BACKGROUND:The soybean cyst nematode (SCN) is a persistent threat to soybean production. SCN populations continually overcome resistant cultivars, causing significant yield losses. Studies conducted with a single reference genome restrict our understanding of intraspecific diversity, masking significant mechanisms of virulence evolution and host adaptation. Here we report a pangenome constructed of nine SCN populations of different pathotypes, including eight newly generated high-fidelity genome assemblies. RESULTS:We detected over 19,000 orthologous gene families and more than 12,000 putative secreted proteins in SCN. Combined, these data indicate substantial diversity across populations. Gene content analysis showed that 35% of gene families were the conserved core, 15% were soft-core, and 48% were accessory. Evidence of rapid evolution was identified in a high portion (40%) of core single-copy genes, most notably inside the protein domains responsible for host recognition and immune modulation. Analysis of gene-family expansion revealed extensive duplication and loss across lineages, suggesting ongoing paralog turnover within SCN populations. Finally, a graph-based pangenome enabled the identification of numerous structural variants within regions under selection. CONCLUSIONS:Our study highlights substantial genetic variation in SCN that is not captured by single-reference analyses. By integrating multiple high-quality assemblies, we show that the SCN genome is highly dynamic, with extensive gene duplication and loss as well as structural variation shaping the differences among nematode populations. Collectively, the SCN pangenome provides a robust resource for studying virulence and adaptation mechanisms in SCN and establishes a genomic foundation for the development of more precise management strategies.

Idiopathic pulmonary fibrosis (IPF) is a progressive, fatal disease lacking clarity on the mechanisms linking epithelial injury to fibrotic remodeling. Here, we identify the microbiota-derived corisin as a potent, multifaceted driver of epithelial injury and pulmonary fibrosis. Leveraging targeted DNA sequencing of bronchoalveolar lavage fluid, we provide the first sequence-based identification of corisin in IPF patients and show that functional depletion of native corisin from patient bronchoalveolar lavage fluid abolishes its proapoptotic activity in alveolar epithelial cells. Synthetic corisin readily penetrates epithelial cells, localizes to mitochondria, and induces apoptosis, cellular senescence, and epithelial-mesenchymal transition, effects validated by single-cell transcriptomic analysis. High-throughput protein-interaction screening identifies the ubiquitin-proteasome system as the primary target, demonstrating that corisin enhances proteasome activity and disrupts epithelial proteostasis. Intracellular expression of native corisin recapitulates these cellular pathologies at concentrations relevant to human disease, confirming its high intrinsic potency. Most critically, transgenic mice constitutively expressing native corisin develop spontaneous, progressive pulmonary fibrosis and exhibit exacerbated injury and increased mortality following bleomycin challenge. Collectively, our findings establish corisin as a microbiota-derived effector that directly couples the collapse of epithelial proteostasis to multimechanistic cell-fate dysregulation and fibrotic remodeling, thereby defining a potent and causal microbial-epithelial axis in the pathogenesis of IPF.

