences in reovirus infection of murine macrophages segregate with polymorphisms in viral outer-capsid protein σ3.

Mammalian orthoreovirus (reovirus) strains type 1 Lang (T1L) and type 3 Dearing-RV (T3D-RV) infect the intestine in mice but differ in the induction of inflammatory responses. T1L infection is associated with the blockade of oral immunological tolerance to newly introduced dietary antigens, whereas T3D-RV is not. T1L infection leads to an increase in infiltrating phagocytes, including macrophages, in gut-associated lymphoid tissues that are not observed in T3D-RV infection. However, the function of macrophages in reovirus intestinal infection is unknown. Using cells sorted from infected intestinal tissue and primary cultures of bone-marrow-derived macrophages (BMDMs), we discovered that T1L infects macrophages more efficiently than T3D-RV. Analysis of T1L × T3D-RV reassortant viruses revealed that the viral S4 gene segment, which encodes outer-capsid protein σ3, is responsible for strain-specific differences in infection of BMDMs. Differences in the binding of T1L and T3D-RV to BMDMs also segregated with the σ3-encoding S4 gene. Paired immunoglobulin-like receptor B (PirB), which serves as a receptor for reovirus, is expressed on macrophages and engages σ3. We found that PirB-specific antibody blocks T1L binding to BMDMs and that T1L binding to PirB-/- BMDMs is significantly diminished. Collectively, our data suggest that reovirus T1L infection of macrophages is dependent on engagement of PirB by viral outer-capsid protein σ3. These findings raise the possibility that macrophages function in the innate immune response to reovirus infection that blocks immunological tolerance to new food antigens.IMPORTANCEMammalian orthoreovirus (reovirus) infects humans throughout their lifespan and has been linked to celiac disease (CeD). CeD is caused by a loss of oral immunological tolerance (LOT) to dietary gluten and leads to intestinal inflammation following gluten ingestion, which worsens with prolonged exposure and can cause malnutrition. There are limited treatment options for CeD. While there are genetic risk factors associated with the illness, triggers for disease onset are not completely understood. Enteric viruses, including reovirus, have been linked to CeD induction. We found that a reovirus strain associated with oral immunological tolerance blockade infects macrophages by virtue of its capacity to bind macrophage receptor PirB. These data contribute to an understanding of the innate immune response elicited by reovirus, which may shed light on how viruses trigger LOT and inform the development of CeD vaccines and therapeutic agents.

Statistical analysis of repertoire data demonstrates the influence of microhomology in V(D)J recombination.

V(D)J recombination generates the diverse B and T cell receptors essential for recognizing a wide array of antigens. This diversity arises from the combinatorial assembly of V(D)J genes and the junctional deletion and insertion of nucleotides. While previous in vitro studies have shown that microhomology--short stretches of sequence homology between gene ends--can bias the recombination process, the extent of microhomology's impact in vivo, particularly in humans, remains unknown. In this paper, we assess how germline-encoded microhomology influences trimming and ligation during V(D)J recombination using statistical inference on previously-published high-throughput TCRα repertoire sequencing data. We find that microhomology increases both trimming and ligation probabilities, making it an important predictor of recombination outcomes. These effects are consistent across different receptor loci and sequence types. Further, we demonstrate that accounting for microhomology effects significantly alters sequence annotation probabilities and rankings, highlighting its practical importance for accurately inferring the V(D)J recombination events that generated an observed sequence. Together, these results enhance our understanding of how microhomologous nucleotides shape the human V(D)J recombination process.

Genetic features of avian influenza (A/H5N8) clade 2.3.4.4b isolated from quail in Egypt

Several genotypes of the highly pathogenic avian influenza (HPAI) virus H5N8 subtype within clade 2.3.4.4b continue to circulate in different species of domestic birds across Egypt. It is believed that quail contribute to virus replication and adaptation to other gallinaceous poultry species and humans. This study provides genetic characterization of the full genome of HPAI H5N8 isolated from quail in Egypt. The virus was isolated from a commercial quail farm associated with respiratory signs. To characterize the genetic features of the detected virus, gene sequencing via Sanger technology and phylogenetic analysis were performed. The results revealed high nucleotide identity with the HPAI H5N8 virus from Egypt, which has multiple basic amino acid motifs PLREKRRKR/GLF at the hemagglutinin (HA) cleavage site. Phylogenetic analysis of the eight gene segments revealed that the quail isolate is grouped with HPAI H5N8 viruses of clade 2.3.4.4b and closely related to the most recent circulating H5N8 viruses in Egypt. Whole-genome characterization revealed amino acid preferences for avian receptors with few mutations, indicating their affinity for human-like receptors and increased virulence in mammals, such as S123P, S133A, T156A and A263T in the HA gene. In addition, the sequencing results revealed a lack of markers associated with influenza antiviral resistance in the neuraminidase and matrix-2 coding proteins. The results of the present study support the spread of HPAIV H5N8 to species other than chickens in Egypt. Therefore, continuous surveillance of AIV in different bird species in Egypt followed by full genomic characterization is needed for better virus control and prevention.

