Phage-host Systems Research Articles

Prediction of strain level phage-host interactions across the Escherichia genus using only genomic information.

Predicting bacteriophage infection of specific bacterial strains promises advancements in phage therapy and microbial ecology. Whether the dynamics of well-established phage-host model systems generalize to the wide diversity of microbes is currently unknown. Here we show that we could accurately predict the outcomes of phage-bacteria interactions at the strain level in natural isolates from the genus Escherichia using only genomic data (area under the receiver operating characteristic curve (AUROC) of 86%). We experimentally established a dataset of interactions between 403 diverse Escherichia strains and 96 phages. Most interactions are explained by adsorption factors as opposed to antiphage systems which play a marginal role. We trained predictive algorithms and pinpoint poorly predicted interactions to direct future research efforts. Finally, we established a pipeline to recommend tailored phage cocktails, demonstrating efficiency on 100 pathogenic E. coli isolates. This work provides quantitative insights into phage-host specificity and supports the use of predictive algorithms in phage therapy.

Pseudo-pac site sequences used by phage P22 in generalized transduction of Salmonella.

Salmonella enterica Serovar Typhimurium (Salmonella) and its bacteriophage P22 are a model system for the study of horizontal gene transfer by generalized transduction. Typically, the P22 DNA packaging machinery initiates packaging when a short sequence of DNA, known as the pac site, is recognized on the P22 genome. However, sequences similar to the pac site in the host genome, called pseudo-pac sites, lead to erroneous packaging and subsequent generalized transduction of Salmonella DNA. While the general genomic locations of the Salmonella pseudo-pac sites are known, the sequences themselves have not been determined. We used visualization of P22 sequencing reads mapped to host Salmonella genomes to define regions of generalized transduction initiation and the likely locations of pseudo-pac sites. We searched each genome region for the sequence with the highest similarity to the P22 pac site and aligned the resulting sequences. We built a regular expression (sequence match pattern) from the alignment and used it to search the genomes of two P22-susceptible Salmonella strains-LT2 and 14028S-for sequence matches. The final regular expression successfully identified pseudo-pac sites in both LT2 and 14028S that correspond with generalized transduction initiation sites in mapped read coverages. The pseudo-pac site sequences identified in this study can be used to predict locations of generalized transduction in other P22-susceptible hosts or to initiate generalized transduction at specific locations in P22-susceptible hosts with genetic engineering. Furthermore, the bioinformatics approach used to identify the Salmonella pseudo-pac sites in this study could be applied to other phage-host systems.

Genomic Analysis of Two Cold-Active Pseudoalteromonas Phages Isolated from the Continental Shelf in the Arctic Ocean.

Cold-active bacteriophages are bacterial viruses that infect and replicate at low temperatures (≤4 °C). Understanding remains limited of how cold-active phage-host systems sustain high viral abundance despite the persistently low temperatures in pelagic sediments in polar seas. In this study, two Pseudoalteromonas phages, ACA1 and ACA2, were isolated from sediment core samples of the continental shelf in the western Arctic Ocean. These phages exhibited successful propagation at a low temperature of 1 °C and displayed typical myovirus morphology with isometric icosahedral heads and contractile tails. The complete genome sequences of phages ACA1 and ACA2 were 36,825 bp and 36,826 bp in size, respectively, sharing almost the same gene content. These are temperate phages encoding lysogeny-related proteins such as anti-repressor, immunity repressor and integrase. The absence of cross-infection between the host strains, which were genomically distinct Pseudoalteromonas species, can likely be attributed to heavy divergence in the anti-receptor apparently mediated by an associated diversity-generating retroelement. HHpred searching identified genes for all of the structural components of a P2-like phage (family Peduoviridae), although the whole of the Peduoviridae family appeared to be divided between two anciently diverged tail modules. In contrast, Blast matching and whole genome tree analysis are dominated by a nonstructural gene module sharing high similarity with Pseudoalteromonas phage C5a (founder of genus Catalunyavirus). This study expands the knowledge of diversity of P2-like phages known to inhabit Peudoalteromonas and demonstrates their presence in the Arctic niche.

Three Phages from a Boreal Lake during Ice Cover Infecting Xylophilus, Caulobacter, and Polaromonas Species.

