Monitoring of hepatitis E virus in wastewater can identify clinically relevant variants.

Hepatitis E virus (HEV) is prevalent worldwide and can cause persistent infection with severe morbidity. Antiviral treatment approaches can lead to the emergence of viral variants encoding escape mutations that may impede viral clearance. The frequency of these variants remains unknown in the human population as well as environment due to limited comprehensive data on HEV diversity. In this study, we investigated the HEV prevalence and diversity of circulating variants in environmental samples, that is, wastewater and rivers from North-Rhine Westphalia, Germany. HEV prevalence could be determined with 73% of samples tested positive for viral RNA via qRT-PCR. Using high-throughput sequencing, we were able to assess the overall genetic diversity in these samples and identified the presence of clinically relevant variants associated with drug resistance. In summary, monitoring variants from environmental samples could provide valuable insights into estimating HEV prevalence and identifying circulating variants that can impact treatment outcome.

Rapid Consumption of Dihydrogen Injected into a Shallow Aquifer by Ecophysiologically Different Microbes.

The envisaged future dihydrogen (H2) economy requires a H2 gas grid as well as large deep underground stores. However, the consequences of an unintended spread of H2 through leaky pipes, wells, or subterranean gas migrations on groundwater resources and their ecosystems are poorly understood. Therefore, we emulated a short-term leakage incident by injecting gaseous H2 into a shallow aquifer at the TestUM test site and monitored the subsequent biogeochemical processes in the groundwater system. At elevated H2 concentrations, an increase in acetate concentrations and a decrease in microbial α-diversity with a concomitant change in microbial ß-diversity were observed. Additionally, microbial H2 oxidation was indicated by temporally higher abundances of taxa known for aerobic or anaerobic H2 oxidation. After H2 concentrations diminished below the detection limit, α- and ß-diversity approached baseline values. In summary, the emulated H2 leakage resulted in a temporally limited change of the groundwater microbiome and associated geochemical conditions due to the intermediate growth of H2 consumers. The results confirm the general assumption that H2, being an excellent energy and electron source for many microorganisms, is quickly microbiologically consumed in the environment after a leakage.

Conditioning Film and Early Biofilm Succession on Plastic Surfaces.

In the context of environmental plastic pollution, it is still under debate if and how the "plastisphere", a plastic-specific microbial community, emerges. In this study, we tested the hypothesis that the first conditioning film of dissolved organic matter (DOM) sorbs selectively to polymer substrates and that microbial attachment is governed in a substrate-dependent manner. We investigated the adsorption of stream water-derived DOM to polyethylene terephthalate (PET), polystyrene (PS), and glass (as control) including UV-weathered surfaces by Fourier-transform ion cyclotron mass spectrometry. Generally, the saturated, high-molecular mass and thus more hydrophobic fraction of the original stream water DOM preferentially adsorbed to the substrates. The UV-weathered polymers adsorbed more polar, hydrophilic OM as compared to the dark controls. The amplicon sequencing data of the initial microbial colonization process revealed a tendency of substrate specificity for biofilm attachment after 24 h and a clear convergence of the communities after 72 h of incubation. Conclusively, the adsorbed OM layer developed depending on the materials' surface properties and increased the water contact angles, indicating higher surface hydrophobicity as compared to pristine surfaces. This study improves our understanding of molecular and biological interactions at the polymer/water interface that are relevant to understand the ecological impact of plastic pollution on a community level.

Secondary structure of DNA released from purified capsids of human parvovirus B19 under moderate denaturing conditions.

Parvovirus B19 (B19V) possesses a linear single-stranded DNA genome of either positive or negative polarity. Due to intramolecular sequence homologies, either strand may theoretically be folded in several alternative ways. Viral DNA, when extracted from virions by several procedures, presents as linear single-stranded and/or linear double-stranded molecules, except when one particular commercial kit is used. This protocol yields DNA with an aberrant electrophoretic mobility in addition to linear double-stranded molecules, but never any single-stranded molecules. This peculiar kind of DNA was found in all plasma or serum samples tested and so we decided to analyse its secondary structure. In line with our results for one- and two-dimensional electrophoresis, mobility shift assays, DNA preparation by an in-house extraction method with moderate denaturing conditions, density gradient ultracentrifugation, DNA digestion experiments and competition hybridization assays, we conclude that (i) the unique internal portions of this distinctive single-stranded molecules are folded into tight tangles and (ii) the two terminal redundant regions are associated with each other, yielding non-covalently closed pseudo-circular molecules stabilized by a short (18 nucleotides) intramolecular stem, whereas the extreme 3'- and 5'-ends are folded back on themselves, forming a structure resembling a twin hairpin. The question arises as to whether this fairly unstable structure represents the encapsidated genome structure. The answer to this question remains quite relevant in terms of comprehending the initiation and end of B19V genome replication.

