Biology subjects

Graf, J.

Publications and source records attributed to Graf, J..

At least 19 recordsLinked to original sources

bioRxiv2026

A rhizarian genome reveals an osmotrophic route to extracellular digestion in eukaryotes

Rhizaria, one of the most species-rich and ecologically important eukaryotic supergroups, accommodates a newly discovered osmotrophic species, yet high-quality genomic resources remain scarce, limiting our understanding of their metabolic diversity and ecological functions. Here, we present the genome of Saccharomycomorpha psychra, the first rhizarian telomere-anchored assembly, to be explored as a model and reference for rhizarian ecology and evolution. The 62 Mb assembly, of which half comprises 22 telomere-to-telomere scaffolds, shows a BUSCO completeness of 94.2%, encodes 17,680 genes, and provides the genomic foundation for investigating rhizarian ecology and evolution. The genome of osmotrophic Saccharomycomorpha psychra reveals a functionally integrated secretome of 1,015 proteins dominated by carbohydrate-active enzymes (CAZymes), proteases, lipases, and oxidoreductases. Taken together with 303 predicted high-confidence membrane transporters of a total of 680, skewed toward H-coupled secondary carriers, these features constitute the genomic signature of an extracellular digestive strategy convergent with saprotrophic fungi. Phenotypic MicroArrayTM assays confirmed active utilization of 17 carbon sources, including all six C5 pentose sugars tested, consistent with the predicted arabinose and ribokinase pathways among the most highly expressed metabolic genes in the transcriptome. These findings demonstrate that osmotrophic saprotrophy has evolved independently in Rhizaria with enzymatic solutions that closely parallel those of evolutionary distant fungal decomposers, highlighting the power of ecological context over phylogenetic heritage in shaping extracellular metabolic architecture.

genomics↗

bioRxiv2026

Succession and Shifting Identities in Freshwater, Built Environment Biofilm Communities

Biofilms on aquaculture infrastructure harbor diverse microbial communities that may influence water quality and fish health, yet the temporal dynamics of these communities remain poorly characterized. Here, we used 16S rRNA gene amplicon sequencing to profile biofilm communities on concrete raceway surfaces across an 80-day rainbow trout (Oncorhynchus mykiss) indoor hatch-house production period. One hundred twenty-three wall swab samples from 19 raceways at six time points (9, 23, 38, 53, 65, and 80 days) were analyzed after stringent quality control. Beta diversity analyses revealed that biofilm communities at each time point were significantly distinct (PERMANOVA, p < 0.001 for all pairwise comparisons), with early communities exhibiting greater variability than late-stage biofilms. Total bacterial load increased approximately 2.5-fold from early to late stages (qPCR, p < 0.001). Differential abundance testing (ANCOM-BC) identified 57 differentially abundant genera between early-and late-stage biofilms, and random forest classification distinguished early from late communities with over 93% test accuracy. A clear successional trajectory emerged: early biofilms were dominated by pioneer taxa including Pseudomonas, Caulobacter, and Flavobacterium; mid-succession communities featured predatory Bdellovibrio and the methylotroph Methylotenera; and mature biofilms were enriched in saprophytic Saprospiraceae and Haliscomenobacter, polysaccharide-degrading Verrucomicrobiaceae, and cooperative predatory myxobacteria. Flavobacterium columnare, a pathogen of concern in aquaculture, was detected at low levels throughout the production period. These results demonstrate predictable ecological succession in freshwater built environment biofilms and provide a foundation for understanding the role of surface-associated microbial communities in hatchery management.

microbiology↗

bioRxiv2026

Spontaneous network activity links misrouted interneuron migration to cortical maturation abnormalities