Although the important role of microbes in freshwater is well understood, studies on phage-host systems in such environments during ice cover are completely lacking. Here, we describe the isolation and characterization of three new bacteriophages infecting Xylophilus sp., Caudobacter sp., and Polaromonas sp. from freshwater samples taken under the ice cover of Lake Konnevesi, Finland. Lumi, Kuura, and Tiera bacteriophages have tailed icosahedral virions and double-stranded DNA. Lumi is a siphophage with a genome of 80,496 bp, and Kuura and Tiera are podophages, and their genomes are 43,205 and 45,327 bp in length, resembling viruses in the class Caudoviricetes. Their host ranges were very limited among the winter-isolated bacterial strains from Konnevesi, each infecting only their own hosts. They can infect efficiently at 4 °C, showing that they are adapted to living in lake water under ice cover. Analysis of the viral genome sequences showed that a significant number of the gene products of each virus are unique, indicating that there is unexplored viral diversity in freshwaters. To our knowledge, Lumi and Tiera are the first phages isolated on the Xylophilus sp. and Polaromonas sp. strains, allowing their exploitation in further studies of freshwater bacterial-phage interactions.

The Life Cycle Transitions of Temperate Phages: Regulating Factors and Potential Ecological Implications.

Phages are viruses that infect bacteria. They affect various microbe-mediated processes that drive biogeochemical cycling on a global scale. Their influence depends on whether the infection is lysogenic or lytic. Temperate phages have the potential to execute both infection types and thus frequently switch their infection modes in nature, potentially causing substantial impacts on the host-phage community and relevant biogeochemical cycling. Understanding the regulating factors and outcomes of temperate phage life cycle transition is thus fundamental for evaluating their ecological impacts. This review thus systematically summarizes the effects of various factors affecting temperate phage life cycle decisions in both culturable phage-host systems and natural environments. The review further elucidates the ecological implications of the life cycle transition of temperate phages with an emphasis on phage/host fitness, host-phage dynamics, microbe diversity and evolution, and biogeochemical cycles.

Bacteriophages and their microbial hosts in terrestrial biotopes of Antarctica

AbstractVirus diversity in Antarctic biotopes remains understudied. Here, we describe bacteriophages isolated from terrestrial environments, provide data on their natural bacterial hosts and study phage-host systems. Six bacterial isolates (FCKU 539, FCKU 533, FCKU 534, FCKU 538, FCKU 542 and FCKU 540) were recovered and characterized. Isolated bacteria belonged to Pseudomonas genus (Pseudomonas sp., Pseudomonas fluorescens, Pseudomonas putida) with optimal cultivation temperatures of 16–28°C. These bacteria and previously described Bacillus subtilis FCKU 537 were used for analysing virus-host interactions. Six lytic phages were isolated and named P. fluorescens Antarctic virus 1 (PFAV1), P. fluorescens Antarctic virus 2 (PFAV2), P. fluorescens Antarctic virus 3 (PFAV3), P. putida Antarctic virus 4 (PPAV4), Pseudomonas sp. Antarctic virus 5 (PSAV5) and B. subtilis Antarctic virus 6 (BSAV6) in relation to their natural hosts. According to electron microscopy data, these phages belonged to Caudovirales order. Cross-inoculation demonstrated high specificity of all Antarctic phages, which infected only their initial hosts at moderate temperatures. PFAV2 and PFAV3 phages also infected laboratory Pseudomonas savastanoi and P. fluorescens isolates. This paper adds new data on the occurrence and diversity of viruses and their respective bacterial hosts in soil biotopes of Antarctica.

Characterization of CRISPR Spacer and Protospacer Sequences in Paenibacillus larvae and Its Bacteriophages.

The bacterium Paenibacillus larvae is the causative agent of American foulbrood, the most devastating bacterial disease of honeybees. Because P. larvae is antibiotic resistant, phages that infect it are currently used as alternative treatments. However, the acquisition by P. larvae of CRISPR spacer sequences from the phages could be an obstacle to treatment efforts. We searched nine complete genomes of P. larvae strains and identified 714 CRISPR spacer sequences, of which 384 are unique. Of the four epidemiologically important P. larvae strains, three of these have fewer than 20 spacers, while one strain has over 150 spacers. Of the 384 unique spacers, 18 are found as protospacers in the genomes of 49 currently sequenced P. larvae phages. One P. larvae strain does not have any protospacers found in phages, while another has eight. Protospacer distribution in the phages is uneven, with two phages having up to four protospacers, while a third of phages have none. Some phages lack protospacers found in closely related phages due to point mutations, indicating a possible escape mechanism. This study serve a point of reference for future studies on the CRISPR-Cas system in P. larvae as well as for comparative studies of other phage–host systems.