Influence of IS256 on Genome Variability and Formation of Small-Colony Variants in Staphylococcus aureus.

Staphylococcus aureus has acquired resistance to nearly all antibiotics used in clinical practice. Whereas some resistance mechanisms are conferred by uptake of resistance genes, others evolve by mutation. In this study, IS256 has been shown to play a role, e.g., in S. aureus strains displaying intermediate resistance to vancomycin (VISA). To characterize the IS256 insertion sites in the genomes of two closely related sequence type 247 (ST247) VISA strains, all insertions were mapped in both VISA and a susceptible control strain. The results showed that the three ST247 strains contained the highest number so far of IS256 insertions for all sequenced S. aureus strains. Furthermore, in contrast to the case with the other IS elements in these genomes, the IS256 insertion sites were not identical in the closely related strains, indicating a high transposition frequency of IS256 When IS256 was introduced into a laboratory strain which was then cultured in the presence of antibiotics, it was possible to isolate small-colony variants (SCVs) that possessed IS256 insertions in guaA and hemY that displayed increased resistance to vancomycin and aminoglycosides, respectively. For these clones, a very rapid reversion to the wild type that resembled the fast reversion of clinical SCVs was observed. The reversion was caused by excision of IS256 in a small number of fast-growing clones that quickly outcompeted the SCVs in broth cultures. In conclusion, the presence of IS256 confers a strong genomic plasticity that is useful for adaptation to antibiotic stress.

A novel rhabdovirus isolated from the straw-colored fruit bat Eidolon helvum, with signs of antibodies in swine and humans.

UNLABELLED: Bats have been implicated as reservoirs of emerging viruses. Bat species forming large social groups and roosting in proximity to human communities are of particular interest. In this study, we sampled a colony of ca. 350,000 individuals of the straw-colored fruit bat Eidolon helvum in Kumasi, the second largest city of Ghana. A novel rhabdovirus (Kumasi rhabdovirus [KRV]) was isolated in E. helvum cell cultures and passaged to Vero cells as well as interferon-competent human and primate cells (A549 and MA104). Genome composition was typical for a rhabdovirus. KRV was detected in 5.1% of 487 animals, showing association with the spleen but not the brain. Antibody prevalence was 11.5% by immunofluorescence and 6.4% by plaque reduction virus neutralization test (PRNT). Detection throughout 3 sampling years was pronounced in both annual wet seasons, of which only one overlaps the postparturition season. Juvenile bats showed increased viral prevalence. No evidence of infection was obtained in 1,240 female mosquitos (6 different genera) trapped in proximity to the colony to investigate potential vector association. Antibodies were found in 28.9% (5.4% by PRNT) of 107 swine sera but not in similarly large collections of sheep, goat, or cattle sera. The antibody detection rate in human subjects with occupational exposure to the bat colony was 11% (5/45 persons), which was significantly higher than in unexposed adults (0.8% [1/118]; chi square, P < 0.001). KRV is a novel bat-associated rhabdovirus potentially transmitted to humans and swine. Disease associations should be investigated. IMPORTANCE: Bats are thought to carry a huge number of as-yet-undiscovered viruses that may pose epidemic threats to humans and livestock. Here we describe a novel dimarhabdovirus which we isolated from a large colony of the straw-colored fruit bat Eidolon helvum in Ghana. As these animals are exposed to humans and several livestock species, we looked for antibodies indicating infection in humans, cattle, swine, sheep, and goats. Signs of infection were found in swine and humans, with increased antibody findings in humans who are occupationally exposed to the bat colony. Our data suggest that it is worthwhile to look for diseases caused by the novel virus in humans and livestock.

Epithelial cell lines of the cotton rat (Sigmodon hispidus) are highly susceptible in vitro models to zoonotic Bunya-, Rhabdo-, and Flaviviruses.