Cortical inhibitory neurons (CIN) populate the neocortex and hippocampus by extensive tangential migration. This process is highly vulnerable to genetic and environmental disturbances and is linked to neuropsychiatric disorders. However, the mechanisms by which transient migratory abnormalities translate into persistent functional deficits remain poorly understood. Here, we utilized a conditional Cxcr4 knockout to investigate the consequences of disrupted migratory guidance - a feature of genetic schizophrenia models. This demonstrated that despite migrating in ectopic cortical layers, CIN quantitatively colonized the neocortex until birth, whereas limbic regions developed a permanent deficit in CIN numbers. Furthermore, CIN failed to populate the neocortical marginal zone, a transient reservoir for late-born CIN destined for superficial cortical layers. Consequently, the layering and molecular identities of CIN, as well as the synaptic connectivity and spontaneous activity of the neuronal network, were significantly altered in the early postnatal neocortex. Although abnormal CIN layering was gradually compensated before maturity, functional differences persisted, as evidenced by facilitated propagation of sensory stimuli between cortical areas. These results demonstrate that CIN migration is an instructive process critical for activity-driven early postnatal network maturation and CIN identity, thus providing a mechanistic link between migratory guidance and the integrity of mature circuits.

neuroscience↗

bioRxiv2025

From Biofilms to Biocontrol: Aquaculture Biofilms as a Reservoir of Pseudomonas spp. with Activity Against Fish and Human Pathogens

The antimicrobial resistance crisis, driven by excessive use of antimicrobials in medical and agricultural settings, has spurred the search for alternative strategies. In the present study, we investigated surface biofilms from a rainbow trout aquaculture facility to identify bacteria with inhibitory activity against fish and human pathogens. A total of 906 isolates were obtained, of which 478 were taxonomically classified using full-length 16S rRNA gene sequencing, revealing Pseudomonas, Aeromonas, and Flavobacterium as the most abundant genera. Twenty-nine isolates, predominantly novel Pseudomonas species, inhibited one or more fish pathogen strains, with two P. aphyarum strains inhibiting every fish pathogen tested. All human pathogen strains were inhibited by at least one trout farm isolate. A biofilm assay identified strains capable of excluding F. columnare from established biofilms. Numerous strains reduced biofilm colonization, with P. aphyarum and P. idahonensis strains exhibiting the greatest reduction. Genome analysis and biosynthetic gene cluster (BGC) identification from a subset of isolates revealed shared and unique clusters with strong potential for antimicrobial production, correlating in part with observed inhibition patterns. Metabolomics profiling identified a suite of strain-specific siderophores that may mediate biofilm disruption. Additionally, co-culture of these isolates with the human pathogen Pseudomonas aeruginosa enhanced the production of specialized metabolites with reported antimicrobial activities. Collectively, these findings highlight trout farm biofilms as a promising source of bacteria that could be developed as probiotics or investigated for novel antimicrobial discovery. ImportanceDisease outbreaks in aquaculture facilities pose significant threats to food security and often rely on antibiotic treatments, contributing to antimicrobial resistance. This study identifies Pseudomonas isolates from native biofilms of a trout aquaculture facility that are capable of disrupting biofilms and inhibiting a wide range of fish and human pathogens. These Pseudomonas isolates produce a variety of siderophores that may mediate biofilm disruption, offering a promising avenue for developing probiotic applications or the discovery of new antimicrobials to improve aquaculture health and reduce reliance on antibiotics. The discovery of these biofilm-disrupting bacteria holds potential significance for combating antimicrobial resistance in both aquaculture and clinical settings.

microbiology↗

bioRxiv2025

Integration of Multiomic and Multi-phenotypic Data Identifies Biological Pathways Associated with Physical Fitness

Unraveling the complex associations between human phenotypes and molecular pathways can pave the way to improved health and performance, but faces a fundamental challenge: the measurable genes, proteins, and metabolites vastly outnumber the participants in even the largest studies, yielding spurious correlations. To address this imbalance, we have developed a bioinformatic framework and computational approach ("PhenoMol") to discover biological drivers of phenotypic characteristics that integrates all available phenotypic data predictive of outcomes and reduces multi-omic data dimensionality by generating "expression circuits" via graph theory constrained by prior biological knowledge of molecular interactions. We applied PhenoMol to analyze causal patterns and predict elite physical performance in a healthy cohort with deep physiological, physical, behavioral, cognitive, and molecular characterization. PhenoMol outperforms regression models based on equivalent analytic methodologies that do not employ network biology for dimensionality reduction. The PhenoMol software is provided for future studies.

