Nanoscale Investigation on Pseudomonas aeruginosa Biofilm Formed on Porous Silicon using Atomic Force Microscopy ASHWIN KANNAN,SUBBALAKSHMI LATHA KARUMANCHI,VINATHA KRISHNA,KOTHAI THIRUVENGADAM, SUBRAMANIAM RAMALINGAM, AND PENNATHUR GAUTAM Centre for Biotechnology, Anna University, Chennai, India Summary: Colonization of surfaces by bacterial cells results in the formation of biofilms. There is a need to study the factors that are important for formation of biofilms since biofilms have been implicated in the failure of semiconductor devices and implants. In the present study, the adhesion force of biofilms (formed by Pseudomonas aeruginosa) on porous silicon substrates of varying surface roughness was quantified using atomic force microscopy (AFM). The experiments were carried out to quantify the effect of surface roughness on the adhesion force of biofilm. The results show that the adhesion force increased from 1.5 Æ 0.5 to 13.2 Æ 0.9 nN with increase in the surface roughness of silicon substrate. The results suggest that the adhesion force of biofilm is affected by surface roughness of substrate. SCANNING 36:551–553, 2014. © 2014 Wiley Periodicals, Inc. Key words: Pseudomonas aeruginosa, biofilm, atomic force microscopy, porous silicon, surface roughness Bacterial attachment to surfaces is affected by factors like surface topography, temperature, surface character- istics of the material and cellular characteristics of the bacteria (Katsikogianni and Missirlis 2004). Coloniza- tion of surfaces by bacterial cells leads to the formation of biofilms. Biofilms are aggregates of microorganisms embedded in a matrix of extracellular polymeric material (Colvin et al. 2011). Biofilm formation is a multistep process initiated by bacterial adhesion to solid surfaces (Dunne 2002). Colonization of surfaces by bacteria promotes the formation of micro-colonies resulting in formation of biofilms. This is followed by their growth and maturation (Dunne 2002). Biofilms thus formed, help in immobilization of the cells and also function as a protective barrier against antibiotics (Colvin et al. 2011). Formation of biofilms has been the main cause for failure of implants since the bacterial cells within the biofilm matrix exibit enhanced tolerance to the effect of antimicrobial agents. Formation of biofilms has also been reported to cause heavy losses in industries due to biocorrosion (Katsikogianni and Missirlis 2004). They have also been implicated in water contamination leading to a multitude of diseases. Hence there is a need to study the factors that are crucial for biofilm formation. The adherence of hippocampal cells on silicon wafers with nano-scale morphology was studied using atomic force microscopy (AFM) and it was found that the frictional force on the boundary around the cells increased due to accumulation of proteins (Ma et al. 2005). Fan et al. (2002) studied the effect of surface roughness on the adherence and viability of central neural cells and reported that cell adhesion and viability were significantly affected by surface rough- ness (on silicon wafers). The effect of nanostructured surfaces (on bacterial adhesion and subsequent biofilm formation) has been studied using confocal microscopy (Singh et al. 2011). Singh et al. (2011) reported that bacterial adhesion and biofilm formation were en- hanced on surfaces with roughness upto 20 nm and that a further increase in surface roughness caused a decrease in bacterial adhesion and inhibited biofilm formation. Oh et al. (2009) studied the formation of biofilms on different substrates (aluminium, steel, rubber, and polypropylene) using AFM and suggested that surface roughness (of the substrates) could have an effect on biofilm adhesion force (or tip-biofilm interaction force). The main drawback of this work was that surface roughness parameters (characterizing surface roughness) for each of the substrates were not evaluated. The other drawback was that substrates Contract grant sponsor: Department of Biotechnology.Contract grant sponsor: Department of Science & Technology. Conflict of Interest: None. Address for reprints: P. Gautam, Centre for Biotechnology, Anna University, Chennai, 25, India E-mail: pgautam@annauniv.edu Received 19 March 2014; Revised 20 April 2014; Accepted with revision 5 May 2014 DOI: 10.1002/sca.21148 Published online 10 July 2014 in Wiley Online Library (wileyonlinelibrary.com). SCANNING VOL. 36, 551–553 (2014) © 2014 Wiley Periodicals, Inc.