Mutant and Recombinant Phages Selected from In Vitro Coevolution Conditions Overcome Phage-Resistant Listeria monocytogenes.

Bacteriophages (phages) are currently available for use by the food industry to control the foodborne pathogen Listeria monocytogenes Although phage biocontrols are effective under specific conditions, their use can select for phage-resistant bacteria that repopulate phage-treated environments. Here, we performed short-term coevolution experiments to investigate the impact of single phages and a two-phage cocktail on the regrowth of phage-resistant L. monocytogenes and the adaptation of the phages to overcome this resistance. We used whole-genome sequencing to identify mutations in the target host that confer phage resistance and in the phages that alter host range. We found that infections with Listeria phages LP-048, LP-125, or a combination of both select for different populations of phage-resistant L. monocytogenes bacteria with different regrowth times. Phages isolated from the end of the coevolution experiments were found to have gained the ability to infect phage-resistant mutants of L. monocytogenes and L. monocytogenes strains previously found to be broadly resistant to phage infection. Phages isolated from coinfected cultures were identified as recombinants of LP-048 and LP-125. Interestingly, recombination events occurred twice independently in a locus encoding two proteins putatively involved in DNA binding. We show that short-term coevolution of phages and their hosts can be utilized to obtain mutant and recombinant phages with adapted host ranges. These laboratory-evolved phages may be useful for limiting the emergence of phage resistance and for targeting strains that show general resistance to wild-type (WT) phages.IMPORTANCEListeria monocytogenes is a life-threatening bacterial foodborne pathogen that can persist in food processing facilities for years. Phages can be used to control L. monocytogenes in food production, but phage-resistant bacterial subpopulations can regrow in phage-treated environments. Coevolution experiments were conducted on a Listeria phage-host system to provide insight into the genetic variation that emerges in both the phage and bacterial host under reciprocal selective pressure. As expected, mutations were identified in both phage and host, but additionally, recombination events were shown to have repeatedly occurred between closely related phages that coinfected L. monocytogenes This study demonstrates that in vitro evolution of phages can be utilized to expand the host range and improve the long-term efficacy of phage-based control of L. monocytogenes This approach may also be applied to other phage-host systems for applications in biocontrol, detection, and phage therapy.

How do temperate bacteriophages affect the fitness of Pseudomonas aeruginosa?

The Liverpool Epidemic Strain(LES) of Pseudomonas aeruginosa is a key opportunistic bacterium and a major cause of mortality and morbidity in cystic fibrosis (CF) patients. It has been established to carry distinctive prophages within its genome, providing the host bacterium with advantages through horizontal gene transfer. Several well-known partnerships between prophage and their bacterial hosts have been characterised, however, very little is known about other phage-host systems. This project explores the dynamics between Pseudomonas aeruginosa PAO1 and its phage, determining whether they confer an advantage in the CF lung.

Bacteriophages for Chronic Wound Treatment: from Traditional to Novel Delivery Systems.

The treatment and management of chronic wounds presents a massive financial burden for global health care systems, with significant and disturbing consequences for the patients affected. These wounds remain challenging to treat, reduce the patients’ life quality, and are responsible for a high percentage of limb amputations and many premature deaths. The presence of bacterial biofilms hampers chronic wound therapy due to the high tolerance of biofilm cells to many first- and second-line antibiotics. Due to the appearance of antibiotic-resistant and multidrug-resistant pathogens in these types of wounds, the research for alternative and complementary therapeutic approaches has increased. Bacteriophage (phage) therapy, discovered in the early 1900s, has been revived in the last few decades due to its antibacterial efficacy against antibiotic-resistant clinical isolates. Its use in the treatment of non-healing wounds has shown promising outcomes. In this review, we focus on the societal problems of chronic wounds, describe both the history and ongoing clinical trials of chronic wound-related treatments, and also outline experiments carried out for efficacy evaluation with different phage-host systems using in vitro, ex vivo, and in vivo animal models. We also describe the modern and most recent delivery systems developed for the incorporation of phages for species-targeted antibacterial control while protecting them upon exposure to harsh conditions, increasing the shelf life and facilitating storage of phage-based products. In this review, we also highlight the advances in phage therapy regulation.

