DC Electric Fields Promote Biodegradation of Waterborne Naphthalene in Biofilter Systems.

Biofiltration is a simple and low-cost method for the cleanup of contaminated water. However, the reduced availability of dissolved chemicals to surface-attached degrader bacteria may limit its efficient use at certain hydraulic loadings. When a direct current (DC) electric field is applied to an immersed packed bed, it invokes electrokinetic processes, such as electroosmotic water flow (EOF). EOF is a surface-charge-induced plug-flow-shaped movement of pore fluids. It acts at a nanometer distance above surfaces and allows the change of microscale pressure-driven flow profiles and, hence, the availability of dissolved contaminants to microbial degraders. In laboratory percolation columns, we assessed the effects of a weak DC electric field (E = 0.5 V·cm-1) on the biodegradation of waterborne naphthalene (NAH) by surface-attached Pseudomonas fluorescens LP6a. To vary NAH bioavailability, we used different NAH concentrations (C0 = 2.7, 5.1, or 7.8 × 10-5 mol·L-1) and Darcy velocities typical for biofiltration (U¯ = 0.2-1.2 × 10-4 m·s-1). In DC-free controls, we observed higher specific degradation rates (qc) at higher NAH concentrations. The qc depended on U¯, suggesting bioavailability restrictions depending on the hydraulic residence times. DC fields consistently increased qc and resulted in linearly increasing benefits up to 55% with rising hydraulic loadings relative to controls. We explain these biodegradation benefits by EOF-altered microscale flow profiles allowing for better NAH provision to bacteria attached to the collectors even though the EOF was calculated to be 100-800 times smaller than bulk water flow. Our data suggest that electrokinetic approaches may give rise to future technical applications that allow regulating biodegradation, for example, in response to fluctuating hydraulic loadings.

Mechanisms of heating-electrokinetic co-driven perfluorooctanoic acid (PFOA) adsorption on zeolite.

Slow release of emerging contaminants limits their accessibility from soil to pore water, constraining the treatment efficiency of physio-chemical treatment sites. DC fields mobilize organic contaminants and influence their interactions with geo-matrices such as zeolites. Poor knowledge, however, exists on the joint application of heating and electrokinetic approaches on perfluorooctanoic acid (PFOA) transport in porous media. Here, we investigated electrokinetic PFOA transport in zeolite-filled percolation columns at varying temperatures. Variations of pseudo-second-order kinetic constants (kPSO) were correlated to the liquid viscosity variations (η) and elctroosmotic flow velocities (vEOF). Applying DC fields and elevated temperature significantly (>37%) decreased PFOA sorption to zeolite. A good correlation between η, vEOF, and kPSO was found and used to develop an approach interlinking the three parameters to predict the joint effects of DC fields and temperature on PFOA sorption kinetics. These findings may give rise to future applications for better tailoring PFOA transport in environmental biotechnology.

Effects of Temperature and DC Electric Fields on Perfluorooctanoic Acid Sorption Kinetics to Activated Carbon.

Sorption to activated carbon is a common approach to reducing environmental risks of waterborne perfluorooctanoic acid (PFOA), while effective and flexible approaches to PFOA sorption are needed. Variations in temperature or the use of electrokinetic phenomena (electroosmosis and electromigration) in the presence of external DC electric fields have been shown to alter the contaminant sorption of contaminants. Their role in PFOA sorption, however, remains unclear. Here, we investigated the joint effects of DC electric fields and the temperature on the sorption of PFOA on activated carbon. Temperature-dependent batch and column sorption experiments were performed in the presence and absence of DC fields, and the results were evaluated by using different kinetic sorption models. We found an emerging interplay of DC and temperature on PFOA sorption, which was linked via the liquid viscosity (η) of the electrolyte. For instance, the combined presence of a DC field and low temperature increased the PFOA loading up to 38% in 48 h relative to DC-free controls. We further developed a model that allowed us to predict temperature- and DC field strength-dependent electrokinetic benefits on the drivers of PFOA sorption kinetics (i.e., intraparticle diffusivity and the film mass transfer coefficient). Our insights may give rise to future DC- and temperature-driven applications for PFOA sorption, for instance, in response to fluctuating PFOA concentrations in contaminated water streams.

Host and nonhost bacteria support bacteriophage dissemination along mycelia and abiotic dispersal networks.

