Biology subjects

Chatzinotas, A.

Publications and source records attributed to Chatzinotas, A..

11 recordsLinked to original sources

bioRxiv2026

A new automated pipeline for whole genome shotgun sequencing analysis and hazard characterization of microbial pesticides

Microbial pesticides are increasingly important for sustainable crop protection, yet hazard analysis and risk assessment remain challenging and require specific characterization of properties relevant to both safety and biological activity. Whole-genome sequencing (WGS) can support the identification of microorganisms at high resolution and characterize potential hazards associated with infectivity, pathogenicity, antimicrobial resistance, and toxic metabolite production. However, the routine use of WGS in this context requires accessible, reproducible, and interpretable workflows. Here, we present a publicly available web-based workflow for WGS-supported hazard analysis of microbial biocontrol agents. The workflow accepts assembled genomes as well as short-read, long-read and hybrid sequencing data, and performs genome quality assessment, taxonomic assignment, genome annotation, AMR detection, mobile-element screening, pathogenicity prediction and secondary-metabolite analysis, and compiles the results into an HTML report. For bacterial agents, we implement a transparent rule-based risk-classification module integrating taxonomic identity, PathogenFinder2 predictions, CARD/RGI resistance evidence, WHO priority taxa, and medically important antimicrobial categories. Secondary-metabolite assessment combines antiSMASH with local BLAST searches and EFSA-aligned identity/coverage thresholds to support product-level interpretation of biosynthetic gene clusters. Comparison with the currently used MOpS workflow developed by EFSA demonstrates the added benefits of the proposed workflow for a guided hazard analysis of microbial pesticides. The workflow is intended as a community-accessible pre-assessment tool that complements regulatory platforms by improving transparency, reproducibility, and early identification of potential hazards in microbial biocontrol candidates, envisioned to make WGS an integral part of the risk assessment of microbial pesticides. Key pointsO_LIThe RATION-GUI provides a public, reproducible workflow for whole-genome sequencing- supported hazard characterization of bacterial and fungal microbial pesticides. C_LIO_LIThe workflow integrates genome quality, taxonomy, annotation, antimicrobial resistance, pathogenicity-related evidence, mobile elements, and secondary-metabolite potential. C_LIO_LIDomain-specific outputs are translated into structured hazard indicators, evidence summaries, and recommended follow-up actions without replacing expert regulatory judgment. C_LIO_LICase studies demonstrate how the workflow improves transparency, traceability, and interpretation of genomic evidence for microbial pesticide risk assessment. C_LI

bioinformatics↗

bioRxiv2026

The illusion of diversity: sampling design drives conflicting estimates of soil bacterial richness

Soil microbial diversity is shaped by the spatial scale at which communities are sampled, yet standard sampling practices often homogenize samples, obscuring fine-scale spatial structure and diversity patterns. To better understand how sampling effort, spatial extent, and physical homogenization influence plot-level microbial richness estimates, we sampled 57 forest and grassland sites across three regions in Germany using a 14-core cross-transect design, and performed 16S rRNA gene metabarcoding. We simulated sampling efforts of 1-14 cores and a range of spatial extents, and compared diversity estimates to those from physically homogenized composite samples. Plot-level richness increased continuously with sampling effort and spatial extent, with no evidence of saturation. However, when sequencing depth was held constant, sampling completeness declined with increasing sampling effort, meaning that more diversity is not captured. Composite samples substantially underestimated plot-level richness and altered apparent diversity relationships between ecosystems; individual cores identified forests as richer than grasslands, whereas homogenized samples suggested the opposite relationship. These results demonstrate that sampling effort, spatial extent, and homogenization fundamentally shape soil microbial diversity estimates. Homogenized composite samples cannot substitute for individual cores when the goal is to reliably quantify plot-level richness or compare diversity across ecosystems.

ecology↗

bioRxiv2025

PHI: Prophage-Host Interaction toolkit for automated prediction and comprehensive profiling of prophages and their hosts via Galaxy