Commercial pork production is practiced worldwide and represents a major source of protein for global populations. Pigs, however, are plagued by various diseases that affect their productivity. A common practice is to administer antibiotics in the feed to reduce infections and promote growth. However, antibiotic utilization in pig production has been identified as a source of spread of antibiotic resistance genes, prompting the need for antibiotic alternatives in swine production. Salmonella enterica serotype Choleraesuis and porcine reproductive and respiratory syndrome virus (PRRSV) are two disease agents with a significant impact on the pork industry. In this study, we designed experiments to test the hypothesis that a Bacillus-based direct-fed microbial (DFM) cocktail will alleviate the impact of Salmonella infection alone or in combination with PRRSV. Both single and dual infections resulted in shifts in the cecal microbiota from that of the Control group, with administration of the DFM dampening this effect, especially in the Salmonella-infected group. In the absence of the DFM, the infected pigs exhibited gross changes in the lungs, including tissue hepatization. Significantly, the DFM application suppressed the lesions in the lungs of Salmonella-only infected pigs. Using metagenome-assembled genomes, we found that DFM administration to Salmonella-only infected pigs led to cecal microbiota enriched in the potential to produce immune-stimulating short-chain fatty acids and naturally occurring antimicrobials, including peptides. The putative antimicrobial peptides derived from this study, upon biochemical characterization, could lead to their application as novel antimicrobials in animal agriculture and health.IMPORTANCEAntibiotics, as feed additives, have been integral to commercial pork production. Their use, however, has fostered the spread of antibiotic resistance genes in the environment. In this study, we explored the use of a mixture of naturally occurring bacteria, comprising species of the genus Bacillus, as an alternative to antibiotics in the pig diet. The bacterial mixture reversed disease lesions in the lungs of pigs infected with Salmonella enterica serotype Choleraesuis, a bacterium that causes severe disease in commercial pigs. Our findings suggest that applying the bacterial mixture to the Salmonella-infected pigs shifts the microbes in the gut to a community that is endowed with antimicrobials that mitigate the effects of Salmonella infection. We present data showing the novelty of putative antimicrobials discovered in the present study and postulate that their characterization will yield new antimicrobials that can be used in different sectors of animal production and health. PRRSV was included in the study to model a common bacterial-viral co-infection in swine, as it exacerbates disease severity. This design allowed assessment of whether Bacillus-based DFM could improve outcomes along the gut-lung axis under realistic co-infection conditions.

Polycomb (PcG) bodies are nuclear foci formed by polycomb protein complexes that contain PcG-bound DNA and are implicated in gene regulation during development and differentiation, although their precise molecular function remains unclear. Using tyramide signal amplification sequencing, we provide a comprehensive view of genomic regions associated with PcG bodies, including specific centromeric and telomeric sites. These regions are enriched for the repressive marks H3K27me3 and H3K9me3, depleted of the active mark H3K4me3, and display low chromatin accessibility. We find a high density of replication origins around PcG bodies, consistent with the replication factor origin recognition complex-associated protein (ORCA)/LRWD1 localizing at these sites. ORCA interacts with the polycomb-repressive complex, stabilizes the H3K27 methyltransferase, and facilitates H3K27me3 deposition at specific chromatin sites. Loss of ORCA affects chromatin organization around PcG bodies, leading to decompaction of the repeat regions, and enhanced initiation from replication origins. Our results suggest that ORCA associates with PcG-bound chromatin to maintain a repressive environment that regulates replication origin firing timing.

Recent genomics research has redefined the taxonomy of giraffes (genus Giraffa), identifying four distinct species rather than just one. This new understanding raises concerns about the ancestry of North American giraffe populations in human care (ex situ) and whether they still serve as meaningful conservation assurance populations for wild giraffe taxa. To address this, we performed whole-genome sequencing and analyses of 52 giraffes kept ex situ across North America, comparing them to wild giraffes representing all four recognized species. The analyses-including principal component analysis, admixture estimation, local ancestry inference, and mitochondrial phylogenetics-revealed extensive hybridization in giraffes kept ex situ. Most demonstrated mixed ancestry, primarily between northern and reticulated giraffes, with only a few individuals retaining un-admixed ancestries. Although some wild giraffes are known to be natural hybrids, overall there is strong reproductive isolation among giraffe species in the wild. Thus hybridization across species boundaries and potentially founder misclassification are responsible for the patterns observed ex situ. These findings highlight substantial genetic admixture in captivity, diminishing the conservation value of the current ex situ population. We recommend phasing out hybrid individuals from breeding programs and establishing new conservation-relevant stocks through collaboration with willing African governments and conservation organizations. Success will require coordinated international efforts and updates to global conservation frameworks, building on the formal recognition by the International Union for Conservation of Nature of distinct giraffe species and subspecies, to support taxon-specific conservation strategies that reflect the genetic distinctiveness of giraffe taxa.