BACKGROUND: Small mammals such as bats and rodents have been increasingly recognized as reservoirs of novel potentially zoonotic pathogens. However, few in vitro model systems to date allow assessment of zoonotic viruses in a relevant host context. The cotton rat (Sigmodon hispidus) is a New World rodent species that has a long-standing history as an experimental animal model due to its unique susceptibility to human viruses. Furthermore, wild cotton rats are associated with a large variety of known or potentially zoonotic pathogens. METHODS: A method for the isolation and culture of airway epithelial cell lines recently developed for bats was applied for the generation of rodent airway and renal epithelial cell lines from the cotton rat. Continuous cell lines were characterized for their epithelial properties as well as for their interferon competence. Susceptibility to members of zoonotic Bunya-, Rhabdo-, and Flaviviridae, in particular Rift Valley fever virus (RVFV), vesicular stomatitis virus (VSV), West Nile virus (WNV), and tick-borne encephalitis virus (TBEV) was tested. Furthermore, novel arthropod-derived viruses belonging to the families Bunya-, Rhabdo-, and Mesoniviridae were tested. RESULTS: We successfully established airway and kidney epithelial cell lines from the cotton rat, and characterized their epithelial properties. Cells were shown to be interferon-competent. Viral infection assays showed high-titre viral replication of RVFV, VSV, WNV, and TBEV, as well as production of infectious virus particles. No viral replication was observed for novel arthropod-derived members of the Bunya-, Rhabdo-, and Mesoniviridae families in these cell lines. CONCLUSION: In the current study, we showed that newly established cell lines from the cotton rat can serve as host-specific in vitro models for viral infection experiments. These cell lines may also serve as novel tools for virus isolation, as well as for the investigation of virus-host interactions in a relevant host species.

Marine Phages As Tracers: Effects of Size, Morphology, and Physico-Chemical Surface Properties on Transport in a Porous Medium.

Although several studies examined the transport of viruses in terrestrial systems only few studies exist on the use of marine phages (i.e., nonterrestrial viruses infecting marine host bacteria) as sensitively detectable microbial tracers for subsurface colloid transport and water flow. Here, we systematically quantified and compared for the first time the effects of size, morphology and physicochemical surface properties of six marine phages and two coliphages (MS2, T4) on transport in sand-filled percolated columns. Phage-sand interactions were described by colloidal filtration theory and the extended Derjaguin-Landau-Verwey-Overbeek approach (XDLVO), respectively. The phages belonged to different families and comprised four phages never used in transport studies (i.e., PSA-HM1, PSA-HP1, PSA-HS2, and H3/49). Phage transport was influenced by size, morphology and hydrophobicity in an approximate order of size > hydrophobicity ≥ morphology. Two phages PSA-HP1, PSA-HS2 (Podoviridae and Siphoviridae) exhibited similar mass recovery as commonly used coliphage MS2 and were 7-fold better transported than known marine phage vB_PSPS-H40/1. Differing properties of the marine phages may be used to trace transport of indigenous viruses, natural colloids or anthropogenic nanomaterials and, hence, contribute to better risk analysis. Our results underpin the potential role of marine phages as microbial tracer for transport of colloidal particles and water flow.

Novel Arenavirus Isolates from Namaqua Rock Mice, Namibia, Southern Africa.

Arenaviruses are feared as agents that cause viral hemorrhagic fevers. We report the identification, isolation, and genetic characterization of 2 novel arenaviruses from Namaqua rock mice in Namibia. These findings extend knowledge of the distribution and diversity of arenaviruses in Africa.

Characterization of a novel betacoronavirus related to middle East respiratory syndrome coronavirus in European hedgehogs.

Bats are known to host viruses closely related to important human coronaviruses (HCoVs), such as HCoV-229E, severe-acute respiratory syndrome coronavirus (SARS-CoV), and Middle East respiratory syndrome CoV (MERS-CoV). As RNA viruses may coevolve with their hosts, we sought to investigate the closest sister taxon to bats, the Eulipotyphla, and screened European hedgehogs (Erinaceus europaeus) from Germany for CoV by nested reverse transcriptase PCR. A novel betacoronavirus species in a phylogenetic sister relationship to MERS-CoV and clade c bat CoVs was detected and characterized on the whole-genome level. A total of 58.9% of hedgehog fecal specimens were positive for the novel CoV (EriCoV) at 7.9 log10 mean RNA copies per ml. EriCoV RNA concentrations were higher in the intestine than in other solid organs, blood, or urine. Detailed analyses of the full hedgehog intestine showed the highest EriCoV concentrations in lower gastrointestinal tract specimens, compatible with viral replication in the lower intestine and fecal-oral transmission. Thirteen of 27 (48.2%) hedgehog sera contained non-neutralizing antibodies against MERS-CoV. The animal origins of this betacoronavirus clade that includes MERS-CoV may thus include both bat and nonbat hosts.