bioinformatics↗

bioRxiv2025

Comparison of a long-read amplicon sequencing approach to short-read amplicons for microbiome analysis

Most microbiome studies to date rely on sequencing short amplicons of the 16S rRNA gene on Illuminas platforms. Because of the short read length, sequences often can be identified reliably only to the family or genus levels. Long read sequencing with whole-length 16S rRNA sequencing can improve taxonomic resolution, but often only to the species level. StrainID is an alternative approach that amplifies a large segment of the ribosomal operon, including the entire 16S rRNA gene, internal transcribed spacer, and a portion of the 23S rRNA gene. This longer amplicon is designed to allow ribotype-level classification. Although studies have demonstrated the utility of StrainID for several sample types, it has not yet been validated for saliva. Here, we compared the performance of StrainID to short read amplicons with saliva samples as well as a synthetic mock DNA community and human and mouse fecal samples. Short reads were amplified with primer pairs appropriate for the corresponding sample type, and were classified with two different taxonomic databases. For both saliva and fecal samples, we found that StrainID performed similarly to short reads overall and demonstrated a key benefit with phylogenetic-based beta diversity tests and taxonomic classification. Our results further build on establishing StrainID as a valid method and specifically validate its use with saliva samples. ImportanceThe interpretation of microbiome composition studies is highly dependent on the methodologies chosen during experimental design, which affects factors such as resolution, throughput, cost, and accuracy. StrainID is an approach that can improve resolution while maintaining high-throughput and similar costs to short-read sequencing. The salivary microbiome represents a diverse community of microbes with links to a variety of health conditions and disease states. Closely related strains of bacteria can have drastically different effects on their host. Establishing StrainID as a valid approach for studying the salivary microbiome opens avenues for research that improve upon alternative methods by increasing sensitivity and accuracy compared to traditional short read approaches.

microbiology↗

bioRxiv2025

High-Resolution Spatial Proteomics Characterises Colorectal Cancer Consensus Molecular Subtypes

BackgroundIdentification of the consensus molecular subtypes (CMS) opened significant potential for understanding the tumor biology and intertumoral heterogeneity of colorectal cancer (CRC). However, molecular subtyping in CRC traditionally relies on bulk transcriptomics, therefore, lacks spatial and single-cell level aspect. MethodsWe constructed tissue microarrays using tumor cores from 222 CRC patients. Arrays were stained and imaged using 54 cell identity and cancer hallmark markers, delivering spatially resolved protein profiles of >2 million cells. RNA sequencing data and CMS classification were also available for these patients. After segmentation of cancer, stromal and immune cells, we investigated intratumoral heterogeneity within CMS subtypes using spatially resolved single-cell protein profiling (>2 million cells). We compared cell types, their spatial organization and their expression of cancer hallmark-related proteins in CMS 1-4 subtypes. ResultsWe revealed tissue atlases illustrating the cell types/states, spatial heterogeneity, cellular neighborhoods, cellular network, and single-cell protein profiles of CMS tumors. CMS1 tumors had more CD3+, CD8+, and PD1+ immune cells that were found in the epithelial layer frequently. CMS1 was also associated with higher levels of metabolic reprogramming markers such as upregulated glycolysis. CMS2 showed immune segregation, reactive stroma patterns and higher levels of apoptotic and proliferative signaling proteins. CMS3 exhibited clustered cancer cells with high RIP3 levels, suggesting a pro-inflammatory microenvironment. CMS4 displayed stromal-centric and immune-evasive tumors characterized by decreased HLA-1 levels and regulatory T-cell exclusion from epithelium. ConclusionWe present a spatial protein atlas of CRC at single-cell resolution and demonstrate novel aspects of CMS tumour structures.