Many infants consume both human milk and infant formula (mixed-fed); however, few studies have investigated how mixed feeding affects the gut microbiome composition and metabolic profiles compared to exclusive breastfeeding or formula feeding. Herein, how delivery mode and early nutrition affect the microbiome and metabolome of 6-week-old infants in the STRONG Kids2 cohort was investigated. Fecal samples were collected from exclusively breastfed (BF; n = 25), formula-fed (FF; n = 25) or mixed-fed (MF; n = 25) participants. Within each feeding group, infants were either delivered vaginally (VD; n = 13) or by Cesarean section (CS; n = 12). Feeding mode affects the fecal microbiome diversity, composition, and functional potential, as well as metabolomic profiles regardless of delivery mode. Alpha and beta diversity of MF differed from that of BF (p < 0.05) but were comparable to FF infants. Functional analyses have shown 117 potential metabolic pathways differed between BF and FF, 112 between BF and MF, and 8 between MF and FF infants (p < 0.05, q < 0.10). Fecal metabolomic profiles of MF and FF clustered together and separated from BF infants. In total, 543 metabolites differed between BF and FF, 517 between BF and MF, and 3 between MF and FF (p < 0.05, q < 0.10). Delivery mode affected overall microbial composition (p = 0.022) at the genus level and 24 potential functional pathways, with 16 pathways being higher in VD than CS infants (p < 0.05, q < 0.10). Metabolomic analysis identified 47 differential metabolites between CS and VD, with 39 being lower in CS than VD (p < 0.05, q < 0.10). In summary, fecal microbiota composition and function and metabolite profiles of 6-week-old MF infants are closer to FF than BF infants.

Background Soybean cyst nematode (SCN) poses a persistent challenge to soybean production. SCN populations repeatedly overcome resistant cultivars, resulting in crop yield loss. Reference-genome driven analysis limits our knowledge of intraspecific diversity, impeding our understanding of virulence evolution and host adaptation. Here, we leverage high-fidelity long-read sequencing and comparative analysis to generate a pangenome from nine SCN populations differing in their pathotypes. Results In this study we compared 9 SCN genomes, including 8 new assemblies at the chromosome level. We identified over 19,000 orthologous gene families, with nearly 50% comprising the conserved core genome. A large portion of the core genome (40%) exhibited signatures of more rapid evolution in a positive selection analysis, particularly in domains related to host interaction and immune evasion. Structural variants in genomic regions under selection suggest population-specific haplotypes that may underlie differential virulence. Furthermore, the secretome, comprising ~ 1,400 genes per genome, revealed dynamic effector content across accessions. Conclusions Our study highlights the power of pangenomics in revealing hidden genetic diversity in SCN. The dynamic nature of both core and accessory genomes, shaped by selection and structural rearrangements, illustrates the genomic evolutionary arms race between SCN and soybean. These insights provide a foundational resource for resistance breeding and pathogen surveillance, with broader implications for managing rapidly evolving crop pathogens.

Stochastic nature of gene expression leads to the complex formation of the bacterial transcriptome and proteome. In contrast to typical transcriptome studies, we employ a near wild-type, Syn1.0, of the naturally genome-reduced Mycoplasmas , and the dramatically further genome-reduced JCVI-syn3A thus avoiding additional contributions from many non-essential cellular functions. To aid in profiling the transcriptional landscape within these bacteria, we present a bioinformatic analysis of the genetic sequence motifs implicated in modulating the stochastic gene expression events, coupled with genome-wide short-read (Illumina) and long-read (Oxford Nanopore Technologies and Pacfic Biosciences) RNA sequencing. The bioinformatic analysis coupled with information from structural studies assigns strengths of the Shine-Dalgarno signatures and identifies both transcription initiation and termination sites, leading to predictions of RNA isoforms in Syn1.0 (and related organisms). The long-read and short-read RNA sequencing characterized the predicted transcriptional activity, and the long-read methods provide direct insight into the RNA isoform complexity within Syn1.0. Comparison of the RNA sequencing results with that of the bioinformatic analysis highlights the inability of bioinformatics alone to capture the results of bacterial transcription without including effects of RNA degradation. This study emphasizes the need for comparative analysis and potential dangers of genome reduction, exemplified through the discovery of altered gene expression patterns of JCVI-syn1.0 and JCVI-syn3A, achieved via the union of our transcriptome study with their proteomics data. Analysis of the transcriptomics data sets through a Jupyter notebook allows any genomic region to be easily examined. Table of Content Image:

Plant-microbe symbioses such as the legume-rhizobium mutualism are vital in the web of ecological relationships within both natural and managed ecosystems, influencing primary productivity, crop yield, and ecosystem services. The outcome of these interactions for plant hosts varies quantitatively and can range from highly beneficial to even detrimental depending on natural genetic variation in microbial symbionts. Here, we take a systems genetics approach, harnessing the genetic diversity present in wild rhizobial populations to predict genes and molecular pathways crucial in determining partner quality, i.e., the benefits of symbiosis for legume hosts. We combine traits, dual-RNAseq of both partners from active nodules, pangenomics/pantranscriptomics, and Weighted Gene Co-expression Network Analysis (WGCNA) for a panel of 20 Sinorhizobium meliloti strains that vary in symbiotic partner quality. We find that genetic variation in the nodule transcriptome predicts host plant biomass, and WGCNA reveals networks of genes in plants and rhizobia that are coexpressed and associated with high-quality symbiosis. Presence-absence variation of gene clusters on the symbiosis plasmid (pSymA), validated in planta, is associated with high or low-quality symbiosis and is found within important coexpression modules. Functionally our results point to management of oxidative stress, amino acid and carbohydrate transport, and NCR peptide signaling mechanisms in driving symbiotic outcomes. Our integrative approach highlights the complex genetic architecture of microbial partner quality and raises hypotheses about the genetic mechanisms and evolutionary dynamics of symbiosis.

While the world is aware of America's history of enslavement, the ongoing impact of anti-Black racism in the United States remains underemphasized in health intervention modeling. This Perspective argues that algorithmic bias—manifested in the worsened performance of clinical algorithms for Black vs. white patients—is significantly driven by the failure to model the cumulative impacts of racism-related stress, particularly racial heteroscedasticity. Racial heteroscedasticity refers to the unequal variance in health outcomes and algorithmic predictions across racial groups, driven by differential exposure to racism-related stress. This may be particularly salient for Black Americans, where anti-Black bias has wide-ranging impacts that interact with differing backgrounds of generational trauma, socioeconomic status, and other social factors, promoting unaccounted for sources of variance that are not easily captured with a blanket “race” factor. Not accounting for these factors deteriorates performance for these clinical algorithms for all Black patients. We outline key principles for anti-racist AI governance in healthcare, including: (1) mandating the inclusion of Black researchers and community members in AI development; (2) implementing rigorous audits to assess anti-Black bias; (3) requiring transparency in how algorithms process race-related data; and (4) establishing accountability measures that prioritize equitable outcomes for Black patients. By integrating these principles, AI can be developed to produce more equitable and culturally responsive healthcare interventions. This anti-racist approach challenges policymakers, researchers, clinicians, and AI developers to fundamentally rethink how AI is created, used, and regulated in healthcare, with profound implications for health policy, clinical practice, and patient outcomes across all medical domains.

The increasing global prevalence of diabetic nephropathy poses substantial health and economic burdens. Currently, effective anti-fibrotic therapies for managing kidney fibrosis associated with chronic kidney disease are lacking. This study reveals corisin, a microbiota-derived peptide, as a central driver in the progression of diabetic kidney fibrosis. Corisin levels were found to be markedly elevated in the serum of diabetic chronic kidney disease patients relative to healthy controls, with strong correlations to advanced disease stages and declining renal function. In a murine model of kidney fibrosis, corisin levels were similarly heightened, directly contributing to increased inflammation and worsening fibrosis and renal impairment. Notably, the use of a monoclonal anti-corisin antibody significantly reduced nephropathy severity in diabetic mice. Through molecular dynamics simulations and experimental validation, we demonstrated that corisin interacts with human serum albumin, potentially enhancing its renal accumulation and pathological impact. The pathogenic mechanism of corisin involves the acceleration of cellular senescence and the induction of epithelial-mesenchymal transition and apoptosis in kidney cells. These findings underscore the critical role of corisin in progressive diabetic nephropathy and suggest a promising new target for therapeutic intervention.