Evidence for novel hepaciviruses in rodents.

Hepatitis C virus (HCV) is among the most relevant causes of liver cirrhosis and hepatocellular carcinoma. Research is complicated by a lack of accessible small animal models. The systematic investigation of viruses of small mammals could guide efforts to establish such models, while providing insight into viral evolutionary biology. We have assembled the so-far largest collection of small-mammal samples from around the world, qualified to be screened for bloodborne viruses, including sera and organs from 4,770 rodents (41 species); and sera from 2,939 bats (51 species). Three highly divergent rodent hepacivirus clades were detected in 27 (1.8%) of 1,465 European bank voles (Myodes glareolus) and 10 (1.9%) of 518 South African four-striped mice (Rhabdomys pumilio). Bats showed anti-HCV immunoblot reactivities but no virus detection, although the genetic relatedness suggested by the serologic results should have enabled RNA detection using the broadly reactive PCR assays developed for this study. 210 horses and 858 cats and dogs were tested, yielding further horse-associated hepaciviruses but none in dogs or cats. The rodent viruses were equidistant to HCV, exceeding by far the diversity of HCV and the canine/equine hepaciviruses taken together. Five full genomes were sequenced, representing all viral lineages. Salient genome features and distance criteria supported classification of all viruses as hepaciviruses. Quantitative RT-PCR, RNA in-situ hybridisation, and histopathology suggested hepatic tropism with liver inflammation resembling hepatitis C. Recombinant serology for two distinct hepacivirus lineages in 97 bank voles identified seroprevalence rates of 8.3 and 12.4%, respectively. Antibodies in bank vole sera neither cross-reacted with HCV, nor the heterologous bank vole hepacivirus. Co-occurrence of RNA and antibodies was found in 3 of 57 PCR-positive bank vole sera (5.3%). Our data enable new hypotheses regarding HCV evolution and encourage efforts to develop rodent surrogate models for HCV.

Discovery of a unique novel clade of mosquito-associated bunyaviruses.

Bunyaviruses are the largest known family of RNA viruses, infecting vertebrates, insects, and plants. Here we isolated three novel bunyaviruses from mosquitoes sampled in Côte d'Ivoire, Ghana, and Uganda. The viruses define a highly diversified monophyletic sister clade to all members of the genus Orthobunyavirus and are virtually equidistant to orthobunyaviruses and tospoviruses. Maximal amino acid identities between homologous putative proteins of the novel group and orthobunyaviruses ranged between 12 and 25%. The type isolates, tentatively named Herbert virus (HEBV), Taï virus (TAIV), and Kibale virus (KIBV), comprised genomes with L, M, and S segments of about 7.4 kb, 2.7 kb, and 1.1 kb, respectively. HEBV, TAIV, and KIBV encode the shortest bunyavirus M segments known and did not seem to encode NSs and NSm proteins but contained an elongated L segment with an ∼500-nucleotide (nt) insertion that shows no identity to other bunyaviruses. The viruses replicated to high titers in insect cells but did not replicate in vertebrate cells. The enveloped virions were 90 to 110 nm in diameter and budded at cellular membranes with morphological features typical of the Golgi complex. Viral RNA recovered from infected cells showed 5'-terminal nontemplated sequences of 9 to 22 nt, suggestive of cap snatching during mRNA synthesis, as described for other bunyaviruses. Northern blotting identified RNA species of full and reduced lengths, suggested upon analogy with other bunyaviruses to constitute antigenomic-sense cRNA and transcript mRNAs, respectively. Functional studies will be necessary to determine if this group of viruses constitutes a novel genus in the bunyavirus family.

MuDoGeR: Multi-Domain Genome recovery from metagenomes made easy.