cancer biology↗

bioRxiv2025

Identification and characterization of the surface layer protein AvsA in outer membrane vesicles, antibiotic resistance, and in vivo host colonization in Aeromonas veronii

Outer membrane vesicles (OMVs) are important in bacterial communication and the transfer of virulence factors. In this study, we identified and characterized the surface layer protein (SLP) AvsA (Aeromonas veronii surface protein A) in the OMVs of Aeromonas veronii Hm21, a strain isolated from the medicinal leech Hirudo verbana. The surface layer proteins (SLPs) play critical roles in how bacteria interact with each other and their environments, particularly in mediating antibiotic resistance and facilitating host colonization. Furthermore, we investigate the ability of AvsA to confer protection against antibiotics, affect biofilm formation, and contribute to host colonization, providing insight into antibiotic resistance and two crucial factors contributing to the persistence of the bacteria in its host. Our findings suggest that AvsA enhances antibiotic tolerance, facilitates biofilm development, and is important for successful colonization of the leech digestive tract. These data demonstrate that AvsA performs important roles in a wide range of critical phenotypes. This work provides insights into the functional significance of SLPs in A. veronii and highlights AvsA as a potential target for modulating bacterial colonization and resilience against antibiotics. ImportanceOuter membrane vesicles (OMVs) are important for bacterial communication, pathogenesis, and stress adaptation, yet how this is accomplished remains poorly understood. Here, we identify and characterize a surface layer protein (SLP), AvsA, associated with OMVs in Aeromonas veronii Hm21. Bioinformatic, phylogenetic, and mass spectrometry analyses suggest that Aeromonas veronii ORF M001_06550 encodes a surface layer protein (SLP) with high similarity to a characterized A. hydrophila SLP, supporting its designation as Aeromonas veronii Surface Protein A (AvsA). AvsA forms a paracrystalline layer on bacterial cells and OMVs. Functionally, AvsA contributes to antibiotic resistance, enhances biofilm formation, and is essential for colonization in a symbiotic host, the medicinal leech. These findings highlight a novel role of SLPs in bacterial physiology and host interactions. Given the widespread presence of AvsA homologs, our study provides insights into conserved bacterial mechanisms that may be relevant for both pathogenic and beneficial host-microbe interactions.

microbiology↗

bioRxiv2024

Anti-NMDAR encephalitis alters intrinsic spatiotemporal coding by enhancing neuronal coupling and clustering

Autoimmune anti-NMDA-receptor encephalitis is characterized by severe neuropsychiatric symptoms including memory dysfunction and seizures. However, it remains enigmatic what functional changes at the multi-neuronal level mediate network dysfunction. We used two-photon in vivo recording in a passive-transfer mouse model with patients monoclonal anti-GluN1-autoantibodies during slow-wave sleep-like conditions, a critical phase for memory processing. We find enhanced functional coupling and clustering between hippocampal CA1 pyramidal neurons (PNs), which intrinsically expose the network to hypersynchrony. These connectivity changes are associated with a selective preservation of strong excitatory synapses despite overall reduced excitation, thus enhancing hub-like properties of functionally connected PNs. Furthermore, we find abnormal PN firing characteristics, decreased transmission failure, and increased similarity of spontaneous spatiotemporal activity patterns, all affecting CA1 intrinsic neuronal coding. Collectively, the functional rewiring of hippocampal networks and altered intrinsic information processing provide new mechanistic insights into the NMDAR-hypofunction consequences and pathomechanisms of anti-NMDAR encephalitis symptomatology.

neuroscience↗

bioRxiv2024

Chemogenetic silencing reveals presynaptic Gi/o protein-mediated inhibition of synchronized activity in the developing hippocampus in vivo