BackgroundViruses that infect bacteria, known as bacteriophages or phages, are widespread in nature and play important roles in shaping microbial communities and ecosystem functions. Some phages can integrate into bacterial genomes as "prophages", where they may influence the biology of their host by carrying genes that affect metabolism, virulence, or environmental adaptation. Despite their importance, studying prophages and their interactions with bacterial hosts remains challenging because it typically requires combining many complex computational tools and can be resource-intensive. ResultsIn this study, we introduce the Prophage-Host Interaction Toolkit (PHI), a user-friendly and automated workflow available through the Galaxy platform. PHI brings together multiple established tools into a single, reproducible pipeline that identifies candidate prophages, evaluates their quality, predicts host relationships, and characterizes key functional genes. Importantly, all results are summarized in an interactive report that simplifies interpretation. When applied to a mock community composed of 22 bacteria as a workflow demonstration, PHI detected 41 prophages across 14 hosts, classifying them into high- and medium-quality phage genomes. Host assemblies exhibited > 99 % completeness and < 1 % contamination for most genomes, while DefenseFinder revealed between 3 and 24 antiviral systems per genome. ConclusionsBy removing installation barriers and consolidating the outputs of multiple established tools, PHI lowers the barrier to advanced phage analysis, enabling both specialists and non-experts to explore phage-host interactions and their implications in areas such as microbiome research, biotechnology, and environmental science.

bioinformatics↗

bioRxiv2025

HVRLocator: A Computationally Efficient Tool for Identifying Hypervariable Regions in 16S rRNA Big Datasets

BackgroundAmplicon sequencing of the 16S rRNA gene is widely used to assess microbial diversity due to its cost-effectiveness and efficiency. However, public 16S rRNA datasets often lack standardized metadata, particularly information on the sequenced hypervariable regions or primers used, which are critical for accurate analysis and data reuse. To address this, we present the HVRLocator, a computational tool that reliably identifies sequenced hypervariable regions, enhancing metadata quality and enabling more robust large-scale microbiome studies. ResultsThe HVRLocator tool processed samples at an average rate of 0.147 per minute. Validation confirmed 100% accuracy in predicting alignment positions, correctly matching sequences to the expected primer regions based on literature. We demonstrated how to use the tool to select appropriate and comparable sequences for building a global bacterial database from V4 region amplicons of the 16S rRNA gene. Using HVRLocator, we selected 36,217 valid samples out of 45,882 runs, enabling us to identify cases where metadata incorrectly labeled sequences as targeting the V4 region. ConclusionEven when metadata is available, it can be inaccurate or misleading. HVRLocator offers a reliable and efficient method to identify the exact hypervariable sequenced region, ensuring accurate processing of large-scale 16S rRNA amplicon data. By bypassing inconsistent metadata and literature, it streamlines data curation and enhances the reliability of microbial studies, syntheses, and meta-analyses. Its use is essential for critically evaluating published data and enabling accurate and reproducible research in microbial ecology.

bioinformatics↗

bioRxiv2025

Effects of a novel Paraburkholderia phage IPK on the phenanthrene degradation efficiency of the PAH-degrading strain Paraburkholderia caledonica Bk.

Phages are a major cause of bacterial mortality, affecting bacterial diversity and ecosystem functioning. However, the impact of phage-host interactions in contaminated environments and their role in pollutant biodegradation have been largely overlooked. We isolated and characterized a novel phage from a polycyclic aromatic hydrocarbon (PAH)-contaminated soil that infects the PAH-degrading bacterium Paraburkholderia caledonica Bk, and investigated the effect of different multiplicity of infection (MOI) on the degradation efficiency of phenanthrene. The phage IPK is a temperate phage with a wide pH and temperature tolerance and a burst size of 80 PFC.ml{square}1. The IPK phage was classified as a member of the Caudoviricetes, related to Pseudomonas and Burkholderia phages; however, its low intergenomic similarity indicates that it belongs to a new species. Three AMGs related to amino acid metabolism and to bacterial growth regulation were identified in the phage genome. The highest multiplicity of infection (MOI 10) showed a rapid recovery of the host density abundance and greater phenanthrene degradation than MOIs ranging from 0.01 to 1. This work highlights the critical role of phage-host interactions in modulating pollutant degradation efficiency, which could be a key for improving the establishment of inoculants in bioremediation processes.

microbiology↗

bioRxiv2025

Soil protist diversity enhances prokaryotic diversity, and regulates dominant prokaryotes and the abundance of key nitrogen cycling genes