Diversity and Host Interactions Among Virulent and Temperate Baltic Sea Flavobacterium Phages.

Viruses in aquatic environments play a key role in microbial population dynamics and nutrient cycling. In particular, bacteria of the phylum Bacteriodetes are known to participate in recycling algal blooms. Studies of phage–host interactions involving this phylum are hence important to understand the processes shaping bacterial and viral communities in the ocean as well as nutrient cycling. In this study, we isolated and sequenced three strains of flavobacteria—LMO6, LMO9, LMO8—and 38 virulent phages infecting them. These phages represent 15 species, occupying three novel genera. Additionally, one temperate phage was induced from LMO6 and was found to be competent at infecting LMO9. Functions could be predicted for a limited number of phage genes, mainly representing roles in DNA replication and virus particle formation. No metabolic genes were detected. While the phages isolated on LMO8 could infect all three bacterial strains, the LMO6 and LMO9 phages could not infect LMO8. Of the phages isolated on LMO9, several showed a host-derived reduced efficiency of plating on LMO6, potentially due to differences in DNA methyltransferase genes. Overall, these phage–host systems contribute novel genetic information to our sequence databases and present valuable tools for the study of both virulent and temperate phages.

The run-on oligomer filament enzyme mechanism of SgrAI: Part 1. Assembly kinetics of the run-on oligomer filament

Filament or run-on oligomer formation by metabolic enzymes is now recognized as a widespread phenomenon having potentially unique enzyme regulatory properties and biological roles, and its dysfunction is implicated in human diseases such as cancer, diabetes, and developmental disorders. SgrAI is a bacterial allosteric type II restriction endonuclease that binds to invading phage DNA, may protect the host DNA from off-target cleavage activity, and forms run-on oligomeric filaments with enhanced DNA-cleavage activity and altered DNA sequence specificity. However, the mechanisms of SgrAI filament growth, cooperativity in filament formation, sequestration of enzyme activity, and advantages over other filament mechanisms remain unknown. In this first of a two-part series, we developed methods and models to derive association and dissociation rate constants of DNA-bound SgrAI in run-on oligomers and addressed the specific questions of cooperativity and filament growth mechanisms. We show that the derived rate constants are consistent with the run-on oligomer sizes determined by EM analysis and are most consistent with a noncooperative growth mode of the run-on oligomer. These models and methods are extended in the accompanying article to include the full DNA-cleavage pathway and address specific questions related to the run-on oligomer mechanism including the sequestration of DNA-cleavage activity and trapping of products.

Biogeography and taxonomic overview of terrestrial hot spring thermophilic phages.

Bacterial viruses ("phages") play important roles in the regulation and evolution of microbial communities in most ecosystems. Terrestrial hot springs typically contain thermophilic bacterial communities, but the diversity and impacts of its associated viruses ("thermophilic phages") are largely unexplored. Here, we provide a taxonomic overview of phages that have been isolated strictly from terrestrial hot springs around the world. In addition, we placed 17 thermophilic phage genomes in a global phylogenomic context to detect evolutionary patterns. Thermophilic phages have diverse morphologies (e.g., tailed, filamentous), unique virion structures (e.g., extremely long tailed siphoviruses), and span five taxonomic families encompassing strictly thermophilic phage genera. Within the phage proteomic tree, six thermophilic phage-related clades were identified, with evident genomic relatedness between thermophilic phages and archaeal viruses. Moreover, whole proteome analyses showed clustering between phages that infect distinct host phyla, such as Firmicutes and Deinococcus-Thermus. The potential for discovery of novel phage-host systems in terrestrial hot springs remain mostly untapped, thus additional emphasis on thermophilic phages in ecological prospecting is encouraged to gain insights into the microbial population dynamics of these environments.

Large-scale maps of variable infection efficiencies in aquatic Bacteroidetes phage-host model systems.