The development of neuronal circuits is an activity-dependent process. Research aimed at deciphering the learning rules governing these developmental refinements is increasingly utilizing a class of chemogenetic tools that employ Gi/o protein signaling (Gi-DREADDs) for neuronal silencing. However, their mechanisms of action and inhibitory efficacy in immature neurons with incompletely developed Gi/o signaling are poorly understood. Here, we analyze the impact of Gi/o signaling on cellular and network excitability in the neonatal hippocampus by expressing the Gi-DREADD hM4Di in telencephalic glutamatergic neurons of neonatal mice of both sexes. Using acousto-optic two-photon Ca2+ imaging, we report that activation of hM4Di leads to a complete arrest of spontaneous synchronized activity in CA1 in vitro. Electrophysiological analyses demonstrate that hM4Di-mediated silencing is not accounted for by changes in intrinsic excitability of CA1 pyramidal cells (PCs). Instead, activation of hM4Di robustly restrains synaptic glutamate release by the first postnatal week, effectively reducing recurrent excitation in CA1. In vivo, inhibition through hM4Di potently suppresses early sharp waves (eSPWs) and discontinuous oscillatory network activity in CA1 of head-fixed mice before eye opening. In summary, hM4Di dampens glutamatergic neurotransmission through a presynaptic mechanism sufficient to terminate spontaneous synchronized activity in the neonatal CA1. Our findings have implications for designing and interpreting experiments utilizing Gi-DREADDs in immature neurons and further point to a potential role of Gi/o-dependent endogenous neuromodulators in activity-dependent hippocampal development. Significance StatementRecent advances in understanding how early activity shapes developing brain circuits increasingly rely on Gi/o-dependent inhibitory chemogenetic tools (Gi-DREADDs). However, their single-cellular mechanisms and efficacy in neurons with immature Gi/o signaling are elusive. To close this gap, we analyzed the impact of the Gi-DREADD hM4Di in the neonatal murine hippocampus. We report that hM4Di does not mediate hyperpolarization or shunting but rather acts as a potent presynaptic silencer of glutamatergic neurotransmission in CA1 pyramidal cells. Presynaptic inhibition of excitation suffices to arrest spontaneous synchronized network activity in vitro and in vivo. Our findings provide novel insights into the generative mechanisms of spontaneous synchronized activity and bear relevance for applying chemogenetic silencing at early stages of brain development.

neuroscience↗

bioRxiv2023

The long non-coding RNA LINC00941 modulates MTA2/NuRD occupancy to suppress premature human epidermal differentiation

Numerous long non-coding RNAs (lncRNAs) were shown to have functional impact on cellular processes such as human epidermal homeostasis. However, the mechanism of action for many lncRNAs remains unclear to date. Here, we report that lncRNA LINC00941 regulates keratinocyte differentiation on an epigenetic level through association with the NuRD complex, one of the major chromatin remodelers in cells. We find that LINC00941 interacts with NuRD-associated MTA2 in human primary keratinocytes. LINC00941 perturbation changes MTA2/NuRD occupancy at bivalent chromatin domains in close proximity to transcriptional regulator genes, including the EGR3 gene coding for a transcription factor regulating epidermal differentiation. Notably, LINC00941 depletion resulted in reduced NuRD occupancy at the EGR3 gene locus, increased EGR3 expression in human primary keratinocytes, as well as increased abundance of EGR3-regulated epidermal differentiation genes in cells and human organotypic epidermal tissue. Our results therefore indicate a role for LINC00941/NuRD in repressing EGR3 expression in non-differentiated keratinocytes, consequentially preventing premature differentiation of human epidermal tissue.

molecular biology↗

bioRxiv2022

Pseudomonas apudapuas sp. nov., Pseudomonas fontis sp. nov., Pseudomonas idahonensis sp. nov., and Pseudomonas rubra sp. nov. isolated from in, and around, a rainbow trout farm

