Studies of Black Diamond as an antibacterial surface for Gram Negative bacteria: the interplay between chemical and mechanical bactericidal activity

Abstract

‘Black silicon’ (bSi) samples with surfaces covered in nanoneedles of length ~5 µm were fabricated using a plasma etching process and then coated with a conformal uniform layer of diamond using hot filament chemical vapour deposition to produce ‘black diamond’ (bD) nanostructures. The diamond needles were then chemically terminated with H, O, NH 2 or F using plasma treatment, and the hydrophilicity of the resulting surfaces were assessed using water droplet contact-angle measurements, and scaled in the order O > H ≈NH 2 >F, with the F-terminated surface being superhydrophobic. The effectiveness of these differently terminated bD needles in killing the Gram-negative bacterium E. coli was semi-quantified by Live/Dead staining and fluorescence microscopy, and visualised by environmental scanning electron microscopy. The total number of adhered bacteria was consistent for all the nanostructured bD surfaces at around 50% of the value for the flat diamond control. This, combined with a chemical bactericidal effect of 20–30%, shows that the nanostructured bD surfaces supported significantly fewer viable E. coli than flat surfaces. Moreover, the bD surfaces were particularly effective at preventing the establishment of bacterial aggregates – a precursor to biofilm formation. The percentage of dead bacteria also decreased as a function of hydrophilicity. These results are consistent with a predominantly mechanical mechanism for bacteria death based on the stretching and disruption of the cell membrane, combined with an additional effect from the chemical nature of the surface.

Bactericidal efficiency of micro- and nanostructured surfaces: a critical perspective

“…Similarly no correlation is evident on hydrophilic MNSS as some hydrophilic surfaces were effective against Gram-positive, 38,51 while some were effective against Gramnegative strains, 37,38,45,51,53 and some effective against both types. 37,38,51 Despite high bactericidal effect by some hydrophobic surfaces and hydrophilic surfaces as above, certain hydrophobic 28,55 as well as hydrophilic 36,46,55,[58][59][60][61] MNSS are reported with low or no bactericidal effect. Most of unsuccessful substrates were titanium, 36,46,[58][59][60] and features were pores, 59 tubes, 58 or crystals.…”

“…Moreover, these nanowires were sharper with diameters ranging from 17-42 nm, compared to most of pillar type nanostructures reported in literature that has pillar tip diameters above 100 nm. 28,38,44,45,[54][55][56][57] Several other reports also conrmed that thinner features demonstrate high bactericidal effect on Gram-positive species compared to Gram-negative strains. For an example, 20-40 nm diameter titanium nano tubes exhibited 51-60% BE against S. aureus, 58 and silicon pillars with 21 nm diameter on S. aureus was 90-97% (ref.…”

“…The major similarity of protocols used for these studies is that bacteria are incubated on substrates in a contained space with constant amount of growth media in static or agitated conditions. 26,28,31,33,[36][37][38][39][40][44][45][46][47][48]51,[53][54][55][56][58][59][60][62][63][64] Phosphate buffered saline solution is a commonly used medium to make bacterial suspension. The substrate is immersed in bacterial suspension within a container such as in a microwell plate.…”

“…For the viability experiment, a common step is to rinse the substrates to remove non-adhered or weakly-adhered bacteria from the surface before conducting quantication test such as live dead analysis or colony counting. 31,33,37,38,40,45,47,51,53,55,56,59,63,[66][67][68][69][70][71][72][73][74][75][76][77][78][79][80][81][82][83][84][85] Commonly used rinsing methods are summarised in Table 1. Strikingly, the volumes or owrates used in the rinsing step of these studies are unspeci-ed, and the effect of stresses exerted on the loosely attached bacterial cells have not been accounted for.…”

Hydrophobicity and Adhesion of Aggregated Diamond Particles

“…Aminated (–NH 2 ) diamond surfaces are slightly hydrophobic, whereas fluorinated ones can be superhydrophobic. [ 15 ]…”