Microbes drive ecosystem functioning and their viruses modulate these impacts through mortality, gene transfer and metabolic reprogramming. Despite the importance of virus-host interactions and likely variable infection efficiencies of individual phages across hosts, such variability is seldom quantified. Here, we quantify infection efficiencies of 38 phages against 19 host strains in aquatic Cellulophaga (Bacteroidetes) phage-host model systems. Binary data revealed that some phages infected only one strain while others infected 17, whereas quantitative data revealed that efficiency of infection could vary 10 orders of magnitude, even among phages within one population. This provides a baseline for understanding and modeling intrapopulation host range variation. Genera specific host ranges were also informative. For example, the Cellulophaga Microviridae, showed a markedly broader intra-species host range than previously observed in Escherichia coli systems. Further, one phage genus, Cba41, was examined to investigate nonheritable changes in plating efficiency and burst size that depended on which host strain it most recently infected. While consistent with host modification of phage DNA, no differences in nucleotide sequence or DNA modifications were detected, leaving the observation repeatable, but the mechanism unresolved. Overall, this study highlights the importance of quantitatively considering replication variations in studies of phage-host interactions.

The interactome of Streptococcus pneumoniae and its bacteriophages show highly specific patterns of interactions among bacteria and their phages

Although an abundance of bacteriophages exists, little is known about interactions between their proteins and those of their bacterial hosts. Here, we experimentally determined the phage-host interactomes of the phages Dp-1 and Cp-1 and their underlying protein interaction network in the host Streptococcus pneumoniae. We compared our results to the interaction patterns of E. coli phages lambda and T7. Dp-1 and Cp-1 target highly connected host proteins, occupy central network positions, and reach many protein clusters through the interactions of their targets. In turn, lambda and T7 targets cluster to conserved and essential proteins in E. coli, while such patterns were largely absent in S. pneumoniae. Furthermore, targets in E. coli were mutually strongly intertwined, while targets of Dp-1 and Cp-1 were strongly connected through essential and orthologous proteins in their immediate network vicinity. In both phage-host systems, the impact of phages on their protein targets appears to extend from their network neighbors, since proteins that interact with phage targets were located in central network positions, have a strong topologically disruptive effect and touch complexes with high functional heterogeneity. Such observations suggest that the phages, biological impact is accomplished through a surprisingly limited topological reach of their targets.

Isolation and characterization of phage–host systems from the Baltic Sea ice

In search for sea ice bacteria and their phages from the Baltic Sea ice, two ice samples were collected from land-fast ice in a south-west Finland coastal site in February and March 2011. Bacteria were isolated from the melted sea ice samples and phages were screened from the same samples for 43 purified isolates. Plaque-producing phages were found for 15 bacterial isolates at 3 °C. Ten phage isolates were successfully plaque purified and eight of them were chosen for particle purification to analyze their morphology and structural proteins. Phage 1/32 infecting an isolate affiliated to phylum Bacteroidetes (Flavobacterium sp.) is a siphovirus and six phages infecting isolates affiliated to γ-Proteobacteria (Shewanella sp.) hosts were myoviruses. Cross titrations between the hosts showed that all studied phages are host specific. Phage solutions, host growth and phage infection were tested in different temperatures revealing phage temperature tolerance up to 45 °C, whereas phage infection was in most of the cases retarded above 15 °C. This study is the first to report isolation and cultivation of ice bacteria and cold-active phages from the Baltic Sea ice.

Twelve previously unknown phage genera are ubiquitous in global oceans.

Viruses are fundamental to ecosystems ranging from oceans to humans, yet our ability to study them is bottlenecked by the lack of ecologically relevant isolates, resulting in "unknowns" dominating culture-independent surveys. Here we present genomes from 31 phages infecting multiple strains of the aquatic bacterium Cellulophaga baltica (Bacteroidetes) to provide data for an underrepresented and environmentally abundant bacterial lineage. Comparative genomics delineated 12 phage groups that (i) each represent a new genus, and (ii) represent one novel and four well-known viral families. This diversity contrasts the few well-studied marine phage systems, but parallels the diversity of phages infecting human-associated bacteria. Although all 12 Cellulophaga phages represent new genera, the podoviruses and icosahedral, nontailed ssDNA phages were exceptional, with genomes up to twice as large as those previously observed for each phage type. Structural novelty was also substantial, requiring experimental phage proteomics to identify 83% of the structural proteins. The presence of uncommon nucleotide metabolism genes in four genera likely underscores the importance of scavenging nutrient-rich molecules as previously seen for phages in marine environments. Metagenomic recruitment analyses suggest that these particular Cellulophaga phages are rare and may represent a first glimpse into the phage side of the rare biosphere. However, these analyses also revealed that these phage genera are widespread, occurring in 94% of 137 investigated metagenomes. Together, this diverse and novel collection of phages identifies a small but ubiquitous fraction of unknown marine viral diversity and provides numerous environmentally relevant phage-host systems for experimental hypothesis testing.