During a large-scale bacterial culturing effort of biofilms in the vicinity of a rainbow trout aquaculture facility in Idaho, USA, ten isolates were identified as having pathogen inhibiting activity and were characterized further. These isolates were shown to be Gram negative, rod-shaped bacteria belonging to the genus Pseudomonas. Whole genome comparisons and multi-locus sequence analysis using four housekeeping genes (16S rDNA, gyrA, rpoB, rpoD) showed that these 10 isolates clustered into four distinct species groups. These comparisons also indicated that these isolates were below the established species cutoffs for the genus Pseudomonas. Further phenotypic characterization using API 20NE, API ZYM, and BioLog GENIII assays and chemotaxonomic analysis of cellular fatty acids were carried out. Based on the genomic, physiological, and chemotaxonomic properties of these isolates, we concluded that these strains composed four novel species of Pseudomonas. The proposed names are: Pseudomonas apudapuas sp. nov. consisting of strains ID233, ID386T, and ID387 with ID386T (DSM 114641) as the type strain; Pseudomonas rubra sp. nov. consisting of strains ID291T, ID609, and ID1025 with ID291T (DSM 114640) as the type strain; Pseudomonas idahonensis sp. nov. consisting of strains ID357T and ID1048 with ID357T (DSM 114609) as the type strain; and Pseudomonas fontis sp. nov. consisting of strains ID656T and ID681 with ID656T (DSM 114610) as the type strain.

microbiology↗

bioRxiv2022

Doomed by popularity: The broad use of the Pm8 resistance gene in wheat resulted in hypermutation of the AvrPm8 gene in the powdery mildew pathogen

O_LIThe Pm8 resistance gene against the powdery mildew disease has been introgressed from rye into wheat as part of a large 1BL.1RS chromosomal translocation. Due to its high agronomic value, this translocation has seen continuous global use since the 1960s on large growth areas, even after Pm8 resistance was overcome. This allows studying the effect of long and widespread resistance gene use on a pathogen population. C_LIO_LIUsing genome wide association studies in a global population of wheat mildew isolates, we identified the avirulence effector AvrPm8 specifically recognized by Pm8. C_LIO_LIHaplovariant mining in the global population revealed 17 virulent haplotypes of the AvrPm8 gene that grouped into two categories. The first one from geographically diverse regions comprised two single amino acid polymorphisms at the same position in the AvrPm8 protein, which we confirmed to be crucial for recognition by Pm8. The second category consisted of numerous destructive mutations to the AvrPm8 open reading frame. C_LIO_LIMost individual gain-of-virulence mutations were found in geographically restricted regions, indicating they occurred recently as a consequence of the frequent Pm8 use. We conclude that both standing genetic variation as well as locally occurring new mutations contributed to the global breakdown of the Pm8 resistance gene introgression. C_LI

plant biology↗

bioRxiv2022

Large-Scale, Multi-Year Microbial Community Survey of a Freshwater Trout Aquaculture Facility

Aquaculture is an important tool for solving growing worldwide food demand, but infectious diseases of the farmed animals represent a serious roadblock to continued industry growth. Therefore, it is essential to understand the microbial communities that reside within the built environments of aquaculture facilities to identify reservoirs of bacterial pathogens and potential correlations between commensal species and specific disease agents. Here, we present the results from three years of sampling a commercial rainbow trout aquaculture facility. The sampling was focused on the early-life stage hatchery building and included sampling of the facility source water and outdoor production raceways. We observed that the microbial communities residing on the abiotic surfaces within the hatchery were distinct from those residing on the surfaces of the facility water source as well as the production raceways, despite similar communities in the water column at each location. Within the hatchery building, most of the microbial classes and families within surface biofilms were also present within the water column, suggesting that these biofilms are seeded by a unique subgroup of microbial taxa from the water. Lastly, we detected a common fish pathogen, Flavobacterium columnare, within the hatchery, including at the source water inlet. Importantly, the relative abundance of this pathogen was correlated with clinical disease. Our results characterized the microbial communities in an aquaculture facility, established that the hatchery environment contains a unique community composition, and demonstrated that a specific fish pathogen resides within abiotic surface biofilms and is seeded from the natural source water. ImportanceThe complex microbial consortium residing in the built environment of aquaculture facilities is poorly understood. In this study, we provide a multi-year profile of the surface- and water-associated microbial communities of this biome. The results demonstrated that distinct community structures exist in the water and on surfaces. Furthermore, it was shown that a common and economically impactful bacterial pathogen, F. columnare, is continually introduced via the source water, is widespread within surface biofilms in the hatchery environment, and is likely amplified within these raceways but does not always cause disease despite being present. These results advance our understanding of pathogen localization at fish farms, show the interplay between host and environmental microbiomes, and reveal the importance of microbial community sequencing in aquaculture for identifying potential beneficial and harmful microbes. This study adds to the aquaculture microecology dataset and enhances our ability to understand this environment from a "One Health" perspective.

