Disruption of <i>Mycobacterium smegmatis</i> Biofilms Using Bacteriophages Alone or in Combination with Mechanical Stress

Abstract

Environmental mycobacteria are capable of forming biofilms in low-nutrient environments, and these biofilms may act as reservoirs for opportunistic infections. The purpose of this study was to determine if bacteriophages could disrupt existing biofilms of acid-fast staining Mycobacterium smegmatis. Using the MBEC 96-well plastic peg assay system, M. smegmatis biofilms were created and then tested for their stability in the presence of mycobacteriophages isolated from a Minnesota sphagnum peat bog. All phages tested were lytic and were observed to have weak, intermediate, and strong abilities to disrupt M. smegmatis biofilms. The formation of biofilms was severely impaired in the presence of mycobacteriophages. Phage treatment was also shown to augment M. smegmatis biofilm disruption by mechanical forces of sonication or water flow. Our study shows that, as with biofilms of Gram-positive and Gram-negative bacteria, mycobacterial biofilms are also susceptible to destruction by bacteriophages.

Insights into the genomic features and lifestyle of B1 subcluster mycobacteriophages

“…Only limited reports are available on the antibiofilm activity of mycobacteriophages [37] and given the predicted presence of a depolymerase‐like enzyme in B1 phages, we wanted to test the antibiofilm activity of Phages 1–10 . Liquid‐air interface biofilms of M. smegmatis mc 2 155 were formed in 24‐well plates, and phages were tested for their ability to inhibit and disrupt biofilm (Figure 6a–c).…”

A novel LysinB from an F2 sub-cluster mycobacteriophageRitSun

“…To examine the inhibitory effect of RitSun LysinB on M. smegmatis Mc 2 155 biofilm formation, the Kiefer et al . [48] protocol was followed with minor modifications. Briefly, M. smegmatis cells (OD 600 adjusted to 0.1) were mixed with LysinB in a 24-well plate and incubated at 37°C for 96 hours.…”

“…After completing LysinB treatment, the planktonic cells were aspirated, and the biofilm was air-dried and stained with freshly prepared crystal violet stain (1%) for 30 minutes at 37°C and washed twice with 1X PBS. The CV stain was extracted with 95% ethanol, and the absorbance was measured at 590 nm [48].…”

Insights into the genomic features, lifestyle and therapeutic potential of B1 sub-cluster mycobacteriophages

“…2.9 M. smegmatis biofilm formation and treatment with phages 2.9.1. Bacterial strains and growth media For biofilm experiments, we followed a protocol (28) with minor modifications. M. smegmatis, strain mc 2 155 cells grown to OD600 0.8 were used in 24-well plates with incubation for 96 hours at 37 °C under static condition.…”

“…Infection recurrence, immune system evasion and inability to respond to antibiotic treatment are all characteristics of the diseasecausing mycobacteria and biofilm formation to colonize host cells (7). However, only limited reports are available on the antibiofilm activity of mycobacteriophages (28,64). Since phageencoded depolymerase enzymes are reported to target extracellular or surface polysaccharides, i.e., bacterial capsules, slime layers, biofilm matrix, or lipopolysaccharides (65), we first analyzed the structural proteins of B1 phages using PHYPred to predict the presence of any encoded enzymes that could aid impacting the biofilm.…”