Soil protists play crucial roles in soil microbial food-webs by preying on bacteria and other microorganisms. However, the effect of protist diversity on soil prokaryotic communities remains poorly understood. This study aimed to elucidate how different protist diversity treatments affect the composition and functionality of soil prokaryotic communities. We established soil microcosms with increasingly complex protist communities, including a control without protists, a medium diversity treatment with three small bacterivorous protists, and a high diversity treatment with seven protists of diverse trophic styles and sizes. Over 21 days, we monitored changes in the prokaryotic community using 16S rRNA gene sequencing and assessed the effects on nitrifiers and denitrifiers by qPCR of nitrogen-cycling genes. Protist diversity explained 23 % of the observed prokaryotic community differentiation over time, with the high-diversity treatment causing the greatest divergence from the control. The most abundant prokaryotes were preferentially predated in all protist treatments. Unexpectedly, the absolute abundance of the nirK gene, which is widely distributed among bacterial taxa and thus associated with high functional redundancy, decreased. The differential response of genes with lower distribution and redundancy, such as the bacterial and archaeal amoA and the Nitrospira-associated nxrB genes, to protist diversity indicated selective predation on archaea. High protist diversity systematically enhanced these effects compared to the medium diversity treatment. Overall, protist diversity was positively associated with prokaryotic diversity, which is crucial for maintaining ecosystem stability. These findings highlight the critical role of protist diversity and likely complementary predation in shaping soil prokaryotic communities and their functioning.

ecology↗

bioRxiv2024

Analysis of predation-driven inoculum loss and carbon flow in bioaugmented soils through DNA-SIP

Bioaugmentation is considered as a sustainable and cost-effective methodology to recover contaminated environments, but its outcome is highly variable. Predation is a key top-down control mechanism affecting inoculum establishment, however its effects on this process have received little attention. This study focused on the impact of trophic interactions on bioaugmentation success in two soils with different pollution exposure histories We inoculated a 13C-labelled pollutant-degrading consortium in these soils and tracked the fate of the labelled biomass through stable isotope probing (SIP) of DNA. We identified active bacterial and eukaryotic inoculum-biomass consumers through amplicon sequencing of 16S rRNA and 18S rRNA genes coupled to modified enrichment factor calculation. Inoculation effectively increased PAH removal in short-term polluted soils but not in long-term polluted soils. A decrease in the relative abundance of the inoculated genera was observed already on day 15 in the long-term polluted soil, while growth of these genera was observed in the short-term polluted soil, indicating establishment of the inoculum. In both soils, eukaryotic genera dominated as early incorporators of 13C-labelled biomass, while bacteria incorporated the labelled biomass at the end of the incubation period, probably through cross-feeding. We also found different successional patterns between the two soils. In the short-term polluted soil, Cercozoa and Fungi genera predominated as early incorporators, whereas Ciliophora, Ochrophyta and Amoebozoa were the predominant genera in the long-term polluted soil. Our results showed differences in the inoculum establishment and predator community behaviours, affecting bioaugmentation efficiency. This highlights the need to further study predation effects on inoculum survival to increase the applicability of inoculation-based technologies.

microbiology↗

bioRxiv2021

Phage co-transport with hyphal-riding bacteria fuels bacterial invasion in water-unsaturated microbial ecosystems

Non-motile microbes enter new habitats often by co-transport with motile microorganisms. Here, we report on the ability of hyphal-riding bacteria to co-transport lytic phages and utilize them as weapons during colonization of new water-unsaturated habitats. This is comparable to the concept of biological invasions in macroecology. In analogy to invasion frameworks in plant and animal ecology, we tailored spatially organized, water-unsaturated model microcosms using hyphae of Pythium ultimum as invasion paths and flagellated soil-bacterium Pseudomonas putida KT2440 as carrier for co-transport of Escherichia virus T4. P. putida KT2440 efficiently dispersed along P. ultimum to new habitats and dispatched T4 phages across air gaps transporting {approx} 0.6 phages bacteria-1. No T4 displacement along hyphae was observed in the absence of carrier bacteria. If E. coli occupied the new habitat, T4 co-transport fueled the fitness of invading P. putida KT2440, while the absence of phage co-transport led to poor colonization followed by extinction. Our data emphasize the importance of hyphal transport of bacteria and associated phages in regulating fitness and composition of microbial populations in water-unsaturated systems. As such co-transport mirrors macroecological invasion processes, we recommend hyphosphere systems with motile bacteria and co-transported phages as models for testing hypotheses in invasion ecology.

microbiology↗

bioRxiv2021

OrtSuite - from genomes to prediction of microbial interactions within targeted ecosystem processes