Genome-wide identification and analysis of anthocyanin synthesis-related R2R3-MYB genes in Fragaria pentaphylla

BackgroundMYB transcription factors regulate anthocyanin biosynthesis across numerous plant species. However, comprehensive genome-wide investigations regarding the R2R3-MYB gene family and its involvement in regulating anthocyanin biosynthesis in the red and white fruit color morphs of Fragaria pentaphylla remain scarce.ResultsA total of 101 FpR2R3-MYB genes were identified from the F. pentaphylla genome and were divided into 34 subgroups based on phylogenetic analysis. Gene structure (exon/intron) and protein motifs were particularly conserved among the FpR2R3-MYB genes, especially members within the same subgroup. The FpR2R3-MYB genes were distributed over seven F. pentaphylla chromosomes. Analysis of gene duplication events revealed five pairs of tandem duplication genes and 16 pairs of segmental duplication genes, suggesting that segmental duplications are the major pattern for expansion of the FpR2R3-MYB gene family expansion in F. pentaphylla. Cis-regulatory elements of the FpR2R3-MYB promoters were involved in cellular development, phytohormones, environmental stress and photoresponse. Based on the analysis of the FpR2R3-MYB gene family and transcriptome sequencing (RNA-seq) data, FpMYB9 was identified as a key transcription factor involved in the regulation of anthocyanin synthesis in F. pentaphylla fruits. The expression of FpMYB9 increases significantly during the ripening stage of red fruits, as confirmed by reverse transcription quantitative real-time PCR. In addition, subcellular localization experiments further confirmed the nuclear presence of FpMYB9, supporting its role as a transcription factor involved in anthocyanin biosynthesis.ConclusionOur results showed that the FpR2R3-MYB genes are highly conserved and play important roles in the anthocyanin biosynthesis in F. pentaphylla fruits. Our results also provide a compelling basis for further understanding of the regulatory mechanism underlying the role of FpMYB9 in anthocyanin formation in F. pentaphylla fruits.

Phylogenetic and Morphological Perspectives on Crepidotus subg. Dochmiopus: Exploratively Unveiling Hidden Diversity in China.

Crepidotus subg. Dochmiopus contributes to more than half of Crepidotus species and exhibits highly hidden diversity. However, C. subg. Dochmiopus is challenging to study because the basidiomata of C. subg. Dochmiopus species are usually small and white, inconspicuous interspecific distinctions, and possess a familiar complex. In this study, we utilized a variety of characteristics for species identification, including habitat, presence or absence of a stipe in mature specimens, pileipellis and cheilocystidia patterns, whether the lamellae edges are fimbriated, and other characteristics. Above all, cheilocystidia and pileipellis patterns will be important in C. subg. Dochmiopus research. Based on the present specimens, we constructed a multigene phylogenetic tree (ITS + LSU) and recognized four new species: C. lamellomaculatus sp. nov., C. capitatocystidiatus sp. nov., C. succineus sp. nov., C. clavocystidiatustustus sp. nov. Detailed morphological descriptions, photographs, line drawings and comparisons with closely related taxa for the new species are provided. The current phylogenetic analysis does not support the previously classifications, indicating that the classification of Crepidotus requires re-evaluation. But the existing molecular datasets and species' descriptions are insufficient to fully resolve the classification. Further integration of new gene segments and a comprehensive review of morphological characteristics will reveal a natural classification for Crepidotus.

Genome-Wide Identification and Expression Analysis of the BTB Gene Superfamily Provides Insight into Sex Determination and Early Gonadal Development of Alligator sinensis.