Gray whales (Eschrichtius robustus) are unique as bottom feeding baleen whales and they have long been a conservation concern on both sides of the Pacific, in part because they migrate and disperse farther than any other species on earth. They experienced drastic population size declines due to environmental changes and commercial whaling. Here, we present an improved genome assembly for the gray whale. This genome assembly covers 2.4 Gb divided across 2689 contigs with an N50 of 15Mb. From the new assembly, we identify 75Mb sex-linked contigs and a identify 94.6% of searched genes based on Benchmarking Universal Single-Copy Ortholog score. We use the gray whale assembly to explore the effects of mapping to conspecific vs allospecific reference genomes when estimating genome-wide heterozygosity (H) and runs of homozygosity (ROH). The use of allospecific genomes significantly underestimate both H and ROH burden regardless of genomic distance and assembly quality. Our analyses highlight the importance of using contiguous conspecific assemblies in whale genomics and conservation. ### Competing Interest Statement The authors have declared no competing interest.

We report here the complete genome sequence of a type II methanotrophic bacterium, Methylocystis echinoides strain RIM, isolated from the soil surface at Urbana, Illinois. This genome was obtained via HiFi PacBio Sequel II sequencing.

Abstract Chimeric antigen receptor (CAR)-T cell therapy has been shown to effectively treat solid tumors in syngeneic murine models, but antigen loss and T cell trafficking into the tumors pose major obstacles to its long-term efficacy. Here we explored a combination therapy against advanced stages of colorectal and ovarian syngeneic murine tumor models using a tumor-specific small molecule (PAC-1) that triggers the engagement of the host immune system and synergizes with a Tn-specific adoptively transferred 237 CAR-T immunotherapy. PAC-1 has been shown to induce apoptosis in tumor cells with high expression of procaspase 3, via chelation of inhibitory zinc and release of active executioner caspase 3. PAC-1 also recently completed a phase I trial (NCT02355535) and was shown to be well-tolerated in late-stage cancer patients. RNA sequencing of CT26 colorectal cells stimulated with PAC-1 for 24 hours revealed that PAC-1 induced global changes at the transcriptome level, up-regulating genes involved in immune activation and apoptosis, and down-regulating genes involved in DNA repair, cell cycle regulation, and myc-driven proliferation. Validation of RNAseq data was performed using a combination of RT-qPCR, Western blot, flow cytometry, and co-culture studies. A combination of transcriptome and exome sequencing analyses in murine colorectal cell line CT26 revealed that the PAC-1-mediated down-regulation of DNA repair machinery led to somatic alterations in the genome and a higher mutational load in tumor cells, subsequently presented as neo-antigens by antigen-presenting cells. As a result, host CD8+ cytotoxic T cells were able to infiltrate tumors and reduce the tumor burden. Tn-dependent CARs have been used recently to demonstrate the proof of concept in the targeting of cancer-specific Tn-antigens such as the Tn-MUC1 antigen in humans, and an analogous Tn-OTS8 target in mice. In our murine ID8 ovarian tumor model, untreated mice with disseminated tumors have a median survival of 71 days. As a single agent administered intraperitoneally, PAC-1 extended the median survival to 90 days (p<0.0001), while 237 CAR-T treated mice have a median survival of 103 days (p<0.0001). A combination of the two treatment modalities (PAC-1 administered before 237 CAR-T) further extended the median survival to 145 days (p=0.048). Administering PAC-1 after 237 CAR-T also modestly extended the median survival to 132 days (p=0.06). Taken altogether, our results demonstrate that PAC-1 engages the immune system to treat cancer by inducing expression of pro-inflammatory cytokines in tumor cells allowing the recruitment of CD8+ effector T cells to the tumor microenvironment. Combination strategies that boost T cell trafficking shows great promise to enhance the anti-tumor efficacy and feasibility of adoptive CAR-T therapy against solid tumors. Citation Format: Diana R. Ranoa, Yifei Kang, Jenny Drnevich, Gloria Rendon, Christopher J. Fields, Keith Bailey, Edward J. Roy, Timothy M. Fan, David M. Kranz, Paul J. Hergenrother. Small molecule procaspase activating compound 1 (PAC-1) enhances CAR-T immunotherapy for solid tumors in syngeneic murine models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB075.