The high complexity found in microbial communities makes the identification of microbial interactions challenging. To address this challenge, we present OrtSuite, a flexible workflow to predict putative microbial interactions based on genomic content of microbial communities and targeted to specific ecosystem processes. The pipeline is composed of three user-friendly bash commands. OrtSuite combines ortholog clustering with genome annotation strategies limited to user-defined sets of functions allowing for hypothesis-driven data analysis such as assessing microbial interactions in specific ecosystems. OrtSuite matched, on average, 96 % of experimentally verified KEGG orthologs involved in benzoate degradation in a known group of benzoate degraders. Identification of putative synergistic species interactions was evaluated using the sequenced genomes of an independent study which had previously proposed potential species interactions in benzoate degradation. OrtSuite is an easy to use workflow that allows for rapid functional annotation based on a user curated database and can easily be extended to ecosystem processes where connections between genes and reactions are known. OrtSuite is an open-source software available at https://github.com/mdsufz/OrtSuite.

bioinformatics↗

bioRxiv2020

Nutritional inter-dependencies and a carbazole-dioxygenase are key elements of a bacterial consortium relying on a Sphingomonas for the degradation of the fungicide thiabendazol

BackgroundThiabendazole (TBZ), is a benzimidazole fungicide and anthelminthic whose high persistence and toxicity pose a serious environmental threat. In our quest for environmental mitigation we previously isolated the first TBZ-degrading bacterial consortium and provided preliminary evidence for its composition and the degrading role of a Sphingomonas. Here, we employed a multi-omic approach combined with DNA-stable isotope probing (SIP) to determine the genetic make-up of the key consortium members, to disentangle nutritional and metabolic interdependencies, to identify the transformation pathway of TBZ and to understand the genetic network driving its transformation. ResultsTime-series SIP in combination with amplicon sequencing analysis verified the key role of Sphingomonas in TBZ degradation by assimilating over 80% of the 13C-labelled phenyl moiety of TBZ. Non-target mass spectroscopy (MS) analysis showed the accumulation of thiazole-4-carboxamidine as a single dead-end transformation product and no phenyl-containing derivative, in line with the phenyl moiety assimilation in the SIP analysis. Time series metagenomic analysis of the consortium supplemented with TBZ or succinate led to the assembly of 18 metagenome-assembled genomes (MAGs) with >80% completeness, six (Sphingomonas 3X21F,{gamma} -Proteobacterium 34A, Bradyrhizobiaceae 9B and Hydrogenophaga 19A, 13A, and 23F) being dominant. Meta-transcriptomic and -proteomic analysis suggested that Sphingomonas mobilize a carbazole dioxygenase (car) operon during the initial cleavage of TBZ to thiazole-4-carboxamidine and catechol, the latter is further transformed by enzymes encoded in a catechol ortho-cleavage (cat) operon; both operons being up-regulated during TBZ degradation. Computational docking analysis of the terminal oxygenase component of car, CarAa, showed high affinity to TBZ, comparable to carbazole, reinforcing its high potency for TBZ transformation. These results suggest no interactions between consortium members in TBZ transformation, performed solely by Sphingomonas. In contrast, gene expression network analysis revealed strong interactions between Sphingomonas MAG 3X12F and Hydrogenophaga MAG 23F, with Hydrogenophaga activating its cobalamin biosynthetic pathway and Sphingomonas its cobalamin salvage pathway along TBZ degradation. ConclusionsOur findings suggest interactions between consortium members which align with the "black queen hypothesis": Sphingomonas detoxifies TBZ, releasing consortium members by a toxicant; in return for this, Hydrogenophaga 23F provides cobalamin to the auxotrophic Sphingomonas.

systems biology↗

bioRxiv2019

Resilience trinity: safeguarding ecosystem services across three different time horizons and decision contexts

Ensuring ecosystem resilience is an intuitive approach to safeguard future provisioning of ecosystem services (ES). However, resilience is an ambiguous concept and difficult to operationalize. Focusing on resilience mechanisms, such as diversity, network architectures or adaptive capacity, has recently been suggested as means to operationalize resilience. Still, the focus on mechanisms is not specific enough because the usefulness of a mechanism is context-dependent. We suggest a conceptual framework, resilience trinity, to facilitate management of resilience mechanisms in three distinctive decision contexts and time-horizons. i) reactive, when there is an imminent threat to ES resilience and a high pressure to act, ii) adjustive, when the threat is known in general but there is still time to adapt management, and iii) provident when time horizons are very long and the nature of the threats is uncertain, leading to a low willingness to act. This emphasizes that resilience has different interpretations and implications at different time horizons which however need to be reconciled. The inclusion of time into resilience thinking ensures that longer-term management actions are not missed while urgent threats to ES are given priority.

ecology↗