Genomic analysis of cold-active Colwelliaphage 9A and psychrophilic phage–host interactions

The 104kb genome of cold-active bacteriophage 9A, which replicates in the marine psychrophilic gamma-proteobacterium Colwellia psychrerythraea strain 34H (between -12 and 8°C), was sequenced and analyzed to investigate elements of molecular adaptation to low temperature and phage-host interactions in the cold. Most characterized ORFs indicated closest similarity to gamma-proteobacteria and their phages, though no single module provided definitive phylogenetic grouping. A subset of primary structural features linked to psychrophily suggested that the majority of annotated phage proteins were not psychrophilic; those that were, primarily serve phage-specific functions and may also contribute to 9A's restricted temperature range for replication as compared to host. Comparative analyses suggest ribonucleotide reductase genes were acquired laterally from host. Neither restriction modification nor the CRISPR-Cas system appeared to be the predominant phage defense mechanism of Cp34H or other cold-adapted bacteria; we hypothesize that psychrophilic hosts rely more on the use of extracellular polymeric material to block cell surface receptors recognized by phages. The relative dearth of evidence for genome-specific defenses, genetic transfer events or auxiliary metabolic genes suggest that the 9A-Cp34H system may be less tightly coupled than are other genomically characterized marine phage-host systems, with possible implications for phage specificity under different environmental conditions.

PhiSiGns: an online tool to identify signature genes in phages and design PCR primers for examining phage diversity

BackgroundPhages (viruses that infect bacteria) have gained significant attention because of their abundance, diversity and important ecological roles. However, the lack of a universal gene shared by all phages presents a challenge for phage identification and characterization, especially in environmental samples where it is difficult to culture phage-host systems. Homologous conserved genes (or "signature genes") present in groups of closely-related phages can be used to explore phage diversity and define evolutionary relationships amongst these phages. Bioinformatic approaches are needed to identify candidate signature genes and design PCR primers to amplify those genes from environmental samples; however, there is currently no existing computational tool that biologists can use for this purpose.ResultsHere we present PhiSiGns, a web-based and standalone application that performs a pairwise comparison of each gene present in user-selected phage genomes, identifies signature genes, generates alignments of these genes, and designs potential PCR primer pairs. PhiSiGns is available at (http://www.phantome.org/phisigns/; http://phisigns.sourceforge.net/) with a link to the source code. Here we describe the specifications of PhiSiGns and demonstrate its application with a case study.ConclusionsPhiSiGns provides phage biologists with a user-friendly tool to identify signature genes and design PCR primers to amplify related genes from uncultured phages in environmental samples. This bioinformatics tool will facilitate the development of novel signature genes for use as molecular markers in studies of phage diversity, phylogeny, and evolution.

A Single-Cell Analysis of Virioplankton Adsorption, Infection, and Intracellular Abundance in Different Bacterioplankton Physiologic Categories

Culture studies of phage-host systems have shown that phage proliferation strongly depends on the physiological state of the host, but it is still unclear to what extent this holds true within aquatic ecosystems. We used a combination of flow sorting and electron microscopy to explore how the frequency of bacterial cells with attached viruses (FCAV), of visibly infected cells, and the number of intracellular viruses are distributed within five physiologic categories: cells with high (HNA) and low (LNA) nucleic acid content, with a compromised membrane, in division, and with an intact-looking morphology. FCAV was not different between the cellular physiologic categories, suggesting low influence of host physiology on viral adsorption. Infected cells were found within all the physiologic categories, besides the dividing cells, but showed different levels of new virion production, with the abundance of intracellular viruses ranked as follows: HNA > intact-looking cells > LNA > compromised membrane cells. These results favor the physiological control hypothesis of viral progeny production. The calculation of viral production rate of the HNA and LNA cells show that viral infection of HNA cells likely accounts for the majority of viral production. It also show that cells considered as less active can still act as resources for phages, although they contain much less intracellular phage particles.