Bacteriophages play a crucial role in shaping bacterial communities, yet the mechanisms by which nonmotile bacteriophages interact with their hosts remain poorly understood. This knowledge gap is especially pronounced in structured environments like soil, where spatial constraints and air-filled zones hinder aqueous diffusion. In soil, hyphae of filamentous microorganisms form a network of 'fungal highways' (FHs) that facilitate the dispersal of other microorganisms. We propose that FHs also promote bacteriophage dissemination. Viral particles can diffuse in liquid films surrounding hyphae or be transported by infectable (host) or uninfectable (nonhost) bacterial carriers coexisting on FH networks. To test this, two bacteriophages that infect Pseudomonas putida DSM291 (host) but not KT2440 (nonhost) were used. In the absence of carriers, bacteriophages showed limited diffusion on 3D-printed abiotic networks, but diffusion was significantly improved in Pythium ultimum-formed FHs when the number of connecting hyphae exceeded 20. Transport by both host and nonhost carriers enhanced bacteriophage dissemination. Host carriers were five times more effective in transporting bacteriophages, particularly in FHs with over 30 connecting hyphae. This study enhances our understanding of bacteriophage dissemination in nonsaturated environments like soils, highlighting the importance of biotic networks and bacterial hosts in facilitating this process.

Variovorax sp. strain P1R9 applied individually or as part of bacterial consortia enhances wheat germination under salt stress conditions.

Endophytes isolated from extremophile plants are interesting microbes for improving the stress tolerance of agricultural plants. Here, we isolated and characterized endophytic bacteria showing plant growth-promoting (PGP) traits from plants in two extreme Chilean biomes (Atacama Desert and Chilean Patagonia). Forty-two isolates were characterized as both halotolerant auxin producers (2-51 mg L-1) and 1-aminocyclopropane-1-carboxylate (ACC)-degrading bacteria (15-28 µmol αKB mg protein-1 h-1). The most efficient isolates were tested as single strains, in dual and triple consortia, or in combination with previously reported PGP rhizobacteria (Klebsiella sp. 27IJA and 8LJA) for their impact on the germination of salt-exposed (0.15 M and 0.25 M NaCl) wheat seeds. Interestingly, strain P1R9, identified as Variovorax sp., enhanced wheat germination under salt stress conditions when applied individually or as part of bacterial consortia. Under salt stress, plants inoculated with dual consortia containing the strain Variovorax sp. P1R9 showed higher biomass (41%) and reduced lipid peroxidation (33-56%) than uninoculated plants. Although the underlying mechanisms remain elusive, our data suggest that the application of Variovorax sp. P1R9, alone or as a member of PGP consortia, may improve the salt stress tolerance of wheat plants.

Fungal drops: a novel approach for macro- and microscopic analyses of fungal mycelial growth.

This study presents an inexpensive approach for the macro- and microscopic observation of fungal mycelial growth. The 'fungal drops' method allows to investigate the development of a mycelial network in filamentous microorganisms at the colony and hyphal scales. A heterogeneous environment is created by depositing 15-20 µl drops on a hydrophobic surface at a fixed distance. This system is akin to a two-dimensional (2D) soil-like structure in which aqueous-pockets are intermixed with air-filled pores. The fungus (spores or mycelia) is inoculated into one of the drops, from which hyphal growth and exploration take place. Hyphal structures are assessed at different scales using stereoscopic and microscopic imaging. The former allows to evaluate the local response of regions within the colony (modular behaviour), while the latter can be used for fractal dimension analyses to describe the hyphal network architecture. The method was tested with several species to underpin the transferability to multiple species. In addition, two sets of experiments were carried out to demonstrate its use in fungal biology. First, mycelial reorganization of Fusarium oxysporum was assessed as a response to patches containing different nutrient concentrations. Second, the effect of interactions with the soil bacterium Pseudomonas putida on habitat colonization by the same fungus was assessed. This method appeared as fast and accessible, allowed for a high level of replication, and complements more complex experimental platforms. Coupled with image analysis, the fungal drops method provides new insights into the study of fungal modularity both macroscopically and at a single-hypha level.

Modernizing persistence-bioaccumulation-toxicity (PBT) assessment with high throughput animal-free methods.