microbiology↗

bioRxiv2021

Detecting Flavobacterial Fish Pathogens in the Environment Using High-Throughput Community Analysis

Diseases caused by the fish pathogens Flavobacterium columnare and Flavobacterium psychrophilum are major contributors of preventable losses in the aquaculture industry. The persistent and difficult to control infections caused by these bacteria make timely intervention and prophylactic elimination of pathogen reservoirs important measures to combat these disease-causing agents. In the present study, we present two independent assays for detecting these pathogens in a range of environmental samples. Natural water samples were inoculated with F. columnare and F. psychrophilum cells, and pathogen levels were detected using Illumina MiSeq sequencing and droplet digital PCR. Both detection methods accurately identified pathogen-positive samples and showed good agreement in quantifying each pathogen. Additionally, the real-world application of these approaches was demonstrated using environmental samples collected at a rainbow trout aquaculture facility. These results show that both methods can serve as useful tools for surveillance efforts in aquaculture facilities, where the early detection of these flavobacterial pathogens may direct preventative measures to reduce disease occurrence. ImportanceEarly detection of a deadly disease outbreak in a population can be the difference between mass fatality or mitigated effects. In the present study, we evaluated and compared two techniques for detecting economically impactful aquaculture pathogens. We demonstrate that one of these techniques, 16S rRNA gene sequencing using Illumina MiSeq technology, provides the ability to accurately detect two fish pathogens, F. columnare and F. psychrophilum, while simultaneously profiling the native microbial community. The second technique, droplet digital PCR, is commonly used for pathogen detection, and the results obtained using the assays we designed with this method served to validate those obtained using the MiSeq method. These two methods offer distinct advantages. The MiSeq method pairs pathogen detection and microbial community profiling to answer immediate and long-term fish health concerns, while droplet digital PCR method provides fast and highly sensitive detection that is useful for surveillance and rapid clinical responses.

molecular biology↗

bioRxiv2021

Network instability dynamics drive a transient bursting period in the developing hippocampus in vivo

Spontaneous correlated activity is a universal hallmark of immature neural circuits. However, the cellular dynamics and intrinsic mechanisms underlying network burstiness in the intact developing brain are largely unknown. Here, we use two-photon Ca2+ imaging to comprehensively map the developmental trajectories of spontaneous network activity in hippocampal area CA1 in vivo. We unexpectedly find that network burstiness peaks after the developmental emergence of effective synaptic inhibition in the second postnatal week. We demonstrate that the enhanced network burstiness reflects an increased functional coupling of individual neurons to local population activity. However, pairwise neuronal correlations are low, and network bursts (NBs) recruit CA1 pyramidal cells in a virtually random manner. Using a dynamic systems modeling approach, we reconcile these experimental findings and identify network bi-stability as a potential regime underlying network burstiness at this age. Our analyses reveal an important role of synaptic input characteristics and network instability dynamics for NB generation. Collectively, our data suggest a mechanism, whereby developing CA1 performs extensive input-discrimination learning prior to the onset of environmental exploration.