The BTB gene superfamily is widely distributed among higher eukaryotes and plays a significant role in numerous biological processes. However, there is limited knowledge about the structure and function of BTB genes in the critically endangered species Alligator sinensis, which is endemic to China. A total of 170 BTB genes were identified from the A. sinensis genome, classified into 13 families, and unevenly distributed across 16 chromosomes. Analysis of gene duplication events yielded eight pairs of tandem duplication genes and six pairs of segmental duplication genes. Phylogenetics shows that the AsBTB genes are evolutionarily conserved. The cis-regulatory elements in the AsBTB family promoter region reveal their involvement in multiple biological processes. Protein interaction network analysis indicates that the protein interactions of the AsBTB genes are centered around CLU-3, mainly participating in the regulation of biological processes through the ubiquitination pathway. The expression profile and protein interaction network analysis of AsBTB genes during sex differentiation and early gonadal development indicate that AsBTB genes are widely expressed in this process and involves numerous genes and pathways for regulation. This study provides a basis for further investigation of the role of the BTB gene in sex differentiation and gonadal development in A. sinensis.

Evolution of Influenza A(H3N2) Viruses in Bhutan for Two Consecutive Years, 2022 and 2023.

Influenza A viruses pose a significant public health threat globally and are characterized by rapid evolution of the hemagglutinin (HA) gene causing seasonal epidemics. The aim of this study was to investigate the evolutionary dynamics of A(H3N2) circulating in Bhutan during 2022 and 2023. We analysed 166 whole-genome sequences of influenza A(H3N2) from Bhutan, obtained from the GISAID database. We employed a Bayesian Markov Chain Monte Carlo (MCMC) framework, with a curated global dataset of HA sequences from regions with significant migration links to Bhutan. Phylogenetic, temporal, and phylogeographic analyses were conducted to elucidate the evolutionary dynamics and spatial dissemination of the viruses. Our phylogenetic analysis identified the circulation of influenza A(H3N2) Clade 3C.2a1b.2a.2 in Bhutan during 2022 and 2023, with viruses further classified into three subclades: 2a.3 (39/166), 2a.3a.1 (58/166) and 2a.3b (69/166). The TMRCA estimates suggest that these viral lineages originated approximately 1.93 years prior to their detection. Phylogeographic analysis indicates introductions from the United States in 2022 and Australia in 2023. The mean evolutionary rate across all gene segments was calculated to be 4.42 × 10-3 substitutions per site per year (95% HPD: 3.19 × 10-3 to 5.84 × 10-3), with evidence of purifying selection and limited genetic diversity. Furthermore, reassortment events were rare, with an estimated rate of 0.045 events per lineage per year. Our findings show that primary forces shaping the local evolution of the influenza A(H3N2) in Bhutan are largely stochastic, with only sporadic instances of adaptive change, and thus underscore the importance of continuous surveillance to mitigate the impact of evolving strains.

Comparison of B Cell Variable Region Gene Segment Characteristics in Neuro-autoantibodies.

Autoimmune pediatric neurologic diseases have variable phenotypes and presentations, making diagnosis challenging. The pathologic mechanisms are also distinct, including cell-mediated and Ab-mediated autoimmunity, paraneoplastic syndromes, and postinfectious processes. In recent years a number of studies have described the characteristics of the autoantibodies involved in a number of these diseases. Some of the described Abs use a restricted set of variable gene segments. We sought to compare the Ab characteristics of autoantibodies related to some of the more common disorders to discover whether specific Ab signatures are universally associated with neuroautoimmune diseases. We initially performed a literature review to summarize the Ab characteristics of autoantibodies related to some of the more common disorders, including N-methyl-d-aspartate receptor (NMDAR) and leucine-rich, glioma-inactivated 1 (LGI-1). Next, we performed data analysis from selected studies that sequenced Ig genes to further characterize NMDAR and LGI-1 autoantibodies including CDR3 length distribution, variable gene sequence usage, and isotype use. We found that CDR3 length of NMDAR autoantibodies was normally distributed whereas the CDR3 length distribution of LGI-1 autoantibodies was skewed, suggesting that there is no global structural restriction on types of autoantibodies that can cause encephalitis. We also found that IgG1-IgG3 were the main NMDAR autoantibody isotypes detected, while IgG4 was the major isotype used in autoantibodies from LGI-1 encephalitis. These findings are useful for our understanding of autoimmune encephalitis and will help facilitate better diagnosis and treatment of these conditions in the future.