The assessment of persistence (P), bioaccumulation (B), and toxicity (T) of a chemical is a crucial first step at ensuring chemical safety and is a cornerstone of the European Union's chemicals regulation REACH (Registration, Evaluation, Authorization, and Restriction of Chemicals). Existing methods for PBT assessment are overly complex and cumbersome, have produced incorrect conclusions, and rely heavily on animal-intensive testing. We explore how new-approach methodologies (NAMs) can overcome the limitations of current PBT assessment. We propose two innovative hazard indicators, termed cumulative toxicity equivalents (CTE) and persistent toxicity equivalents (PTE). Together they are intended to replace existing PBT indicators and can also accommodate the emerging concept of PMT (where M stands for mobility). The proposed "toxicity equivalents" can be measured with high throughput in vitro bioassays. CTE refers to the toxic effects measured directly in any given sample, including single chemicals, substitution products, or mixtures. PTE is the equivalent measure of cumulative toxicity equivalents measured after simulated environmental degradation of the sample. With an appropriate panel of animal-free or alternative in vitro bioassays, CTE and PTE comprise key environmental and human health hazard indicators. CTE and PTE do not require analytical identification of transformation products and mixture components but instead prompt two key questions: is the chemical or mixture toxic, and is this toxicity persistent or can it be attenuated by environmental degradation? Taken together, the proposed hazard indicators CTE and PTE have the potential to integrate P, B/M and T assessment into one high-throughput experimental workflow that sidesteps the need for analytical measurements and will support the Chemicals Strategy for Sustainability of the European Union.

Marine Fungi Select and Transport Aerobic and Anaerobic Bacterial Populations from Polycyclic Aromatic Hydrocarbon-Contaminated Sediments.

The organization of microbial communities in marine sediment relies on complex biotic and abiotic interactions. Among them, the interaction between fungi and bacteria plays a crucial role building specific microbial assemblages, resulting in metabolic networks adapted to environmental conditions. The fungal-bacterial interaction (FBI) includes bacterial translocation via fungal mycelia, allowing bacterial dispersion, and ecological niche colonization. In order to demonstrate that the translocation of bacteria through fungal mycelia involves bacterial selection, the mycelia of two fungi isolated from marine coastal sediment, Alternaria destruens F10.81 and Fusarium pseudonygamai F5.76, showing different strategies for uptake of polycyclic aromatic hydrocarbon (PAH), homogenous internalization and vacuole forming respectively, were used to translocate bacteria through hydrophobic hydrocarbon contaminated sediments. A. destruens F10.81 selected four specific bacteria, while bacterial selection by F. pseudonygamai F5.76 was not evident. Among the bacteria selected by A. destruens F10.81, Spirochaeta litoralis, known as strictly anaerobic bacterium, was identified, indicating that A. destruens F10.81 selects and transports both aerobic and anaerobic bacteria. Such a result is consistent with the observed formation of anoxic micro-niches in areas surrounding and affected by fungal hyphae. Our findings provide new insights on the selection and dispersion of bacterial communities by fungi, which are crucial for the organization of microbial communities and their functioning in coastal PAH-contaminated sediments. IMPORTANCE The study provides advances for understanding fungal-bacterial relationships, particularly on the selection and dispersion of bacterial communities by fungi, which are crucial for the organization of microbial communities and their functioning in coastal PAH-contaminated sediments. The transportation of bacteria via fungal hyphae (fungal highway) results in bacterial selection; in particular, fungal hyphae offer adequate conditions for the transport of both aerobic and anaerobic bacteria through hydrophobic patches for the colonization of novel niches.

Mycelial nutrient transfer promotes bacterial co-metabolic organochlorine pesticide degradation in nutrient-deprived environments.

Biotransformation of soil organochlorine pesticides (OCP) is often impeded by a lack of nutrients relevant for bacterial growth and/or co-metabolic OCP biotransformation. By providing space-filling mycelia, fungi promote contaminant biodegradation by facilitating bacterial dispersal and the mobilization and release of nutrients in the mycosphere. We here tested whether mycelial nutrient transfer from nutrient-rich to nutrient-deprived areas facilitates bacterial OCP degradation in a nutrient-deficient habitat. The legacy pesticide hexachlorocyclohexane (HCH), a non-HCH-degrading fungus (Fusarium equiseti K3), and a co-metabolically HCH-degrading bacterium (Sphingobium sp. S8) isolated from the same HCH-contaminated soil were used in spatially structured model ecosystems. Using 13C-labeled fungal biomass and protein-based stable isotope probing (protein-SIP), we traced the incorporation of 13C fungal metabolites into bacterial proteins while simultaneously determining the biotransformation of the HCH isomers. The relative isotope abundance (RIA, 7.1-14.2%), labeling ratio (LR, 0.13-0.35), and the shape of isotopic mass distribution profiles of bacterial peptides indicated the transfer of 13C-labeled fungal metabolites into bacterial proteins. Distinct 13C incorporation into the haloalkane dehalogenase (linB) and 2,5-dichloro-2,5-cyclohexadiene-1,4-diol dehydrogenase (LinC), as key enzymes in metabolic HCH degradation, underpin the role of mycelial nutrient transport and fungal-bacterial interactions for co-metabolic bacterial HCH degradation in heterogeneous habitats. Nutrient uptake from mycelia increased HCH removal by twofold as compared to bacterial monocultures. Fungal-bacterial interactions hence may play an important role in the co-metabolic biotransformation of OCP or recalcitrant micropollutants (MPs).