ABSTRACT The global evolution of SARS-CoV-2 depends in part upon the evolutionary dynamics within individual hosts with varying immune histories. To characterize the within-host evolution of acute SARS-CoV-2 infection, we sequenced saliva and nasal samples collected daily from vaccinated and unvaccinated individuals early during infection. We show that longitudinal sampling facilitates high-confidence genetic variant detection and reveals evolutionary dynamics missed by less-frequent sampling strategies. Within-host dynamics in both unvaccinated and vaccinated individuals appeared largely stochastic; however, in rare cases, minor genetic variants emerged to frequencies sufficient for forward transmission. Finally, we detected significant genetic compartmentalization of viral variants between saliva and nasal swab sample sites in many individuals. Altogether, these data provide a high-resolution profile of within-host SARS-CoV-2 evolutionary dynamics. IMPORTANCE We detail the within-host evolutionary dynamics of SARS-CoV-2 during acute infection in 31 individuals using daily longitudinal sampling. We characterized patterns of mutational accumulation for unvaccinated and vaccinated individuals, and observed that temporal variant dynamics in both groups were largely stochastic. Comparison of paired nasal and saliva samples also revealed significant genetic compartmentalization between tissue environments in multiple individuals. Our results demonstrate how selection, genetic drift, and spatial compartmentalization all play important roles in shaping the within-host evolution of SARS-CoV-2 populations during acute infection.

Brewer's dried yeast has a high nutritional value and has long been utilized by the animal feed industry as a source of protein, B-complex vitamins, and minerals. Brewer's dried yeast is also rich in bioactive compounds and may thereby be used as a functional ingredient, providing benefits beyond that of its nutrient content. Canola meal is a high-fiber ingredient that also has unique properties, especially when it is wetted and dried using a proprietary drying system that creates a "functionalized" canola meal. The objective of this experiment was to evaluate the effects of a yeast-enriched functionalized canola meal (FCM) on apparent total tract digestibility (ATTD) and the fecal quality, metabolite concentrations, and microbiota populations, and immune function of healthy adult dogs. Twelve adult female beagles (body weight [BW] = 7.6 ± 0.7 kg; age = 5.8 ± 1.3) were used in a replicated 4 × 4 Latin square design with 28-d periods. Each experimental period consisted of a 22-d adaptation phase, 5 d of total and fresh fecal collection, and blood collection on the last day. To start, all dogs were fed a basal diet to maintain BW for 14 d. Following fecal and blood collections at baseline (-1 d) to confirm health status, experimental periods began testing the following dietary treatments using a Latin square design experiment: 1) FCM only (no yeast inclusion), 2) FCM + low yeast dose, 3) FCM + medium yeast dose, and 4) FCM + high yeast dose. All treatments were top-dressed onto the basal diet at a rate estimated to be 1% of daily intake (as-is basis). Statistical analysis was performed using the PROC MIXED procedure of SAS with the main effect of treatment and the random effect of dog. Significance was declared at P ≤ 0.05, and trends reported if 0.05 < P ≤ 0.10. Supplementation with yeast-enriched FCM had no significant effect on the ATTD of macronutrients or energy or the fecal characteristics, metabolite concentrations, and microbiota populations of dogs. Additionally, no significant differences were observed in circulating immune cell counts or response to Toll-like receptor agonists among treatments. Our results suggest that the yeast-enriched FCM could be included in canine diets without negatively affecting stool quality, fecal metabolite concentrations, or ATTD. Further research is necessary to determine the effective dose of yeast-enriched FCM, potential mechanisms of action, and other potential implications it has on canine health.