Interactions between bacteriophages (phages) and biofilms remain poorly understood despite the broad implications for microbial ecology, water quality, and microbiome engineering. Here, we demonstrate that lytic coliphage PHH01 can hitchhike on carrier bacteria Bacillus cereus to facilitate its infection of host bacteria, Escherichia coli, in biofilms. Specifically, PHH01 could adsorb onto the flagella of B. cereus, and thus phage motility was increased, resulting in 4.36-fold more effective infection of E. coli in biofilm relative to free PHH01 alone. Moreover, phage infection mitigated interspecies competition and enhanced B. cereus colonization; the fraction of B. cereus in the final biofilm increased from 9% without phages to 43% with phages. The mutualistic relationship between the coliphage and carrier bacteria was substantiated by migration tests on an E. coli lawn: the conjugation of PHH01 and B. cereus enhanced B. cereus colonization by 6.54-fold compared to B. cereus alone (6.15 vs 0.94 cm 2 in 24 h) and PHH01 migration by 5.15-fold compared to PHH01 alone (10.3 vs 2.0 mm in 24 h). Metagenomic and electron microscopic analysis revealed that the phages of diverse taxonomies and different morphologies could be adsorbed by the flagella of B. cereus, suggesting hitchhiking on flagellated bacteria might be a widespread strategy in aquatic phage populations. Overall, our study highlights that hitchhiking behavior in phages can facilitate phage infection of biofilm bacteria, promote carrier bacteria colonization, and thus significantly influence biofilm composition, which holds promise for mediating biofilm functions and moderating associated risks.

Background Microbe–virus interactions have broad implications on the composition, function, and evolution of microbiomes. Elucidating the effects of environmental stresses on these interactions is critical to identify the ecological function of viral communities and understand microbiome environmental adaptation. Heavy metal-contaminated soils represent a relevant ecosystem to study the interplay between microbes, viruses, and environmental stressors. Results Metagenomic analysis revealed that Cr pollution adversely altered the abundance, diversity, and composition of viral and bacterial communities. Host–phage linkage based on CRISPR indicated that, in soils with high Cr contamination, the abundance of phages associated with heavy metal-tolerant hosts increased, as did the relative abundance of phages with broad host ranges (identified as host–phage linkages across genera), which would facilitate transfection and broader distribution of heavy metal resistance genes in the bacterial community. Examining variations along the pollutant gradient, enhanced mutualistic phage–bacterium interactions were observed in the face of greater environmental stresses. Specifically, the fractions of lysogens in bacterial communities (identified by integrase genes within bacterial genomes and prophage induction assay by mitomycin-C) were positively correlated with Cr contamination levels. Furthermore, viral genomic analysis demonstrated that lysogenic phages under higher Cr-induced stresses carried more auxiliary metabolic genes regulating microbial heavy metal detoxification. Conclusion With the intensification of Cr-induced environmental stresses, the composition, replication strategy, and ecological function of the phage community all evolve alongside the bacterial community to adapt to extreme habitats. These result in a transformation of the phage–bacterium interaction from parasitism to mutualism in extreme environments and underscore the influential role of phages in bacterial adaptation to pollution-related stress and in related biogeochemical processes.

Phthalate esters (PAEs) are commonly released from plastic pipes in some water distribution systems. Here, we show that exposure to a low concentration (1–10 μg/L) of three PAEs (dimethyl phthalate (DMP), di-n-hexyl phthalate (DnHP), and di-(2-ethylhexyl) phthalate (DEHP)) promotes Pseudomonas biofilm formation and resistance to free chlorine. At PAE concentrations ranging from 1 to 5 μg/L, genes coding for quorum sensing, extracellular polymeric substances excretion, and oxidative stress resistance were upregulated by 2.7- to 16.8-fold, 2.1- to 18.9-fold, and 1.6- to 9.9-fold, respectively. Accordingly, more biofilm matrix was produced and the polysaccharide and eDNA contents increased by 30.3–82.3 and 10.3–39.3%, respectively, relative to the unexposed controls. Confocal laser scanning microscopy showed that PAE exposure stimulated biofilm densification (volumetric fraction increased from 27.1 to 38.0–50.6%), which would hinder disinfectant diffusion. Biofilm densification was verified by atomic force microscopy, which measured an increase of elastic modulus by 2.0- to 3.2-fold. PAE exposure also stimulated the antioxidative system, with cell-normalized superoxide dismutase, catalase, and glutathione activities increasing by 1.8- to 3.0-fold, 1.0- to 2.0-fold, and 1.2- to 1.6-fold, respectively. This likely protected cells against oxidative damage by chlorine. Overall, we demonstrate that biofilm exposure to environmentally relevant levels of PAEs can upregulate molecular processes and physiologic changes that promote biofilm densification and antioxidative system expression, which enhance biofilm resistance to disinfectants.

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Bacteriophages (phages) are an underutilized biological resource with vast potential for pathogen control and microbiome editing. Phage research and commercialization have increased rapidly in biomedical and agricultural industries, but adoption has been limited elsewhere. Nevertheless, converging advances in DNA sequencing, bioinformatics, microbial ecology, and synthetic biology are now poised to broaden phage applications beyond pathogen control toward the manipulation of microbial communities for defined functional improvements. Enhancements in sequencing combined with network analysis make it now feasible to identify and disrupt microbial associations to elicit desirable shifts in community structure or function, indirectly modulate species abundance, and target hub or keystone species to achieve broad functional shifts. Sequencing and bioinformatic advancements are also facilitating the use of temperate phages for safe gene delivery applications. Finally, integration of synthetic biology stands to create novel phage chassis and modular genetic components. While some fundamental, regulatory, and commercialization barriers to widespread phage use remain, many major challenges that have impeded the field now have workable solutions. Thus, a new dawn for phage-based (chemical-free) precise biocontrol and microbiome editing is on the horizon to enhance, suppress, or modulate microbial activities important for public health, food security, and more sustainable energy production and water reuse.