Water Stress-Driven Changes in Bacterial Cell Surface Properties.

Increased drought intensity and frequency exposes soil bacteria to prolonged water stress. While numerous studies reported on behavioral and physiological mechanisms of bacterial adaptation to water stress, changes in bacterial cell surface properties during adaptation are not well researched. We studied adaptive changes in cell surface hydrophobicity (CSH) after exposure to osmotic (NaCl) and matric stress (polyethylene glycol 8000 [PEG 8000]) for six typical soil bacteria (Bacillus subtilis, Arthrobacter chlorophenolicus, Pseudomonas fluorescens, Novosphingobium aromaticivorans, Rhodococcus erythropolis, and Mycobacterium pallens) covering a wide range of cell surface properties. Additional physicochemical parameters (surface chemical composition, surface charge, cell size and stiffness) of B. subtilis and P. fluorescens were analyzed to understand their possible contribution to CSH development. Changes in CSH caused by osmotic and matric stress depend on strain and stress type. CSH of B. subtilis and P. fluorescens increased with stress intensity, R. erythropolis and M. pallens exhibited a generally high but constant contact angle, while the response of A. chlorophenolicus and N. aromaticivorans depended on growth conditions and stress type. Osmotically driven changes in CSH of B. subtilis and P. fluorescens are accompanied by increasing surface N/C ratio, suggesting an increase in protein concentration within the cell wall. Cell envelope proteins thus presumably control bacterial CSH in two ways: (i) by increases in the relative density of surface proteins due to efflux of cytoplasmic water and subsequent cell shrinkage, and (ii) by destabilization of cell wall proteins, resulting in conformational changes which render the surface more hydrophobic. IMPORTANCE Changes in precipitation frequency, intensity, and temporal distribution are projected to result in increased frequency and intensity of droughts and heavy rainfall events. Prolonged droughts can promote the development of soil water repellency (SWR); this impacts the infiltration and distribution of water in the soil profile, exposing soil microorganisms to water stress. Exposure to water stress has recently been reported to result in increased cell surface hydrophobicity. However, the mechanism of this development is poorly understood. This study investigates the changes in the physicochemical properties of bacterial cell surfaces under water stress as a possible mechanism of increased surface hydrophobicity. Our results improve understanding of the microbial response to water stress in terms of surface properties, the variations in stress response depending on cell wall composition, and its contribution to the development of SWR.

Eggerthella lenta DSM 2243 Alleviates Bile Acid Stress Response in Clostridium ramosum and Anaerostipes caccae by Transformation of Bile Acids.

Bile acids are crucial for the uptake of dietary lipids and can shape the gut-microbiome composition. This latter function is associated with the toxicity of bile acids and can be modulated by bile acid modifying bacteria such as Eggerthella lenta, but the molecular details of the interaction of bacteria depending on bile acid modifications are not well understood. In order to unravel the molecular response to bile acids and their metabolites, we cultivated eight strains from a human intestinal microbiome model alone and in co-culture with Eggerthella lenta in the presence of cholic acid (CA) and deoxycholic acid (DCA). We observed growth inhibition of particularly gram-positive strains such as Clostridium ramosum and the gram-variable Anaerostipes cacae by CA and DCA stress. C. ramosum was alleviated through co-culturing with Eggerthella lenta. We approached effects on the membrane by zeta potential and genotoxic and metabolic effects by (meta)proteomic and metabolomic analyses. Co-culturing with Eggerthella lenta decreased both CA and DCA by the formation of oxidized and epimerized bile acids. Eggerthella lenta also produces microbial bile salt conjugates in a co-cultured species-specific manner. This study highlights how the interaction with other bacteria can influence the functionality of bacteria.