Several computational frameworks and workflows that recover genomes from prokaryotes, eukaryotes and viruses from metagenomes exist. Yet, it is difficult for scientists with little bioinformatics experience to evaluate quality, annotate genes, dereplicate, assign taxonomy and calculate relative abundance and coverage of genomes belonging to different domains. MuDoGeR is a user-friendly tool tailored for those familiar with Unix command-line environment that makes it easy to recover genomes of prokaryotes, eukaryotes and viruses from metagenomes, either alone or in combination. We tested MuDoGeR using 24 individual-isolated genomes and 574 metagenomes, demonstrating the applicability for a few samples and high throughput. While MuDoGeR can recover eukaryotic viral sequences, its characterization is predominantly skewed towards bacterial and archaeal viruses, reflecting the field's current state. However, acting as a dynamic wrapper, the MuDoGeR is designed to constantly incorporate updates and integrate new tools, ensuring its ongoing relevance in the rapidly evolving field. MuDoGeR is open-source software available at https://github.com/mdsufz/MuDoGeR. Additionally, MuDoGeR is also available as a Singularity container.

Identification of Huge Phages from Wastewater Metagenomes.

Huge phages have genomes larger than 200 kilobases, which are particularly interesting for their genetic inventory and evolution. We screened 165 wastewater metagenomes for the presence of viral sequences. After identifying over 600 potential huge phage genomes, we reduced the dataset using manual curation by excluding viral contigs that did not contain viral protein-coding genes or consisted of concatemers of several small phage genomes. This dataset showed seven fully annotated huge phage genomes. The phages grouped into distinct phylogenetic clades, likely forming new genera and families. A phylogenomic analysis between our huge phages and phages with smaller genomes, i.e., less than 200 kb, supported the hypothesis that huge phages have undergone convergent evolution. The genomes contained typical phage protein-coding genes, sequential gene cassettes for metabolic pathways, and complete inventories of tRNA genes covering all standard and rare amino acids. Our study showed a pipeline for huge phage analyses that may lead to new enzymes for therapeutic or biotechnological applications.

A long-term passive sampling approach for wastewater-based monitoring of SARS-CoV-2 in Leipzig, Germany.

Wastewater-based monitoring of SARS-CoV-2 has become a promising and useful tool in tracking the potential spread or dynamics of the virus. Its recording can be used to predict how the potential number of infections in a population will develop. Recent studies have shown that the use of passive samplers is also suitable for the detection of SARS-CoV-2 genome copies (GC) in wastewater. They can be used at any site, provide timely data and may collect SARS-CoV-2 GC missed by traditional sampling methods. Therefore, the aim of this study was to evaluate the suitability of passive samplers for the detection of SARS-CoV-2 GC in wastewater in the long-term at two different scales. Polyethylene-based plastic passive samplers were deployed at the city-scale level of Leipzig at 13 different locations, with samples being taken from March 2021 to August 2022. At the smaller city district level, three types of passive samplers (cotton-cloth, unravelled polypropylene plastic rope and polyethylene-based plastic strips) were used and sampled on a weekly basis from March to August 2022. The results are discussed in relation to wastewater samples taken at the individual passive sampling point. Our results show that passive samplers can indicate at a city-scale level an accurate level of positive infections in the population (positive-rate: 86 %). On a small-scale level, the use of passive samplers was also feasible and effective to detect SARS-CoV-2 GC easily and cost-effectively, mirroring a similar trend to that at a city-scale level. Thus, this study demonstrated that passive samplers provide reproducible SARS-CoV-2 GC signals from wastewater and a time-integrated measurement of the sampled matrix with greater sensitivity compared to wastewater. We thus recommend the use of passive samplers as an alternative method for wastewater-based epidemiology. Passive samplers can in particular be considered for a better estimation of infections compared to incidence levels.

Changes of the bacterial composition in duodenal fluid from patients with liver cirrhosis and molecular bacterascites.

Small intestinal bacterial overgrowth and compositional changes of intestinal microbiota are pathomechanistic factors in liver cirrhosis leading to bacterial translocation and infectious complications. We analyzed the quantity and composition of duodenal bacterial DNA (bactDNA) in relation to bactDNA in blood and ascites of patients with liver cirrhosis. Duodenal fluid and corresponding blood and ascites samples from 103 patients with liver cirrhosis were collected. Non-liver disease patients (n = 22) served as controls. BactDNA was quantified by 16S-rRNA gene-based PCR. T-RFLP and 16S-rRNA amplicon sequencing were used to analyze bacterial composition. Duodenal bacterial diversity in cirrhosis was distinct to controls showing significantly higher abundances of Streptococcus, Enterococcus and Veillonella. Patients with bactDNA positive ascites revealed reduced spectrum of core microbiota with Streptococcus as key player of duodenal community and higher prevalence of Granulicatella proving presence of cirrhosis related intestinal dysbiosis. Regarding duodenal fluid bactDNA quantification, no significant differences were found between patients with cirrhosis and controls. Additionally, percentage of subjects with detectable bactDNA in blood did not differ between patients and controls. This study evaluated the diversity of bacterial DNA in different body specimens with potential implications on understanding how intestinal bacterial translocation may affect infectious complications in cirrhosis.

