Subdiffusive motion of bacteriophage in mucosal surfaces increases the frequency of bacterial encounters

Abstract

Bacteriophages (phages) defend mucosal surfaces against bacterial infections. However, their complex interactions with their bacterial hosts and with the mucus-covered epithelium remain mostly unexplored. Our previous work demonstrated that T4 phage with Hoc proteins exposed on their capsid adhered to mucin glycoproteins and protected mucus-producing tissue culture cells in vitro. On this basis, we proposed our bacteriophage adherence to mucus (BAM) model of immunity. Here, to test this model, we developed a microfluidic device (chip) that emulates a mucosal surface experiencing constant fluid flow and mucin secretion dynamics. Using mucus-producing human cells and Escherichia coli in the chip, we observed similar accumulation and persistence of mucus-adherent T4 phage and nonadherent T4Δhoc phage in the mucus. Nevertheless, T4 phage reduced bacterial colonization of the epithelium >4,000-fold compared with T4Δhoc phage. This suggests that phage adherence to mucus increases encounters with bacterial hosts by some other mechanism. Phages are traditionally thought to be completely dependent on normal diffusion, driven by random Brownian motion, for host contact. We demonstrated that T4 phage particles displayed subdiffusive motion in mucus, whereas T4Δhoc particles displayed normal diffusion. Experiments and modeling indicate that subdiffusive motion increases phage-host encounters when bacterial concentration is low. By concentrating phages in an optimal mucus zone, subdiffusion increases their host encounters and antimicrobial action. Our revised BAM model proposes that the fundamental mechanism of mucosal immunity is subdiffusion resulting from adherence to mucus. These findings suggest intriguing possibilities for engineering phages to manipulate and personalize the mucosal microbiome.BAM | virus | mucus | subdiffusion | search strategy I n all animals, mucosal surfaces provide critical immunological services by both protecting against invading bacterial pathogens and supporting large communities of commensal microorganisms (1, 2). Being exposed to the environment, mucosal surfaces are also the infection sites for many important bacterial diseases, including acute diarrhea and cystic fibrosis in humans. This, combined with their accessibility, make mucosal surfaces attractive venues for phage therapy; that is, the use of bacteriophages (phages) to treat and clear bacterial infections (3,4). Clinical success so far has been erratic (5). The complexities and dynamics of the mucus layer are rarely considered, and the activity of phages therein is mostly unknown. Not surprisingly, phages effective in vitro do not consistently reduce mucosal bacterial host levels in vivo (6, 7). An understanding of the interactions between phages and their bacterial hosts within the relevant physiological environment is critical for consistent success of phage therapy applications.The multilayered mucus is composed primarily of gel-forming mucin glycoproteins that are continually secreted by the underlying epithelium (8). The mucin...

Bacteriophages targeting adherent invasive Escherichia coli strains as a promising new treatment for Crohn's disease

“…[37][38][39][40] Also, recent evidence showed that some bacteriophages bind mucus, which could affect their persistence in the GIT. 41,42 Using CEABAC10 mice, we showed that the cocktail of bacteriophages strongly decreased intestinal colonisation by strain LF82, suggesting that bacteriophages were able to reach the epithelial surface bearing the CEACAM6 receptor of strain LF82. This strong decrease was observed with both CEABAC10 and conventional mice in the absence and presence of antibiotic pressure, respectively.…”

Stress as a Normal Cue in the Symbiotic Environment

“…Within the scaffolding provided by host-derived nutrients, competition among members of the microbiota may make the community more resilient to external perturbation by pathogens, or chemical agents [54]. Bacteriocins [55], antimicrobials [56], reactive oxygen species [57], and bacteriophage [58, 59] are stresses derived from microbes in the host environment that can structure the competition for nutrients among communties. The production of the bacteriocin coproporphyrin III by a human skin-specific Propionibacterium sp .…”

“…Hydrogen peroxide produced by Streptococcus oligofermentans counteracts lactic acid production by Streptococcus mutans in the oral microbiota [57], suggesting that the two species may interact within the diffusive limits of their environment. Bacteriophage-mucin binding interactions have been shown to structure colonization of mucosal layers in the mammalian gut [58, 59], and the accumulation of bacteriophage in sputum or EPS may enhance the protective function of this barrier towards antimicrobials or other chemical stresses [61]. In addition, a functional CRISPR foreign DNA defense system is required for Xenorhabdus nematophila to colonize the gut of its nematode host [62] (Figure 2), suggesting that bacteriophage may be a stressful attribute of the nematode gut microbiota.…”

Viruses and the origin of microbiome selection and immunity

“…Colonization by additional bacterial species and their associated phage continued in the SML until niche space was filled and a community was formed. Mutualistic bacteria continued to protect the metazoan host through competitive exclusion of potential pathogens (Hibbing et al, 2010) while phage provided immune protection as proposed in the bacteriophage adherence to mucus (BAM) model (Barr et al, 2013(Barr et al, , 2015. In the BAM model, phage adhering to mucus provide the host with immune protection against invading pathogens (Figure 1a).…”

“…The LCMA secreted mucins from epithelial tissue, generating an SML that selected for bacteria and phage with mucin-binding properties. Phage provided the LCMA with an external microbial selective in which phage bound to mucus via hypervariable domains protect the metazoan host from invading bacteria (BAM) (Barr et al, 2013(Barr et al, , 2015. In addition to attracting bacteria and phage, mucins also increased the rate of contact with eukaryotic viruses resulting in the development of mutualistic symbiosis that provided the LCMA with immune protection.…”