Design and construction of 3D printed devices to investigate active and passive bacterial dispersal on hydrated surfaces.

BACKGROUND: To disperse in water-unsaturated environments, such as the soil, bacteria rely on the availability and structure of water films forming on biotic and abiotic surfaces, and, especially, along fungal mycelia. Dispersal along such "fungal highways" may be driven both by mycelial physical properties and by interactions between bacteria and fungi. However, we still do not have a way to disentangle the biotic and abiotic elements. RESULTS: We designed and 3D printed two devices establishing stable liquid films that support bacteria dispersal in the absence of biotic interactions. The thickness of the liquid film determined the presence of hydraulic flow capable of transporting non-motile cells. In the absence of flow, only motile cells can disperse in the presence of an energy source. Non-motile cells could not disperse autonomously without flow but dispersed as "hitchhikers" when co-inoculated with motile cells. CONCLUSIONS: The 3D printed devices can be used as an abiotic control to study bacterial dispersal on hydrated surfaces, such as plant roots and fungal hyphae networks in the soil. By teasing apart the abiotic and biotic dimensions, these 3D printed devices will stimulate further research on microbial dispersal in soil and other water-unsaturated environments.

Phase-specific stable isotope fractionation effects during combined gas-liquid phase exchange and biodegradation.

Stable isotope fractionation of toluene under dynamic phase exchange was studied aiming at ascertaining the effects of gas-liquid partitioning and biodegradation of toluene stable isotope composition in liquid-air phase exchange reactors (Laper). The liquid phase consisted of a mixture of aqueous minimal media, a known amount of a mixture of deuterated (toluene-d) and non-deuterated toluene (toluene-h), and bacteria of toluene degrading strain Pseudomonas putida KT2442. During biodegradation experiments, the liquid and air-phase concentrations of both toluene isotopologues were monitored to determine the observable stable isotope fractionation in each phase. The results show a strong fractionation in both phases with apparent enrichment factors beyond -800‰. An offset was observed between enrichment factors in the liquid and the gas phase with gas-phase values showing a stronger fractionation in the gas than in the liquid phase. Numerical simulation and parameter fitting routine was used to challenge hypotheses to explain the unexpected experimental data. The numerical results showed that either a very strong, yet unlikely, fractionation of the phase exchange process or a - so far unreported - direct consumption of gas phase compounds by aqueous phase microorganisms could explain the observed fractionation effects. The observed effect can be of relevance for the analysis of volatile contaminant biodegradation using stable isotope analysis in unsaturated subsurface compartments or other environmental compartment containing a gas and a liquid phase.

pH Distribution along Growing Fungal Hyphae at Microscale.

Creating unique microenvironments, hyphal surfaces and their surroundings allow for spatially distinct microbial interactions and functions at the microscale. Using a microfluidic system and pH-sensitive whole-cell bioreporters (Synechocystis sp. PCC6803) attached to hyphae, we spatially resolved the pH along surfaces of growing hyphae of the basidiomycete Coprinopsis cinerea. Time-lapse microscopy analysis of ratiometric fluorescence signals of >2400 individual bioreporters revealed an overall pH drop from 6.3 ± 0.4 (n = 2441) to 5.0 ± 0.3 (n = 2497) within 7 h after pH bioreporter loading to hyphal surfaces. The pH along hyphal surfaces varied significantly (p < 0.05), with pH at hyphal tips being on average ~0.8 pH units lower than at more mature hyphal parts near the entrance of the microfluidic observation chamber. Our data represent the first dynamic in vitro analysis of surface pH along growing hyphae at the micrometre scale. Such knowledge may improve our understanding of spatial, pH-dependent hyphal processes, such as the degradation of organic matter or mineral weathering.

Impact of Fungal Hyphae on Growth and Dispersal of Obligate Anaerobic Bacteria in Aerated Habitats.