neuroscience↗

bioRxiv2021

Host adaptation through hybridization: Genome analysis of triticale powdery mildew reveals unique combination of lineage-specific effectors

The emergence of new fungal pathogens through hybridization represents a serious challenge for agriculture. Hybridization between the wheat mildew (Blumeria graminis f.sp. tritici) and rye mildew (B.g. f.sp. secalis) pathogens have led to the emergence of a new mildew form (B.g. f.sp. triticale) growing on triticale, a man-made amphiploid crop derived from crossing rye and wheat which was originally resistant to the powdery mildew disease. The identification of the genetic basis of host-adaptation in triticale mildew has been hampered by the lack of a reference genome. Here we report the 141.4 Mb reference assembly of B.g. triticale isolate THUN-12 derived from long-read sequencing and genetic map-based scaffolding. All eleven B.g. triticale chromosomes were assembled from telomere-to-telomere and revealed that 19.7% of the hybrid genome was inherited from the rye mildew parental lineage. We identified lineage-specific regions in the hybrid, inherited from the rye or wheat mildew parental lineages, that harbour numerous bona fide candidate effectors. We propose that the combination of lineage-specific effectors in the hybrid genome is crucial for host-adaptation, allowing the fungus to simultaneously circumvent the immune systems contributed by wheat and rye in the triticale crop. In line with this we demonstrate the functional transfer of the SvrPm3 effector from wheat to triticale mildew, a virulence effector that specifically suppresses resistance of the wheat Pm3 allelic series. This transfer is the likely underlying cause for the observed poor effectiveness of several Pm3 alleles against triticale mildew and exemplifies the negative implications of pathogen hybridizations on resistance breeding.

genomics↗

bioRxiv2021

Standing genetic variation of the AvrPm17 avirulence gene in powdery mildew limits the effectiveness of an introgressed rye resistance gene in wheat

Introgressions of chromosomal segments from related species into wheat are important sources of resistance against fungal diseases. The durability and effectiveness of introgressed resistance genes upon agricultural deployment is highly variable - a phenomenon that remains poorly understood as the corresponding fungal avirulence genes are largely unknown. Until its breakdown, the Pm17 resistance gene introgressed from rye to wheat provided broad resistance against powdery mildew (Blumeria graminis). Here, we used QTL mapping to identify the corresponding wheat mildew avirulence effector AvrPm17. It is encoded by two paralogous genes that exhibit signatures of re-occurring gene conversion events and are members of a mildew sub-lineage specific effector cluster. Extensive haplovariant mining in wheat mildew and related sub-lineages identified several ancient virulent AvrPm17 variants that were present as standing genetic variation in wheat powdery mildew prior to the Pm17 introgression, thereby paving the way for the rapid breakdown of the Pm17 resistance. QTL mapping in mildew identified a second genetic component likely corresponding to an additional resistance gene present on the 1AL.1RS translocation carrying Pm17. This gene remained previously undetected due to suppressed recombination within the introgressed rye chromosomal segment. We conclude that the initial effectiveness of 1AL.1RS was based on simultaneous introgression of two genetically linked resistance genes. Our results demonstrate the relevance of pathogen-based genetic approaches to disentangle complex resistance loci in wheat. We propose that identification and monitoring of avirulence gene diversity in pathogen populations becomes an integral part of introgression breeding to ensure effective and durable resistance in wheat. Significance StatementDomesticated and wild wheat relatives provide an important source of new immune receptors for wheat resistance breeding against fungal pathogens. The durability of these resistance genes is variable and difficult to predict, yet it is crucial for effective resistance breeding. We identified a fungal effector protein recognised by an immune receptor introgressed from rye to wheat. We found that variants of the effector allowing the fungus to overcome the resistance are ancient. They were already present in the wheat powdery mildew gene pool before the introgression of the immune receptor and are therefore responsible for the rapid resistance breakdown. Our study demonstrates that the effort to identify new resistance genes should be accompanied by studies of avirulence genes on the pathogen side.

plant biology↗