Polyphasic Analysis Reveals Potential Petroleum Hydrocarbon Degradation and Biosurfactant Production by Rare Biosphere Thermophilic Bacteria From Deception Island, an Active Antarctic Volcano.

Extreme temperature gradients in polar volcanoes are capable of selecting different types of extremophiles. Deception Island is a marine stratovolcano located in maritime Antarctica. The volcano has pronounced temperature gradients over very short distances, from as high as 100°C in the fumaroles to subzero next to the glaciers. These characteristics make Deception a promising source of a variety of bioproducts for use in different biotechnological areas. In this study, we isolated thermophilic bacteria from sediments in fumaroles at two geothermal sites on Deception Island with temperatures between 50 and 100°C, to evaluate the potential capacity of these bacteria to degrade petroleum hydrocarbons and produce biosurfactants under thermophilic conditions. We isolated 126 thermophilic bacterial strains and identified them molecularly as members of genera Geobacillus, Anoxybacillus, and Brevibacillus (all in phylum Firmicutes). Seventy-six strains grew in a culture medium supplemented with crude oil as the only carbon source, and 30 of them showed particularly good results for oil degradation. Of 50 strains tested for biosurfactant production, 13 showed good results, with an emulsification index of 50% or higher of a petroleum hydrocarbon source (crude oil and diesel), emulsification stability at 100°C, and positive results in drop-collapse, oil spreading, and hemolytic activity tests. Four of these isolates showed great capability of degrade crude oil: FB2_38 (Geobacillus), FB3_54 (Geobacillus), FB4_88 (Anoxybacillus), and WB1_122 (Geobacillus). Genomic analysis of the oil-degrading and biosurfactant-producer strain FB4_88 identified it as Anoxybacillus flavithermus, with a high genetic and functional diversity potential for biotechnological applications. These initial culturomic and genomic data suggest that thermophilic bacteria from this Antarctic volcano have potential applications in the petroleum industry, for bioremediation in extreme environments and for microbial enhanced oil recovery (MEOR) in reservoirs. In addition, recovery of small-subunit rRNA from metagenomes of Deception Island showed that Firmicutes is not among the dominant phyla, indicating that these low-abundance microorganisms may be important for hydrocarbon degradation and biosurfactant production in the Deception Island volcanic sediments.

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.

Transport of marine tracer phage particles in soil.

Marine phages have been applied to trace ground- and surface water flows. Yet, information on their transport in soil and related particle intactness is limited. Here we compared the breakthrough of two lytic marine tracer phages (Pseudoalteromonas phages PSA-HM1 and PSA-HS2) with the commonly used Escherichia virus T4 in soil- and sand-filled laboratory percolation columns. All three phages showed high mass recoveries in the effluents and a higher transport velocity than non-reactive tracer Br-. Comparison of effluent gene copy numbers (CN) to physically-determined phage particle counts or infectious phage counts showed that PSA-HM1 and PSA-HS2 retained high phage particle intactness (Ip > 81%), in contrast to T4 (Ip < 36%). Our data suggest that marine phages may be applied in soil to mimic the transport of (bio-) colloids or anthropogenic nanoparticles of similar traits. Quantitative PCR (qPCR) thereby allows for highly sensitive quantification and thus for the detection of even highly diluted marine tracer phages in environmental samples.

Phage co-transport with hyphal-riding bacteria fuels bacterial invasion in a water-unsaturated microbial model system.

Nonmotile microorganisms often enter new habitats by co-transport with motile microorganisms. Here, we report that also lytic phages can co-transport with hyphal-riding bacteria and facilitate bacterial colonization of a new habitat. 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 ≈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 seems analogous to macroecological invasion processes, hyphosphere systems with motile bacteria and co-transported phages could be useful models for testing hypotheses in invasion ecology.