Mycobacterium fortuitum biofilm formation on water distribution pipe materials

摘要

Nontuberculous mycobacteria (NTM), containing several opportunistically pathogenicrnspecies, are present in many water distribution systems, including those in healthcarernfacilities. Since previous work has shown that NTM are more resistant to chlorinerndisinfectant than are most other water bacteria, the potential exists that increasingrnconcentrations of chlorine or chloramine result in the selection for NTM in waterrnsystems. However, little is known about NTM populations in water distribution systems.rnThis study compared the effect of different substrata (copper, glass, polycarbonate,rnpolyvinyl chloride (PVC), and stainless steel) on Mycobacterium fortuitum biofilmrnstructure under high and low nutrient conditions. Biofilms were grown in a 24-well platernin batch mode with shaking for 3 days at 35°C in either autoclaved tap water (lowrnnutrient) or modified R2A broth (high nutrient). M. fortuitum biofilms were then stainedrnwith Sybr Green I and examined using a Zeiss Axioplan 2 with an apatome attachment.rnThree-dimensional image stacks of biofilms on each material were collected with anrnAxioCamHRm and the AxioVision software (Zeiss, v. 4.2 ), then analyzed in thernprogram COMSTAT for maximum and average thickness, total biomass, andrnsurface:biomass ratio. For all materials, the biofilm was thicker when grown in nutrientrnmedium than in tap water, ranging from 1.69 (± 0.85) μm maximum thickness on copperrnto 15.3 (± 5.80) μm on polycarbonate. M. fortuitum biofilm developed in tap waterrnreached a maximum thickness of 0.78 (± 0.43), 3.85 (± 1.73), and 4.61 (± 2.49) μm onrncopper, polycarbonate, and PVC respectively. The biofilm was thicker and containedrnmore biomass when grown on stainless steel, polycarbonate or PVC than when grown onrncopper or glass. Under both nutrient conditions, the biofilm had the highestrnsurface:biomass ratio on PVC. However, all materials, except copper, supported therndevelopment of microcolonies in tap water within the 3-day period. Copper substraterndeveloped small microcolonies in modified R2A medium but contained only individualrnbacterial cells after incubation in tap water. In conclusion, our results a) show thatrnquantification of biofilm structural features provides a novel means for comparing therneffects of materials on biofilm formation, and b) suggest that pipe material and nutrientrnconditions both play a role in limiting (copper, low nutrient) or enhancing (plastics,rnstainless steel, high nutrient) NTM biofilm formation in water systems. Further studies