Anoxic microsites arising in fungal biofilms may foster the presence of obligate anaerobes. Here, we analyzed whether and to which degree hyphae of Coprinopsis cinerea thriving in oxic habitats enable the germination, growth, and dispersal of the obligate anaerobic soil bacterium Clostridium acetobutylicum. Time-resolved optical oxygen mapping, microscopy, and metabolite analysis revealed the formation and persistence of anoxic circum hyphal niches, allowing for spore germination, growth, and fermentative activity of the obligate anaerobe in an otherwise inhabitable environment. Hypoxic liquid films containing 80% ± 10% of atmospheric oxygen saturation around single air-exposed hyphae thereby allowed for efficient clostridial dispersal amid spatially separated (>0.5 cm) anoxic sites. Hyphae hence may serve as good networks for the activity and spatial organization of obligate anaerobic bacteria in oxygenated heterogeneous environments such as soil. IMPORTANCE Although a few studies have reported on the presence of anoxic microniches in fungal biofilms, knowledge of the effects of fungal oxygen consumption on bacterial-fungal interactions is limited. Here, we demonstrate the existence and persistence of oxygen-free zones in air-exposed mycelia enabling spore germination, growth, fermentative activity, and dispersal of the obligate anaerobe. Our study points out a previously overlooked role of aerobic fungi in creating and bridging anoxic microniches in ambient oxic habitats. Air-exposed hyphae hence may act as a scaffold for activity and dispersal of strictly anaerobic microbes. Given the short-term tolerance of strict anaerobes to oxygen and reduced oxygen content in the mycosphere, hyphae can promote spatial organization of both obligate anaerobic and aerobic bacteria. Such finding may be important for a better understanding of previously observed co-occurrences of aerobes and anaerobes in well-aerated habitats such as upland soils.

Efficient removal of trifluoroacetic acid from water using surface-modified activated carbon and electro-assisted desorption.

Trifluoroacetic acid (TFA) is a very persistent, very mobile substance (vPvM) with potential toxicity, and causes increasing environmental concerns worldwide. Conventional wastewater treatment strategies are inefficient for selective TFA removal in the presence of inorganic anions. Here we show that surface defunctionalized activated carbon felt (DeACF) carrying anion exchange sites exhibits an outstanding adsorption efficiency towards TFA thanks to introduced electrostatic attraction and enhanced interactions between hydrophobic carbon surface and CF3 moieties (qmax = 30 mg/g, Kd = (840 ± 80) L/kg at cTFA = 3.4 mg/L in tap water). Flow-cell experiments demonstrated a strongly favored TFA uptake by DeACF from tap water over Cl- and SO42- but a remarkable co-adsorption of the inorganic water contaminant NO3-. Electro-assisted TFA desorption using 10 mM Na2SO4 as electrolyte and oxidized ACF as anode showed high recoveries of ≥ 87% at low cell voltages (< 1.1 V). Despite an initial decrease in TFA adsorption capacity (by 33%) caused by partial surface oxidation of DeACF after the 1st ad-/desorption cycle, the system stability was fully maintained over the next 4 cycles. Such electro-assisted 'trap&release' approach for TFA removal can be exploited for on-site regenerable adsorption units and as a pre-concentration step combined with degradation technologies.

Draft Genome Sequences of Two Sphingobium Species Associated with Hexachlorocyclohexane (HCH) Degradation Isolated from an HCH-Contaminated Soil.

The draft genome sequences of two Sphingobium strains that are hexachlorocyclohexane (HCH) degraders are presented. The strains were isolated from HCH-contaminated soil in Kitengela, Kenya. Both genomes possess the lin genes responsible for HCH degradation and gene clusters for degradation of other xenobiotic compounds.

Mycelia-Assisted Isolation of Non-Host Bacteria Able to Co-Transport Phages.

Recent studies have demonstrated that phages can be co-transported with motile non-host bacteria, thereby enabling their invasion of biofilms and control of biofilm composition. Here, we developed a novel approach to isolate non-host bacteria able to co-transport phages from soil. It is based on the capability of phage-carrying non-host bacteria to move along mycelia out of soil and form colonies in plaques of their co-transported phages. The approach was tested using two model phages of differing surface hydrophobicity, i.e., hydrophobic Escherichia virus T4 (T4) and hydrophilic Pseudoalteromonas phage HS2 (HS2). The phages were mixed into soil and allowed to be transported by soil bacteria along the mycelia of Pythium ultimum. Five phage-carrying bacterial species were isolated (Viridibacillus sp., Enterobacter sp., Serratia sp., Bacillus sp., Janthinobacterium sp.). These bacteria exhibited phage adsorption efficiencies of ≈90-95% for hydrophobic T4 and 30-95% for hydrophilic HS2. The phage adsorption efficiency of Viridibacillus sp. was ≈95% for both phages and twofold higher than T4-or HS2-adsorption to their respective hosts, qualifying Viridibacillus sp. as a potential super carrier for phages. Our approach offers an effective and target-specific way to identify and isolate phage-carrying bacteria in natural and man-made environments.